Methods and systems for promoting plant growth

Abstract

The present disclosure provides methods and systems for production of biostimulants that promote growth in plants. Embodiments described include methods of making a biostimulant composition in a biore

Description

METHODS AND SYSTEMS FOR PROMOTING PLANT GROWTHCROSS-REFERENCE

[0001] This application claims the benefit of U.S. Patent Application No. 18 / 541,671, filed December 15, 2023, U.S. Patent Application No. 18 / 541,917, filed December 15, 2023, U.S. Provisional Application No. 63 / 509,263 filed June 20, 2023, U.S. Provisional Application No. 63 / 510,615 filed June 27, 2023, U.S. Provisional Application No. 63 / 610,535, filed December 15, 2023, each of which is entirely incorporated herein by reference. This application claims the benefit of U.S. Provisional Application No. 63 / 613,548, filed December 21, 2023, the content of which is entirely incorporated herein by reference. This application claims the benefit of U.S. Provisional Application No. 63 / 613,560, filed December 21, 2023, the content of which is entirely incorporated herein by.BACKGROUND

[0002] The disclosure is generally related to biostimulant compositions and methods of using such biostimulant compositions to promote plant growth.

[0003] Promoting efficient production of food crops and other crops is an important goal for environmental and economic reasons. Plant growth promoting products sourced from organic materials can help to enhance crop growth, improve the efficacy of agricultural products, such as fertilizers, and reduce the environmental impacts of synthetic fertilizers and climate change. There exists a need for plant growth promoting biostimulant compositions that use abundant and available organic feedstocks.SUMMARY

[0004] In an aspect, the present disclosure provides a method of making a biostimulant composition having a plant growth promoting property, the method comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; (iii) maintaining a concentration of the microbial strain throughout the duration of time in at least the first container at at least 80% of a concentration of the microbial strain at the beginning of the duration of time; and (iv) collectingat least a portion of the product outflow stream as the biostimulant composition, wherein the biostimulant composition has the plant growth promoting property; wherein the duration of time is at least 5 days; and wherein the microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the microbial strain in the first container.

[0005] In some embodiments, the maintaining of step (b)(iii) comprises maintaining the concentration of the microbial strain at at least 1x103CFU / ml. In some embodiments, the maintaining of step (b)(iii) comprises providing nutrients in the first container that selectively promote growth of the microbial strain and sustain the established population. In some embodiments, the maintaining of step (b)(iii) comprises applying selective pressure in at least the first container. In some embodiments, the selective pressure comprises conditions that reduce the growth rate of microbes present in the first container relative to the growth rate of the microbial strain in the first container. In some embodiments, the selective pressure comprises conditions that shifts an ionic concentration of the working fluid. In some embodiments, the microbial strain, microbes of the microbial consortium, or any combination thereof may be enriched following the shift in the ionic concentration of the working fluid. In some embodiments, the selective pressure increases an activity of microbes within the microbial consortium. In some embodiments, the activity comprises solubilization of an inorganic substrate. In some embodiments, the inorganic substrate comprises phosphate or zinc.

[0006] In some embodiments, the biostimulant composition comprises an amount of the microbial strain. In some embodiments, the amount of the microbial strain is effective to impart the plant growth promoting property to the biostimulant composition. In some embodiments, the amount of the microbial strain is 1x102to 1x106CFU / ml.

[0007] In some embodiments, the aqueous feedstock further comprises an organic substrate. In some embodiments, the organic substrate comprises manure, lignocellulosic material, wastewater biosolids, food waste, energy crops, yeast, guano, agricultural waste, algae, or any combination thereof. In some embodiments, the biostimulant composition comprises digestion products produced by digestion of the organic substrate by the microbial strain and / or the microbial consortium. In some embodiments, the biostimulant composition comprises metabolites produced by the microbial strain and / or the microbial consortium. In some embodiments, the metabolites and / or digestion products in the biostimulant composition are present in an amount effective to impart the plant growth promoting property to the biostimulant composition. In some embodiments, the biostimulant composition retains the plant growth promoting property after filter sterilization. In some embodiments, the plant growth promotingproperty comprises an ability to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof. In some embodiments, the plant growth promoting property comprises an ability to increase nitrogen use efficiency, zinc solubilization, or phosphate solubilization.

[0008] In another aspect, the present disclosure provides a method of making a biostimulant composition having a desired plant growth promoting property, the method comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the desired plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as the biostimulant composition; wherein the established population of the microbial strain is not present in an aqueous feedstock or any other input in the bioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the established population of the microbial strain in the first container.

[0009] In another aspect, the present disclosure provides a method, comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having a plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the amount of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of a plant not administered the biostimulant composition.

[0010] In some embodiments, the nutrient uptake is measured by one or more of analysis of biomass, depletion method, or nutrient balance method, or by an assay as described in Example 4. In some embodiments, the nutrient uptake comprises macronutrients comprising nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof. In some embodiments, the nutrient uptake comprises micronutrients comprising boron, zinc, manganese, iron, copper, or a combination thereof.

[0011] In another aspect, the present disclosure provides a method, comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having a plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the biostimulant composition is configured to promote a plant growth property of a plant administered the biostimulant composition as compared to that of a plant not administered the biostimulant composition.

[0012] In some embodiments, the biostimulant composition is configured to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof. In some embodiments, the total biomass of the plant administered the biostimulant composition is increased by at least 5% as compared to the plant not administered the biostimulant composition. In some embodiments, the total biomass of the plant administered the biostimulant composition is increased by at least 10% as compared to the plant not administered the biostimulant composition. In some embodiments, an average leaf area of the plant administered the biostimulant composition is increased by at least 10% compared the plant not administered the biostimulant composition. In some embodiments, an average leaf area of the plant administered the biostimulant composition is increased by at least 25% compared the plant not administered the biostimulant composition.

[0013] In some embodiments, the established population of the microbial strain comprises a zinc solubilizing microbial strain. In some embodiments, administering the biostimulant composition to the plant increases an average leaf area by at least 25% as compared to an average leaf area of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition to the plant increases an average leaf area by at least 10% compared to an average leaf area of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition to a plant increases a nitrogen content, a phosphorous content, a potassium content, or any combination thereof by at least 20 ug compared to that of a plant not administered the biostimulant composition.

[0014] In some embodiments, the established population of the microbial strain comprises a phosphate solubilizing microbial strain. In some embodiments, administering the biostimulant composition to one or more plants increases a crop yield by at least 5% as compared to a crop yield of one or more plants not administered the biostimulant composition. In some embodiments, administering the biostimulant composition to one or more plants increases a dry biomass by at least 25% compared a dry biomass of one or more plants not administered the biostimulant composition.In some embodiments, administering the biostimulant composition to a plant increases an average leaf area by at least 10% compared to an average leaf area of a plant not administered the biostimulant composition. In some embodiments, the established population of the microbial strain comprises a nitrogen use efficiency -promoting microbial strain. In some embodiments, administering the biostimulant composition to the plant increases an average leaf area by at least 50% compared to an average leaf area of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition to the plant increases a dry shoot weight, a dry root weight, a stem diameter, or any combination thereof by at least 10% compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition to the plant increases a leaf chlorophyll content by at least 10% compared to that of a plant not administered the biostimulant composition.

[0015] In some embodiments, the established population of the strain in (b) is incubated for a duration of time. In some embodiments, the duration of time is at least 5 days.

[0016] In some embodiments, the method further comprises in (b), operating the bioreactor system for the duration of time by: (iii) maintaining a concentration of the microbial strain throughout the duration of time in at least the first container at at least 80% of a concentration of the microbial strain at the beginning of the duration of time. In some embodiments, themaintaining of step (b)(iii) comprises maintaining the concentration of the microbial strain at at least 1x103CFU / ml. In some embodiments, the maintaining of step (b)(iii) comprises providing nutrients in the first container that selectively promote growth of the microbial strain and sustain the established population. In some embodiments, the maintaining of step (b)(iii) comprises applying selective pressure in at least the first container.

[0017] In some embodiments, the selective pressure comprises conditions that reduce the growth rate of microbes present in the first container relative to the growth rate of the microbial strain in the first container. In some embodiments, the selective pressure comprises conditions that shifts an ionic concentration of the working fluid. In some embodiments, the microbial strain, microbes of the microbial consortium, or any combination thereof may be enriched following the shift in the ionic concentration of the working fluid. In some embodiments, the desired plant growth promoting property comprises an ability to promote uptake of a nutrient by one or more plants, increase bioavailability of a nutrient in soil, recruit microbes having the desired plant growth promoting property to root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof. In some embodiments, the nutrient comprises macronutrients. In some embodiments, the macronutrients comprises nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof. In some embodiments, the nutrient comprises micronutrients.

[0018] In some embodiments, the micronutrients comprises boron, zinc, manganese, iron, copper, or any combination thereof.

[0019] In some embodiments, the microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time. In some embodiments, the microbial strain is present in the aqueous feedstock or any other input into the bioreactor system during the duration of time. In some embodiments, the method further comprises, before (a), adding an inoculum of the microbial strain to the bioreactor system, wherein the inoculum of the microbial strain produces an initial population of the microbial strain of at least 0.5x104CFU / ml in at least one container. In some embodiments,, before adding the inoculum of the microbial strain, the concentration of the microbial strain is less than 1x102CFU / ml. In some embodiments, the aqueous feedstock further comprises an organic material at least partially digestible by microbes present in at least one of the containers. In some embodiments,, before the transferring of step (b)(i), the organic material had been partially digested by microbesendogenous to the organic material. In some embodiments, the method further comprises digesting the organic material in two or more serially connected containers before the transferring of step (b)(i).

[0020] In some embodiments, the organic material comprises manure and / or material produced by microbial digestion of manure. In some embodiments, the aqueous feedstock further comprises an inorganic material. In some embodiments, the inorganic material comprises rock phosphate particles. In some embodiments, prior to the transferring of step (b)(i), the rock phosphate particles had been partially digested by microbes present in the aqueous feedstock. In some embodiments, the method further comprises partially digesting the rock phosphate particles in two or more serially connected containers before the transferring of step (b)(i). In some embodiments, the microbial consortium comprises at least 1x105CFU / ml. In some embodiments, the microbial consortium comprises microbes derived from manure and from rock phosphate particles.

[0021] In some embodiments, the operating of step (b) further comprises producing microbial metabolites that have the plant growth promoting property. In some embodiments, the transferring of step (b)(i), the transferring of step (b)(ii), and the collecting of step (b) are performed continuously throughout the duration of time. In some embodiments, the transferring of step (b), the transferring of step (b), and the collecting of step (b) are performed periodically throughout the duration of time. In some embodiments, the method further comprises adding one or more carbon sources to at least one container of the bioreactor system.

[0022] In some embodiments, the one or more carbon sources are comprised in the aqueous feedstock. In some embodiments, the bioreactor system comprises a clarifier container comprising a clarifier working fluid. In some embodiments, the method further comprises further comprising separating a supernatant portion of the clarifier working fluid from a floc portion of the clarifier working fluid within the clarifier container. In some embodiments, the separating comprises gravity separation.

[0023] In some embodiments, the method further comprises folding the floc portion of the clarifier working fluid. In some embodiments, the folding further comprises releasing a population of the microbial strain into the supernatant portion without introducing floc solids into the supernatant portion. In some embodiments, the folding is performed by folding wipers in a bottom portion of the clarifier container. In some embodiments, the operating further comprises transferring the floc portion from the clarifier container to an earlier container in the bioreactor system. In some embodiments, the product outflow stream comprises the supernatantportion of the clarifier working fluid. In some embodiments, the method further comprises producing at least 1x104CFU / ml of the microbial strain in the product outflow stream.

[0024] In some embodiments, the bioreactor system comprises the first container comprising a volume of a first working fluid, a second container comprising a volume of a second working fluid, and a third container comprising a volume of a third working fluid. In some embodiments, at least one of the first working fluid, the second working fluid, or the third working fluid comprises a pH buffering system. In some embodiments, the method further comprises maintaining the pH of at least one of the first working fluid, the second working fluid, or the third working fluid between 6 and 9 throughout the duration of time. In some embodiments, the first container comprises an outlet port fluidly connected to an inlet port of the second container and the second container comprises an outlet port fluidly connected to an input port of the third container.

[0025] In some embodiments, the third container comprises an outlet port fluidly connected to a clarifier container. In some embodiments, the method further comprises maintaining the volume of each of the first working fluid, the second working fluid, and the third working fluid constant throughout the duration of time. In some embodiments, step (b) comprises operating the bioreactor system in a hydraulically balanced manner. In some embodiments, the transferring of step (b), the transferring of step (b, and the collecting of step (b) are driven by gravity.

[0026] In some embodiments, the operating comprises maintaining a flow rate that results in a hydraulic retention time of at least 5 days. In some embodiments, the operating comprises maintaining the product outflow stream at a flow rate of at least 100 gallons per day. In some embodiments, the volume of working fluid in each of the two or more containers is at least 100 gallons. In some embodiments, at least one of the two or more containers is a fluidized bed reactor. In some embodiments, at least one of the two or more containers is a packed bed reactor. In some embodiments, the method further comprises maintaining at least one of the two or more containers under aerobic conditions. In some embodiments, the method further comprises maintaining at least one of the two or more containers under microaerobic conditions.

[0027] In some embodiments, the bioreactor system is operated continuously for at least 90 days. In some embodiments, the aqueous feedstock does not comprise the microbial strain at a concentration higher than 10 CFU / ml. In some embodiments, the microbial strain is not added to the bioreactor system during the duration of time at a concentration that is higher than 10 CFU / ml. In some embodiments, the bioreactor system comprises at least one container placed in the series before the first container. In some embodiments, the volume of working fluid of one of the two or more containers comprises the microbial consortium, wherein each microbialconsortium is distinct from all of the microbial consortia in other working fluids. In some embodiments, the method further comprises producing a population of sporulated bacteria in the product outflow stream. In some embodiments, the method further comprises producing a population of the microbial strain in the product outflow stream that is sporulated. In some embodiments, the population of the microbial strain that is sporulated comprises at least 1x102CFU / ml. In some embodiments, the method further comprises adding an additional population of the microbial strain to the biostimulant product.

[0028] In some embodiments, the plant growth promoting property does not comprise nitrogen use efficiency, zinc solubilization, or phosphate solubilization. In some embodiments, the microbial strain is a nitrogen use efficiency-promoting microbial strain. In some embodiments, the nitrogen use efficiency-promoting microbial strain is one that performs nitrogen fixation, promotes nitrogen fixation in the tissues of plants, recruits nitrogen fixers to the root zones or other tissues of plants, or increases organic nitrogen content and / or mineralization of organic nitrogen in soil. In some embodiments, the nitrogen use efficiency-promoting microbial strain is positive for a nifH gene. In some embodiments, the nitrogen use efficiency-promoting microbial strain is one that promotes plant growth in a nitrogen-poor growth medium. In some embodiments, the nitrogen-poor growth medium comprises nitrate at less than 10 ppm.

[0029] In some embodiments, the nitrogen use efficiency-promoting microbial strain is of the genus Kosakonia, Klebsiella, Rahnella, Kluyvera, Enterobacter, Achromobacter, Microbacterium, Gluconobacter, Methylobacterium, Pseudomonas, Pantoea, Azospirillum, Azocarus, Herbaspirillum, Burkholderia, Cyanobacteria, Bacillus, Paenibacillus, or any combination thereof. In some embodiments, the nitrogen use efficiency -promoting microbial strain is of the species Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enterobacter spp., Achromobacter marplatensis, Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Paenibacillus borealis, Bacillus megaterium (Priestia megaterium), or Paenibacillus sonchi. In some embodiments, the nitrogen use efficiencypromoting microbial strain is the strain deposited under ATCC Accession No. PTA-127654 (MS3907), the strain deposited under ATCC Accession No. PTA-127653 (MS3900), the strain deposited under ATCC Accession No. PTA-127655 (MS4921), or the strain deposited under ATCC Accession No. PTA-127652 (MS2748). In some embodiments, the nitrogen use efficiency-promoting microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration of greater than 100 CFU / ml.

[0030] In some embodiments, the microbial strain is a zinc solubilizing bacterial strain. In some embodiments, the zinc-solubilizing bacterial strain is of the genus Bacillus. In some embodiments, the zinc-solubilizing bacterial strain is of the species Bacillus safensis or Bacillus megaterium.

[0031] In some embodiments, the zinc-solubilizing bacterial strain is one of the following: (a) a Bacillus safensis strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 1; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 4; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 7; (b) a Bacillus megaterium strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 2; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 5; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 8; or (c) a Bacillus megaterium strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 3; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 6; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 9.

[0032] In some embodiments, the zinc-solubilizing bacterial strain is the Bacillus safensis strain deposited under ATCC Accession No. PTA-127681, the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127683, or the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127682. In some embodiments, the microbial strain is a phosphate solubilizing bacterial strain. In some embodiments, the phosphate solubilizing bacterial strain is of the genus Bacillus. In some embodiments, the phosphate solubilizing bacterial strain is of the species Bacillus amyloliquefaciens or Bacillus licheniformis . In some embodiments, the phosphate solubilizing bacterial strain is one of the following: (a) a Bacillus amyloliquefaciens strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 10; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 12; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 14; or (b) a Bacillus licheniformis strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 11; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 13; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 15.

[0033] In some embodiments, the phosphate solubilizing bacterial strain is the Bacillus amyloliquefaciens strain deposited under ATCC Accession No. PTA-127657 or the Bacillus licheniformis strain deposited under ATCC Accession No. PTA-127656. In some embodiments, the phosphate solubilizing bacterial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration of greater than 100 CFU / ml.

[0034] In an aspect, the present disclosure provides a biostimulant composition made by the method described herein.

[0035] In another aspect, the present disclosure provides a method of promoting plant growth comprising contacting a plant, seed, or plant growth medium with the biostimulant composition described herein.

[0036] In an aspect, the present disclosure provides a bioreactor system comprising: (i) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property, wherein a concentration of the microbial strain in the first working fluid is at least 100 times higher than a concentration of the microbial strain in the aqueous feedstock stream or in any other input into the bioreactor system; (ii) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and (iii) a product outflow stream in fluid communication with the product outflow stream port.

[0037] In another aspect, the present disclosure provides a bioreactor system comprising: (i) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property; (ii) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and (iii) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of the plant not administered the biostimulant composition.

[0038] In another aspect, the present disclosure provides a bioreactor system comprising: (i) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbialconsortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property; (ii) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and (iii) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain is configured to promote a plant growth promoting property of a plant administered the biostimulant composition as compared to the plant not administered the biostimulant composition.

[0039] In some embodiments, the desired plant growth promoting property comprises an ability to promote uptake of a nutrient by one or more plants, increase bioavailability of a nutrient in soil, recruit microbes having the desired plant growth promoting property to root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof. In some embodiments, the nutrient comprises macronutrients. In some embodiments, the macronutrients comprises nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof. In some embodiments, the nutrient comprises micronutrients. In some embodiments, the micronutrients comprises boron, zinc, manganese, iron, copper, or any combination thereof.

[0040] In some embodiments, the bioreactor system is a continuous flow bioreactor system and the stream of the aqueous feedstock is a continuous stream. In some embodiments, each of the volume of the working fluid is constant. In some embodiments, each of the first container and the one or more additional containers comprises a concentration of the microbial strain that remains at least 1x104CFU / ml during operation of the bioreactor system. In some embodiments, the aqueous feedstock and any other input into the bioreactor system does not comprise the population of the microbial strain or does not comprise a concentration of the microbial strain at level higher than 100 CFU / ml. In some embodiments, the microbial consortium comprises at least 1x104CFU / ml. In some embodiments, the aqueous feedstock further comprises an organic material digestible by microbes present in the container.

[0041] In some embodiments, the organic material comprises manure or material derived from manure. In some embodiments, the aqueous feedstock further comprises rock phosphateparticles. In some embodiments, the microbial consortium comprises microbes derived from manure and rock phosphate particles. In some embodiments, the container comprising the product outflow stream port is a clarifier container configured to separate a portion of a working fluid in the clarifier container into a supernatant portion and a floc portion. In some embodiments, the clarifier container comprises one or more floc folding flights configured to agitate settled floc in the clarifier container without resuspending solids in the floc portion into the supernatant portion. In some embodiments, the system further comprises a floc return stream that flows from the clarifier to an earlier container in the series.

[0042] In some embodiments, the product outflow stream comprises the supernatant portion. In some embodiments, the product outflow stream comprises at least 1x104CFU / ml of the microbial strain. In some embodiments, the product outflow stream comprises at least 1x102CFU / ml of a sporulated form of the microbial strain. In some embodiments, the product outflow stream comprises a total dry weight of 0.2 to 2.5 mg / ml. In some embodiments, the product outflow stream has a chemical oxygen demand between 80 to 500 mg / L. In some embodiments, the product outflow stream has an electrical conductivity between 0.1 and 3.0 mS / cm. In some embodiments, at least the first container comprises a mixer configured to aerate the first working fluid. In some embodiments, the first working fluid and / or the working fluid in at least one of the one or more additional containers comprises aerobic conditions. In some embodiments, the microbial strain is a nitrogen use efficiency-promoting microbial strain.

[0043] In some embodiments, the nitrogen use efficiency-promoting microbial strain is one that performs nitrogen fixation, promotes nitrogen fixation in the tissues of plants, recruits nitrogen fixers to the root zones or other tissues of plants, or increases organic nitrogen content and / or mineralization of organic nitrogen in soil. In some embodiments, the nitrogen use efficiencypromoting microbial strain is positive for a nifH gene. In some embodiments, the nitrogen use efficiency-promoting microbial strain is one that promotes plant growth in a nitrogen-poor growth medium. In some embodiments, the nitrogen-poor growth medium comprises nitrate at less than 10 ppm.

[0044] In some embodiments, the nitrogen use efficiency-promoting microbial strain is of the genus Kosakonia, Klebsiella, Rahnella, Kluyvera, Enterobacter, Achromobacter, Microbacterium, Gluconobacter, Methylobacterium, Pseudomonas, Pantoea, Azospirillum, Azocarus, Herbaspirillum, Burkholderia, Cyanobacteria, Bacillus, Paenibacillus, or any combination thereof. In some embodiments, the nitrogen use efficiency -promoting microbial strain is of the species Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enterobacter spp., Achromobacter marplatensis,Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Paenibacillus borealis, Bacillus megaterium (Priestia megaterium), or Paenibacillus sonchi. In some embodiments, the nitrogen use efficiencypromoting microbial strain is the strain deposited under ATCC Accession No. PTA-127654 (MS3907), the strain deposited under ATCC Accession No. PTA-127653 (MS3900), the strain deposited under ATCC Accession No. PTA-127655 (MS4921), or the strain deposited under ATCC Accession No. PTA-127652 (MS2748).

[0045] In some embodiments, the microbial strain is a phosphate solubilizing bacterial strain. In some embodiments, the phosphate solubilizing bacterial strain is of the genus Bacillus. In some embodiments, the phosphate solubilizing bacterial strain is of the species Bacillus amyloliquefaciens or Bacillus licheniformis . In some embodiments, the phosphate solubilizing bacterial strain is one of the following: (a) a Bacillus amyloliquefaciens strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 10; (ii) (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 12; and (iii) (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 14; or (b) a Bacillus licheniformis strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 11; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 13; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 15. In some embodiments, the phosphate solubilizing bacterial strain is the Bacillus amyloliquefaciens strain deposited under ATCC Accession No. PTA-127657 or the Bacillus licheniformis strain deposited under ATCC Accession No. PTA-127656.

[0046] In some embodiments, the microbial strain is a zinc solubilizing bacterial strain. In some embodiments, the zinc-solubilizing bacterial strain is of the genus Bacillus. In some embodiments, the zinc-solubilizing bacterial strain is of the species Bacillus safensis or Bacillus megaterium. In some embodiments, the zinc-solubilizing bacterial strain is one of the following: (a) a Bacillus safensis strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 1; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 4; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 7; (b) a Bacillus megaterium strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 2; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 5; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 8; or (c) a Bacillus megaterium strain having one or more of the following: (i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 3; (ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 6; and (iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 9. In someembodiments, the zinc-solubilizing bacterial strain is the Bacillus safensis strain deposited under ATCC Accession No. PTA-127681, the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127683, or the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127682.

[0047] In some embodiments, the nitrogen use efficiency-promoting microbial strain is one of the following: (a) a Bacillus megaterium strain having one or more of the following: a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 16; a gyrB gene sequence at least 95% identical to SEQ ID NO: 20; and a rpoB gene sequence at least 95% identical to SEQ ID NO: 24; (b) a Paenibacillus borealis strain having one or more of the following: a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 17; a gyrB gene sequence at least 95% identical to SEQ ID NO: 21; and a rpoB gene sequence at least 95% identical to SEQ ID NO: 25; (c) a Paenibacillus sonchi strain having one or more of the following: a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 18; a gyrB gene sequence at least 95% identical to SEQ ID NO: 22; and a rpoB gene sequence at least 95% identical to SEQ ID NO: 26; or (d) a Bacillus megaterium strain having one or more of the following: a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 19; a gyrB gene sequence at least 95% identical to SEQ ID NO: 23; and a rpoB gene sequence at least 95% identical to SEQ ID NO: 27.

[0048] In an aspect, the present disclosure provides a biostimulant composition made by the system described herein.

[0049] In an aspect, the present disclosure provides A method of promoting plant growth comprising contacting a plant, seed, or plant growth medium with the biostimulant composition described herein.

[0050] Additional aspects and advantages of the present disclosure will become readily apparent to those skilled in this art from the following detailed description, wherein only illustrative embodiments of the present disclosure are shown and described. As will be realized, the present disclosure is capable of other and different embodiments, and its several details are capable of modifications in various obvious respects, all without departing from the disclosure.Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.INCORPORATION BY REFERENCE

[0001] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. To theextent publications and patents or patent applications incorporated by reference contradict the disclosure contained in the specification, the specification is intended to supersede and / or take precedence over any such contradictory material.BRIEF DESCRIPTION OF THE DRAWINGS

[0051] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings (also “Figure” and “FIG.” herein), of which:

[0052] Figure 1 depicts a schematic labeling of components of a ZST system with a single reactor.

[0053] Figure 2 is an exemplary schematic emphasizing the floc flights present in the clarifier of the NTS 2.0 system.

[0054] Figure 3 is an exemplary schematic depicting the PST system.

[0055] Figure 4 depicts a serialized PST system (PST-sIP) with a series of reactors and inputs to the bioreactor system.

[0056] Figure 5 depicts a serialized ZST system featuring labeling of a series of reactors and other components.

[0057] Figure 6 is an exemplary schematic labelling the parts of a NTS bioreactor system.

[0058] Figure 7 depicts another serialized ZST system featuring a complete mixed reactor and a series of fluidized bed reactors.

[0059] Figure 8 depicts another serialized ZST system featuring a complete mixed reactor and a series of packed bed reactors with scaffolding.

[0060] Figure 9 is a graph showing the comparison of solubilized Zn2+ (in mg / L) in solution between the ZST 2.0 fluidized bed reactor (FBR) and the ZST 2.0 packed bed reactor (PBR) across different phases.

[0061] Figure 10 is a graph showing the comparison of dissolved oxygen (in mg / L) between the ZST fluidized bed reactor (FBR) and the ZST packed bed reactor (PBR) across different phases.

[0062] Figures 11A-11B are a series of graphs showing measures of Zn uptake and plant growth in navy beans treated with MAP fertilizer coated with one of four different treatments: water (UTC), Circa™ Zn, MS4666, and MS4666 with Circa™ Zn. FIG. 11A shows the amount of zinc uptake (μg / plant) for navy beans. FIG. 11B shows total biomass, shoot dry weights, and root dry weights across conditions.

[0063] Figure 12 is a graph showing results of measuring zinc uptake per corn plant treated with MAP fertilizer coated with one of four different treatments: water (UTC), Circa™ Zn, MS4666, and MS4666 with Circa™ Zn.

[0064] Figures 13A-13B are a series of graphs showing evaluation of Zn uptake and crop growth promotion in corn treated with MAP fertilizer coated with one of four different treatments: water (UTC), Circa™ Zn, MS4666, and MS4666 with Circa™ Zn. FIG. 13A shows results of measuring zinc uptake per plant for each condition. FIG. 13B shows total biomass, shoot dry weights, and root dry weights across conditions.

[0065] Figure 14 is a graph showing the uptake of nitrogen, phosphorus, and potassium (μg / plant) in com treated with MAP fertilizer coated with one of four different treatments: water (UTC), Circa™ Zn (CZN), MS4666, and MS4666 with CZN. For each macronutrient condition, left to right bars are: UTC, CZN, MS4666, MS4666 + CZN.

[0066] Figure 15 is a graph showing plant growth promotion (measured as dry weights (g)) in com fertilized with MAP fertilizer coated with one of four different treatments: water (UTC), Circa™ Zn, MS4666, and MS4666 with Circa™ Zn. Dry weights were measured for the total biomass, shoot, and root. For each treatment condition, left to right bars are: root, stem, and total biomass.

[0067] Figure 16 shows the differences in Arabidopsis growth (in average leaf area(cm2)) across intact and filter-sterilized (F / S) solutions for all ZST systems. UTC and Accomplish LM are used as controls.

[0068] Figures 17A-17C depict effects of different treatments on the dry biomass of corn. Percentages above the bars denote the percent change from UTC. An asterisk denotes significant difference from UTC. FIG. 17A shows the percent change of total dry biomass from UTC across multiple treatments. FIG. 17B depicts the average shoot dry biomass. FIG. 17C depicts the average root dry biomass.

[0069] Figures 18A-18C are a series of graphs showing the effects of the loading input components and supernatant (SPN) of the ZST system on plant growth production in com. FIG. 18A shows total biomass treated with loading inputs and ZST 1.5 SPN. FIG. 18B shows total biomass with fertilizer, loading inputs plus Circa™ Zn. and ZST 1.5 SPN plus Circa™ Zn. FIG. 18C shows total biomass with of Circa™ Zn alone, ZST 1.5 SPN alone, and ZST 1.5 SPN plus Circa™ Zn.

[0070] Figures 19A-19B are a series of graphs depicting zinc uptake (in μg / plant) in com after treatment with ZST SPN or isolate input (MS4666) compared to UTC and Circa™ Zn (CZN). FIG. 19A shows zinc uptake from the input into the ZST system (MS4666) and ZST SPNcompared to UTC and CZN alone. FIG. 19B shows zinc uptake from the isolate input into the ZST system in combination with CZN (MS4666 CZN) and from ZST 1.5 SPN with CZN.

[0071] Figure 20 shows a series of graphs depicting total corn grain yield (in g) across different ZST systems, untreated control (UTC), and Circa™ Zn (CZN) fertilizer alone.

[0072] Figure 21 depicts the total zinc (ug) measured per ear of corn across different ZST systems, untreated control (UTC), and Circa™ Zn (CZN) fertilizer alone and in combination with ZST systems.

[0073] Figures 22A-22B are a series of graphs depicting the total macronutrients (nitrogen, phosphorus, and potassium) measured in mg per ear of corn across different ZST systems, untreated control (UTC), and Circa™ Zn (CZN) fertilizer alone. FIG. 22A shows results for ZST systems without CZn. FIG. 22B shows results for CZN alone and ZST systems with CZn. For each condition, left to right bars are: UTC, ZST 1.5, ZST 2.0 FBR, and ZST 2.0 PBR.

[0074] Figures 23A-23B are tables showing macronutrient and micronutrient content in the corn grain across ZST treatment systems using an untreated control (UTC) without CZN fertilizer and all ZST treatments (FIG. 23 A) and using CZN fertilizer and all ZST treatments (FIG. 23B)

[0075] Figures 24A-24C are a series of graphs with harvest yield for three vegetables: broccoli (FIG. 24A), cabbage (FIG. 24B), and lettuce (FIG. 24C), measured as boxes / acre.

[0076] Figures 25A-25C show a series of graphs with plant growth promotion results of MS4666. Treatment with MS4666 shows superior com yield compared to that from untreated controls (FIGs. 25A-25B). Treatment with MS4666 at a rate of 2.0 qt / acre shows superior soybean yield compared to that from untreated controls (FIG. 25C).

[0077] Figure 26 shows a distance-based neighbor joining phylogenetic tree based on 16s rRNA gene.

[0078] Figure 27 shows results from a zinc solubilization capacity in vitro assay between the target isolate MS4666, PST Floc, and ZST-1.0 Batch solutions using PST Floc (Batch-A) or PST Floc with MS4666 (Batch-B).

[0079] Figure 28 shows a graph depicting concentrations of total bacteria and Zn-solubilizers across ZST- 1.5 process and base product.

[0080] Figure 29 shows measurements of Zn2+ made soluble from ZST-1.5 Reactor 1 (Rl) and ZST-1.5 Base Product. * indicates p < 0.0001.

[0081] Figure 30 shows concentration of Zn2+ found in ZST-2.0 FBR and ZST-2.5 reactors and base product.

[0082] Figure 31 shows results measuring Zn2+ made soluble in ZET experiment from each treatment condition.

[0083] Figures 32A-32B depict experimental results of MS2839 and MS 1835 spore counts following seven or fourteen days of incubation. Treatments were separated based on normal feeding (e.g., N Feed) or supplemental feeding (e.g., S Feed). FIG. 32A shows results of MS2839 and FIG. 32B shows results of MS 1835.

[0084] FIGs. 33A-33B depicts box plot target counts of MS 1835 (FIG. 33 A) or MS2839 (FIG. 33B). Treatments used either base product of the P2 digestion system or water.

[0085] FIGs. 34A-34B depicts box plot target counts of MS 1835 (FIG. 34A) or MS2839 (FIG. 34B). Treatments used either liquid or powder spore preparation as the inoculation medium.

[0086] FIGs. 35A-35B depicts box plot target counts of MS 1835 (FIG. 35A) or MS2839 (FIG.35B). Treatments comprised either treatments with additional supplemented feeding or treatments that received normal feeding.

[0087] Figure 36 is a graph showing the plant growth production of Arabidopsis (measured in average leaf area, cm2) for UTC (untreated control) and PST supernatant.

[0088] Figure 37 is a graph depicting corn shoot dry biomass (in grams) across UTC and PST 2.0 sIP. Results are from experiments with com which was fertilized with PST 2.0 sIP coated monoammonium phosphate (MAP).

[0089] Figure 38 is a graph depicting corn shoot dry biomass (in grams) across UTC and PST 2.0 sIP. Results are from experiments in which corn was treated via in-furrow application.

[0090] Figures 39A-39C are graphs showing the effects of treatment of corn with MAP coated with PST-sIP supernatant, PST supernatant, or no coating (UTC) on plant height (FIG. 39A), leaf area (FIG. 39B), and leaf chlorophyll content (FIG. 39C).

[0091] Figure 40 is a graph depicting uptake of macronutrients, such as nitrogen, sulfur, phosphorus, potassium, magnesium, and calcium, in com shoots (represented as %UTC, mg / shoot) after treatment with PST-sIP product at 2 qt / t rate. All results are from com which was fertilized with MAP coated with PST-sIP product.

[0092] Figure 41 is a graph depicting uptake of micronutrients, such as boron, zinc, manganese, copper, and iron in com shoots (represented as % UTC, μg / shoot) after treatment with PST-sIP product at 2 qt / t rate. All results are from com which was fertilized with MAP coated with PST- sIP product.

[0093] Figure 42 is a graph showing uptake of macronutrients in corn shoots (represented as %UTC, mg / shoot) across PST-sIP. Results are from an experiment in com in which the treatments were applied in-furrow at planting. Bars from left to right for each system are:nitrogen (N), sulfur (S), phosphorus (P), potassium (K), magnesium (Mg), and calcium (Ca). All results are from corn which was treated with PST-sIP product in-furrow at 4qt / acre application rate.

[0094] Figure 43 is a graph showing uptake of micronutrients in com shoots (represented as % UTC, μg / shoot) across PST-sIP. Results are from a com experiment in which the treatments were applied in-furrow at planting. Bars from left to right for each system are: boron (B), zinc (Zn), manganese (Mn), iron (Fe), and copper (Cu). All results are from com which was treated with PST-sIP product in-furrow at 4qt / acre application rate.

[0095] Figures 44A-44C are graphs showing the effects of treating corn in-furrow with PST-sIP supernatant, PST supernatant, or untreated control on leaf area (FIG. 44A), stem diameter (FIG. 44B), and leaf chlorophyll content (FIG. 44C).

[0096] Figure 45 is a graph showing MS2839 spore count (cfu / ml) in supernatant for different ratios of floc. Bars from left to right for each percent floc condition are: Time Zero, 1 HR, 6 HR, 24 HR, and 48 HR post-inoculation.

[0097] Figure 46 is a graph showing MS2839 counts (cfu / ml) from PST-sIP digestion system over time in supernatant (BP) and WB.

[0098] Figure 47 is a graph showing FIG. 27 shows counts of MS2839 (cfu / ml) from PwST-sIP digestion system over time with or without floc-folding flights. T.L. denotes trend-line for each condition.

[0099] Figure 48 shows an exemplary schematic of a NTS system with packed bed reactors.

[0100] Figure 49 shows an exemplary schematic of a NTS system with fluidized bed reactors.

[0101] Figure 50 is an exemplary schematic of a NTS 2.0 system with floc flights present in the clarifier.

[0102] Figures 51A-51B are a series of graphs showing leaf chlorophyll contents across treatment conditions. FIG. 51A shows results from V5 growth stage in corn. FIG. 51B shows results from V8 growth stage in com.

[0103] Figure 52 is a graph depicting the photosynthetic quantum yield across treatment conditions tested in corn at V9 growth stage. NTS treatments at both applications rates and isolates showed increased photosystem light capture efficiency.

[0104] Figure 53 is a graph depicting photosynthetic electron transport rate across treatment conditions in corn at V9 growth stage.

[0105] Figure 54 is a graph depicting the number of corn tillers per com plant across treatment conditions, measured at V5 growth stage.

[0106] Figure 55 is a graph depicting corn plant height across treatment conditions, measured at V7 growth stage.

[0107] Figure 56 is a graph depicting the stem diameter across treatment conditions.

[0108] Figure 57 is a graph depicting the stomatai conductance (the rate of CO2 and H2O gas exchange) across treatment conditions in corn, at V9 growth stage.

[0109] Figure 58 is a graph depicting the transpiration rate (efficiency of water movement into and through the plant) across treatment conditions, measured at V9 growth stage in corn.

[0110] Figure 59 is a graph depicting the corn dry biomass, measured as dry shoot weight, across treatment conditions.

[0111] Figure 60 is a graph depicting results of acetylene reduction in corn root / basal stem, measured across treatment conditions.

[0112] Figure 61 shows results from the sorghum grass study testing shake flasks using isolates with different aeration. All treatments showed significantly more leaf area than that of untreated control (UTC). ANOVA student’s t-test (p<0.05) Levels not connected by same letter are significantly different.

[0113] Figure 62 shows average pixel shoot area when shake flasks performance is grouped by aeration conditions.

[0114] Figure 63 shows average performance of shake flasks when different ratios of PST WB are used. Shake flask treatments were grouped based on the ratio of PST WB used under both aerobic and anaerobic conditions.

[0115] Figures 64A-64B show exemplary schematics of the NTS batch systems. FIG. 64A shows the prototype 1 (e.g., PT1) system. FIG. 64B shows the prototype 2 (e.g., PT2) system.

[0116] Figure 65 shows the effects of products from the NTS batch systems (e.g., PT1 and PT2) on ethylene production. PT1 product shows the greatest ethylene peak compared to that from UTC and PT2.

[0117] Figures 66A-66B show two graphs with results from a plant growth promotion test of the PT1 and PT2. Products of the NTs batch system were applied at 2 qt / acre or 4 qt / acre. On both Day 10 (FIG. 66A) and Day 14 (FIG. 66B), PT2 showed the greatest leaf area compared to that from PT1.

[0118] Figure 67 shows results from an Arabidopsis plant growth promotion rockwool test for NTS 1.1, 1.2, 1.3 and 1.4 lines with three reactors. All treatments showed numerically greater average leaf area measurement from the UTC with all having significant plant growth promotion (p=0.0008).

[0119] Figure 68 shows results across NTS 1.0 systems with either three or four reactors in the system. There was a general increase of performance in the output solutions when the retention time of the system was extended by about 42.86%.

[0120] Figure 69 shows the plant growth promotion of intact NTS solutions and metabolites. Intact NTS 1.0 system solutions and their metabolites showed increased average leaf area compared to that from UTC plants.

[0121] Figure 70 shows Arabidopsis plant growth in NTS 2.2 and NTS 2.3 systems with intact solution or metabolites. All NTS 2.0 system treatments showed greater average leaf area than untreated control.

[0122] Figures 71A-71B show Arabidopsis plant growth in NTS 1.4 and NTS 1.5 systems with intact solution or metabolites. FIG. 71A shows the results from intact solution treatments. FIG. 71B shows the results from metabolite treatments. All NTS system treatments showed greater average leaf area than untreated control, with NTS 1.4 BP intact and metabolite showing the greatest plant growth promotion.DETAILED DESCRIPTION

[0123] While various embodiments of the invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions may occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed.

[0124] Described herein are systems and methods that employ microbial digestion of various feedstocks. A system of the present disclosure may comprise a continuous system capable of serialized isolate production (e.g., sIP system). The isolate production of the sIP system can occur within a mixed consortium of microbes. The target isolates of a sIP bioreactor system may become enriched in the microbial environment and may demonstrate improved efficacy and functionality. A targeted functionality may be zinc solubilization of bound zinc from soil or fertilizer and improved nutrient uptake in plants. A targeted functionality may be phosphate solubilization of insoluble phosphate from soil or fertilizer and improved nutrient uptake in plants. A targeted functionality may be nitrogen use efficiency and / or nitrogen fixation from soil or fertilizer and improved nutrient uptake in plants. A target isolate may possess commercially valuable properties and can be introduced into a continuous (e.g., serialized) reactor system comprised of a complex microbial consortia that has been modified for functionality (e.g., for zinc solubilization, phosphate solubilization, nitrogen use efficiency, etc.). Without wishing tobe bound by theory, a target isolate may provide a performance benefit to a microbial community of the digestions systems described herein, providing a chemical and / or functional synergistic relationship as it grows in the system. A bioreactor system described herein may comprise a zinc solubilization technology (ZST) system, with a target function to enrich a population or populations of zinc-solubilizing microbes, metabolites, or any combination thereof. A bioreactor system described herein may comprise a phosphate solubilization technology (PST) system, with a target function to enrich a population or populations of phosphate-solubilizing microbes, metabolites, or any combination thereof. A bioreactor system described herein may comprise a nitrogen use efficiency (NUE) system, with a target function to enrich a population or population of nitrogen use efficiency and / or nitrogen fixation microbes, metabolites, or any combination thereof.

[0125] The products of digestion methods and systems described herein can comprise microbes and metabolites produced by microbial digestion of feedstock substrates. In some embodiments, the products of digestion methods and systems described herein can comprise biostimulant compositions that have plant growth promoting properties when applied to horticultural items such as plants, plant parts (e.g., shoots, stems, leaves, buds, flowers, leaf axils, roots), seeds, or a medium in which plants grow (e.g., soil). In some embodiments, methods and systems described herein are arranged to selectively promote growth of microbes that have a desired plant growth promoting property themselves or that produce metabolites that have the desired plant growth promoting property, such that the biostimulant product has the desired plant growth promoting property. Applications of the products of the bioreactor systems described herein may be on dryfertilizers, applied in conjunction with the application of fertilizers, in formulations with additional components including liquid fertilizers, micronutrient coating formulations, foliar applications, or any combination thereof. Applications of the products of the bioreactor systems described herein may be to a part of a plant, such as a shoot, a stem, a leaf, a lateral bud, a terminal bud, a flower, a leaf axil, a root (e.g., a primary root, a lateral root, a root hair, a root cap), or any combination thereof. These and other features of embodiments disclosed herein are described in more detail below.I. Definitions

[0126] In the following description, certain specific details are set forth in order to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the embodiments provided may be practiced without these details. Unless the context requires otherwise, throughout the specification and claims which follow, the word “comprise”and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense, that is, as “including, but not limited to.” As used in this specification and the appended claims, the singular forms “a,” “an,” and “the” comprise plural referents unless the content clearly dictates otherwise. It should also be noted that the term “or” is generally employed in its sense including “and / or” unless the content clearly dictates otherwise. Further, headings provided herein are for convenience only and do not interpret the scope or meaning of the claimed embodiments.

[0127] The term “about” or “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, e.g., the limitations of the measurement system. For example, “about” can mean within 1 or more than 1 standard deviation, per the practice in the given value. Where particular values are described in the application and claims, unless otherwise stated the term “about” should be assumed to mean an acceptable error range for the particular value.

[0128] The term “culturing”, as used herein, can refer to the propagation of organisms on or in media of various kinds. Non-limiting examples of suitable media comprise tryptic soy agar (TSA), zinc agar, phosphate solubilizing medium which contain insoluble forms of phosphate, nutrient medium, lysogeny broth (LB medium), plate count agar, or any combination thereof.

[0129] As used herein, the term “enriched culture” of an isolated microbial strain can refer to a microbial culture wherein the total microbial population of the culture contains a percentage of a target isolated strain. An enriched culture may comprise a percentage of a target isolated strain and a population of microbes enriched for a particular functionality (e.g., zinc solubilization, nitrogen use efficiency, phosphate solubilization). In some embodiments, an enriched culture may comprise a percentage of a target isolated strain, a population of microbes enriched for a particular functionality, and metabolites enriched for a particular functionality (e.g., zinc solubilization, nitrogen use efficiency, phosphate solubilization). The enriched culture may comprise a percentage of a total bacteria population in a container of bioreactor system described herein. The enriched culture may comprise a percentage of a total bacteria population in an output product (e.g., biostimulant) described herein. An enriched culture may comprise an increased amount of a target isolated strain and / or a target population of microbes compared to a total microbial population of a culture. An enriched culture may comprise a growing population of a target isolated strain and a population of microbes enriched for a particular functionality (e.g., zinc solubilization, nitrogen use efficiency, phosphate solubilization) over a time period. In some embodiments, an enriched culture can refer to a microbial culture wherein the totalmicrobial population of the culture contains at least about 0.001%, at least about 0.01%, at least about 0.05%, at least about 0.1%, at least about 0.5%, at least about 1%, at least about 1.5%, at least about 2%, at least about 2.5%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 75%, at least about 80%, or at least about 90% of a target isolated strain, a population of microbes enriched for a particular functionality, metabolites enriched for a particular functionality, or any combination thereof. In some embodiments, an enriched culture can refer to a microbial culture wherein the total microbial population of the culture contains at most about 90%, at most about 80%, at most about 75%, at most about 70%, at most about 60%, at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2.5%, at most about 2%, at most about 1.5%, at most about 1%, or at most about 0.5% of a target isolated strain, a population of microbes enriched for a particular functionality, metabolites enriched for a particular functionality, or any combination thereof. In some embodiments, an enriched culture can refer to a microbial culture wherein the total microbial population of the culture contains from about 0.5% to about 75% of a target isolated strain, a population of microbes enriched for a particular functionality, metabolites enriched for a particular functionality, or any combination thereof. In some embodiments, an enriched culture can refer to a microbial culture wherein the total microbial population of the culture contains from about 0.5% to about 1%, about 0.5% to about 2%, about 0.5% to about 3%, about 0.5% to about 5%, about 0.5% to about 10%, about 0.5% to about 15%, about 0.5% to about 20%, about 0.5% to about 25%, about 0.5% to about 50%, about 0.5% to about 60%, about 0.5% to about 75%, about 1% to about 2%, about 1% to about 3%, about 1% to about 5%, about 1% to about 10%, about 1% to about 15%, about 1% to about 20%, about 1% to about 25%, about 1% to about 50%, about 1% to about 60%, about 1% to about 75%, about 2% to about 3%, about 2% to about 5%, about 2% to about 10%, about 2% to about 15%, about 2% to about 20%, about 2% to about 25%, about 2% to about 50%, about 2% to about 60%, about 2% to about 75%, about 3% to about 5%, about 3% to about 10%, about 3% to about 15%, about 3% to about 20%, about 3% to about 25%, about 3% to about 50%, about 3% to about 60%, about 3% to about 75%, about 5% to about 10%, about 5% to about 15%, about 5% to about 20%, about 5% to about 25%, about 5% to about 50%, about 5% to about 60%, about 5% to about 75%, about 10% to about 15%, about 10% to about 20%, about 10% to about 25%, about 10% to about 50%, about 10% to about 60%, about 10% to about 75%, about 15% to about 20%, about 15% to about 25%,about 15% to about 50%, about 15% to about 60%, about 15% to about 75%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about 75%, about 25% to about 50%, about 25% to about 60%, about 25% to about 75%, about 50% to about 60%, about 50% to about 75%, or about 60% to about 75% of a target isolated strain, a population of microbes enriched for a particular functionality, metabolites enriched for a particular functionality, or any combination thereof.

[0130] The term “composition” as used herein can refer to a combination of an active agent (e.g., a microbial strain described herein) and at least one other compound, carrier, or composition, which can be inert (for example, a detectable agent or liquid carrier) or active, such as, but not limited to, a fertilizer, nutrient, or pesticide. A microbial composition refers to a composition comprising at least one microbial species. A composition may comprise microbial metabolites generated in a microbial consortium of a bioreactor system described herein.

[0131] An “effective amount”, as used herein, can refer to an amount sufficient to effect beneficial and / or desired results. An effective amount can be administered in one or more administrations. The expression “effective microorganism” used herein in reference to a microorganism is intended to mean that the subject strain exhibits a degree of promotion of plant health, growth and / or yield, at a statistically significant level, that of an untreated control. In some instances, the expression “an effective amount” is used herein in reference to that quantity of microbial treatment which can be used to obtain a beneficial or desired result relative to that occurring in an untreated control under suitable conditions of treatment as described herein. For example, the expression “an agriculturally effective amount” is used herein in reference to that quantity of microbial treatment which can be used to obtain an agriculturally beneficial or desired result relative to that occurring in an untreated control under suitable conditions of treatment as described herein. The effective amount of an agricultural formulation or composition that may be applied for the improvement of plant health, growth and / or yield, can be readily determined.

[0132] A “carrier” as used herein can refer to a substance or a composition that support the survival of the microbes. Such carriers may be either organic or non-organic.

[0133] “Percentage of sequence identity”, as used herein, can be determined by comparing two optimally locally aligned sequences over a comparison window defined by the length of the local alignment between the two sequences. The amino acid sequence in the comparison window may comprise additions or deletions (e.g., gaps or overhangs) as compared to the reference sequence (which does not comprise additions or deletions) for optimal alignment of the two sequences.

[0134] Local alignment between two sequences may comprise segments of each sequence that are deemed to be sufficiently similar according to a criterion that depends on the algorithm used to perform the alignment (e.g., BLAST). The percentage of sequence identity is calculated by determining the number of positions at which the identical nucleic acid base or amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison and multiplying the result by 100. Optimal alignment of sequences for comparison may be conducted by the local homology algorithm of Smith and Waterman (Add. APL. Math. 2:482, 1981), by the global homology alignment algorithm of Needleman and Wunsch (J Mol. Biol. 48:443, 1970), by the search for similarity method of Pearson and Lipman (Proc. Natl. Acad. Sci. USA 85:2444, 1988), by heuristic implementations of these algorithms (NCBI BLAST, WU-BLAST, BLAT, SIM, BLASTZ), or by inspection. Given that two sequences have been identified for comparison, GAP and BESTFIT may be employed to determine their optimal alignment. Typically, the default values of 5.00 for gap weight and 0.30 for gap weight length are used. The term “substantial sequence identity” between polynucleotide or polypeptide sequences refers to polynucleotide or polypeptide comprising a sequence that has at least about 50% sequence identity, at least about 60% sequence identity, at least about 70% sequence identity, at least about 80% sequence identity, at least about 85% sequence identity, at least about 90% sequence identity, at least about 95% sequence identity, at least about 96% sequence identity, at least about 97% sequence identity, at least about 98% sequence identity, or at least about 99% sequence identity compared to a reference sequence using the programs. In addition, pairwise sequence homology or sequence similarity, as used, refers to the percentage of residues that are similar between two aligned sequences . Families of amino acid residues having similar side chains have been well defined in the art. These families comprise amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Query nucleic acid and amino acid sequences can be searched against subject nucleic acid or amino acid sequences residing in public or proprietary databases. Such searches can be done using the National Center for Biotechnology Information Basic Local Alignment Search Tool (NCBI BLAST v 2.18) program. The NCBI BLAST program is available on the internet from the National Center for Biotechnology Information (blast.ncbi.nlm.nih.gov / Blast.cgi). Typically thefollowing parameters for NCBI BLAST can be used: Filter options set to “default”, the Comparison Matrix set to “BLOSUM62”, the Gap Costs set to “Existence: 11, Extension: 1”, the Word Size set to 3, the Expect (E threshold) set to le-3, and the minimum length of the local alignment set to 50% of the query sequence length. Sequence identity and similarity may also be determined using GenomeQuest™ software (Gene-IT, Worcester Mass. USA).

[0135] The term “plant growth promotion” (PGP) can refer to processes that can promote plant health, growth, yield, or any combination thereof. In some embodiments, PGP can encompass a wide range of improved plant properties, including but not limited to, improved nitrogen fixation, improved phosphate uptake, improved zinc uptake, improved root development, increased leaf area, increased plant yield, increased uptake of macronutrients, increased uptake of micronutrients, increased seed germination, enhancing seed germination, enhancing early plant development, improving root growth, improving shoot growth, improving plant height, increasing nutrient uptake, mitigating transplant shock, improving plant reproduction, improving soil microbial activity, increased photosynthesis, increased abundance of functional enzymes, increased dry biomass, or an increase in the accumulated biomass of the plant. In some embodiments, the microbial strains, isolates, cultures, compositions, or synthetic consortia as described herein improve nutrient uptake, plant health and vigor, improve root development, increase leaf area, increase plant yield, increased uptake of macronutrients, increased uptake of micronutrients, increase seed germination, increased abundance of functional enzymes, increased dry biomass, or an increase in the accumulated biomass of the plant. In some embodiments, the microbial strains, isolates, cultures, or compositions as described herein increase the size or mass of a plant or parts thereof, as compared to a control plant, or a plant that has not been treated with a substance, or parts thereof or as compared to a predetermined standard. In some embodiments, the microbial strains, isolates, cultures, compositions, or synthetic consortia as described herein improve the health, vigor and yield of a plant, as compared to a control plant or a plant that has not been treated with a substance, but also can survive and multiply in microhabitats associated with the root surface.

[0136] Microbes described herein as “zinc-solubilizing microbes” and “zinc-solubilizing bacteria” or “zinc solubilizers” may refer to a microbe or bacterium that either (a) has a measurable increase in zinc made soluble relative to a control in an assay that comprises suspending adding a test solution that comprises the microbe or bacterium to media with insoluble, inorganic zinc, incubating the media, and measuring an amount of insoluble zinc released into solution and made soluble using an instrument such as a Inductively CouplesPlasma (ICP) Atomic Emission Spectrometer and comparing to known standards, or (b) makes a visible halo when grown in a colony on a plate with insoluble zinc.

[0137] Microbes described herein as “phosphate-solubilizing microbes” and “phosphate- solubilizing bacteria” or “phosphate solubilizers” may refer to a microbe or bacterium that either (a) has a measurable increase in phosphate made soluble relative to a control in an assay that comprises suspending adding a test solution that comprises the microbe or bacterium to media with insoluble, inorganic phosphate (e.g., tricalcium phosphate (Ca3(PO4)2), incubating the media, and measuring an amount of insoluble phosphate released into solution and made soluble using an instrument such as a SEAL AQ400 and comparing to known standards, or (b) makes a visible halo when grown in a colony on a plate with insoluble phosphate.

[0138] Microbes described herein as “nitrogen use efficiency microbes” and “nitrogen use efficiency bacteria” may refer a microbe or bacterium that (a) has a measurable increase in nitrogen fixation (conversion of molecular dinitrogen (N2) into ammonia (NH3)) relative to a control in an assay that comprises adding a test solution that comprises the microbe or bacterium to media, incubating the media, and measuring an amount of molecular dinitrogen and ammonia using an instrument such as a SEAL AQ400 and comparing to known standards, (b) is identified via an acetylene reduction assay as described herein, (c) is identified based on nifh gene quantification (e.g., the microbe may have a nifti gene), (c) recruits nitrogen fixers to the root zones or other tissues of plants, increases organic nitrogen content, in the soil, or increases mineralization of organic nitrogen in the soil, or (d) enables plant growth in nitrogen concentrations that normally reduce growth compared to that of a control (e.g., non-nitrogen use efficiency microbe).

[0139] As used herein, the term “yield” can refer to the amount of harvestable plant material or plant-derived product, and is normally defined as the measurable produce of economic value of a crop.

[0140] For crop plants, “yield” can also mean the amount of harvested material per acre or unit of production. Yield may be defined in terms of quantity or quality. The harvested material may vary from crop to crop, for example, it may be seeds, above ground biomass, roots, fruits, cotton fibers, any other part of the plant, any plant-derived product which is of economic value, or any combination thereof.

[0141] In some embodiments, microbial strains, isolates, cultures, compositions, biostimulant compositions, or any combination thereof described herein may be used to promote improved crop or product quality, including food quality. Improving crop quality may comprise improving characteristics that make a crop or product more marketable such as, for example, a desiredcolor, size, or shape. Improving crop quality may also including making a food product with desired nutritional characteristics, nutraceutical characteristics, pharmaceutical characteristics, or any combination thereof. Improving crop or product quality may comprise adding valuable attributes that may increase price. Improving crop or product quality may comprise increasing value per harvest unit, such as, for example, providing increased oil, sugar, starch, or protein yield per unit weight of harvested crop or product.

[0142] In some embodiments, the microbial strains, isolates, cultures and compositions according to the embodiments of this application can lead to plant growth promotion or plant growth improvement that is an at least 5% increase, at least 10% increase, at least 25% increase, at least 50% increase, at least 75% increase, or at least a 100% increase in the property being measured. In some embodiments, the microbial strains, isolates, cultures and compositions according to the embodiments of this application lead to plant growth promotion or plant growth improvement that is at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or a 100% increase in the property being measured. In some embodiments, the microbial strains, isolates, cultures and compositions of this application may produce an above stated percentage increase in zinc uptake, an above stated increase in zinc solubilization capacity, an above stated percentage increase in phosphate uptake, an above stated increase in phosphate solubilization capacity, an above stated percentage increase in nitrogen fixation, an above stated increase in nitrogen content, an above stated increase in nitrogen acquisition, an above stated increase in nitrogen uptake, or an above stated increase in total root weight, leaf area, or plant product yield (e.g., an above stated percentage increase in plant product weight).

[0143] A “control plant”, as used herein, can provide a reference point for measuring changes in phenotype of the subject plant, and may be any suitable plant cell, seed, plant component, plant tissue, plant organ or whole plant. A control plant may include, but is not limited to, (a) a plant which is genetically identical to the subject plant but which is not exposed to the same treatment (e.g., inoculant treatment) as the subject plant or (b) the subject plant itself, under conditions in which it has not been exposed to a particular treatment such as, for example, an inoculant or combination of inoculants, a biostimulant composition, other chemicals, or any combination thereof. A treated plant may comprise a plant that has had an inoculum of a microbe and / or a biostimulant composition as described herein applied to any part of the plant (e.g., seed, stem,root, shoot, leaf, or any combination thereof). A treated plant may comprise a plant that has had an inoculum of a microbe or a biostimulant composition as described herein applied using an infurrow application. A treated plant may comprise a plant that has had an inoculum of a microbe and / or a biostimulant composition as described herein applied using a side-dress application, a broadcast application, a y-drop application, a tape application, a fertigation application, or any combination of applications thereof. A treated plant may comprise a plant which has had an inoculum of a microbe or a biostimulant composition as described herein applied to the seed of the plant. A treated plant may comprise a plant that has had an inoculum of a microbe or a biostimulant composition as described herein applied to the soil. An untreated plant may comprise a plant that that has not had an inoculum of a microbe or a biostimulant composition as described herein applied directly or indirectly.

[0144] “Inoculant” as used herein can refer to any culture or preparation that comprises at least one microorganism. In some embodiments, an inoculant (sometimes referred to as a microbial inoculant or a soil inoculant) is an agricultural addition that uses beneficial microbes (including but not limited to endophytes) to promote plant health, growth and / or yield. Many of the microbes suitable for use in an inoculant form symbiotic relationships with the target crops where both parties benefit (mutualism). For example, an isolated microbial strain as described herein may benefit from carbon sources from the roots of a plant and the plant may benefit from metabolites generated by metabolism of the microbe. Without wishing to be bound by theory, a plant may be colonized by the isolate, and the colonization of the roots may block plant pathogens from accessing the roots and / or provide plant growth and / or health benefits to the plant. An inoculant (e.g., inoculum of a microbe / microbial strain) can be added at one time point during an operating of a bioreactor system process. An inoculant (e.g., inoculum of a microbe / microbial strain) can be added at two or more time points during an operating of a bioreactor system process.

[0145] The term “serialized isolate production”, (sIP), can refer to specialized manipulated continuous serialized reactors that may enable the growth and enrichment of the microbes, isolates, target isolates, and / or microorganisms as described herein.

[0146] The term “reactor” can refer to a container of a system described herein that may hold a volume of fluid. The terms “reactor” and “container” can be used interchangeably.

[0147] The term “floc” can refer to a mass formed by the aggregation of a number of fine suspended particles. Particles can comprise biological particles, non-biological particles, or a combination thereof. For example, a floc can comprise organic materials recovered from a feedstock, waste, wastewater, sludge material, or any combination thereof of a fluid used in abioreactor system. A floc can comprise biosolids and / or particles from digestion products of organic materials. Floc can comprise an aggregated mass of microorganisms (e.g., bacteria).

[0148] The term “whole broth” (WB) can refer to a blend of supernatant and floc at a ratio for use in the technologies as described herein. A whole broth may comprise microbial populations (e.g., zinc-solubilizing microbes, phosphate solubilizing microbes, nitrogen use efficiencypromoting microbes), enzymes, fungi, biosolids, or any combination thereof. For example, bacterial genera of a whole broth may comprise Haliscomenobacler. I. ew ine Ila. Caldilinea, Terrimonas, Acidobacleriiim. Lewinella cohaerens, Thauera phenylacetica, Thauera mechernichensis, Solitalea canadensis, Nitrospira moscoviensis, Kosakonia, Klebsiella, Rahnella, Kluyvera, Enterobacter, Achromobacter, Microbacterium, Gluconobacter, Methylobacterium, Pseudomonas, Pantoea, Azospirillum, Azocarus, Herbaspirillum, Burkholderia, Cyanobacteria, Bacillus, Paenibacillus, Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enterobacter spp., Achromobacter marplatensis, Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enterobacter spp., Achromobacter marplatensis, Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Paenibacillus borealis, Bacillus megaterium (Priestia megaterium), Bacillus amyloliquefaciens, Bacillus licheniformis, Paenibacillus sonchi, or any combination thereof. A whole broth may have plant growth promotion properties. For example, a whole broth may have zinc-solubilization capacity, phosphate-solubilization capacity, nitrogen-fixation capacity, or any combination thereof.

[0149] The terms “microbial consortium” or “microbial population” can refer to a group of microorganisms in an environment. Consortiums may be endosymbiotic or ectosymbiotic. Microorganisms in a microbial consortium can include, but are not limited to, bacteria, fungi, yeasts, lichens, algae, protozoa, archaea, molds, or any combination thereof.

[0150] The terms “supernatant,” “biostimulant product,” or “base product” can refer to the final product of the bioreactor system. The supernatant can be measured for amount of a microbial isolate, number of members within a microbial consortium, or types and amount of microbial metabolites with plant growth promotion capacity.

[0151] The term “load rate” can refer to a rate at which a source material is introduced into a bioreactor system. In some embodiments, load rate may refer to “organic load rate” or “hydraulic load rate”. An organic load rate can comprise a rate at which feedstock (e.g., organicfeedstock) is introduced into the system. A hydraulic load rate can comprise a rate at which a hydraulic source is introduced into the system.

[0152] The term “internal recycle rate” can refer to a rate at which a working fluid is recycled within a phase space.

[0153] The term “hydraulic feed rate” can refer to a rate at which working fluid is transferred between phase spaces.

[0154] The term “hydraulic dwell time” can refer to an amount of time that a working fluid is present in a phase space.

[0155] The term “working fluid” can refer to a fluid substance supporting and transporting biology and nutrients through a system of containers. For example, a working fluid may comprise organic materials, microorganisms (e.g., microbes and / or metabolites), biosolids, macronutrients, micronutrients, organic nutrients, inorganic nutrients, or any combination thereof. A working fluid can comprise a solution that flows throughout a bioreactor system and may provide an enriched environment for microbes of the bioreactor system.

[0156] Whenever the term “at least,” “greater than,” or “greater than or equal to” precedes the first numerical value in a series of two or more numerical values, the term “at least,” “greater than” or “greater than or equal to” can apply to each of the numerical values in that series of numerical values. For example, greater than or equal to 1, 2, or 3 is equivalent to greater than or equal to 1, greater than or equal to 2, or greater than or equal to 3.

[0157] Whenever the term “at most,” “no more than,” “less than,” or “less than or equal to” precedes the first numerical value in a series of two or more numerical values, the term “at most,” “no more than,” “less than,” or “less than or equal to” can apply to each of the numerical values in that series of numerical values. For example, less than or equal to 3, 2, or 1 is equivalent to less than or equal to 3, less than or equal to 2, or less than or equal to 1.II. Bioreactor systems for Production of Isolates

[0158] The present disclosure provides systems (e.g., bioreactor systems) with conditions to produce biostimulant products with plant-growth promoting capabilities (e.g., improved zinc solubilization capacity, improved phosphate solubilization capacity, promotion of efficient use of nitrogen).

[0159] Embodiments of systems and methods described herein can produce biostimulant products that may have a multi-modal way of promoting zinc uptake in plants. Embodiments of systems and methods described herein can produce biostimulant products that may have a multimodal way of promoting phosphate uptake in plants. Embodiments of systems and methodsdescribed herein can produce biostimulant products that may have a multi-modal way of promoting efficient use of nitrogen by plants. Biostimulant products produced by embodiments described herein may be used to promote plant growth by applying the products to horticultural items such as plants, plant parts, seeds plant growth media (e.g., soil, fertilizer), or any combination thereof.

[0160] FIGs. 1-8 schematically illustrate examples of bioreactor systems 100 with conditions (e.g., microbes) that produce biostimulant products that may have a multi-modal way of promoting zinc solubilization, phosphate solubilization, and / or efficient use of nitrogen by plants.

[0161] For example, in FIG. 6., the system 100 comprises a water tank 141 that comprises city water 142 and RO water 143. The system 100 also comprises a first reactor 111, a second reactor 112, a third reactor 113, and / or a clarifier container 120 connected sequentially in which feedstock can continuously flow and microbial consortia as described herein can be grown. The first reactor, second reactor, and / or third reactor may be packed bed reactors with a scaffolding within the reactors or may be fluidized bed reactors without the scaffolding. The water tank 141 is coupled to the first reactor 111 and provides continuous flow of water to the first reactor 111. The system 100 can also comprise an input channel that flows inputs into the bioreactor system. In some cases, the inputs into the bioreactor system 130 are flown into the first reactor through the water tank. In other cases, the input composition can be added to the first reactor. The inputs, as described herein, may comprise one or more of water 140, a microbial inoculum 134, nutrients 133, and / or a digestion substrate (e.g., aqueous organic feedstock).

[0162] In some embodiments, a reactor tank circulates working fluid within itself to recycle working fluid, wherein reactor tanks can comprise outflow pipes to circulate and recycle working fluid within each tank. Ports and piping between tanks can assist in transferring working fluid to adjacent reactor tanks. A working fluid in a final clarifier of a system may transfer from the reactor tank to a clarifier may produce a supernatant (e.g., base product). Working fluid flows through the serialized reactor system which can aid in selective growth of the added isolate and other microbes present that have the same property as the added isolate. Base product from the clarifier can be accessed and further analyzed for microbial composition. In a bioreactor system described herein, a working fluid may flow from a mixing chamber through at least one reaction and to a clarifier container.

[0163] Parameters of the bioreactor system, such as flow rate and the solids content of the feedstock (e.g., organic feedstock), may be varied to achieve desired properties in the outflow biostimulant base product. In some embodiments, the hydraulic source is water. In someembodiments, the hydraulic source is a base product of another system. In some embodiments, the hydraulic source is a combination of water and a base product of another system. Water from the hydraulic source can be added to the feedstock (e.g., organic feedstock) of the bioreactor system to make an aqueous organic feedstock.

[0164] In some aspects, provided herein is a method of making a biostimulant composition. The biostimulant composition may comprise a plant growth promoting property (e.g., a desired plant growth promoting property). A bioreactor system may be provided. A bioreactor system comprising one or more containers (e.g., two or more containers) may be provided. A bioreactor system comprising one or more containers (e.g., two or more containers) arranged in a series may be provided. A container of the one or more containers (e.g., two or more containers) may comprise a volume of a working fluid. In some embodiments, each container of the one or more containers (e.g., two or more containers) may comprise a volume of a working fluid. The one or more containers (e.g., two or more containers) may comprise a first container, a second container, a third container, a fourth container, and the like. A container of the bioreactor system (e.g., a first container of the bioreactor system) may comprise a population of a microbial strain. The population of a microbial strain may comprise an established population of a microbial strain. The population of a microbial strain (e.g., the established population of a microbial strain) may have a plant growth promoting property (e.g., a desired plant growth promoting property).

[0165] In some embodiments, the bioreactor system may be operated for a duration of time. Operating the bioreactor system can comprise transferring an aqueous feedstock in a container (e.g., a first container). The aqueous feedstock can comprise a microbial consortium. Operating the bioreactor system can comprise transferring a portion of working fluid of a container (e.g., of the one or more containers) of the bioreactor system. Working fluid may be transferred out of a container of the two or more containers into a subsequent container. For example, a portion of working fluid may be transferred out of a second container into a third container. Working fluid may be transferred out of a container of the two or more containers as a product outflow stream. Operating the bioreactor system can comprise collecting at least a portion of working fluid. For example, at least a portion of the product outflow stream may be collected as the biostimulant composition.

[0166] The population of the microbial strain (e.g., the established population of the microbial strain) may not be present in the aqueous feedstock. The population of the microbial strain (e.g., the established population of the microbial strain) may not be present in the aqueous feedstock during the duration of time of operation. The population of the microbial strain (e.g., the established population of the microbial strain) may not be present in any other input to thebioreactor system (e.g., to a container of the bioreactor system). The population of the microbial strain (e.g., the established population of the microbial strain) may not be present in any other input to the bioreactor system (e.g., to a container of the bioreactor system) during the duration of time of operation. Any other input may comprise nutrients and / or digestion substrates (e.g., manure, biosolids, food waste, energy crops, or any combination thereof). In some embodiments, the established population of a microbial strain may not be present in the aqueous feedstock and / or any other input to the bioreactor system during a duration of operation at a concentration that is higher than 1% of the concentration of the established population of the microbial strain in the first container.

[0167] In some embodiments, operating the bioreactor system can comprise maintaining a concentration of the microbial strain. The concentration of the microbial strain may be maintained for the duration of time at which the bioreactor system is operating. A concentration of the microbial strain may be maintained in a container (e.g., a first container) at at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or greater than about 95% of a concentration of the microbial strain at the beginning of the duration of time. A concentration of the microbial strain may be maintained in a container (e.g., a first container) at at most about 95%, at most about 90%, at most about 85%, at most about 80%, at most about 75%, at most about 70%, or less than about 70% of a concentration of the microbial strain at the beginning of the duration of time. For example, a concentration of the microbial strain may be 1000 CFU / ml at the beginning of the duration of time and the concentration of the microbial strain may be maintained at 800 CFU / ml in a first container of the bioreactor system.

[0168] As an example, the present disclosure provides a method of making a biostimulant composition having a desired plant growth promoting property, the method comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the desired plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as the biostimulant composition; wherein the established population of the microbial strain is not present in an aqueous feedstock or any other input in thebioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the established population of the microbial strain in the first container.

[0169] As another example, the present disclosure provides a method of making a biostimulant composition having a desired plant growth promoting property, the method comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the desired plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; (iii) maintaining a concentration of the microbial strain throughout the duration of time in at least the first container at at least 80% of a concentration of the microbial strain at the beginning of the duration of time; and (iv) collecting at least a portion of the product outflow stream as the biostimulant composition; wherein the duration of time is at least 5 days; and wherein the microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the microbial strain in the first container.

[0170] Maintaining the concentration of the microbial strain may comprise maintaining the concentration at at least about 10 CFU / ml, at least about 1x102CFU / ml, at least about 5x102CFU / ml, at least about 1x103CFU / ml, at least about 5x103CFU / ml, at least about 1x104CFU / ml, at least about 5x104CFU / ml, at least about 1x105CFU / ml, or greater than about 1x105CFU / ml. Maintaining the concentration of the microbial strain may comprise maintaining the concentration at at most about 1x105CFU / ml, 5x104CFU / ml, 1x104CFU / ml, 5x103CFU / ml, 1x103CFU / ml, 5x102CFU / ml, 1x102CFU / ml, 10 CFU / ml, or less than about 10 CFU / ml.

[0171] In some embodiments, maintaining the concentration of the microbial strain may comprise providing nutrients. The nutrients may be provided in a container (e.g., a first container) of the bioreactor system. The nutrients may selectively promote growth of the microbial strain and / or sustain the established population. In some embodiments, maintaining the concentration of the microbial strain may comprise applying selective pressure. The selective pressure may comprise a selective pressure described herein. In some embodiments, the selective pressure may be applied in at least one container of the bioreactor system (e.g., in at least the first container). The selective pressure may comprise conditions that reduce a growth rate of microbes in a container (e.g., in the first container). The growth rate of microbes in thecontainer may be reduced relative to a growth rate of the microbial strain (e.g., the established population of the microbial strain).

[0172] In some embodiments, the biostimulant composition can comprise at least a portion of the established population of the microbial strain. In some embodiments, the biostimulant composition can comprise an amount of the microbial strain. The amount of the microbial strain may be a concentration of the microbial strain. The amount of the microbial strain can be at least about 1 CFU / ml, at least about 10 CFU / ml, at least about 100 CFU / ml, at least about 1x103CFU / ml, at least about 1x104CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, at least about 1x107CFU / ml, at least about 1x108CFU / ml, at least about 1x109CFU / ml, at least about 1x1010CFU / ml, or greater than about 1x1010CFU / ml. The amount of the microbial strain can be at most about 1x1010CFU / ml, at most about 1x109CFU / ml, at most about 1x108CFU / ml, at most about 1x107CFU / ml, at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 1x104CFU / ml, at most about 1x103CFU / ml, at most about 100 CFU / ml, at most about 10 CFU / ml, at most about 1 CFU / ml, or less than about 1 CFU / ml.

[0173] The established population of the microbial strain (e.g., at least a portion of the established population of the microbial strain) may be configured to promote a plant growth promoting property (e.g., increase nutrient uptake of a plant). Administering the biostimulant composition to a plant may promote a plant growth promoting property (e.g., increase a nutrient uptake) of the plant compared to that of a plant not administered the biostimulant composition.

[0174] As an example, the present disclosure provides a method, comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having a plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the amount of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of a plant not administered the biostimulant composition.

[0175] As another example, the present disclosure provides a method, comprising: (a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises anestablished population of a microbial strain having a plant growth promoting property; (b) operating the bioreactor system for a duration of time by: (i) transferring into the first container an aqueous feedstock comprising a microbial consortium; (ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and (iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the biostimulant composition is configured to promote a plant growth promoting property of a plant administered the biostimulant composition as compared to that of a plant not administered the biostimulant composition. The biostimulant composition can be capable of promoting a plant growth promoting property.

[0176] The nutrient uptake of the plant may be measured by an assay including, but not limited to, analysis of biomass, depletion method, or nutrient balance method. The nutrient uptake may comprise macronutrients. The macronutrients can comprise nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or any combination thereof. The nutrient uptake may comprise micronutrients. The micronutrients can comprise boron, zinc, manganese, iron, copper, or a combination thereof. In some embodiments, the nutrient uptake may comprise a combination of macronutrients and micronutrients.

[0177] A plant growth promoting property may comprise any plant growth promoting property described herein. For example a plant growth promoting property can comprise an ability to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof. In some embodiments, the plant growth promoting property (e.g., desired plant growth promoting property) may not comprise nitrogen use efficiency, zinc solubilization, phosphate solubilization, or any combination thereof.

[0178] A plant administered the biostimulant composition described herein may have an increased biomass compared to the plant not administered the biostimulant composition. For example, a biomass (e.g., a total biomass) of a plant administered the biostimulant composition described herein can be increased by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, or greater than about 25% as compared to that of a plant not administered thebiostimulant composition. A biomass (e.g., a total biomass) of a plant administered the biostimulant composition described herein can be increased by at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition. In some embodiments, a biomass (e.g., a total biomass) of a plant administered the biostimulant composition described herein can be increased from about 1% to about 20% as compared to that of a plant not administered the biostimulant composition. In some embodiments, a biomass (e.g., a total biomass) of a plant administered the biostimulant composition described herein can be increased from about 1% to about 2%, about 1% to about 3%, about 1% to about 4%, about 1% to about 5%, about 1% to about 6%, about 1% to about 7%, about 1% to about 8%, about 1% to about 10%, about 1% to about 12%, about 1% to about 15%, about 1% to about 20%, about 2% to about 3%, about 2% to about 4%, about 2% to about 5%, about 2% to about 6%, about 2% to about 7%, about 2% to about 8%, about 2% to about 10%, about 2% to about 12%, about 2% to about 15%, about 2% to about 20%, about 3% to about 4%, about 3% to about 5%, about 3% to about 6%, about 3% to about 7%, about 3% to about 8%, about 3% to about 10%, about 3% to about 12%, about 3% to about 15%, about 3% to about 20%, about 4% to about 5%, about 4% to about 6%, about 4% to about 7%, about 4% to about 8%, about 4% to about 10%, about 4% to about 12%, about 4% to about 15%, about 4% to about 20%, about 5% to about 6%, about 5% to about 7%, about 5% to about 8%, about 5% to about 10%, about 5% to about 12%, about 5% to about 15%, about 5% to about 20%, about 6% to about 7%, about 6% to about 8%, about 6% to about 10%, about 6% to about 12%, about 6% to about 15%, about 6% to about 20%, about 7% to about 8%, about 7% to about 10%, about 7% to about 12%, about 7% to about 15%, about 7% to about 20%, about 8% to about 10%, about 8% to about 12%, about 8% to about 15%, about 8% to about 20%, about 10% to about 12%, about 10% to about 15%, about 10% to about 20%, about 12% to about 15%, about 12% to about 20%, or about 15% to about 20% as compared to that of a plant not administered the biostimulant composition.

[0179] A plant administered the biostimulant composition described herein may have an increased average leaf area compared to the plant not administered the biostimulant composition. For example, an average leaf area of a plant administered the biostimulant composition described herein can be increased by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulantcomposition. An average leaf area of a plant administered the biostimulant composition described herein can be increased by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0180] In some embodiments, an average leaf area of a plant administered the biostimulant composition described herein can be increased from about 1% to about 50% as compared to that of a plant not administered the biostimulant composition. In some embodiments, an average leaf area of a plant administered the biostimulant composition described herein can be increased from about 1% to about 2%, about 1% to about 3%, about 1% to about 4%, about 1% to about 5%, about 1% to about 10%, about 1% to about 15%, about 1% to about 20%, about 1% to about 25%, about 1% to about 30%, about 1% to about 40%, about 1% to about 50%, about 2% to about 3%, about 2% to about 4%, about 2% to about 5%, about 2% to about 10%, about 2% to about 15%, about 2% to about 20%, about 2% to about 25%, about 2% to about 30%, about 2% to about 40%, about 2% to about 50%, about 3% to about 4%, about 3% to about 5%, about 3% to about 10%, about 3% to about 15%, about 3% to about 20%, about 3% to about 25%, about 3% to about 30%, about 3% to about 40%, about 3% to about 50%, about 4% to about 5%, about 4% to about 10%, about 4% to about 15%, about 4% to about 20%, about 4% to about 25%, about 4% to about 30%, about 4% to about 40%, about 4% to about 50%, about 5% to about 10%, about 5% to about 15%, about 5% to about 20%, about 5% to about 25%, about 5% to about 30%, about 5% to about 40%, about 5% to about 50%, about 10% to about 15%, about 10% to about 20%, about 10% to about 25%, about 10% to about 30%, about 10% to about 40%, about 10% to about 50%, about 15% to about 20%, about 15% to about 25%, about 15% to about 30%, about 15% to about 40%, about 15% to about 50%, about 20% to about 25%, about 20% to about 30%, about 20% to about 40%, about 20% to about 50%, about 25% to about 30%, about 25% to about 40%, about 25% to about 50%, about 30% to about 40%, about 30% to about 50%, or about 40% to about 50% as compared to that of a plant not administered the biostimulant composition.

[0181] In some embodiments, the established population of the microbial strain can comprise a zinc solubilizing microbial strain. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a zinc solubilizing microbial strain) can be increased by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared tothat of a plant not administered the biostimulant composition. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a zinc solubilizing microbial strain) can be increased by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0182] In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a zinc solubilizing microbial strain) to a plant can increase a macronutrient content (e.g., a nitrogen content, a phosphorous content, a potassium content, or any combination thereof) by at least about 1 μg, at least about 2 μg, at least about 3 μg, at least about 4 μg, at least about 5 μg, at least about 10 μg, at least about 15 μg, at least about 20 μg, at least about 25 μg, at least about 30 μg, at least about 40 μg, at least about 50 μg, or greater than about 50 μg as compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a zinc solubilizing microbial strain) to a plant can increase a macronutrient content (e.g., a nitrogen content, a phosphorous content, a potassium content, or any combination thereof) by at most about 50 μg, at most about 40 μg, at most about 30 μg, at most about 25 μg, at most about 20 μg, at most about 15 μg, at most about 10 μg, at most about 5 μg, at most about 4 μg, at most about 3 μg, at most about 2 μg, at most about 1 μg, or less than about 1 μg as compared to that of a plant not administered the biostimulant composition.

[0183] In some embodiments, the established population of the microbial strain can comprise a phosphate solubilizing microbial strain. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) can be increased by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulant composition. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) can be increased by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0184] In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) to a plant can increase a crop yield by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) to a plant can increase a crop yield by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0185] In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) to a plant can increase a dry biomass by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a phosphate solubilizing microbial strain) to a plant can increase a dry biomass by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0186] In some embodiments, the established population of the microbial strain can comprise a nitrogen use efficiency-promoting microbial strain. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiencypromoting microbial strain) can be increased by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, or greater than about 80% as compared to that of a plant not administered the biostimulant composition. An average leaf area of a plant administered the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiency -promoting microbial strain) can be increased by at most about 80%, at most about 70%, at most about 60%, at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, atmost about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0187] In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiency-promoting microbial strain) to a plant can increase a dry shoot weight, a dry root weight, a stem diameter, or any combination thereof by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiency-promoting microbial strain) to a plant can increase a dry shoot weight, a dry root weight, a stem diameter, or any combination thereof by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0188] In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiency-promoting microbial strain) to a plant can increase a leaf chlorophyll content by at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or greater than about 50% as compared to that of a plant not administered the biostimulant composition. In some embodiments, administering the biostimulant composition (e.g., a biostimulant comprising a nitrogen use efficiency -promoting microbial strain) to a plant can increase a leaf chlorophyll content by at most about 50%, at most about 40%, at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, or less than about 1% as compared to that of a plant not administered the biostimulant composition.

[0189] In some embodiments, prior to (a) of a method described herein, an inoculum of the microbial strain may be added to the bioreactor system (e.g., may be added to a container of the bioreactor system). An inoculum of the microbial strain may be added to one or more containers of the bioreactor system (e.g., a first container, a second container, a third container, a fourth container, a fifth container, or any combination thereof). Adding the inoculum of the microbial strain may produce an initial population of the microbial strain. The initial population of the microbial strain may be present at a concentration in at least one container of the bioreactorsystem. For example, an initial population of the microbial strain may be present at at least about 10 CFU / ml, at least about 100 CFU / ml, at least about 0.5x103CFU / ml, at least about 1x103CFU / ml, at least about 0.5x104CFU / ml, at least about 1x104CFU / ml, at least about 0.5x105CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, or greater than about 1x106CFU / ml in at least one container. An initial population of the microbial strain may be present at at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 0.5x105CFU / ml, at most about 1x104CFU / ml, at most about 0.5x104CFU / ml, at most about 1x103CFU / ml, at most about 0.5x103CFU / ml, at most about 100 CFU / ml, at most about 10 CFU / ml, or less than about 10 CFU / ml.

[0190] The aqueous feedstock may comprise an organic material. The organic material can be at least partially digestible by microbes (e.g., microbes present in at least one container of the bioreactor system). In some embodiments, the organic material may have been partially digested by microbes endogenous to the organic material. The organic material may have been partially digested prior to a transferring of step (b)(i) of a method described herein. In some embodiments, a method described herein may further comprise digesting the organic material in one or more containers. In some embodiments, a method described herein may further comprise digesting the organic material in serially connected containers (e.g., two or more serially connected containers). The digesting of the organic material can occur before the transferring of step (b)(i) of a method described herein. The organic material may comprise an organic material described herein.

[0191] The aqueous feedstock may comprise an inorganic material. The inorganic material can comprise rock phosphate (e.g., rock phosphate particles). In some embodiments, the inorganic material (e.g., rock phosphate particles) may have been partially digested by microbes (e.g., microbes present in the aqueous feedstock). The inorganic material may have been partially digested by microbes (e.g., microbes present in the aqueous feedstock). In some embodiments, a method described herein may further comprise digesting the inorganic material (e.g., rock phosphate particles) in one or more containers. In some embodiments, a method described herein may further comprise digesting the inorganic material (e.g., rock phosphate particles) in serially connected containers (e.g., two or more serially connected containers). The digesting of the inorganic material (e.g., rock phosphate particles) can occur before the transferring of step (b)(i) of a method described herein. In some embodiments, digestion (e.g., partial digestion) of the organic material and inorganic material can occur at the same time. In some embodiments, digestion (e.g., partial digestion) of the organic material and inorganic material can occur at different times. In some embodiments, digestion (e.g., partial digestion) of the organic materialand inorganic material can occur in the same container (e.g., a same container of the bioreactor system). For example, the organic material and inorganic material may be digested or partially digested in a first container. In some embodiments, digestion (e.g., partial digestion) of the organic material and inorganic material can occur in different containers (e.g., different containers of the bioreactor system). For example, the organic material may be digested or partially digested in a first container and the inorganic material may be digested or partially digested in a second container.

[0192] The microbial consortium of the aqueous feedstock may be present at a concentration. In some embodiments, the microbial consortium can comprise at least about 100 CFU / ml, at least about 1x103CFU / ml, at least about 1x104CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, at least about 1x107CFU / ml, at least about 1x108CFU / ml, at least about 1x109CFU / ml, at least about 1x1010CFU / ml, or greater than about 1x1010CFU / ml. In some embodiments, the microbial consortium can comprise at most about 1x1010CFU / ml, at most about 1x109CFU / ml, at most about 1x108CFU / ml, at most about 1x107CFU / ml, at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 1x104CFU / ml, at most about 1x103CFU / ml, at most about 100 CFU / ml, or less than about 100 CFU / ml. The microbial consortium may comprise microbes derived from an organic material, an inorganic material, or any combination thereof. In some embodiments, the microbial consortium may comprise microbes derived manure, rock phosphate particles, or any combination thereof.

[0193] In some embodiments, a method described herein may further comprise producing metabolites. The metabolites may be produced from microbes of the bioreactor system. For example, metabolites may be produced from the microbial consortium, the population of the microbial strain (e.g., the established population of the microbial strain), or any combination thereof. In some embodiments, the metabolites (e.g., microbial metabolites) may have a plant growth promoting property. The metabolites (e.g., microbial metabolites) may have two or more plant growth promoting properties. In some embodiments, the metabolites (e.g., microbial metabolites) may have the same plant growth promoting property as the established population of the microbial strain. For example, the metabolites (e.g., microbial metabolites) and the established population of the microbial strain may increase nutrient uptake of a plant.

[0194] In some embodiments, the transferring of step (b)(i), the transferring of step (b)(ii), the collecting of step (b), or any combination thereof of the method described herein may be performed continuously. The transferring and / or collecting of the method described herein can be performed continuously over the duration of time. In some embodiments, the transferring of step (b)(i), the transferring of step (b)(ii), the collecting of step (b), or any combination thereofof the method described herein may be performed periodically. The transferring and / or collecting of the method described herein can be performed periodically over the duration of time.

[0195] A method described herein may further comprise adding one or more carbon sources to a container (e.g., at least one container of the bioreactor system). The carbon source may comprise any carbon sources described herein. In some embodiments, a carbon source (e.g., one or more carbon sources) may be comprised in the aqueous feedstock.

[0196] A bioreactor system described herein can comprise a clarifier (e.g., a clarifier container). The clarifier (e.g., a clarifier container) may comprise a working fluid (e.g., a clarifier working fluid). A method described herein can further comprise separating portions of the clarifier working fluid. In some embodiments, a method described herein comprises separating a supernatant portion of the clarifier working fluid from a floc portion of the clarifier working fluid. The separating can occur within the clarifier container. A method described herein may further comprise folding the floc portion of the clarifier working fluid. The folding may comprise releasing a population of the microbial strain. The population of the microbial strain may release into the supernatant portion (e.g., the supernatant portion of the clarifier working fluid). While the folding can release the population of the microbial strain into a supernatant portion of the clarifier working fluid, the folding may not introduce solid from the floc portion (e.g., floc solids) into the supernatant portion of the clarifier working fluid.

[0197] In some embodiments, the method can further comprise transferring a floc portion. The floc portion may be transferred from the clarifier container to a container of bioreactor system. The floc portion may be reintroduced into an earlier container in the bioreactor system by transferring the floc portion from the clarifier container to the earlier container. For example, the floc portion may be transferred from the clarifier container to a first container, second container, third container, fourth container, fifth container, sixth container, or any combination thereof of the bioreactor system. The product outflow stream of the bioreactor system can comprise the supernatant portion of the clarifier working fluid. The product outflow stream may be collected from the bioreactor system.

[0198] In some embodiments, a method of making a biostimulant composition is performed using a bioreactor system that comprises two or more containers arranged in a series. A series of containers can comprise 2, 3, 4, 5, 6, 7 or more containers. The containers may be the same or different. A series of containers may be fluidly connected to the other containers in the series. A series of containers may be arranged such that fluid can flow from a first container to a second container and from a second container to a third container, and so on from each subsequentcontainer to a next container in the series. Referring to a container as a “first container” does not necessarily indicate that the referenced container is the most upstream container in a series of container. In some embodiments, additional containers in a series are upstream of a container referred to as a “first container.” In some embodiments, a container that is the most upstream container in a series of fluidly connected containers does not have another container upstream of the container. In some embodiments, a container that is the most upstream container in a series receives an inflow stream such as, for example, a feedstock stream. In some embodiments, a feedstock stream comes from a mixing container, which may be referred to as a mixing tank or complete mix reactor. In some embodiments, components that are placed in a mixing container have a shorter dwell time than working fluids in subsequent containers in the system. In some embodiments, a container that is the most downstream container in a series is a clarifier, which may be configured to separate a supernatant portion of a working fluid from a floc portion of the working fluid. The most downstream container may not be a clarifier and may not be configured to separate different portions of a working fluid. In some embodiments, the most downstream container has an outlet port through which a product outflow stream may exit the most downstream container. Containers arranged in a series may have inflows other than those coming from the neighboring upstream container and may have outflows other than those coming from the neighboring downstream container.

[0199] In some embodiments, a reactor or a series of reactors (e.g., serialized assembly of reactors) functions to contribute to the growth of one or more microbes having desired plant growth promoting properties and / or to the production of digestion products having plant growth promoting properties. In some embodiments, the operation of a bioreactor system may lead to growth of one or more microbes having a desired plant growth promoting effect. The one or more microbes may be one or more isolated microbes added separately as an inoculum to the bioreactor system. The one or more microbes may also be input into the system as part of a feed material that comprises a mixture of microbes. The one or more microbes may be endogenous to an organic material such as, for example, a manure, a plant, a lignocellulosic material, or an algae. The one or more microbes may also be endogenous to other types of feed materials, such as rock phosphate. In some embodiments, endogenous microbes are those microbes naturally present in feedstock material (e.g., a manure, a plant, a lignocellulosic material, or an algae). These microbes may naturally reside in a closed system and / or are present in the ecosystem of the feedstock material.

[0200] In some embodiments, the bioreactor system may be operated in a hydraulically balanced manner. A hydraulically balanced manner can comprise working fluid transferring from onecontainer of the system to another container to a further container at an equal rate. In some embodiments, the transferring and / or collecting of the methods described herein may be driven by gravity. The outlet port of a container may be higher than an inlet port of another container so that the working fluid transfers via gravity between containers of the bioreactor system. In some embodiments, the opening of an outlet port of a first container is at the same level as the top of the working fluid or slightly below the level of the working fluid (i.e., the bottom of the opening of the outlet port may be 1, 2, 3, or 4 cm below the level of the working fluid) so that the rate of flow of liquid into the first container can be equaled by the rate of flow of liquid out of the first container. The first container may be fluidly connected to a second container, e.g., by a pipe or other conduit connected to the outlet port of the first container. The pipe or other conduit may connect to an inlet port of a second container. The inlet port of the second container may be below the level of the outlet port of the first container such that the working fluid from the first container may flow by gravity alone into the second container. The second container may have an outlet port with an opening that is at the same level or is at least partially below the level of the working fluid in the second container (i.e., the bottom of the opening of the outlet port may be 1, 2, 3, or 4 cm below the level of the working fluid) so that the rate of flow of liquid into the first container can be equaled by the rate of flow of liquid out of the first container solely due to the effect of gravity. Additional containers (e.g., a third, fourth, fifth, sixth, seventh, or more containers) can have inlet and outlet ports arranged in the same way, with each subsequent container having inlet and outlet ports arranged such that flow between containers can be continuous and driven solely by gravity, i.e., the inlet and outlet ports of each subsequent container in a series of containers is at a level lower than the outlet port of the previous container. In some embodiments, all the inlet and outlet ports of containers in the system are open at all times to allow continuous flow driven solely by gravity (i.e., without any pumps needed to drive flow from any container to a subsequent container in the system). Operating the bioreactor system can comprise maintaining a flow rate. In some embodiments, a flow rate may be maintained to result in a hydraulic retention time of at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 15 days, at least about 20 days, at least about 25 days, at least about 30 days, or greater than about 30 days. In some embodiments, a flow rate may be maintained to result in a hydraulic retention time of at most about 30 days, at most about 25 days, at most about 20 days, at most about 15 days, at most about 10 days, at most about 9 days, at most about 8 days, at most about 7 days, atmost about 6 days, at most about 5 days, at most about 4 days, at most about 3 days, at most about 2 days, at most about 1 day, or less than about 1 day.

[0201] Operating the bioreactor system can comprise maintaining the product outflow stream. The product outflow stream may be maintained at a flow rate of at least about 5 gallons per day, at least about 10 gallons per day, at least about 25 gallons per day, at least about 50 gallons per day, at least about 75 gallons per day, at least about 100 gallons per day, at least about 150 gallons per day, at least about 250 gallons per day, at least about 500 gallons per day, at least about 1,000 gallons per day, or greater than about 1,000 gallons per day. The product outflow stream may be maintained at a flow rate of at most about 1,000 gallons per day, at most about 500 gallons per day, at most about 250 gallons per day, at most about 150 gallons per day, at most about 100 gallons per day, at most about 75 gallons per day, at most about 50 gallons per day, at most about 25 gallons per day, at most about 10 gallons per day, at most about 5 gallons per day, or less than about 5 gallons per day.

[0202] The bioreactor system may operate continuously for a duration of time. In some embodiments, the bioreactor system may be operated continuously for at least about 5 days, at least about 10 days, at least about 20 days, at least about 30 days, at least about 40 days, at least about 50 days, at least about 60 days, at least about 70 days, at least about 80 days, at least about 90 days, at least about 100 days, at least about 150 days, at least about 200 days, or greater than about 200 days. In some embodiments, the bioreactor system may be operated continuously for at most about 200 days, at most about 150 days, at most about 100 days, at most about 90 days, at most about 80 days, at most about 70 days, at most about 60 days, at most about 50 days, at most about 40 days, at most about 30 days, at most about 20 days, at most about 10 days, at most about 5 days, or less than about 5 days.

[0203] In some embodiments, the methods described herein further comprise producing bacteria. The bacteria can be sporulated bacteria and the sporulated bacteria may be in the product outflow stream. The methods may further comprise producing a population of the microbial strain (e.g., an amount of the microbial strain) in the product outflow stream. The population of the microbial strain in the product outflow stream may be sporulated. An amount (e.g., a concentration) of the population of the microbial strain in the product outflow stream may be sporulated. In some embodiments, a population of the microbial strain that is sporulated may comprise at least about 100 CFU / ml, at least about 1x103CFU / ml, at least about 1x104CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, at least about 1x107CFU / ml, at least about 1x108CFU / ml, at least about 1x109CFU / ml, at least about 1x1010CFU / ml, or greater than about 1x1010CFU / ml. In some embodiments, a population of themicrobial strain that is sporulated may comprise at most about 1x1010CFU / ml, at most about 1x109CFU / ml, at most about 1x108CFU / ml, at most about 1x107CFU / ml, at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 1x104CFU / ml, at most about 1x103CFU / ml, at most about 100 CFU / ml, or less than about 100 CFU / ml. In some embodiments, the methods described herein can further comprise adding an additional population of the microbial strain to the biostimulant composition (e.g., the biostimulant product).

[0204] In an exemplary system of the present disclosure, a series of reactors functions to contribute to the growth of an inoculum of a microbe (e.g., isolate) having desired plant growth promoting properties. A series of reactors (e.g., serialized assembly of reactors) may also function to contribute to the production of microbial metabolites having desired plant growth promoting properties. The bioreactor system described herein can enrich an inoculum of a target microbe, a population of microbes within a microbial consortium with plant-growth promotion properties (e.g., zinc solubilization, phosphate solubilization, nitrogen use efficiency), metabolites produced by the enriched population of the microbial consortium with plant-growth promotion properties (e.g., zinc solubilization, phosphate solubilization, nitrogen use efficiency), or any combination thereof. This system provides added benefits to other bioreactor systems in that it can target a functional community of microbes and / or metabolites with specific functionality and enrich and / or maintain the community in the bioreactor system. The system can comprise two reactor chambers, three reactor chambers, four reactor chambers, five reactor chambers, or more reactor chambers (e.g., containers or chambers). Without wishing to be bound by theory, the serialized reactors enable the growth and enrichment of proprietary specialist target microbes with optimal plant growth promoting properties. The system may direct a flow of working fluid comprising an inoculum, carbon source, and / or nutrient source from an input feedstock (e.g., organic feedstock) to produce a base product (BP). A hydraulic source can flow into a reactor via in-flow port to comprise a first working fluid in a reactor tank. A substance providing selective pressure can flow into a reactor via in-flow port to comprise a first working fluid in a reactor tank. A hydraulic source and a substance providing selective pressure can flow into a reactor via in-flow port to comprise a first working fluid in a reactor tank. A hydraulic source may input into a first reactor or any reactor of the system. In some embodiments, a hydraulic source may input (e.g., flow) into a tank or container prior to a first reactor. In some embodiments, the container may comprise a “complete mixed reactor” (CMR). Other inputs into a system described herein may flow into any reactor of the system, including but not limited to a first reactor, a second reactor, a third reactor, or any other reactor following a first reactor. In some embodiments, the bioreactor system can comprise at least one containerprior to a first container (e.g., at least one container in the series of container before the first container).

[0205] In some embodiments, a method of making a biostimulant composition is described herein as including operating a bioreactor system for a duration of time. It is contemplated that the bioreactor system may be operated both before and after the duration of time in the same way as during the duration of time. Some embodiments of methods described herein may describe steps taking place during a particular duration of time, such as, for example, 1 day, 2 days, 3 days, 4 days, 5 days, 10 days, 15 days, 30 days, 60 days, 90 days, 120 days, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 1 year, 2 years, 3 years, 4 years, 5 years or more. Those durations of time described herein may be a fraction of a larger duration of time during which the bioreactor system is operating continuously or mostly continuously.

[0206] A digestion process to produce the biostimulant may be performed in a bioreactor system that comprises a tank, a container, a vessel (e.g., reactor), or a series of tanks, a series of containers, or a series of vessels (e.g., reactors) through which the feedstock continuously flows. A reactor can be a fluidly connected container, system, vessel, or tank in which microbial consortia including the microbes, isolates, and / or microorganisms as described herein can be grown. A reactor can be separate or continuous. A reactor may be a physical containment system arranged in a discrete order to favor growth of particular microbes. Types of reactors can include, but are not limited to, fluidized-bed reactors (FBRs) or packed-bed reactors (PBRs).

[0207] In an aspect, the present disclosure provides a bioreactor system. The bioreactor system can comprise a stream of an aqueous feedstock. The stream may be in fluid communication with a container (e.g., a first container). The container can comprise a volume of working fluid. For example, the stream may be in fluid communication with a first container of the bioreactor system comprising a volume of a first working fluid. The aqueous feedstock can comprise a microbial consortium, wherein the microbial consortium can comprise a microbial consortium described herein. The working fluid (e.g., the first working fluid) may comprise a microbial strain. The microbial strain can be a population of a microbial strain and may comprise a plant growth promoting property (e.g., a desired plant growth promoting property). A first container of the bioreactor system may comprise a mixer. The mixer can be configured to aerate the working fluid (e.g., the first working fluid) in a container.

[0208] In some embodiments, a concentration of the microbial strain may be higher in the working fluid than in the aqueous feedstock. In some embodiments, a concentration of the microbial strain may be higher in the working fluid than in any other input to the bioreactor. For example, a concentration of the microbial strain in the first working fluid may be at least about 2times higher, at least about 5 times higher, at least about 10 times higher, at least about 25 times higher, at least about 50 times higher, at least about 100 times higher, at least about 150 times higher, at least about 200 times higher, at least about 250 times higher, at least about 500 times higher, or greater than about 500 times higher than a concentration of the microbial strain in the aqueous feedstock stream or in any other input into the bioreactor system. A concentration of the microbial strain in the first working fluid may be at most about 500 times higher, at most about 250 times higher, at most about 200 times higher, at most about 150 times higher, at most about 100 times higher, at most about 50 times higher, at most about 25 times higher, at most about 10 times higher, at most about 5 times higher, at most about 2 times higher, or less than about 2 times higher than a concentration of the microbial strain in the aqueous feedstock stream or in any other input into the bioreactor system.

[0209] The bioreactor system described herein may comprise one or more additional containers (e.g., one or more containers in addition to a first container). The container can be arranged in a series. In some embodiments, the series of containers can comprise a first container, a second container, a third container, a fourth container, a fifth container, a sixth container, and any additional container(s). One or more additional containers in the bioreactor system may comprise a volume of working fluid. In some embodiments, each container of the one or more additional containers comprises a volume of working fluid. The one or more additional containers may be in fluid communication with a container of the series in the bioreactor system. In some embodiments, at least one of the containers of the bioreactor system can comprise a product outflow stream port. In some embodiments, each container of the series of containers may comprise a product outflow stream port. A product outflow stream (e.g., stream of the volume of working fluid of a container) can be in fluid communication with the product outflow stream port.

[0210] As an example, the present disclosure provides a bioreactor system comprising: (a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property, wherein a concentration of the microbial strain in the first working fluid is at least 100 times higher than a concentration of the microbial strain in the aqueous feedstock stream or in any other input into the bioreactor system; (b) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the oneor more additional containers comprises a product outflow stream port; and (c) a product outflow stream in fluid communication with the product outflow stream port.

[0211] In some embodiments, the product outflow stream may comprise a biostimulant composition. The biostimulant composition can comprise an amount of the microbial strain, wherein the amount of the microbial strain can be an amount described herein. The biostimulant composition can comprise a plant growth promoting property (e.g., a desired plant growth promoting property). In some embodiments, the amount of microbial strain can be configured to promote a plant growth promoting property (e.g., an ability to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof).

[0212] As another example, a bioreactor system can comprise: (a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property; (b) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and (c) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain is configured to promote a plant growth promoting property of a plant administered the biostimulant composition as compared to the plant not administered the biostimulant composition. The microbial strain and / or the biostimulant composition described herein can be capable of promoting a plant growth promoting property.

[0213] As another example, a bioreactor system can comprise: (a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property; (b) one or more additional containers arranged in a series that comprises thefirst container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and (c) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain at least a portion of the population of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of the plant not administered the biostimulant composition. In some embodiments, the microbial strain and / or the biostimulant composition described herein may be capable of increasing nutrient uptake of a plant.

[0214] The bioreactor system can be a continuous flow system. A continuous flow system can comprise a system wherein the flow of fluid is not interrupted. The stream of aqueous feedstock can be a continuous stream. In some embodiments, the bioreactor system can comprise a periodic flow, wherein there can be interruptions to the flow stream. In some embodiments, as a continuous flow bioreactor system, the volume of working fluid may be constant.

[0215] In some embodiments, the bioreactor system can comprise the clarifier container as described herein. The clarifier container can be configured to separate a portion of a working fluid in the clarifier container into a supernatant portion and a floc portion. In some embodiments, the product outflow stream of the system can comprise the supernatant portion.

[0216] In some embodiments, the product outflow stream comprises at least about 100 CFU / ml, at least about 1x103CFU / ml, at least about 1x104CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, at least about 1x107CFU / ml, at least about 1x108CFU / ml, at least about 1x109CFU / ml, at least about 1x1010CFU / ml, or greater than about 1x1010CFU / ml of the microbial strain. In some embodiments, the product outflow stream comprises at most about 1x1010CFU / ml, at most about 1x109CFU / ml, at most about 1x108CFU / ml, at most about 1x107CFU / ml, at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 1x104CFU / ml, at most about 1x103CFU / ml, at most about 100 CFU / ml, or less than about 100 CFU / ml of the microbial strain.

[0217] In some embodiments, the product outflow stream comprises at least about 100 CFU / ml, at least about 1x103CFU / ml, at least about 1x104CFU / ml, at least about 1x105CFU / ml, at least about 1x106CFU / ml, at least about 1x107CFU / ml, at least about 1x108CFU / ml, at least about 1x109CFU / ml, at least about 1x1010CFU / ml, or greater than about 1x1010CFU / ml of a sporulated form of the microbial strain. In some embodiments, the product outflow streamcomprises at most about 1x1010CFU / ml, at most about 1x109CFU / ml, at most about 1x108CFU / ml, at most about 1x107CFU / ml, at most about 1x106CFU / ml, at most about 1x105CFU / ml, at most about 1x104CFU / ml, at most about 1x103CFU / ml, at most about 100 CFU / ml, or less than about 100 CFU / ml of a sporulated form of the microbial strain.

[0218] In some embodiments, the product outflow stream may comprise a total dry weight. The total dry weight of the product outflow stream may be at least about 0.05 mg / ml, at least about 0.1 mg / ml, at least about 0.2 mg / ml, at least about 0.3 mg / ml, at least about 0.4 mg / ml, at least about 0.5 mg / ml, at least about 1.0 mg / ml, at least about 1.5 mg / ml, at least about 2.0 mg / ml, at least about 2.5 mg / ml, at least about 3.0 mg / ml, at least about 3.5 mg / ml, at least about 4.0 mg / ml, at least about 5.0 mg / ml, or greater than about 5.0 mg / ml. The total dry weight of the product outflow stream may be at most about 5.0 mg / ml, at most about 4.0 mg / ml, at most about 3.5 mg / ml, at most about 3.0 mg / ml, at most about 2.5 mg / ml, at most about 2.0 mg / ml, at most about 1.5 mg / ml, at most about 1.0 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, at most about 0.05 mg / ml, or less than about 0.05 mg / ml. The total dry weight of the product outflow stream may be between about 0.05 mg / ml to about 4 mg / ml. The total dry weight of the product outflow stream may be between about 0.05 mg / ml to about 0.1 mg / ml, about 0.05 mg / ml to about 0.2 mg / ml, about 0.05 mg / ml to about 0.3 mg / ml, about 0.05 mg / ml to about 0.4 mg / ml, about 0.05 mg / ml to about 0.5 mg / ml, about 0.05 mg / ml to about 1 mg / ml, about 0.05 mg / ml to about 1.5 mg / ml, about 0.05 mg / ml to about 2 mg / ml, about 0.05 mg / ml to about 2.5 mg / ml, about 0.05 mg / ml to about 3 mg / ml, about 0.05 mg / ml to about 4 mg / ml, about 0.1 mg / ml to about 0.2 mg / ml, about 0.1 mg / ml to about 0.3 mg / ml, about 0.1 mg / ml to about 0.4 mg / ml, about 0.1 mg / ml to about 0.5 mg / ml, about 0.1 mg / ml to about 1 mg / ml, about 0.1 mg / ml to about 1.5 mg / ml, about 0.1 mg / ml to about 2 mg / ml, about 0.1 mg / ml to about 2.5 mg / ml, about 0.1 mg / ml to about 3 mg / ml, about 0.1 mg / ml to about 4 mg / ml, about 0.2 mg / ml to about 0.3 mg / ml, about 0.2 mg / ml to about 0.4 mg / ml, about 0.2 mg / ml to about 0.5 mg / ml, about 0.2 mg / ml to about 1 mg / ml, about 0.2 mg / ml to about 1.5 mg / ml, about 0.2 mg / ml to about 2 mg / ml, about 0.2 mg / ml to about 2.5 mg / ml, about 0.2 mg / ml to about 3 mg / ml, about 0.2 mg / ml to about 4 mg / ml, about 0.3 mg / ml to about 0.4 mg / ml, about 0.3 mg / ml to about 0.5 mg / ml, about 0.3 mg / ml to about 1 mg / ml, about 0.3 mg / ml to about 1.5 mg / ml, about 0.3 mg / ml to about 2 mg / ml, about 0.3 mg / ml to about 2.5 mg / ml, about 0.3 mg / ml to about 3 mg / ml, about 0.3 mg / ml to about 4 mg / ml, about 0.4 mg / ml to about 0.5 mg / ml, about 0.4 mg / ml to about 1 mg / ml, about 0.4 mg / ml to about 1.5 mg / ml, about 0.4 mg / ml to about 2 mg / ml, about 0.4 mg / ml to about 2.5 mg / ml, about 0.4 mg / ml to about 3 mg / ml, about 0.4mg / ml to about 4 mg / ml, about 0.5 mg / ml to about 1 mg / ml, about 0.5 mg / ml to about 1.5 mg / ml, about 0.5 mg / ml to about 2 mg / ml, about 0.5 mg / ml to about 2.5 mg / ml, about 0.5 mg / ml to about 3 mg / ml, about 0.5 mg / ml to about 4 mg / ml, about 1 mg / ml to about 1.5 mg / ml, about 1 mg / ml to about 2 mg / ml, about 1 mg / ml to about 2.5 mg / ml, about 1 mg / ml to about 3 mg / ml, about 1 mg / ml to about 4 mg / ml, about 1.5 mg / ml to about 2 mg / ml, about 1.5 mg / ml to about 2.5 mg / ml, about 1.5 mg / ml to about 3 mg / ml, about 1.5 mg / ml to about 4 mg / ml, about 2 mg / ml to about 2.5 mg / ml, about 2 mg / ml to about 3 mg / ml, about 2 mg / ml to about 4 mg / ml, about 2.5 mg / ml to about 3 mg / ml, about 2.5 mg / ml to about 4 mg / ml, or about 3 mg / ml to about 4 mg / ml.

[0219] In some embodiments, the product outflow stream may comprise a chemical oxygen demand. The chemical oxygen demand of the product outflow stream may be at least about 10 mg / L, at least about 20 mg / L, at least about 30 mg / L, at least about 40 mg / L, at least about 50 mg / L, at least about 60 mg / L, at least about 70 mg / L, at least about 80 mg / L, at least about 90 mg / L, at least about 100 mg / L, at least about 250 mg / L, at least about 500 mg / L, at least about 750 mg / L, at least about 1000 mg / L, or greater than about 1000 mg / L. The chemical oxygen demand of the product outflow stream may be at most about 1000 mg / L, at most about 750 mg / L, at most about 500 mg / L, at most about 250 mg / L, at most about 100 mg / L, at most about 90 mg / L, at most about 80 mg / L, at most about 70 mg / L, at most about 60 mg / L, at most about 50 mg / L, at most about 40 mg / L, at most about 30 mg / L, at most about 20 mg / L, at most about 10 mg / L, or less than about 10 mg / L. The chemical oxygen demand of the product outflow stream may be between about 10 mg / L to about 1,000 mg / L. The chemical oxygen demand of the product outflow stream may be between about 10 mg / L to about 20 mg / L, about 10 mg / L to about 30 mg / L, about 10 mg / L to about 40 mg / L, about 10 mg / L to about 50 mg / L, about 10 mg / L to about 80 mg / L, about 10 mg / L to about 100 mg / L, about 10 mg / L to about 150 mg / L, about 10 mg / L to about 200 mg / L, about 10 mg / L to about 500 mg / L, about 10 mg / L to about 750 mg / L, about 10 mg / L to about 1,000 mg / L, about 20 mg / L to about 30 mg / L, about 20 mg / L to about 40 mg / L, about 20 mg / L to about 50 mg / L, about 20 mg / L to about 80 mg / L, about 20 mg / L to about 100 mg / L, about 20 mg / L to about 150 mg / L, about 20 mg / L to about 200 mg / L, about 20 mg / L to about 500 mg / L, about 20 mg / L to about 750 mg / L, about 20 mg / L to about 1,000 mg / L, about 30 mg / L to about 40 mg / L, about 30 mg / L to about 50 mg / L, about 30 mg / L to about 80 mg / L, about 30 mg / L to about 100 mg / L, about 30 mg / L to about 150 mg / L, about 30 mg / L to about 200 mg / L, about 30 mg / L to about 500 mg / L, about 30 mg / L to about 750 mg / L, about 30 mg / L to about 1,000 mg / L, about 40 mg / L to about 50 mg / L, about 40 mg / L to about 80 mg / L, about 40 mg / L to about 100 mg / L, about 40 mg / L to about 150 mg / L, about 40mg / L to about 200 mg / L, about 40 mg / L to about 500 mg / L, about 40 mg / L to about 750 mg / L, about 40 mg / L to about 1,000 mg / L, about 50 mg / L to about 80 mg / L, about 50 mg / L to about 100 mg / L, about 50 mg / L to about 150 mg / L, about 50 mg / L to about 200 mg / L, about 50 mg / L to about 500 mg / L, about 50 mg / L to about 750 mg / L, about 50 mg / L to about 1,000 mg / L, about 80 mg / L to about 100 mg / L, about 80 mg / L to about 150 mg / L, about 80 mg / L to about 200 mg / L, about 80 mg / L to about 500 mg / L, about 80 mg / L to about 750 mg / L, about 80 mg / L to about 1,000 mg / L, about 100 mg / L to about 150 mg / L, about 100 mg / L to about 200 mg / L, about 100 mg / L to about 500 mg / L, about 100 mg / L to about 750 mg / L, about 100 mg / L to about 1,000 mg / L, about 150 mg / L to about 200 mg / L, about 150 mg / L to about 500 mg / L, about 150 mg / L to about 750 mg / L, about 150 mg / L to about 1,000 mg / L, about 200 mg / L to about 500 mg / L, about 200 mg / L to about 750 mg / L, about 200 mg / L to about 1,000 mg / L, about 500 mg / L to about 750 mg / L, about 500 mg / L to about 1,000 mg / L, or about 750 mg / L to about 1,000 mg / L.

[0220] In some embodiments, the product outflow stream may comprise an electrical conductivity. The electrical conductivity of the product outflow stream may be at least about 0.01 mS / cm, at least about 0.05 mS / cm, at least about 0.1 mS / cm, at least about 0.5 mS / cm, at least about 1.0 mS / cm, at least about 1.5 mS / cm, at least about 2.0 mS / cm, at least about 2.5 mS / cm, at least about 3.0 mS / cm, at least about 4.0 mS / cm, at least about 5.0 mS / cm, at least about 10.0 mS / cm, or greater than about 10.0 mS / cm. The electrical conductivity of the product outflow stream may be at most about 10.0 mS / cm, at most about 5.0 mS / cm, at most about 4.0 mS / cm, at most about 3.0 mS / cm, at most about 2.5 mS / cm, at most about 2.0 mS / cm, at most about 1.5 mS / cm, at most about 1.0 mS / cm, at most about 0.5 mS / cm, at most about 0.1 mS / cm, at most about 0.05 mS / cm, at most about 0.01 mS / cm, or less than about 0.01 mS / cm. The electrical conductivity of the product outflow stream may be between about 0.01 mS / cm to about 10 mS / cm. The electrical conductivity of the product outflow stream may be between about 0.01 mS / cm to about 0.05 mS / cm, about 0.01 mS / cm to about 0.1 mS / cm, about 0.01 mS / cm to about 0.5 mS / cm, about 0.01 mS / cm to about 1 mS / cm, about 0.01 mS / cm to about 1.5 mS / cm, about 0.01 mS / cm to about 2 mS / cm, about 0.01 mS / cm to about 2.5 mS / cm, about 0.01 mS / cm to about 3 mS / cm, about 0.01 mS / cm to about 4 mS / cm, about 0.01 mS / cm to about 5 mS / cm, about 0.01 mS / cm to about 10 mS / cm, about 0.05 mS / cm to about 0.1 mS / cm, about 0.05 mS / cm to about 0.5 mS / cm, about 0.05 mS / cm to about 1 mS / cm, about 0.05 mS / cm to about 1.5 mS / cm, about 0.05 mS / cm to about 2 mS / cm, about 0.05 mS / cm to about 2.5 mS / cm, about 0.05 mS / cm to about 3 mS / cm, about 0.05 mS / cm to about 4 mS / cm, about 0.05 mS / cm to about 5 mS / cm, about 0.05 mS / cm to about 10 mS / cm, about 0.1 mS / cm to about 0.5 mS / cm,about 0.1 mS / cm to about 1 mS / cm, about 0.1 mS / cm to about 1.5 mS / cm, about 0.1 mS / cm to about 2 mS / cm, about 0.1 mS / cm to about 2.5 mS / cm, about 0.1 mS / cm to about 3 mS / cm, about 0.1 mS / cm to about 4 mS / cm, about 0.1 mS / cm to about 5 mS / cm, about 0.1 mS / cm to about 10 mS / cm, about 0.5 mS / cm to about 1 mS / cm, about 0.5 mS / cm to about 1.5 mS / cm, about 0.5 mS / cm to about 2 mS / cm, about 0.5 mS / cm to about 2.5 mS / cm, about 0.5 mS / cm to about 3 mS / cm, about 0.5 mS / cm to about 4 mS / cm, about 0.5 mS / cm to about 5 mS / cm, about 0.5 mS / cm to about 10 mS / cm, about 1 mS / cm to about 1.5 mS / cm, about 1 mS / cm to about 2 mS / cm, about 1 mS / cm to about 2.5 mS / cm, about 1 mS / cm to about 3 mS / cm, about 1 mS / cm to about 4 mS / cm, about 1 mS / cm to about 5 mS / cm, about 1 mS / cm to about 10 mS / cm, about1.5 mS / cm to about 2 mS / cm, about 1.5 mS / cm to about 2.5 mS / cm, about 1.5 mS / cm to about 3 mS / cm, about 1.5 mS / cm to about 4 mS / cm, about 1.5 mS / cm to about 5 mS / cm, about 1.5 mS / cm to about 10 mS / cm, about 2 mS / cm to about 2.5 mS / cm, about 2 mS / cm to about 3 mS / cm, about 2 mS / cm to about 4 mS / cm, about 2 mS / cm to about 5 mS / cm, about 2 mS / cm to about 10 mS / cm, about 2.5 mS / cm to about 3 mS / cm, about 2.5 mS / cm to about 4 mS / cm, about2.5 mS / cm to about 5 mS / cm, about 2.5 mS / cm to about 10 mS / cm, about 3 mS / cm to about 4 mS / cm, about 3 mS / cm to about 5 mS / cm, about 3 mS / cm to about 10 mS / cm, about 4 mS / cm to about 5 mS / cm, about 4 mS / cm to about 10 mS / cm, or about 5 mS / cm to about 10 mS / cm.

[0221] In some embodiments, reactors may be arranged so that fluid can flow from an outflow port (e.g., a product outflow port) of a reactor into an adjacent reactor or tank. Fluid from near the top of the working fluid in a reactor may flow into the next reactor continuously. Fluid from the middle of a reactor may flow into the next reactor continuously. Fluid from the bottom of a reactor may flow into the next reactor continuously. Fluid may also be reintroduced from any outflow source into the same reactor. In some embodiments, an outflow port (e.g., a product outflow port) is between 0.1 and 35 inches below the top of the working fluid within a reactor. In some embodiments, an outflow port (e.g., a product outflow port) is at least about 1 inch, at least about 2 inches, at least about 5 inches, at least about 10 inches, at least about 20 inches, at least about 50 inches, at least about 100 inches, at least about 250 inches, at least about 500 inches, at least about 750 inches, at least about 1,000 inches, at least about 2,500 inches, at least about 5,000 inches, at least about 7,500 inches, or at least about 10,000 inches below the top of the working fluid within a reactor. In some embodiments, an outflow port (e.g., a product outflow port) is at most about 10,000 inches, at most about 7,500 inches, at most about 5,000 inches, at most about 2,500 inches, at most about 1,000 inches, at most about 750 inches, at most about 500 inches, at most about 250 inches, at most about 100 inches, at most about 50 inches, at most about 20 inches, at most about 10 inches, at most about 5 inches, at most about 2 inches,or at most about 1 inch below the top of the working fluid within a reactor. In some embodiments, the rate of outflowing product from a bioreactor system may match the rate of inflowing feedstock, providing for a hydraulically balanced flow throughout the system. A reactor within the system may have a unique, stable microbial consortium with distinct physiological characteristics and digestion capabilities as compared to consortia in other tanks in the system. A reactor within the system may have the same microbial consortium with similar physiological characteristics and digestion capabilities as another reactor within the system. Each reactor within the system may have the same volume capacity. Each reactor within the system may have a different volume capacity. The bioreactor system may comprise at least two reactors. The bioreactor system may comprise 2 reactors, 3 reactors, 4 reactors, 5 reactors, 6 reactors, 7 reactors, 8 reactors, 9 reactors, 10 reactors, or more reactors. Reactors of a bioreactor system may be arranged as a serialized assembly of reactors. A serialized assembly of reactors may have conduits (e.g., ports or outputs) connecting each reactor to an adjacent reactor and / or container. A serialized assembly of reactors may have a continuous flow of working fluid through each reactor to the adjacent reactor.

[0222] In some embodiments, a reactor may have a single in-flow port and a single outflow port. In some embodiments, a reactor may have two or more in-flow ports and outflow ports. In some embodiments, a reactor may have a single in-flow port and two or more outflow ports. In some embodiments, a reactor may have two or more in-flow ports and a single outflow port. A reactor may have another in-flow port to provide a carbon source and / or consortium inoculum. An inflow port may be present at any location of a reactor of the bioreactor system. An in-flow port may be present at the top of the reactor or at the bottom of the reactor. An outflow port may be present at the top of the reactor or at the bottom of the reactor. In some embodiments, the outflow ports or in-flow ports described herein comprise one or more pipes, one or more pumps, one or more ventilations, or other systems for transferring fluid from one vessel to another.

[0223] A reactor may have a single fluid connection. A reactor may have two or more fluid connections. The fluid connections may be present at the top of the working fluid in each reactor or near the top of the working fluid in each reactor. In some embodiments, the reactor may have flow from the bottom of the container back to the top to prevent build-up of sludge in the bottom of the reactor. In some embodiments, a reactor may have one or more stirrers in the bottom of the container. In some embodiments, a reactor may have one or more wipers in the bottom of the container. The wipers may stir the feedstock and prevent clogging within the reactor. The wipers may fold floc and ease floc return. The wipers (e.g., folding wipers) can fold the floc portion of the clarifier working fluid in a bottom portion of the clarifier container.

[0224] In some embodiments, the reactors may comprise one or more packed bed reactors. In some embodiments, each of the packed bed reactors has an open cell design to allow free movement of working fluid. A fixed media (e.g., scaffolding) may be secured to the inside of each packed bed reactor. The fixed media comprises materials that increase the contact surface area for the communities of microbes with working fluid. The fixed media also provides a stable platform for anchoring biofilm. The packed bed reactors may be packed with scaffolding to increase surface area within the reactor. Without wishing to be bound by theory, the increased surface area provided by the scaffolding can provide increased contact for biofilm (e.g., microbes of a microbial consortium, an established microbial strain, or any combination thereof) to grow. The scaffolding within the reactor may increase biofilm. The packed bed reactor may improve contact between the biofilm and substrates within the reactor. The fixed media can be of several types, including durable plastic, polyvinyl chloride (PVC), metal, metal alloy, glass, glass compounds, fiberglass, any suitably robust inert material, or any combination thereof. The design and configuration of the fixed media can assume various geometric patterns that allow working fluid to freely move through each packed bed reactor and prevents fouling. In some embodiments, free flow supports can control hydraulic shearing which in time promotes even distribution of working fluid. In some embodiments, the fixed media is dispersed throughout a cross sectional area of each packed bed reactor.

[0225] The scaffolding may comprise one or more tubes, one or more rings, or other packing materials. In some embodiments, the packed bed reactors provided herein may comprise a bundle of tubes or columns. In some embodiments, the scaffolding may comprise hexagonal, grid-like, perforated tubing, or any combination thereof. Without wishing to be bound by theory, hexagonal, grid-like, and / or perforated scaffolding can increase the surface area and the flow through the columns within the container. In some embodiments, the tubes or columns of the scaffolding can comprise a diameter between 0.25 and 50 inches. In some embodiments, the scaffolding can comprise a diameter of at least about 0.20 inches, at least about 0.25 inches, at least about 0.3 inches, at least about 0.4 inches, at least about 0.5 inches, at least about 0.6 inches, at least about 0.7 inches, at least about 0.8 inches, at least about 0.9 inches, at least about 1 inch, at least about 2 inches, at least about 3 inches, at least about 4 inches, at least about 5 inches, at least about 10 inches, at least about 15 inches, at least about 20 inches, at least about 25 inches, at least about 30 inches, at least about 35 inches, at least about 40 inches, at least about 45 inches, at least about 50 inches, at least about 55 inches, at least about 60 inches, at least about 65 inches, at least about 70 inches, or at least about 75 inches. In some embodiments, the scaffolding can comprise a diameter of at most about 75 inches, at most about 70 inches, atmost about 65 inches, at most about 60 inches, at most about 55 inches, at most about 50 inches, at most about 45 inches, at most about 40 inches, at most about 35 inches, at most about 30 inches, at most about 25 inches, at most about 20 inches, at most about 15 inches, at most about 10 inches, at most about 5 inches, at most about 4 inches, at most about 3 inches, at most about 2 inches, at most about 1 inch, at most about 0.9 inches, at most about 0.8 inches, at most about 0.7 inches, at most about 0.6 inches, at most about 0.5 inches, at most about 0.4 inches, at most about 0.3 inches, at most about 0.25 inches, or at most about 0.2 inches. Without wishing to be bound by theory, a system with packed bed reactors may improve production of bacterial isolates or other microbes. In some embodiments, reactors without scaffolding (i.e., reactors that are not packed bed reactors) improve production of bacterial isolates or other microbes or improve digestion of digestible substrates.

[0226] In some embodiments, the reactors may comprise one or more fluidized bed reactors. In fluidized bed reactors, solid particles may be circulated within working fluid of the reactors, which may provide a surface for microbial colonization. Such particles may comprise, for example, particles of an inorganic substrate such as rock phosphate particles. In some embodiments, the fluidized bed reactors may be the same volume. In some embodiments, the fluidized bed reactors may be different volumes. In some embodiments, the fluidized bed reactors increase uniformity of particle mixing within the bioreactor system. The solid material of the fluidized bed reactor can have intrinsic fluid-like properties and allow for a more complete mixing. Reduction or elimination of radial and axial concentration gradients can provide for better fluid-solid contact and can achieve better uniformity of particle mixing. In some embodiments, the fluidized bed reactors increase the uniformity of temperature gradients within the bioreactor system. Without wishing to be bound by theory, the open container of the fluidized bed reactor can provide for a reduction in isolated hot or cold spots that may develop during mixing of a working fluid in the container, allowing for a uniform temperature distribution of the fluid. Hot spots can be areas of localized temperature changes during heating of a working fluid. Cold spots can be areas of localized temperature changes during cooling of a working fluid. The reduction in isolated temperature changes from the configuration of the fluidized bed reactor can provide for a more uniform temperature distribution of the fluid.

[0227] The flow rate of the bioreactor system may be chosen to allow for sufficient dwell time of a working fluid within each of the reactors for a stable and unique microbial consortium to form within each of the reactors. In some embodiments, working fluid in each reactor is continuously recycled at a rate ratio in a range of approximately 25: 1 to 35: 1, 25 to 35 gallons per minute of the recycle rate to one gallon per minute of the hydraulic feed rate. In someembodiments, working fluid in each reactor is continuously recycled at a rate ratio of at least about 10: 1, at least about 12: 1, at least about 14: 1, at least about 16: 1, at least about 18: 1, at least about 20: 1, at least about 22: 1, at least about 24: 1, at least about 26: 1, at least about 28: 1, at least about 30: 1, at least about 32: 1, at least about 34: 1, at least about 36: 1, at least about 38: 1, at least about 40: 1, at least about 45: 1, or at least about 50: 1. In some embodiments, working fluid in each reactor is continuously recycled at a rate ratio of at most about 50: 1 at most about 45:1, at most about 40: 1, at most about 38: 1, at most about 36: 1, at most about 34: 1, at most about 32: 1, at most about 30: 1, at most about 28: 1, at most about 26: 1, at most about 24: 1, at most about 22: 1, at most about 20: 1, at most about 18: 1, at most about 16: 1, at most about 14: 1, at most about 12: 1, or at most about 10: 1. Working fluid may be recycled by one or more pumps to prevent solids settling and to provide sufficient velocity and hydraulic shear to prevent excessive buildup and sloughing of biofilm. A digestion system provided herein may comprise a first flow rate, second flow rate, third flow rate, fourth flow rate, fifth flow rate, sixth flow rate, or seventh flow rate.

[0228] Reactors can be maintained at specific temperatures which may aid in digestion and growth of microbial consortia within the system. In some embodiments, a temperature of a reactor is at least about 15°C, at least about 20°C, at least about 21 °C, at least about 22°C, at least about 23°C, at least about 24°C, at least about 25°C, at least about 26°C, at least about 27°C, at least about 28°C, at least about 29°C, at least about 30°C, at least about 35°C, at least about 40°C, at least about 45°C, or at least about 50°C. In some embodiments, a temperature of a reactor is at most about 50°C, at most about 45°C, at most about 40°C, at most about 35°C, at most about 30°C, at most about 29°C, at most about 28°C, at most about 27°C, at most about 26°C, at most about 25°C, at most about 24°C, at most about 23°C, at most about 22°C, at most about 21 °C, at most about 20°C, or at most about 15°C.

[0229] In some embodiments, a temperature of a reactor is about 15°C to about 45°C. In some embodiments, a temperature of a reactor is about 15°C to about 20°C, about 15°C to about 22°C, about 15°C to about 24°C, about 15°C to about 26°C, about 15°C to about 28°C, about 15°C to about 30°C, about 15°C to about 32°C, about 15°C to about 34°C, about 15°C to about 36°C, about 15°C to about 40°C, about 15°C to about 45°C, about 20°C to about 22°C, about 20°C to about 24°C, about 20°C to about 26°C, about 20°C to about 28°C, about 20°C to about 30°C, about 20°C to about 32°C, about 20°C to about 34°C, about 20°C to about 36°C, about 20°C to about 40°C, about 20°C to about 45°C, about 22°C to about 24°C, about 22°C to about 26°C, about 22°C to about 28°C, about 22°C to about 30°C, about 22°C to about 32°C, about 22°C to about 34°C, about 22°C to about 36°C, about 22°C to about 40°C, about 22°C to about 45°C,about 24°C to about 26°C, about 24°C to about 28°C, about 24°C to about 30°C, about 24°C to about 32°C, about 24°C to about 34°C, about 24°C to about 36°C, about 24°C to about 40°C, about 24°C to about 45°C, about 26°C to about 28°C, about 26°C to about 30°C, about 26°C to about 32°C, about 26°C to about 34°C, about 26°C to about 36°C, about 26°C to about 40°C, about 26°C to about 45°C, about 28°C to about 30°C, about 28°C to about 32°C, about 28°C to about 34°C, about 28°C to about 36°C, about 28°C to about 40°C, about 28°C to about 45°C, about 30°C to about 32°C, about 30°C to about 34°C, about 30°C to about 36°C, about 30°C to about 40°C, about 30°C to about 45°C, about 32°C to about 34°C, about 32°C to about 36°C, about 32°C to about 40°C, about 32°C to about 45°C, about 34°C to about 36°C, about 34°C to about 40°C, about 34°C to about 45°C, about 36°C to about 40°C, about 36°C to about 45°C, or about 40°C to about 45°C.

[0230] In some embodiments, the high aeration of the working fluid of a bioreactor system described herein can help enrich microbes with a targeted functionality (e.g., phosphate solubilizers, zinc solubilizers, nitrogen use efficiency). Reactors may be maintained under aerobic, microaerobic, or anaerobic conditions. The series of reactors in a bioreactor system may have different aerobic conditions. The series of reactors in a bioreactor system may have the same aerobic conditions. In some embodiments, a reactor may have the same aerobic condition as an adjacent reactor. In some embodiments, a reactor may have a different aerobic condition than an adjacent reactor. In some embodiments, a bioreactor system may have aerobic, microaerobic, anaerobic conditions, or any combination thereof.

[0231] In some embodiments, aerobic conditions comprise conditions with a dissolved oxygen measurement of greater than 2 mg / L. In some embodiments, aerobic conditions comprise conditions with a dissolved oxygen measurement of at least about 2 mg / L mg / L, at least about 3 mg / L mg / L, at least about 4 mg / L, at least about 5 mg / L, at least about 6 mg / L, at least about 7 mg / L, at least about 8 mg / L, at least about 9 mg / L, at least about 10 mg / L, at least about 12 mg / L, at least about 14 mg / L, at least about 15 mg / L, or greater than about 15 mg / L. In some embodiments, aerobic conditions comprise conditions with a dissolved oxygen measurement from about 2 mg / L to about 15 mg / L. In some embodiments, aerobic conditions comprise conditions with a dissolved oxygen measurement from about 2 mg / L to about 3 mg / L, about 2 mg / L to about 4 mg / L, about 2 mg / L to about 5 mg / L, about 2 mg / L to about 6 mg / L, about 2 mg / L to about 7 mg / L, about 2 mg / L to about 8 mg / L, about 2 mg / L to about 9 mg / L, about 2 mg / L to about 10 mg / L, about 2 mg / L to about 12 mg / L, about 2 mg / L to about 14 mg / L, about 2 mg / L to about 15 mg / L, about 3 mg / L to about 4 mg / L, about 3 mg / L to about 5 mg / L, about 3 mg / L to about 6 mg / L, about 3 mg / L to about 7 mg / L, about 3 mg / L to about 8 mg / L, about 3mg / L to about 9 mg / L, about 3 mg / L to about 10 mg / L, about 3 mg / L to about 12 mg / L, about 3 mg / L to about 14 mg / L, about 3 mg / L to about 15 mg / L, about 4 mg / L to about 5 mg / L, about 4 mg / L to about 6 mg / L, about 4 mg / L to about 7 mg / L, about 4 mg / L to about 8 mg / L, about 4 mg / L to about 9 mg / L, about 4 mg / L to about 10 mg / L, about 4 mg / L to about 12 mg / L, about 4 mg / L to about 14 mg / L, about 4 mg / L to about 15 mg / L, about 5 mg / L to about 6 mg / L, about 5 mg / L to about 7 mg / L, about 5 mg / L to about 8 mg / L, about 5 mg / L to about 9 mg / L, about 5 mg / L to about 10 mg / L, about 5 mg / L to about 12 mg / L, about 5 mg / L to about 14 mg / L, about 5 mg / L to about 15 mg / L, about 6 mg / L to about 7 mg / L, about 6 mg / L to about 8 mg / L, about 6 mg / L to about 9 mg / L, about 6 mg / L to about 10 mg / L, about 6 mg / L to about 12 mg / L, about 6 mg / L to about 14 mg / L, about 6 mg / L to about 15 mg / L, about 7 mg / L to about 8 mg / L, about 7 mg / L to about 9 mg / L, about 7 mg / L to about 10 mg / L, about 7 mg / L to about 12 mg / L, about 7 mg / L to about 14 mg / L, about 7 mg / L to about 15 mg / L, about 8 mg / L to about 9 mg / L, about 8 mg / L to about 10 mg / L, about 8 mg / L to about 12 mg / L, about 8 mg / L to about 14 mg / L, about 8 mg / L to about 15 mg / L, about 9 mg / L to about 10 mg / L, about 9 mg / L to about 12 mg / L, about 9 mg / L to about 14 mg / L, about 9 mg / L to about 15 mg / L, about 10 mg / L to about 12 mg / L, about 10 mg / L to about 14 mg / L, about 10 mg / L to about 15 mg / L, about 12 mg / L to about 14 mg / L, about 12 mg / L to about 15 mg / L, or about 14 mg / L to about 15 mg / L. In some embodiments, aerobic conditions comprise conditions with a dissolved oxygen measurement of between 2 mg / L and 10 mg / L. In some embodiments, microaerobic conditions comprise conditions with a dissolved oxygen measurement of less than 2 mg / L. In some embodiments, microaerobic conditions comprise conditions with a dissolved oxygen measurement of at most about 1.99 mg / L, at most about 1.8 mg / L, at most about 1.6 mg / L, at most about 1.5 mg / L, at most about 1.4 mg / L, at most about 1.3 mg / L, at most about 1.2 mg / L, at most about 1.1 mg / L, at most about 1 mg / L, at most about 0.9 mg / L, at most about 0.8 mg / L, at most about 0.7 mg / L, at most about 0.6 mg / L, at most about 0.5 mg / L, at most about 0.4 mg / L, at most about 0.3 mg / L, at most about 0.2 mg / L, at most about 0.1 mg / L, or less than about 0.1 mg / L but not 0 mg / L. In some embodiments, microaerobic conditions comprise conditions with a dissolved oxygen measurement from about 0.1 mg / L to about 1.99 mg / L. In some embodiments, microaerobic conditions comprise conditions with a dissolved oxygen measurement from about 0.1 mg / L to about 0.2 mg / L, about 0.1 mg / L to about 0.3 mg / L, about 0.1 mg / L to about 0.4 mg / L, about 0.1 mg / L to about 0.5 mg / L, about 0.1 mg / L to about 0.8 mg / L, about 0.1 mg / L to about 1 mg / L, about 0.1 mg / L to about 1.2 mg / L, about 0.1 mg / L to about 1.4 mg / L, about 0.1 mg / L to about 1.6 mg / L, about 0.1 mg / L to about 1.8 mg / L, about 0.1 mg / L to about 1.99 mg / L, about 0.2 mg / L to about 0.3 mg / L, about 0.2 mg / L to about 0.4 mg / L, about 0.2 mg / L to about0.5 mg / L, about 0.2 mg / L to about 0.8 mg / L, about 0.2 mg / L to about 1 mg / L, about 0.2 mg / L to about 1.2 mg / L, about 0.2 mg / L to about 1.4 mg / L, about 0.2 mg / L to about 1.6 mg / L, about 0.2 mg / L to about 1.8 mg / L, about 0.2 mg / L to about 1.99 mg / L, about 0.3 mg / L to about 0.4 mg / L, about 0.3 mg / L to about 0.5 mg / L, about 0.3 mg / L to about 0.8 mg / L, about 0.3 mg / L to about 1 mg / L, about 0.3 mg / L to about 1.2 mg / L, about 0.3 mg / L to about 1.4 mg / L, about 0.3 mg / L to about 1.6 mg / L, about 0.3 mg / L to about 1.8 mg / L, about 0.3 mg / L to about 1.99 mg / L, about 0.4 mg / L to about 0.5 mg / L, about 0.4 mg / L to about 0.8 mg / L, about 0.4 mg / L to about 1 mg / L, about 0.4 mg / L to about 1.2 mg / L, about 0.4 mg / L to about 1.4 mg / L, about 0.4 mg / L to about 1.6 mg / L, about 0.4 mg / L to about 1.8 mg / L, about 0.4 mg / L to about 1.99 mg / L, about 0.5 mg / L to about 0.8 mg / L, about 0.5 mg / L to about 1 mg / L, about 0.5 mg / L to about 1.2 mg / L, about 0.5 mg / L to about 1.4 mg / L, about 0.5 mg / L to about 1.6 mg / L, about 0.5 mg / L to about 1.8 mg / L, about 0.5 mg / L to about 1.99 mg / L, about 0.8 mg / L to about 1 mg / L, about 0.8 mg / L to about 1.2 mg / L, about 0.8 mg / L to about 1.4 mg / L, about 0.8 mg / L to about 1.6 mg / L, about 0.8 mg / L to about 1.8 mg / L, about 0.8 mg / L to about 1.99 mg / L, about 1 mg / L to about 1.2 mg / L, about 1 mg / L to about 1.4 mg / L, about 1 mg / L to about 1.6 mg / L, about 1 mg / L to about 1.8 mg / L, about 1 mg / L to about 1.99 mg / L, about 1.2 mg / L to about 1.4 mg / L, about 1.2 mg / L to about 1.6 mg / L, about 1.2 mg / L to about 1.8 mg / L, about 1.2 mg / L to about 1.99 mg / L, about 1.4 mg / L to about 1.6 mg / L, about 1.4 mg / L to about 1.8 mg / L, about 1.4 mg / L to about 1.99 mg / L, about 1.6 mg / L to about 1.8 mg / L, about 1.6 mg / L to about 1.99 mg / L, or about 1.8 mg / L to about 1.99 mg / L. In some embodiments, anaerobic conditions comprise conditions with a dissolved oxygen measurement of 0 mg / L.

[0232] In some embodiments, the reactors comprise high-speed mixers to move a working fluid and increase aeration. The mixers may facilitate an aerobic digestion and enrich microbial communities with a target functionality (e.g., phosphate solubilizing microbes). The high-speed mixers can use a speed comprising at least about 1000 RPM, 1100 RPM, 1200 RPM, 1300 RPM, 1400 RPM, 1500 RPM, 1600 RPM, 1700 RPM, 1800 RPM, 1900 RPM, 2000 RPM, or greater than about 2000 RPM.

[0233] Reactors of a bioreactor system described herein may comprise a working volume used to hold a volume of working fluid. A working volume of a reactor of a bioreactor system described herein may be at least about 5 gallons, at least about 10 gallons, at least about 20 gallons, at least about 50 gallons, at least about 75 gallons, at least about 100 gallons, at least about 250 gallons, at least about 500 gallons, at least about 750 gallons, at least about 1,000 gallons, at least about 2,000 gallons, at least about 3,000 gallons, at least about 4,000 gallons, at least about 5,000 gallons, at least about 7,500 gallons, at least about 10,000 gallons, at leastabout 15,000 gallons, at least about 20,000 gallons, at least about 50,000 gallons, or more than about 50,000 gallons. A working volume of a reactor of a bioreactor system described herein may be at most about 50,000 gallons, at most about 20,000 gallons, at most about 15,000 gallons, at most about 10,000 gallons, at most about 7,500 gallons, at most about 5,000 gallons, at most about 4,000 gallons, at most about 3,000 gallons, at most about 2,000 gallons, at most about 1,000 gallons, at most about 750 gallons, at most about 500 gallons, at most about 250 gallons, at most about 100 gallons, at most about 75 gallons, at most about 50 gallons, at most about 20 gallons, at most about 10 gallons, at most about 5 gallons, or less than about 5 gallons.

[0234] Reactors may be maintained at different pH levels within a bioreactor system. Reactors may be maintained at the same pH levels within a bioreactor system. The pH of a reactor in a bioreactor system may be at least about 4.0, at least about 4.5, at least about 5.0, at least about 5.5, at least about 6.0, at least about 6.5, at least about 7.0, at least about 7.5, at least about 8.0, at least about 8.5, at least about 9.0, at least about 9.5, or at least about 10.0. In some embodiments, the pH of a reactor in a bioreactor system may be at most about 10.0, at most about 9.5, at most about 9.0, at most about 8.5, at most about 8.0, at most about 7.5, at most about 7.0, at most about 6.5, at most about 6.0, at most about 5.5, at most about 5.0, at most about 4.5, or at most about 4.0.

[0235] In some embodiments, the pH of a reactor in a bioreactor system may be about 3 to about 9. In some embodiments, the pH of a reactor in a bioreactor system may be about 3 to about 3.5, about 3 to about 4, about 3 to about 4.5, about 3 to about 5, about 3 to about 5.5, about 3 to about 6, about 3 to about 6.5, about 3 to about 7, about 3 to about 7.5, about 3 to about 8, about 3 to about 9, about 3.5 to about 4, about 3.5 to about 4.5, about 3.5 to about 5, about 3.5 to about 5.5, about 3.5 to about 6, about 3.5 to about 6.5, about 3.5 to about 7, about 3.5 to about 7.5, about 3.5 to about 8, about 3.5 to about 9, about 4 to about 4.5, about 4 to about 5, about 4 to about 5.5, about 4 to about 6, about 4 to about 6.5, about 4 to about 7, about 4 to about 7.5, about 4 to about 8, about 4 to about 9, about 4.5 to about 5, about 4.5 to about 5.5, about 4.5 to about 6, about 4.5 to about 6.5, about 4.5 to about 7, about 4.5 to about 7.5, about 4.5 to about 8, about 4.5 to about 9, about 5 to about 5.5, about 5 to about 6, about 5 to about 6.5, about 5 to about 7, about 5 to about 7.5, about 5 to about 8, about 5 to about 9, about 5.5 to about 6, about 5.5 to about 6.5, about 5.5 to about 7, about 5.5 to about 7.5, about 5.5 to about 8, about 5.5 to about 9, about 6 to about 6.5, about 6 to about 7, about 6 to about 7.5, about 6 to about 8, about 6 to about 9, about 6.5 to about 7, about 6.5 to about 7.5, about 6.5 to about 8, about 6.5 to about 9, about 7 to about 7.5, about 7 to about 8, about 7 to about 9, about 7.5 to about 8, about 7.5 to about 9, or about 8 to about 9.

[0236] In some embodiments, the reactors may comprise distribution components (e.g., a distribution ring) that is subsurface of the discharge volume. The distribution component can reduce the amount of surface disruption, and / or keep the environment in the reactor anaerobic. The environment of the reactor may be anaerobic, microaerobic, or aerobic. Without wishing to be bound by theory, the microaerobic conditions of the reactors of the bioreactor system may enrich certain microbes (e.g., zinc-solubilization microbes, phosphate-solubilization microbes, and / or microbes with nitrogen use efficiency capability) in the microbial consortium of the system.

[0237] A working fluid may comprise a fluidic substance that moves through a bioreactor system as described herein. A working fluid can comprise solid components, liquid components, gaseous components, or any combination thereof. A working fluid can comprise microbial consortia, isolated microbes or inoculum of a microbial strain (e.g., target isolates), additional organic and / or materials, or any combination thereof. The mixture of microbial consortia, isolated microbes (e.g., target isolates), additional organic and / or materials within a working fluid may allow for the expansion of microbes or act as a culture for an inoculum of a microbe to grow. A working fluid may comprise a pH, viscosity, temperature, surface tension, adhesion, and / or volume that enhances the growth and / or functioning of microbes or microorganisms. In some embodiments, a volume of working fluid within each reactor may be continuously replenished and drawn from. In some embodiments, a volume of working fluid within each reactor is replenished and drawn from in batches (e.g., discontinuously). In some embodiments, a working fluid in a first reactor may comprise a first working fluid. In some embodiments, a working fluid in a second reactor may comprise a second working fluid. In some embodiments, a working fluid in a third reactor may comprise a third working fluid. In some embodiments, a working fluid in a fourth reactor may comprise a fourth working fluid. In some embodiments, a working fluid in a fifth reactor may comprise a fifth working fluid. In some embodiments, at least one of the first working fluid, the second working fluid, or the third working fluid comprises a pH buffering system. A method described herein can further comprise maintaining the pH of at least one of the first working fluid, the second working fluid, the third working fluid, the fourth working fluid, or the fifth working fluid at at least about 4.0, at least about 4.5, at least about 5.0, at least about 5.5, at least about 6.0, at least about 6.5, at least about 7.0, at least about 7.5, at least about 8.0, at least about 8.5, at least about 9.0, at least about 9.5, or at least about 10.0. A method described herein can further comprise maintaining the pH of at least one of the first working fluid, the second working fluid, the third working fluid, the fourth working fluid, or the fifth working fluid at at most about 10.0, at most about 9.5, at most about9.0, at most about 8.5, at most about 8.0, at most about 7.5, at most about 7.0, at most about 6.5, at most about 6.0, at most about 5.5, at most about 5.0, at most about 4.5, or at most about 4.0.In some embodiments, the methods described herein can comprise maintaining the pH of at least one of the first working fluid, the second working fluid, the third working fluid, the fourth working fluid, or the fifth working fluid between about 3 to about 10. In some embodiments, the methods described herein can comprise maintaining the pH of at least one of the first working fluid, the second working fluid, the third working fluid, the fourth working fluid, or the fifth working fluid between about 3 to about 4, about 3 to about 4.5, about 3 to about 5, about 3 to about 6, about 3 to about 6.5, about 3 to about 7, about 3 to about 7.5, about 3 to about 8, about 3 to about 8.5, about 3 to about 9, about 3 to about 10, about 4 to about 4.5, about 4 to about 5, about 4 to about 6, about 4 to about 6.5, about 4 to about 7, about 4 to about 7.5, about 4 to about 8, about 4 to about 8.5, about 4 to about 9, about 4 to about 10, about 4.5 to about 5, about4.5 to about 6, about 4.5 to about 6.5, about 4.5 to about 7, about 4.5 to about 7.5, about 4.5 to about 8, about 4.5 to about 8.5, about 4.5 to about 9, about 4.5 to about 10, about 5 to about 6, about 5 to about 6.5, about 5 to about 7, about 5 to about 7.5, about 5 to about 8, about 5 to about 8.5, about 5 to about 9, about 5 to about 10, about 6 to about 6.5, about 6 to about 7, about 6 to about 7.5, about 6 to about 8, about 6 to about 8.5, about 6 to about 9, about 6 to about 10, about 6.5 to about 7, about 6.5 to about 7.5, about 6.5 to about 8, about 6.5 to about 8.5, about6.5 to about 9, about 6.5 to about 10, about 7 to about 7.5, about 7 to about 8, about 7 to about 8.5, about 7 to about 9, about 7 to about 10, about 7.5 to about 8, about 7.5 to about 8.5, about7.5 to about 9, about 7.5 to about 10, about 8 to about 8.5, about 8 to about 9, about 8 to about 10, about 8.5 to about 9, about 8.5 to about 10, or about 9 to about 10.

[0238] In some embodiments, a reactor (e.g., container) can comprise a temperature. The temperature of the reactor can provide conditions suitable for growth and / or survival of a microbial strain. In some embodiments, one or more microbes described herein can be psychrophiles, mesophiles, thermophiles, or any combination thereof. In some embodiments, one or more reactors can comprise mesophilic temperature conditions. In some embodiments, one or more reactors (e.g., containers) of a bioreactor system described herein may comprise a temperature of at least about 20°C, at least about 25°C, at least about 30°C, at least about 31°C, at least about 32°C, at least about 33°C, at least about 34°C, at least about 35°C, at least about 36°C, at least about 37°C, at least about 38°C, at least about 39°C, at least about 40°C, at least about 45°C, at least about 50°C, or greater than about 50°C. In some embodiments, the temperature can be at most about 45°C. In some embodiments, the temperature can be maintained in a mesophilic range (e.g., less than 45°C). In some embodiments, the temperaturemay not be thermophilic (e.g., greater than 45 °C). In some embodiments, thermophilic conditions are avoided in one or more containers of a bioreactor system described herein. In some embodiments, one or more reactors (e.g., containers) of a bioreactor system described herein may comprise a temperature of at most about 50°C, at most about 45°C, at most about 40°C, at most about 39°C, at most about 38°C, at most about 37°C, at most about 36°C, at most about 35°C, at most about 34°C, at most about 33°C, at most about 32°C, at most about 31°C, at most about 30°C, at most about 25°C, at most about 20°C, or less than about 20°C. In some embodiments, one or more reactors (e.g., containers) of a bioreactor system described herein may comprise a temperature between about 25 °C to about 40 °C. In some embodiments, one or more reactors (e.g., containers) of a bioreactor system described herein may comprise a temperature between about 25 °C to about 30 °C, about 25 °C to about 31 °C, about 25 °C to about 32 °C, about 25 °C to about 32 °C, about 25 °C to about 34 °C, about 25 °C to about 35 °C, about 25 °C to about 36 °C, about 25 °C to about 37 °C, about 25 °C to about 38 °C, about 25 °C to about 39 °C, about 25 °C to about 40 °C, about 30 °C to about 31 °C, about30 °C to about 32 °C, about 30 °C to about 32 °C, about 30 °C to about 34 °C, about 30 °C to about 35 °C, about 30 °C to about 36 °C, about 30 °C to about 37 °C, about 30 °C to about 38 °C, about 30 °C to about 39 °C, about 30 °C to about 40 °C, about 31 °C to about 32 °C, about 31 °C to about 32 °C, about 31 °C to about 34 °C, about 31 °C to about 35 °C, about31 °C to about 36 °C, about 31 °C to about 37 °C, about 31 °C to about 38 °C, about 31 °C to about 39 °C, about 31 °C to about 40 °C, about 32 °C to about 32 °C, about 32 °C to about34 °C, about 32 °C to about 35 °C, about 32 °C to about 36 °C, about 32 °C to about 37 °C, about 32 °C to about 38 °C, about 32 °C to about 39 °C, about 32 °C to about 40 °C, about32 °C to about 34 °C, about 32 °C to about 35 °C, about 32 °C to about 36 °C, about 32 °C to about 37 °C, about 32 °C to about 38 °C, about 32 °C to about 39 °C, about 32 °C to about 40 °C, about 34 °C to about 35 °C, about 34 °C to about 36 °C, about 34 °C to about 37 °C, about 34 °C to about 38 °C, about 34 °C to about 39 °C, about 34 °C to about 40 °C, about35 °C to about 36 °C, about 35 °C to about 37 °C, about 35 °C to about 38 °C, about 35 °C to about 39 °C, about 35 °C to about 40 °C, about 36 °C to about 37 °C, about 36 °C to about 38 °C, about 36 °C to about 39 °C, about 36 °C to about 40 °C, about 37 °C to about 38 °C, about 37 °C to about 39 °C, about 37 °C to about 40 °C, about 38 °C to about 39 °C, about 38 °C to about 40 °C, or about 39 °C to about 40 °C. In some embodiments, no reactor of the bioreactor system comprises thermophilic conditions (e.g., a temperature above about 50°C or between 55 °C and 60 °C).

[0239] In continuous operation, each reactor may have a constant volume of working fluid therein, with the rate of fluid flowing into each reactor matching the rate of fluid flowing out of each reactor. As each reactor may comprise a different microbial consortium and have different conditions from other reactors, the working fluid within each reactor may be considered to be distinct from working fluids within the other reactors. The total volume of working fluid within a bioreactor system may be referred to herein as the “total working volume” of the bioreactor system.

[0240] A working fluid may be maintained within a reactor 110. A maintained working fluid may be a working fluid as it rests in a reactor 110 of a bioreactor system. Without wishing to be bound by theory, maintenance of a working fluid may support enrichment and / or growth of a population of the microbial strain or another component of a working fluid as described herein. In some embodiments, the volume of working fluid within the reactor 110 may be maintained at 5 gallons. In some embodiments, the volume of working fluid within the reactor may be maintained at least about 1 gallon, at least about 2 gallons, at least about 3 gallons, at least about 4 gallons, at least about 5 gallons, at least about 6 gallons, at least about 7 gallons, at least about 10 gallons, at least about 15 gallons, at least about 20 gallons, at least about 25 gallons, at least about 30 gallons, at least about 50 gallons, at least about 75 gallons, at least about 100 gallons, at least about 250 gallons, at least about 500 gallons, at least about 1,000 gallons, at least about 2,500 gallons, at least about 5,000 gallons, at least about 10,000 gallons, at least about 12,000 gallons, at least about 15,000 gallons, or more than about 15,000 gallons. In some embodiments, the volume of working fluid within the reactor may be maintained at most about 15,000 gallons, at most about 12,000 gallons, at most about 10,000 gallons, at most about 5,000 gallons, at most about 2,500 gallons, at most about 1,000 gallons, at most about 500 gallons, at most about 250 gallons, at most about 100 gallons, at most about 75 gallons, at most about 50 gallons, at most about 30 gallons, at most about 25 gallons, at most about 20 gallons, at most about 15 gallons, at most about 10 gallons, at most about 7 gallons, at most about 6 gallons, at most about 5 gallons, at most about 4 gallons, at most about 3 gallons, at most about 2 gallons, at most about 1 gallon, or less than about 1 gallon.

[0241] In some embodiments, the volume of working fluid within the reactor may be maintained from about 5 gallons to about 10,000 gallons. In some embodiments, the volume of working fluid within the reactor may be maintained from about 5 gallons to about 10 gallons, about 5 gallons to about 25 gallons, about 5 gallons to about 50 gallons, about 5 gallons to about 75 gallons, about 5 gallons to about 100 gallons, about 5 gallons to about 150 gallons, about 5 gallons to about 500 gallons, about 5 gallons to about 1,000 gallons, about 5 gallons to about2,500 gallons, about 5 gallons to about 5,000 gallons, about 5 gallons to about 10,000 gallons, about 10 gallons to about 25 gallons, about 10 gallons to about 50 gallons, about 10 gallons to about 75 gallons, about 10 gallons to about 100 gallons, about 10 gallons to about 150 gallons, about 10 gallons to about 500 gallons, about 10 gallons to about 1,000 gallons, about 10 gallons to about 2,500 gallons, about 10 gallons to about 5,000 gallons, about 10 gallons to about 10,000 gallons, about 25 gallons to about 50 gallons, about 25 gallons to about 75 gallons, about 25 gallons to about 100 gallons, about 25 gallons to about 150 gallons, about 25 gallons to about 500 gallons, about 25 gallons to about 1,000 gallons, about 25 gallons to about 2,500 gallons, about 25 gallons to about 5,000 gallons, about 25 gallons to about 10,000 gallons, about 50 gallons to about 75 gallons, about 50 gallons to about 100 gallons, about 50 gallons to about 150 gallons, about 50 gallons to about 500 gallons, about 50 gallons to about 1,000 gallons, about 50 gallons to about 2,500 gallons, about 50 gallons to about 5,000 gallons, about 50 gallons to about 10,000 gallons, about 75 gallons to about 100 gallons, about 75 gallons to about 150 gallons, about 75 gallons to about 500 gallons, about 75 gallons to about 1,000 gallons, about 75 gallons to about 2,500 gallons, about 75 gallons to about 5,000 gallons, about 75 gallons to about 10,000 gallons, about 100 gallons to about 150 gallons, about 100 gallons to about 500 gallons, about 100 gallons to about 1,000 gallons, about 100 gallons to about 2,500 gallons, about 100 gallons to about 5,000 gallons, about 100 gallons to about 10,000 gallons, about 150 gallons to about 500 gallons, about 150 gallons to about 1,000 gallons, about 150 gallons to about 2,500 gallons, about 150 gallons to about 5,000 gallons, about 150 gallons to about 10,000 gallons, about 500 gallons to about 1,000 gallons, about 500 gallons to about 2,500 gallons, about 500 gallons to about 5,000 gallons, about 500 gallons to about 10,000 gallons, about 1,000 gallons to about 2,500 gallons, about 1,000 gallons to about 5,000 gallons, about 1,000 gallons to about 10,000 gallons, about 2,500 gallons to about 5,000 gallons, about 2,500 gallons to about 10,000 gallons, or about 5,000 gallons to about 10,000 gallons.

[0242] A first container may comprise an outlet port. The outlet port can be fluidly connected to an inlet port of another container of the bioreactor system (e.g., a second container). The second container may comprise an outlet port. The outlet port of the second container can be fluidly connected to an inlet port of another container of the bioreactor system (e.g., a third container). In some embodiments, an inlet port of a container (e.g., a first container) may be at a higher level of the container than an outlet port of the container. A container (e.g., a third container) can comprise an outlet port fluidly connected to a clarifier container. The outlet port of a container may be fluidly connected to an input port of the clarifier container. A method described herein may further comprise maintaining the volume of one or more working fluids for the duration oftime (e.g., for the duration of time of operating the bioreactor system. For example, the method may further comprise maintaining the volume of a first working fluid, a second working fluid, a third working fluid, a fourth working fluid, a fifth working fluid, or any combination thereof throughout the duration of time.

[0243] In some embodiments, the working fluid in the reactor 110 may be recirculated (e.g., recycled) within the reactor of the bioreactor system 100. Working fluid may be recirculated via a conduit or output of a reactor and reintroduced into the same reactor. Without wishing to be bound by theory, recirculation of a working fluid within a reactor may support enrichment and / or growth of a population of the microbial strain or another component of a working fluid as described herein. In some embodiments, the rate of recirculation within the reactor may be between 4-9 gallons / min. In some embodiments, the rate of recirculation within the reactor 110 may be at least about 0.5 gallon / min, at least about 1 gallon / min, at least about 2 gallons / min, at least about 3 gallons / min, at least about 3.5 gallons / min, at least about 4 gallons / min, at least about 4.5 gallons / min, at least about 5 gallons / min, at least about 5.5 gallons / min, at least about 6 gallons / min, at least about 6.5 gallons / min, at least about 7 gallons / min, at least about 7.5 gallons / min, at least about 8 gallons / min, at least about 8.5 gallons / min, at least about 9 gallons / min, at least about 10 gallons / min, at least about 11 gallons / min, at least about 13 gallons / min, at least about 15 gallons / min, at least about 20 gallons / min, at least about 25 gallons / min, at least about 30 gallons / min, at least about 40 gallons / min, or at least about 50 gallons / min.

[0244] In some embodiments, at least a portion of the working fluid from one reactor may be transferred to another reactor (e.g., the subsequent reactor). In some embodiments, at least a portion of the first working fluid may be transferred to the second reactor. In some embodiments, at least a portion of the second working fluid may be transferred to the third reactor. In some embodiments, at least a portion of the third working fluid may be transferred to the fourth reactor. In some embodiments, at least a portion of the fourth working fluid may be transferred to the fifth reactor. In some embodiments, a working fluid may be mixed in a reactor (e.g., chamber or container) prior to a first reactor. In some embodiments, the working fluid in each reactor may be distinct from the working fluid in other reactors in the bioreactor system. Distinct working fluids may comprise different microbial populations. The different microbial populations may comprise different microbes, (e.g., bacteria, fungi, algae, or any combination thereof). Distinct working fluids may comprise different concentrations of a target isolate. Distinct working fluids may comprise different concentrations of a carbon source and / or a nitrogen source. Distinct working fluids may comprise different microbial populations, differentcombinations and / or concentrations of a target isolate, different combinations and / or concentrations of a carbon source, different combinations and / or concentrations of a nitrogen source, or any combination thereof. The working fluid in a reactor of a bioreactor system may be similar to a working fluid of a different reactor of the bioreactor system.

[0245] The working fluid within each reactor may comprise different enzymes, which may be produced by microbes within the working fluid. An enzyme within a working fluid may comprise a dehydrogenase, a hydrogenase, an oxidase, a catalase, a peroxidase, a phenol o- hydroxylase, a dextransucrase, an aminotransferase, a rhodanese, a carboxylesterase, a lipase, a phosphatase, a nuclease, a phytase, an aryl sulphatase, an amylase, a cellulase, an inulase, a xylanase, a dextranase, a levanase, a poly-galacturonase, a glucosidase, a galactosidase, an invertase, a peptidase, an asparaginase, a glutaminase, an amidase, a urease, an aspartate decarboxylase, a glutamate decarboxylase an aromatic amino acid decarboxylase, or any combination thereof. An enzyme within a working fluid may comprise nitrogenase, 1- aminocyclopropane-1 -carboxylate deaminase (e.g., ACC-deaminase), quinoprotein glucose dehydrogenase (e.g., PQQ or quinone), gluconate 2-dehydrogenase, cellulase, endo-l,3(4)-P- glucanase, pectin lyase, or any combination thereof. The working fluid within each reactor may comprise different concentrations of enzymes. The working fluid within each reactor may comprise a different average abundance of an enzyme. An enzyme may be present at an average abundance between 0.001% to 1%. An enzyme may be present at an average abundance of less than 0.001%. An enzyme may be present at an average abundance of greater than 1%. In some embodiments, an enzyme may be present at an average abundance of at least about 0.0001%, at least about 0.001%, at least about 0.01%, at least about 0.02%, at least about 0.03%, at least about 0.04%, at least about 0.05%, at least about 0.06%, at least about 0.07%, at least about 0.08%, at least about 0.09%, at least about 0.1%, at least about 0.2%, at least about 0.3%, at least about 0.4%, at least about 0.5%, at least about 0.6%, at least about 0.7%, at least about 0.8%, at least about 0.9%, at least about 1%, at least about 2%, at least about 3%, at least about 4%, or at least about 5%. In some embodiments, an enzyme may be present at an average abundance of at most about 5%, at most about 4%, at most about 3%, at most about 2%, at most about 1%, at most about 0.9%, at most about 0.8%, at most about 0.7%, at most about 0.6%, at most about 0.5%, at most about 0.4%, at most about 0.3%, at most about 0.2%, at most about 0.1%, at most about 0.09%, at most about 0.08%, at most about 0.07%, at most about 0.06%, at most about 0.05%, at most about 0.04%, at most about 0.03%, at most about 0.02%, at most about 0.01%, at most about 0.001%, or at most about 0.0001%. In some embodiments, an enzyme may be present at an average abundance of between about 0.0001% and about 0.001%,about 0.0001% and about 0.01%, about 0.0001% and about 0.01%, about 0.0001% and about 0.02%, about 0.0001% and about 0.03%, about 0.0001% and about 0.05%, about 0.0001% and about 0.06%, about 0.0001% and about 0.07%, about 0.0001% and about 0.08%, about 0.0001% and about 0.09%, about 0.0001% and about 0.1%, 0.0001% and about 0.2%, 0.0001% and about 0.3%, 0.0001% and about 0.4%, 0.0001% and about 0.5%, 0.0001% and about 0.6%, 0.0001% and about 0.7%, 0.0001% and about 0.8%, 0.0001% and about 0.9%, 0.0001% and about 1%, 0.0001% and about 2%, 0.0001% and about 3%, 0.0001% and about 4%, 0.0001% and about 5%, about 0.001% and about 0.01%, about 0.001% and about 0.02%, about 0.001% and about 0.03%, about 0.001% and about 0.05%, about 0.001% and about 0.06%, about 0.001% and about 0.07%, about 0.001% and about 0.08%, about 0.001% and about 0.09%, about 0.001% and about 0.1%, 0.001% and about 0.2%, 0.001% and about 0.3%, 0.001% and about 0.4%, 0.001% and about 0.5%, 0.001% and about 0.6%, 0.001% and about 0.7%, 0.001% and about 0.8%, 0.001% and about 0.9%, 0.001% and about 1%, 0.001% and about 2%, 0.001% and about 3%, 0.001% and about 4%, 0.001% and about 5%, about 0.01% and about 0.02%, about 0.01% and about 0.03%, about 0.01% and about 0.05%, about 0.01% and about 0.06%, about 0.01% and about 0.07%, about 0.01% and about 0.08%, about 0.01% and about 0.09%, about 0.01% and about 0.1%, 0.01% and about 0.2%, 0.01% and about 0.3%, 0.01% and about 0.4%, 0.01% and about 0.5%, 0.01% and about 0.6%, 0.01% and about 0.7%, 0.01% and about 0.8%, 0.01% and about 0.9%, 0.01% and about 1%, 0.01% and about 2%, 0.01% and about 3%, 0.01% and about 4%, 0.01% and about 5%, about 0.02% and about 0.03%, about 0.02% and about 0.05%, about 0.02% and about 0.06%, about 0.02% and about 0.07%, about 0.02% and about 0.08%, about 0.02% and about 0.09%, about 0.02% and about 0.1%, 0.02% and about 0.2%, 0.02% and about 0.3%, 0.02% and about 0.4%, 0.02% and about 0.5%, 0.02% and about 0.6%, 0.02% and about 0.7%, 0.02% and about 0.8%, 0.02% and about 0.9%, 0.02% and about 1%, 0.02% and about 2%, 0.02% and about 3%, 0.02% and about 4%, 0.02% and about 5%, about 0.03% and about 0.05%, about 0.03% and about 0.06%, about 0.03% and about 0.07%, about 0.03% and about 0.08%, about 0.03% and about 0.09%, about 0.03% and about 0.1%, 0.03% and about 0.2%, 0.03% and about 0.3%, 0.03% and about 0.4%, 0.03% and about 0.5%, 0.03% and about 0.6%, 0.03% and about 0.7%, 0.03% and about 0.8%, 0.03% and about 0.9%, 0.03% and about 1%, 0.03% and about 2%, 0.03% and about 3%, 0.03% and about 4%, 0.05% and about 5%, about 0.05% and about 0.06%, about 0.05% and about 0.07%, about 0.05% and about 0.08%, about 0.05% and about 0.09%, about 0.05% and about 0.1%, 0.05% and about 0.2%, 0.05% and about 0.3%, 0.05% and about 0.4%, 0.05% and about 0.5%, 0.05% and about 0.6%, 0.05% and about 0.7%, 0.05% and about 0.8%, 0.05% and about 0.9%, 0.05% and about 1%, 0.05% andabout 2%, 0.05% and about 3%, 0.05% and about 4%, 0.05% and about 5%, about 0.06% and about 0.07%, about 0.06% and about 0.08%, about 0.06% and about 0.09%, about 0.06% and about 0.1%, 0.06% and about 0.2%, 0.06% and about 0.3%, 0.06% and about 0.4%, 0.06% and about 0.5%, 0.06% and about 0.6%, 0.06% and about 0.7%, 0.06% and about 0.8%, 0.06% and about 0.9%, 0.06% and about 1%, 0.06% and about 2%, 0.06% and about 3%, 0.06% and about 4%, 0.06% and about 5%, about 0.07% and about 0.08%, about 0.07% and about 0.09%, about 0.07% and about 0.1%, 0.07% and about 0.2%, 0.07% and about 0.3%, 0.07% and about 0.4%, 0.07% and about 0.5%, 0.07% and about 0.6%, 0.07% and about 0.7%, 0.07% and about 0.8%, 0.07% and about 0.9%, 0.07% and about 1%, 0.07% and about 2%, 0.07% and about 3%, 0.07% and about 4%, 0.07% and about 5%, about 0.08% and about 0.09%, about 0.08% and about 0.1%, 0.08% and about 0.2%, 0.08% and about 0.3%, 0.08% and about 0.4%, 0.08% and about 0.5%, 0.08% and about 0.6%, 0.08% and about 0.7%, 0.08% and about 0.8%, 0.08% and about 0.9%, 0.08% and about 1%, 0.08% and about 2%, 0.08% and about 3%, 0.08% and about 4%, 0.08% and about 5%, about 0.09% and about 0.1%, 0.09% and about 0.2%, 0.09% and about 0.3%, 0.09% and about 0.4%, 0.09% and about 0.5%, 0.09% and about 0.6%, 0.09% and about 0.7%, 0.09% and about 0.8%, 0.09% and about 0.9%, 0.09% and about 1%, 0.09% and about 2%, 0.09% and about 3%, 0.09% and about 4%, 0.09% and about 5%, 0.1% and about 0.2%, 0.1% and about 0.3%, 0.1% and about 0.4%, 0.1% and about 0.5%, 0.1% and about 0.6%, 0.1% and about 0.7%, 0.1% and about 0.8%, 0.1% and about 0.9%, 0.1% and about 1%, 0.1% and about 2%, 0.1% and about 3%, 0.1% and about 4%, 0.1% and about 5%, 0.2% and about 0.3%, 0.2% and about 0.4%, 0.2% and about 0.5%, 0.2% and about 0.6%, 0.2% and about 0.7%, 0.2% and about 0.8%, 0.2% and about 0.9%, 0.2% and about 1%, 0.2% and about 2%, 0.2% and about 3%, 0.2% and about 4%, 0.2% and about 5%, 0.3% and about 0.4%, 0.3% and about 0.5%, 0.3% and about 0.6%, 0.3% and about 0.7%, 0.3% and about 0.8%, 0.3% and about 0.9%, 0.3% and about 1%, 0.3% and about 2%, 0.3% and about 3%, 0.3% and about 4%, 0.3% and about 5%, 0.4% and about 0.5%, 0.4% and about 0.6%, 0.4% and about 0.7%, 0.4% and about 0.8%, 0.4% and about 0.9%, 0.4% and about 1%, 0.4% and about 2%, 0.4% and about 3%, 0.4% and about 4%, 0.4% and about 5%, 0.5% and about 0.6%, 0.5% and about 0.7%, 0.5% and about 0.8%, 0.5% and about 0.9%, 0.5% and about 1%, 0.5% and about 2%, 0.5% and about 3%, 0.5% and about 4%, 0.5% and about 5%, 0.6% and about 0.7%, 0.6% and about 0.8%, 0.6% and about 0.9%, 0.6% and about 1%, 0.6% and about 2%, 0.6% and about 3%, 0.6% and about 4%, 0.6% and about 5%, 0.7% and about 0.8%, 0.7% and about 0.9%, 0.7% and about 1%, 0.7% and about 2%, 0.7% and about 3%, 0.7% and about 4%, 0.7% and about 5%, 0.8% and about 0.9%, 0.8% and about 1%, 0.8% and about 2%, 0.8% and about 3%, 0.8% and about 4%, 0.8% and about5%, 0.9% and about 1%, 0.9% and about 2%, 0.9% and about 3%, 0.9% and about 4%, 0.9% and about 5%, 1% and about 2%, 1% and about 3%, 1% and about 4%, 1% and about 5%, 2% and about 3%, 2% and about 4%, 2% and about 5%, 3% and about 4%, 3% and about 5%, or 4% and about 5%.

[0246] The working fluid within each reactor may comprise enzymes with different enzymatic activity. Enzymatic activity may include, but is not limited to, nitrogen fixation, ammonia production, phosphate solubilization, zinc solubilization, and / or cell wall lysing.

[0247] A working fluid may comprise different digestion products from working fluid within other reactors of the system. In some embodiments, a working fluid may comprise digestion products from a feedstock (e.g., aqueous organic feedstock) and microbial consortium at least partially derived from a previous working fluid.

[0248] The pH of a working fluid within each reactor may be different from working fluid in other reactors. The pH of a working fluid within each reactor may be the same. The pH of a working fluid may be less than 6. The pH of a working fluid may be greater than 6. The pH of a working fluid may be in a range from 2 to 11. The pH of a working fluid may be at least about 2, at least about 2.5, at least about 3, at least about 3.5, at least about 4, at least about 4.5, at least about 5, at least about 5.5, at least about 6, at least about 6.5, at least about 7, at least about 7.5, at least about 8, at least about 8.5, at least about 9, at least about 9.5, at least about 10, at least about 10.5, or at least about 11. The pH of a working fluid may be at most about 11, at most about 10.5, at most about 10, at most about 9.5, at most about 9, at most about 8.5, at most about 8, at most about 7.5, at most about 7, at most about 6.5, at most about 6, at most about 5.5, at most about 5, at most about 4.5, at most about 4, at most about 3.5, at most about 3, at most about 2.5, or at most about 2.

[0249] The system 100 may comprise a pH sensor 191, pH controller, and / or a buffer addition system 190 to detect and / or control the pH in the reactor so that the pH is maintained at a threshold (e.g., pH 7). The buffer addition system 190 may be automatic. For example, if the pH of the working fluid in the first reactor is below the threshold value, an automatic base (e.g., 3M of NaOH) may be added to the first reactor until the pH reaches the threshold value (e.g., at least about 7).

[0250] In some cases, fluid (e.g., working fluid) can flow in a hydraulically balanced manner. The clarifier container 120 produces biostimulant products or digestion products (e.g., base products) 150. In some embodiments, a pH in a reactor of bioreactor system 100 may be between 4.0-9.0. In some embodiments, the pH in the reactor may be at least about 3.0, at least about 3.5, at least about 4.0, at least about 4.5, at least about 5.0, at least about 5.5, at least about6.0, at least about 6.5, at least about 7.0, at least about 7.5, at least about 8.0, at least about 8.5, at least about 9.0, at least about 9.5, or at least about 10.0.

[0251] In the first reactor, the working fluid may be agitated at a rate that allows heavier or undigested solids to settle to the bottom. An outlet at the top of the first reactors may allow the fluid to flow into the second reactor. An outlet at the bottom of the first reactor may transfer the settled solids back into a reactor tank. Each of the reactors in the series of reactors may have submerged scaffolding that provide a surface for biofilm growth. The scaffolding can be referred to as “fixed media substrates”. The flow of fluid from reactor to reactor may comprise a plug flow model, in which particles of an input fluid have the same velocity and direction of motion. In some embodiments, the flow of fluid in the bioreactor system is driven by gravity. In some embodiments, the flow of fluid in the bioreactor system is driven by a pump. The outflow from the top of the last reactor, may be used to create a product. Products of the methods and systems as described herein can be biostimulants.

[0252] Fluid (e.g., working fluid) from an outflow port (e.g., a product outflow port) of the first reactor 161 can flow into the second reactor 112 continuously. Fluid from an outflow port (e.g., a product outflow port) of the second reactor 112 can flow into the third reactor 113 continuously. Fluid from an outflow port (e.g., a product outflow port) of the third reactor 113 can flow into the clarifier container 120 continuously.

[0253] The outflow port may be positioned on the top, middle, and / or bottom of a reactor. Additionally, each reactor or clarifier container may comprise another outflow port for reintroducing fluid back into the same reactor or clarifier container, and may be pumped back to just below the surface of the same reactor to maintain homogeneous conditions within the working solutions. In some cases, the working fluid from each reactor is recirculated within each reactor from the bottom of the reactor back to just below the surface of the working solution to maintain a homogeneous environment for fermentation. For example, fluid from the first reactor 111 may be reintroduced back into the first reactor 111. Biosolids (e.g., floc) may be generated through the process. In some embodiments, floc can be a flocculated mass of microorganisms, extracellular polymeric substance (EPS) and adsorbed organic and inorganic material. A flocculated mass can comprise an aggregated mass of microorganisms, extracellular polymeric substance (EPS) and adsorbed organic and inorganic material. In some cases, the floc may be returned manually back to a first reactor 111. In some cases, the floc may be returned to any container of the bioreactor system 100.

[0254] In some embodiments, the bioreactor system comprises a clarifier container or clarifier tank (CLF) 120. In some embodiments, the clarifier container is the final container in abioreactor system. In some embodiments, the clarifier container is not the final container in a bioreactor system. The clarifier may comprise a single in-flow port and a single outflow port. The clarifier may comprise a single in-flow port and two or more outflow ports. In some embodiments, the clarifier comprises floc-folding wipes which rotate and release microbes that have been immobilized in the floc without introducing solids in the supernatant. The flocfolding wipers may move a working fluid in the clarifier to re-suspend microbes within the working fluid. In some embodiments, the microbes and / or an amount of target isolate strain may be re-suspended in the solution in the clarifier and transferred to the supernatant. In some cases, the clarifier container 120 may comprise an outflow port (e.g., a product outflow port) for reintroducing fluid back to the first reactor 111. In some embodiments, the clarifier further comprises a flow line to return floc to the first reactor. The flow line may comprise a conduit from the clarifier to a container or combination of containers of the bioreactor system (e.g., a first container, a second container, a third container, a fourth container, a fifth container, a sixth container, or any combination thereof). The clarifier may return flow to any container (e.g., a first container, a second container, a third container, a fourth container, a fifth container, a sixth container, or any combination thereof) of a bioreactor system to provide a recirculation of working fluid. The working fluid recirculated from the clarifier may comprise a different microbial community (e.g., different amounts of microbes, different types of microbes) than a working fluid of another container in the bioreactor system (e.g., a first working fluid, a second working fluid, a third working fluid, a fourth working fluid, a fifth working fluid, and / or a sixth working fluid). Without wishing to be bound by theory, the recirculation of flow from the clarifier to a container of the bioreactor system may help enrich a microbial community of a microbial consortium of a bioreactor system by providing working fluid from the clarifier to a different point (e.g., container) of the system. The recirculated working fluid may comprise organic materials, microbes of a microbial consortium, a target isolate, metabolites, or any combination thereof.

[0255] In the clarifier, a floc portion of a working fluid (e.g., a clarifier working fluid) may separate from a supernatant portion of a working fluid. The floc-folding wipers of the clarifier may help in separating the working fluid of the clarifier. In some embodiments, the separating may comprise gravity separation. The floc may settle on the bottom of the clarifier and the supernatant may be collected.

[0256] In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation of at least about 0.5 L / day, at least about 1.0 L / day, at leastabout 1.5 L / day, at least about 2.0 L / day, at least about 2.5 L / day, at least about 3.0 L / day, at least about 3.5 L / day, at least about 4.0 L / day, at least about 4.1 L / day, at least about 4.2 L / day, at least about 4.3 L / day, at least about 4.4 L / day, at least about 4.5 L / day, at least about 4.51 L / day, at least about 4.52 L / day, at least about 4.53 L / day, at least about 4.54 L / day, at least about 4.55 L / day, at least about 4.56 L / day, at least about 4.57 L / day, at least about 4.58 L / day, at least about 4.59 L / day, at least about 4.6 L / day, at least about 4.7 L / day, at least about 4.8 L / day, at least about 4.9 L / day, at least about 5.0 L / day, at least about 6.0 L / day, at least about 7.0 L / day, at least about 8.0 L / day, at least about 9.0 L / day, or at least about 10.0 L / day. In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation of at least about 10 L / day, at least about 50 L / day, at least about 100 L / day, at least about 200 L / day, at least about 300 L / day, at least about 400 L / day, at least about 500 L / day, at least about 750 L / day, at least about 1000 L / day, at least about 2500 L / day, at least about 5000 L / day, or greater than about 5000 L / day. In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation of at most about 5000 L / day, at most about 2500 L / day, at most about 1000 L / day, at most about 750 L / day, at most about 500 L / day, at most about 400 L / day, at most about 300 L / day, at most about 200 L / day, at most about 100 L / day, at most about 50 L / day, at most about 10 L / day, or less than about 10 L / day.

[0257] In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation of at most about 10.0 L / day, at most about 9.0 L / day, at most about 8.0 L / day, at most about 7.0 L / day, at most about 6.0 L / day, at most about 5.5 L / day, at most about 5.0 L / day, at most about 4.9 L / day, at most about 4.8 L / day, at most about 4.7 L / day, at most about 4.6 L / day, at most about 4.59 L / day, at most about 4.58 L / day, at most about 4.57 L / day, at most about 4.56 L / day, at most about 4.55 L / day, at most about 4.54 L / day, at most about 4.53 L / day, at most about 4.52 L / day, at most about 4.51 L / day, at most about 4.5 L / day, at most about 4.4 L / day, at most about 4.3 L / day, at most about 4.2 L / day, at most about 4.1 L / day, at most about 4.0 L / day, at most about 3.5 L / day, at most about 3.0 L / day, at most about 2.5 L / day, at most about 2.0 L / day, at most about 1.5 L / day, at most about 1.0 L / day, or at most about 0.5 L / day.

[0258] In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation between about 1 L / day to about 1,000 L / day. In someembodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation between at most about 1,000 L / day. In some embodiments, the supernatant from the clarifier container may be continuously collected, and a portion of the floc at the bottom of the clarifier can be returned to the first reactor at a rate of recirculation between about 1 L / day to about 5 L / day, about 1 L / day to about 10 L / day, about 1 L / day to about 25 L / day, about 1 L / day to about 50 L / day, about 1 L / day to about 100 L / day, about 1 L / day to about 200 L / day, about 1 L / day to about 300 L / day, about 1 L / day to about 400 L / day, about 1 L / day to about 500 L / day, about 1 L / day to about 750 L / day, about 1 L / day to about 1,000 L / day, about 5 L / day to about 10 L / day, about 5 L / day to about 25 L / day, about 5 L / day to about 50 L / day, about 5 L / day to about 100 L / day, about 5 L / day to about 200 L / day, about 5 L / day to about 300 L / day, about 5 L / day to about 400 L / day, about 5 L / day to about 500 L / day, about 5 L / day to about 750 L / day, about 5 L / day to about 1,000 L / day, about 10 L / day to about 25 L / day, about 10 L / day to about 50 L / day, about 10 L / day to about 100 L / day, about 10 L / day to about 200 L / day, about 10 L / day to about 300 L / day, about 10 L / day to about 400 L / day, about 10 L / day to about 500 L / day, about 10 L / day to about 750 L / day, about 10 L / day to about 1,000 L / day, about 25 L / day to about 50 L / day, about 25 L / day to about 100 L / day, about 25 L / day to about 200 L / day, about 25 L / day to about 300 L / day, about 25 L / day to about 400 L / day, about 25 L / day to about 500 L / day, about 25 L / day to about 750 L / day, about 25 L / day to about 1,000 L / day, about 50 L / day to about 100 L / day, about 50 L / day to about 200 L / day, about 50 L / day to about 300 L / day, about 50 L / day to about 400 L / day, about 50 L / day to about 500 L / day, about 50 L / day to about 750 L / day, about 50 L / day to about 1,000 L / day, about 100 L / day to about 200 L / day, about 100 L / day to about 300 L / day, about 100 L / day to about 400 L / day, about 100 L / day to about 500 L / day, about 100 L / day to about 750 L / day, about 100 L / day to about 1,000 L / day, about 200 L / day to about 300 L / day, about 200 L / day to about 400 L / day, about 200 L / day to about 500 L / day, about 200 L / day to about 750 L / day, about 200 L / day to about 1,000 L / day, about 300 L / day to about 400 L / day, about 300 L / day to about 500 L / day, about 300 L / day to about 750 L / day, about 300 L / day to about 1,000 L / day, about 400 L / day to about 500 L / day, about 400 L / day to about 750 L / day, about 400 L / day to about 1,000 L / day, about 500 L / day to about 750 L / day, about 500 L / day to about 1,000 L / day, or about 750 L / day to about 1,000 L / day.

[0259] In some embodiments, the clarifier may comprise a floc flight system (e.g., floc-folding flights) as shown in FIG. 2 that can improve the concentration of the added specific isolate(s) in the base product. With the addition of the floc flights in the clarifier, the concentration of a target isolate in the base product may be maintained at a higher concentration for a longerperiod. In the clarifier, floc may settle or become immobilized in the bottom of the clarifier. Slow, floc-folding flights may release immobilized amount(s) of the target isolate strain in the floc. The released target isolate can then be returned (e.g., be reinoculated) into the bioreactor system. The floc-folding flights may re-suspend an amount of target isolate into the supernatant (e.g., base product). In some embodiments, the floc may return back to the system at a concentration of least about a 0.1% v / v rate per day, at least about a 0.5% v / v rate per day, at least about a 1.0% v / v rate per day, at least about a 1.1% v / v rate per day, at least about a 1.2% v / v rate per day, at least about a 1.3% v / v rate per day, at least about a 1.4% v / v rate per day, at least about a 1.5% v / v rate per day, at least about a 1.6% v / v rate per day, at least about a 1.7% v / v rate per day, at least about a 1.8% v / v rate per day, at least about a 1.9% v / v rate per day, at least about a 2.0% v / v rate per day, at least about a 2.1% v / v rate per day, at least about a 2.2% v / v rate per day, at least about a 2.3% v / v rate per day, at least about a 2.4% v / v rate per day, at least about a 2.5% v / v rate per day, at least about a 2.6% v / v rate per day, at least about a 2.7% v / v rate per day, at least about a 2.8% v / v rate per day, at least about a 2.9% v / v rate per day, at least about a 3.0% v / v rate per day, at least about a 4.0% v / v rate per day, at least about a 5.0% v / v rate per day, at least about a 6.0% v / v rate per day, at least about a 7.0% v / v rate per day, at least about a 8.0% v / v rate per day, at least about a 9.0% v / v rate per day, or at least about a 10.0% v / v rate per day.

[0260] In some embodiments, the floc may return back to the system at most about a concentration of 10.0% v / v rate per day, 9.0% v / v rate per day, 8.0% v / v rate per day, 7.0% v / v rate per day, 6.0% v / v rate per day, 5.0% v / v rate per day, 4.0% v / v rate per day, 3.0% v / v rate per day, 2.9% v / v rate per day, 2.8% v / v rate per day, 2.7% v / v rate per day, 2.6% v / v rate per day, 2.5% v / v rate per day, 2.4% v / v rate per day, 2.3% v / v rate per day, 2.2% v / v rate per day, 2.1% v / v rate per day, 2.0% v / v rate per day, 1.9% v / v rate per day, 1.8% v / v rate per day, 1.7% v / v rate per day, 1.6% v / v rate per day, 1.5% v / v rate per day, 1.4% v / v rate per day, 1.3% v / v rate per day, 1.2% v / v rate per day, 1.1% v / v rate per day, 1.0% v / v rate per day, 0.5% v / v rate per day, or 0.1% v / v rate per day.

[0261] In some embodiments, the floc may return back to the system at a concentration of about 0.1 %v / v rate per day to about 15 %v / v rate per day. In some embodiments, the floc may return back to the system at a concentration of about 0.1% v / v rate per day to about 0.5% v / v rate per day, about 0.1% v / v rate per day to about 1% v / v rate per day, about 0.1% v / v rate per day to about 1.5% v / v rate per day, about 0.1% v / v rate per day to about 2% v / v rate per day, about 0.1% v / v rate per day to about 3% v / v rate per day, about 0.1% v / v rate per day to about 4% v / v rate per day, about 0.1% v / v rate per day to about 5% v / v rate per day, about 0.1% v / v rate perday to about 7% v / v rate per day, about 0.1% v / v rate per day to about 10% v / v rate per day, about 0.1% v / v rate per day to about 12% v / v rate per day, about 0.1% v / v rate per day to about 15% v / v rate per day, about 0.5% v / v rate per day to about 1% v / v rate per day, about 0.5% v / v rate per day to about 1.5% v / v rate per day, about 0.5% v / v rate per day to about 2% v / v rate per day, about 0.5% v / v rate per day to about 3% v / v rate per day, about 0.5% v / v rate per day to about 4% v / v rate per day, about 0.5% v / v rate per day to about 5% v / v rate per day, about 0.5% v / v rate per day to about 7% v / v rate per day, about 0.5% v / v rate per day to about 10% v / v rate per day, about 0.5% v / v rate per day to about 12% v / v rate per day, about 0.5% v / v rate per day to about 15% v / v rate per day, about 1% v / v rate per day to about 1.5% v / v rate per day, about 1% v / v rate per day to about 2% v / v rate per day, about 1% v / v rate per day to about 3% v / v rate per day, about 1% v / v rate per day to about 4% v / v rate per day, about 1% v / v rate per day to about 5% v / v rate per day, about 1% v / v rate per day to about 7% v / v rate per day, about 1% v / v rate per day to about 10% v / v rate per day, about 1% v / v rate per day to about 12% v / v rate per day, about 1% v / v rate per day to about 15% v / v rate per day, about 1.5% v / v rate per day to about 2% v / v rate per day, about 1.5% v / v rate per day to about 3% v / v rate per day, about 1.5% v / v rate per day to about 4% v / v rate per day, about 1.5% v / v rate per day to about 5% v / v rate per day, about 1.5% v / v rate per day to about 7% v / v rate per day, about 1.5% v / v rate per day to about 10% v / v rate per day, about 1.5% v / v rate per day to about 12% v / v rate per day, about 1.5% v / v rate per day to about 15% v / v rate per day, about 2% v / v rate per day to about 3% v / v rate per day, about 2% v / v rate per day to about 4% v / v rate per day, about 2% v / v rate per day to about 5% v / v rate per day, about 2% v / v rate per day to about 7% v / v rate per day, about 2% v / v rate per day to about 10% v / v rate per day, about 2% v / v rate per day to about 12% v / v rate per day, about 2% v / v rate per day to about 15% v / v rate per day, about 3% v / v rate per day to about 4% v / v rate per day, about 3% v / v rate per day to about 5% v / v rate per day, about 3% v / v rate per day to about 7% v / v rate per day, about 3% v / v rate per day to about 10% v / v rate per day, about 3% v / v rate per day to about 12% v / v rate per day, about 3% v / v rate per day to about 15% v / v rate per day, about 4% v / v rate per day to about 5% v / v rate per day, about 4% v / v rate per day to about 7% v / v rate per day, about 4% v / v rate per day to about 10% v / v rate per day, about 4% v / v rate per day to about 12% v / v rate per day, about 4% v / v rate per day to about 15% v / v rate per day, about 5% v / v rate per day to about 7% v / v rate per day, about 5% v / v rate per day to about 10% v / v rate per day, about 5% v / v rate per day to about 12% v / v rate per day, about 5% v / v rate per day to about 15% v / v rate per day, about 7% v / v rate per day to about 10% v / v rate per day, about 7% v / v rate per day to about 12% v / v rate per day, about 7% v / v rate per day to about 15% v / v rate per day, about 10% v / v rate per day to about 12% v / v rate per day,about 10% v / v rate per day to about 15% v / v rate per day, or about 12% v / v rate per day to about 15% v / v rate per day.

[0262] In some embodiments, the floc may return back to the system at a concentration of about 0.1% v / v rate per day to about 3% v / v rate per day. In some embodiments, the floc may return back to the system at a concentration of about 0.1% v / v rate per day to about 0.5% v / v rate per day, about 0.1% v / v rate per day to about 1% v / v rate per day, about 0.1% v / v rate per day to about 1.25% v / v rate per day, about 0.1% v / v rate per day to about 1.5% v / v rate per day, about 0.1% v / v rate per day to about 1.75% v / v rate per day, about 0.1% v / v rate per day to about 2% v / v rate per day, about 0.1% v / v rate per day to about 2.25% v / v rate per day, about 0.1% v / v rate per day to about 2.5% v / v rate per day, about 0.1% v / v rate per day to about 2.75% v / v rate per day, about 0.1% v / v rate per day to about 3% v / v rate per day, about 0.5% v / v rate per day to about 1% v / v rate per day, about 0.5% v / v rate per day to about 1.25% v / v rate per day, about 0.5% v / v rate per day to about 1.5% v / v rate per day, about 0.5% v / v rate per day to about 1.75% v / v rate per day, about 0.5% v / v rate per day to about 2% v / v rate per day, about 0.5% v / v rate per day to about 2.25% v / v rate per day, about 0.5% v / v rate per day to about 2.5% v / v rate per day, about 0.5% v / v rate per day to about 2.75% v / v rate per day, about 0.5% v / v rate per day to about 3% v / v rate per day, about 1% v / v rate per day to about 1.25% v / v rate per day, about 1% v / v rate per day to about 1.5% v / v rate per day, about 1% v / v rate per day to about 1.75% v / v rate per day, about 1% v / v rate per day to about 2% v / v rate per day, about 1% v / v rate per day to about 2.25% v / v rate per day, about 1% v / v rate per day to about 2.5% v / v rate per day, about 1% v / v rate per day to about 2.75% v / v rate per day, about 1% v / v rate per day to about 3% v / v rate per day, about 1.25% v / v rate per day to about 1.5% v / v rate per day, about 1.25% v / v rate per day to about 1.75% v / v rate per day, about 1.25% v / v rate per day to about 2% v / v rate per day, about 1.25% v / v rate per day to about 2.25% v / v rate per day, about 1.25% v / v rate per day to about 2.5% v / v rate per day, about 1.25% v / v rate per day to about 2.75% v / v rate per day, about 1.25% v / v rate per day to about 3% v / v rate per day, about 1.5% v / v rate per day to about 1.75% v / v rate per day, about 1.5% v / v rate per day to about 2% v / v rate per day, about 1.5% v / v rate per day to about 2.25% v / v rate per day, about 1.5% v / v rate per day to about 2.5% v / v rate per day, about 1.5% v / v rate per day to about 2.75% v / v rate per day, about 1.5% v / v rate per day to about 3% v / v rate per day, about 1.75% v / v rate per day to about 2% v / v rate per day, about 1.75% v / v rate per day to about 2.25% v / v rate per day, about 1.75% v / v rate per day to about 2.5% v / v rate per day, about 1.75% v / v rate per day to about 2.75% v / v rate per day, about 1.75% v / v rate per day to about 3% v / v rate per day, about 2% v / v rate per day to about 2.25% v / v rate per day, about 2% v / v rate per day to about 2.5% v / v rate per day, about 2% v / v rate perday to about 2.75% v / v rate per day, about 2% v / v rate per day to about 3% v / v rate per day, about 2.25% v / v rate per day to about 2.5% v / v rate per day, about 2.25% v / v rate per day to about 2.75% v / v rate per day, about 2.25% v / v rate per day to about 3% v / v rate per day, about 2.5% v / v rate per day to about 2.75% v / v rate per day, about 2.5% v / v rate per day to about 3% v / v rate per day, or about 2.75% v / v rate per day to about 3% v / v rate per day.

[0263] In some embodiments, as solids accumulate over time in the clarifier container, a range of at least about 20-25% solids v / v may be maintained in the bioreactor system. In some embodiments, additional floc may be harvested from the bioreactor system and removed. In some embodiments, at least about 5% solids v / v, at least about 10% solids v / v, at least about 15% solids v / v, at least about 16% solids v / v, at least about 17% solids v / v, at least about 18% solids v / v, at least about 19% solids v / v, at least about 20% solids v / v, at least about 21% solids v / v, at least about 22% solids v / v, at least about 23% solids v / v, at least about 24% solids v / v, at least about 25% solids v / v, at least about 26% solids v / v, at least about 27% solids v / v, at least about 28% solids v / v, at least about 29% solids v / v, at least about 30% solids v / v, at least about 35% solids v / v, at least about 40% solids v / v, at least about 45% solids v / v, or at least about 50% solids v / v may be maintained in the bioreactor system.

[0264] In some embodiments, at most about 50% solids v / v, at most about 45% solids v / v, at most about 40% solids v / v, at most about 35% solids v / v, at most about 30% solids v / v, at most about 29% solids v / v, at most about 28% solids v / v, at most about 27% solids v / v, at most about 26% solids v / v, at most about 25% solids v / v, at most about 24% solids v / v, at most about 23% solids v / v, at most about 22% solids v / v, at most about 21% solids v / v, at most about 20% solids v / v, at most about 19% solids v / v, at most about 18% solids v / v, at most about 17% solids v / v, at most about 16% solids v / v, at most about 15% solids v / v, at most about 14% solids v / v, at most about 13% solids v / v, at most about 12% solids v / v, at most about 11% solids v / v, at most about 10% solids v / v, or at most about 5% solids v / v may be maintained in the bioreactor system.

[0265] In some embodiments, about 0.1% solids v / v to about 60% solids v / v may be maintained in the bioreactor system. In some embodiments, about 0.1% solids v / v to about 1% solids v / v, about 0.1% solids v / v to about 5% solids v / v, about 0.1% solids v / v to about 10% solids v / v, about 0.1% solids v / v to about 15% solids v / v, about 0.1% solids v / v to about 20% solids v / v, about 0.1% solids v / v to about 25% solids v / v, about 0.1% solids v / v to about 30% solids v / v, about 0.1% solids v / v to about 35% solids v / v, about 0.1% solids v / v to about 40% solids v / v, about 0.1% solids v / v to about 50% solids v / v, about 0.1% solids v / v to about 60% solids v / v, about 1% solids v / v to about 5% solids v / v, about 1% solids v / v to about 10% solids v / v, about 1% solids v / v to about 15% solids v / v, about 1% solids v / v to about 20% solids v / v, about 1%solids v / v to about 25% solids v / v, about 1% solids v / v to about 30% solids v / v, about 1% solids v / v to about 35% solids v / v, about 1% solids v / v to about 40% solids v / v, about 1% solids v / v to about 50% solids v / v, about 1% solids v / v to about 60% solids v / v, about 5% solids v / v to about 10% solids v / v, about 5% solids v / v to about 15% solids v / v, about 5% solids v / v to about 20% solids v / v, about 5% solids v / v to about 25% solids v / v, about 5% solids v / v to about 30% solids v / v, about 5% solids v / v to about 35% solids v / v, about 5% solids v / v to about 40% solids v / v, about 5% solids v / v to about 50% solids v / v, about 5% solids v / v to about 60% solids v / v, about 10% solids v / v to about 15% solids v / v, about 10% solids v / v to about 20% solids v / v, about10% solids v / v to about 25% solids v / v, about 10% solids v / v to about 30% solids v / v, about10% solids v / v to about 35% solids v / v, about 10% solids v / v to about 40% solids v / v, about10% solids v / v to about 50% solids v / v, about 10% solids v / v to about 60% solids v / v, about15% solids v / v to about 20% solids v / v, about 15% solids v / v to about 25% solids v / v, about15% solids v / v to about 30% solids v / v, about 15% solids v / v to about 35% solids v / v, about15% solids v / v to about 40% solids v / v, about 15% solids v / v to about 50% solids v / v, about15% solids v / v to about 60% solids v / v, about 20% solids v / v to about 25% solids v / v, about20% solids v / v to about 30% solids v / v, about 20% solids v / v to about 35% solids v / v, about20% solids v / v to about 40% solids v / v, about 20% solids v / v to about 50% solids v / v, about20% solids v / v to about 60% solids v / v, about 25% solids v / v to about 30% solids v / v, about25% solids v / v to about 35% solids v / v, about 25% solids v / v to about 40% solids v / v, about25% solids v / v to about 50% solids v / v, about 25% solids v / v to about 60% solids v / v, about30% solids v / v to about 35% solids v / v, about 30% solids v / v to about 40% solids v / v, about30% solids v / v to about 50% solids v / v, about 30% solids v / v to about 60% solids v / v, about35% solids v / v to about 40% solids v / v, about 35% solids v / v to about 50% solids v / v, about35% solids v / v to about 60% solids v / v, about 40% solids v / v to about 50% solids v / v, about40% solids v / v to about 60% solids v / v, or about 50% solids v / v to about 60% solids v / v.

[0266] In some embodiments, about 18% solids v / v to about 29% solids v / v. In some embodiments, about 18% solids v / v to about 19% solids v / v, about 18% solids v / v to about 20% solids v / v, about 18% solids v / v to about 21% solids v / v, about 18% solids v / v to about 22% solids v / v, about 18% solids v / v to about 23% solids v / v, about 18% solids v / v to about 24% solids v / v, about 18% solids v / v to about 25% solids v / v, about 18% solids v / v to about 26% solids v / v, about 18% solids v / v to about 27% solids v / v, about 18% solids v / v to about 28% solids v / v, about 18% solids v / v to about 29% solids v / v, about 19% solids v / v to about 20% solids v / v, about 19% solids v / v to about 21% solids v / v, about 19% solids v / v to about 22% solids v / v, about 19% solids v / v to about 23% solids v / v, about 19% solids v / v to about 24%solids v / v, about 19% solids v / v to about 25% solids v / v, about 19% solids v / v to about 26% solids v / v, about 19% solids v / v to about 27% solids v / v, about 19% solids v / v to about 28% solids v / v, about 19% solids v / v to about 29% solids v / v, about 20% solids v / v to about 21% solids v / v, about 20% solids v / v to about 22% solids v / v, about 20% solids v / v to about 23% solids v / v, about 20% solids v / v to about 24% solids v / v, about 20% solids v / v to about 25% solids v / v, about 20% solids v / v to about 26% solids v / v, about 20% solids v / v to about 27% solids v / v, about 20% solids v / v to about 28% solids v / v, about 20% solids v / v to about 29% solids v / v, about 21% solids v / v to about 22% solids v / v, about 21% solids v / v to about 23% solids v / v, about 21% solids v / v to about 24% solids v / v, about 21% solids v / v to about 25% solids v / v, about 21% solids v / v to about 26% solids v / v, about 21% solids v / v to about 27% solids v / v, about 21% solids v / v to about 28% solids v / v, about 21% solids v / v to about 29% solids v / v, about 22% solids v / v to about 23% solids v / v, about 22% solids v / v to about 24% solids v / v, about 22% solids v / v to about 25% solids v / v, about 22% solids v / v to about 26% solids v / v, about 22% solids v / v to about 27% solids v / v, about 22% solids v / v to about 28% solids v / v, about 22% solids v / v to about 29% solids v / v, about 23% solids v / v to about 24% solids v / v, about 23% solids v / v to about 25% solids v / v, about 23% solids v / v to about 26% solids v / v, about 23% solids v / v to about 27% solids v / v, about 23% solids v / v to about 28% solids v / v, about 23% solids v / v to about 29% solids v / v, about 24% solids v / v to about 25% solids v / v, about 24% solids v / v to about 26% solids v / v, about 24% solids v / v to about 27% solids v / v, about 24% solids v / v to about 28% solids v / v, about 24% solids v / v to about 29% solids v / v, about 25% solids v / v to about 26% solids v / v, about 25% solids v / v to about 27% solids v / v, about 25% solids v / v to about 28% solids v / v, about 25% solids v / v to about 29% solids v / v, about 26% solids v / v to about 27% solids v / v, about 26% solids v / v to about 28% solids v / v, about 26% solids v / v to about 29% solids v / v, about 27% solids v / v to about 28% solids v / v, about 27% solids v / v to about 29% solids v / v, or about 28% solids v / v to about 29% solids v / v may be maintained in the bioreactor system.

[0267] Inputs into bioreactor systems may comprise one or more of water, a microbial inoculum (e.g., an inoculum of a microbe or an inoculum of a microbial strain), nutrients (e.g., one or more sources of carbon, nitrogen, phosphorous, etc.), and a digestion substrate. In some embodiments, loading inputs into a reactor can comprise a carbon source, a nitrogen source, a flour, an isolate, or any combination thereof. In some embodiments, the loading inputs comprise recycled floc from the system. In some embodiments, the loading inputs comprise a whole broth (WB). The inoculum of a microbe as described herein may metabolize the carbon source.Metabolism of carbon by the inoculum of the microbe may comprise transfer of carbon-based moieties of the carbon source to substrates in the working fluid.

[0268] Digestion substrates an input stream into a bioreactor system may comprise, for example, organic materials that can be digested by microbes in the bioreactor system. Such organic materials may comprise, for example, manure, lignocellulosic material, wastewater biosolids, food waste, energy crops, yeast, guano, agricultural waste, algae, or any combination thereof. The manure may be chicken manure, cow manure, horse manure, sheep manure, alpaca manure, rabbit manure, and / or pig manure. In some embodiments, the manure is a mixture of one, two, three, or more manures. Digestion substrates input into bioreactor systems may have been subject to a partial digestion before being input into the system. Thus, the input into the system may comprise products of digestion of an original digestion substrate by microbes endogenous to the original digestion substrate, as well as digestible materials still present in the input. In some embodiments, digestion substrates in an input stream may comprise an inorganic substrate. The inorganic substrate may comprise, for example, sand, vermiculite, perlite, and / or pumice. In some embodiments, the inorganic substrates comprises a mineral. In some embodiments, the inorganic substrate comprises rock phosphate. In some embodiments, an input stream may comprise a source of zinc such as, for example, an insoluble form of zinc.

[0269] In some embodiments, the input composition to the bioreactor system 100 may comprise a carbon source 131. The carbon source may be glucose, malate, gluconic acid, lactose, sucrose, pyruvate, other simple sugars, or any combination thereof. In some cases, the carbon source may be added to the bioreactor system on a first day of a digestion process. In some cases, the carbon source may be added to the bioreactor system on a second day, third day, fourth day, or any day following a first day of a digestion process. The carbon source (e.g., glucose) may be added to the reactor to maintain a concentration range of 0.2%-3.0% w / v of carbon based on the total volume of the working fluid of the system. In some embodiments, the concentration of carbon that may be maintained can be based on the total concentration of all components of the working fluid in the at a point during the digestion process of the bioreactor system 100.

[0270] In some embodiments, the carbon source (e.g., glucose, malate) may be added to a reactor to maintain a concentration of carbon source in the working fluid of at least about 0.1%, at least about 0.25%, at least about 0.5%, at least about 0.75%, at least about 1.0%, at least about 1.25%, at least about 1.5%, at least about 1.75%, at least about 2.0%, at least about 2.5%, at least about 3.0%, at least about 5.0%, at least about 7.5%, or at least about 10.0% w / v. In some embodiments, the carbon source (e.g., glucose, malate) may be added to the first reactor to maintain a concentration of carbon source in the working fluid of at most about 10.0%, at mostabout 7.5%, at most about 5.0%, at most about 3.0%, at most about 2.5%, at most about 2.0%, at most about 1.75%, at most about 1.5%, at most about 1.25%, at most about 1.0%, at most about 0.75%, at most about 0.5%, at most about 0.25%, or at most about 0.1% w / v.

[0271] In some embodiments, the carbon source (e.g., glucose) may be added to a reactor to maintain a concentration of carbon source in the working fluid of about 0.1% w / v to about 5% w / v. In some embodiments, the carbon source (e.g., glucose) may be added to a reactor to maintain a concentration of carbon source in the working fluid of about 0.1% w / v to about 0.25% w / v, about 0.1% w / v to about 0.5% w / v, about 0.1% w / v to about 0.75% w / v, about 0.1% w / v to about 1% w / v, about 0.1% w / v to about 1.25% w / v, about 0.1% w / v to about 1.5% w / v, about 0.1% w / v to about 1.75% w / v, about 0.1% w / v to about 2% w / v, about 0.1% w / v to about 2.5% w / v, about 0.1% w / v to about 3% w / v, about 0.1% w / v to about 5% w / v, about 0.25% w / v to about 0.5% w / v, about 0.25% w / v to about 0.75% w / v, about 0.25% w / v to about 1% w / v, about 0.25% w / v to about 1.25% w / v, about 0.25% w / v to about 1.5% w / v, about 0.25% w / v to about 1.75% w / v, about 0.25% w / v to about 2% w / v, about 0.25% w / v to about 2.5% w / v, about 0.25% w / v to about 3% w / v, about 0.25% w / v to about 5% w / v, about 0.5% w / v to about 0.75% w / v, about 0.5% w / v to about 1% w / v, about 0.5% w / v to about 1.25% w / v, about 0.5% w / v to about 1.5% w / v, about 0.5% w / v to about 1.75% w / v, about 0.5% w / v to about 2% w / v, about 0.5% w / v to about 2.5% w / v, about 0.5% w / v to about 3% w / v, about 0.5% w / v to about 5% w / v, about 0.75% w / v to about 1% w / v, about 0.75% w / v to about 1.25% w / v, about 0.75% w / v to about 1.5% w / v, about 0.75% w / v to about 1.75% w / v, about 0.75% w / v to about 2% w / v, about 0.75% w / v to about 2.5% w / v, about 0.75% w / v to about 3% w / v, about 0.75% w / v to about 5% w / v, about 1% w / v to about 1.25% w / v, about 1% w / v to about 1.5% w / v, about 1% w / v to about 1.75% w / v, about 1% w / v to about 2% w / v, about 1% w / v to about 2.5% w / v, about 1% w / v to about 3% w / v, about 1% w / v to about 5% w / v, about 1.25% w / v to about 1.5% w / v, about 1.25% w / v to about 1.75% w / v, about 1.25% w / v to about 2% w / v, about 1.25% w / v to about 2.5% w / v, about 1.25% w / v to about 3% w / v, about 1.25% w / v to about 5% w / v, about 1.5% w / v to about 1.75% w / v, about 1.5% w / v to about 2% w / v, about 1.5% w / v to about 2.5% w / v, about 1.5% w / v to about 3% w / v, about 1.5% w / v to about 5% w / v, about 1.75% w / v to about 2% w / v, about 1.75% w / v to about 2.5% w / v, about 1.75% w / v to about 3% w / v, about 1.75% w / v to about 5% w / v, about 2% w / v to about 2.5% w / v, about 2% w / v to about 3% w / v, about 2% w / v to about 5% w / v, about 2.5% w / v to about 3% w / v, about 2.5% w / v to about 5% w / v, or about 3% w / v to about 5% w / v.

[0272] In some embodiments, the input composition to the bioreactor system 100 may comprise a nitrogen source 132. The nitrogen source may be ammonium sulfate, ammonium chloride,ammonium nitrate, sodium nitrate, yeast extract, yeast, or any combination thereof. In some cases, the nitrogen source may be added to the bioreactor system on a first day of a digestion process. In some cases, the nitrogen source may be added to the bioreactor system on a second day, third day, fourth day, or any day following a first day of a digestion process. The nitrogen source (e.g., ammonium sulfate) may be added to the reactor to maintain a concentration range of 0.02-0.2% w / v of nitrogen based on the total volume of the working fluid of the system. In some embodiments, the concentration of nitrogen that may be maintained can be based on the total concentration of all components of the working fluid in the at a point during the digestion process of the bioreactor system 100.

[0273] In some embodiments, the nitrogen source (e.g., ammonium sulfate) may be added to the reactor to maintain a concentration of nitrogen source in the working fluid of at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.02% w / v, at least about 0.03% w / v, at least about 0.04% w / v, at least about 0.05% w / v, at least about 0.055% w / v, at least about 0.06% w / v, at least about 0.065% w / v, at least about 0.07% w / v, at least about 0.075% w / v, at least about 0.1% w / v, at least about 0.125% w / v, at least about 0.15% w / v, at least about 0.175% w / v, at least about 0.2% w / v, at least about 0.225% w / v, at least about 0.25% w / v, at least about 0.275% w / v, at least about 0.3% w / v, at least about 0.4% w / v, at least about 0.5% w / v, at least about 0.75% w / v, or at least about 1.0% w / v. In some embodiments, the nitrogen source (e.g., ammonium sulfate) may be added to the reactor to maintain a concentration of nitrogen source in the working fluid of at most about 1.0% w / v, at most about 0.75% w / v, at most about 0.5% w / v, at most about 0.4% w / v, at most about 0.3% w / v, at most about 0.275% w / v, at most about 0.25% w / v, at most about 0.225% w / v, at most about 0.2% w / v, at most about 0.175% w / v, at most about 0.15% w / v, at most about 0.125% w / v, at most about 0.1% w / v, at most about 0.075% w / v, at most about 0.07% w / v, at most about 0.065% w / v, at most about 0.06% w / v, at most about 0.055% w / v, at most about 0.05% w / v, at most about 0.04% w / v, at most about 0.03% w / v, at most about 0.02% w / v, at most about 0.01% w / v, or at most about 0.005% w / v.

[0274] In some embodiments, the nitrogen source (e.g., ammonium sulfate) may be added to the reactor to maintain a concentration of nitrogen source in the working fluid of about 0.03% w / v to about 1% w / v. In some embodiments, the nitrogen source (e.g., ammonium sulfate) may be added to the first reactor to maintain a concentration of nitrogen source in the working fluid of about 0.25% w / v to about 0.03% w / v, about 0.25% w / v to about 0.04% w / v, about 0.25% w / v to about 0.05% w / v, about 0.25% w / v to about 0.1% w / v, about 0.25% w / v to about 0.125% w / v, about 0.25% w / v to about 0.15% w / v, about 0.25% w / v to about 0.175% w / v, about 0.25% w / v to about 0.2% w / v, about 0.25% w / v to about 0.5% w / v, about 0.25% w / v to about 0.75% w / v,about 0.25% w / v to about 1% w / v, about 0.03% w / v to about 0.04% w / v, about 0.03% w / v to about 0.05% w / v, about 0.03% w / v to about 0.1% w / v, about 0.03% w / v to about 0.125% w / v, about 0.03% w / v to about 0.15% w / v, about 0.03% w / v to about 0.175% w / v, about 0.03% w / v to about 0.2% w / v, about 0.03% w / v to about 0.5% w / v, about 0.03% w / v to about 0.75% w / v, about 0.03% w / v to about 1% w / v, about 0.04% w / v to about 0.05% w / v, about 0.04% w / v to about 0.1% w / v, about 0.04% w / v to about 0.125% w / v, about 0.04% w / v to about 0.15% w / v, about 0.04% w / v to about 0.175% w / v, about 0.04% w / v to about 0.2% w / v, about 0.04% w / v to about 0.5% w / v, about 0.04% w / v to about 0.75% w / v, about 0.04% w / v to about 1% w / v, about 0.05% w / v to about 0.1% w / v, about 0.05% w / v to about 0.125% w / v, about 0.05% w / v to about 0.15% w / v, about 0.05% w / v to about 0.175% w / v, about 0.05% w / v to about 0.2% w / v, about 0.05% w / v to about 0.5% w / v, about 0.05% w / v to about 0.75% w / v, about 0.05% w / v to about 1% w / v, about 0.1% w / v to about 0.125% w / v, about 0.1% w / v to about 0.15% w / v, about 0.1% w / v to about 0.175% w / v, about 0.1% w / v to about 0.2% w / v, about 0.1% w / v to about 0.5% w / v, about 0.1% w / v to about 0.75% w / v, about 0.1% w / v to about 1% w / v, about 0.125% w / v to about 0.15% w / v, about 0.125% w / v to about 0.175% w / v, about 0.125% w / v to about 0.2% w / v, about 0.125% w / v to about 0.5% w / v, about 0.125% w / v to about 0.75% w / v, about 0.125% w / v to about 1% w / v, about 0.15% w / v to about 0.175% w / v, about 0.15% w / v to about 0.2% w / v, about 0.15% w / v to about 0.5% w / v, about 0.15% w / v to about 0.75% w / v, about 0.15% w / v to about 1% w / v, about 0.175% w / v to about 0.2% w / v, about 0.175% w / v to about 0.5% w / v, about 0.175% w / v to about 0.75% w / v, about 0.175% w / v to about 1% w / v, about 0.2% w / v to about 0.5% w / v, about 0.2% w / v to about 0.75% w / v, about 0.2% w / v to about 1% w / v, about 0.5% w / v to about 0.75% w / v, about 0.5% w / v to about 1% w / v, or about 0.75% w / v to about 1% w / v.

[0275] In some cases, the quantities added of the carbon and / or nitrogen sources to the system may be relative to the retention time of the system and can be adjusted accordingly if the flow rate changes. For example, a bioreactor system with a longer retention time may have a larger amount of a carbon source and / or nitrogen source added compared to an amount of a carbon source and / or nitrogen source added to a system with a short retention time.

[0276] In some embodiments, the input composition to the bioreactor system 100 may comprise micronutrients 133. Micronutrients may comprise important elements for microorganisms to support physiological functions. The micronutrients may comprise less than about 1%, less than about 2%, less than about 3%, less than about 4%, or less than about 5% of a dry weight of a plant. Elemental micronutrients may comprise boron, zinc, manganese, copper, chlorine, molybdenum, or any combination thereof. The addition of micronutrients or compounds thatprovide micronutrients may enrich a population of the microbial strain and / or microbes of a microbial consortium of a working fluid of a bioreactor system as described herein. Addition of inorganic nutrients may enrich a population of the microbial strain and / or microbes of a microbial consortium of a working fluid of a bioreactor system as described herein. The inorganic nutrients may comprise potassium chloride (KC1), dipotassium phosphate (K2HPO4), magnesium sulfate (MgSCU), or any combination thereof.

[0277] Potassium chloride may be added into the reactor at a concentration range of at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.015% w / v, at least about 0.02% w / v, at least about 0.025% w / v, at least about 0.03% w / v, at least about 0.035% w / v, at least about 0.04% w / v, at least about 0.045% w / v, at least about 0.05% w / v, at least about 0.055% w / v, at least about 0.06% w / v, at least about 0.07% w / v, at least about 0.08% w / v, at least about 0.09% w / v, or at least about 0.1% w / v. Potassium chloride may be added into the reactor at a concentration range of at most about 0.005% w / v, at most about 0.01% w / v, at most about 0.015% w / v, at most about 0.02% w / v, at most about 0.025% w / v, at most about 0.03% w / v, at most about 0.035% w / v, at most about 0.04% w / v, at most about 0.045% w / v, at most about 0.05% w / v, at most about 0.055% w / v, at most about 0.06% w / v, at most about 0.07% w / v, at most about 0.08% w / v, at most about 0.09% w / v, or at most about 0.1% w / v.

[0278] Dipotassium phosphate may be added into the reactor at a concentration range of at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.015% w / v, at least about 0.02% w / v, at least about 0.025% w / v, at least about 0.03% w / v, at least about 0.035% w / v, at least about 0.04% w / v, at least about 0.045% w / v, at least about 0.05% w / v, at least about 0.055% w / v, at least about 0.06% w / v, at least about 0.07% w / v, at least about 0.08% w / v, at least about 0.09% w / v, or at least about 0.1% w / v. Dipotassium phosphate may be added into the reactor at a concentration range of at most about 0.005% w / v, at most about 0.01% w / v, at most about 0.015% w / v, at most about 0.02% w / v, at most about 0.025% w / v, at most about 0.03% w / v, at most about 0.035% w / v, at most about 0.04% w / v, at most about 0.045% w / v, at most about 0.05% w / v, at most about 0.055% w / v, at most about 0.06% w / v, at most about 0.07% w / v, at most about 0.08% w / v, at most about 0.09% w / v, or at most about 0.1% w / v.

[0279] Magnesium sulfate may be added into the reactor at a concentration range of at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.015% w / v, at least about 0.02% w / v, at least about 0.025% w / v, at least about 0.03% w / v, at least about 0.035% w / v, at least about 0.04% w / v, at least about 0.045% w / v, at least about 0.05% w / v, at least about 0.055% w / v, at least about 0.06% w / v, at least about 0.07% w / v, at least about 0.08% w / v, at least about 0.09% w / v, or at least about 0.1% w / v. Magnesium sulfate may be added into the reactor at aconcentration range of at most about 0.005% w / v, at most about 0.01% w / v, at most about 0.015% w / v, at most about 0.02% w / v, at most about 0.025% w / v, at most about 0.03% w / v, at most about 0.035% w / v, at most about 0.04% w / v, at most about 0.045% w / v, at most about 0.05% w / v, at most about 0.055% w / v, at most about 0.06% w / v, at most about 0.07% w / v, at most about 0.08% w / v, at most about 0.09% w / v, or at most about 0.1% w / v.

[0280] In some embodiments, micronutrients may be added on a first day, a second day, a third day, a fourth day, and / or a later day of a digestion process using bioreactor system 100 based on the total volume of the working fluid of the system .

[0281] In some embodiments, a concentration of range of 0.1-5.0% v / v of floc of a Phosphate Solubilizing Technology (PST / PwST), PST WB, may also be added as an input to the reactor on the span of the hydraulic retention time of the bioreactor system. A hydraulic retention time may comprise an amount of time a working fluid is maintained in a container (e.g., is not transferred out) of a bioreactor system as described herein. In some embodiments, the concentration range of PST WB input to the system may be at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.2% v / v, at least about 0.3% v / v, at least about 0.4% v / v, at least about 0.5% v / v, at least about 0.6% v / v, at least about 0.7% v / v, at least about 0.8% v / v, at least about 0.9% v / v, at least about 1.0% v / v, at least about 1.5% v / v, at least about 2.0% v / v, at least about 2.5% v / v, at least about 3.0% v / v, at least about 3.5% v / v, at least about 4.0% v / v, at least about 4.5% v / v, at least about 5.0% v / v, at least about 5.5% v / v, at least about 6.0% v / v, at least about 6.5% v / v, at least about 7.0% v / v, at least about 7.5% v / v, at least about 10.0% v / v, at least about 12.5% v / v, at least about 15.0% v / v. In some embodiments, the concentration range of PST WB input to the system may be at most about 0.05% v / v, at most about 0.1% v / v, at most about 0.2% v / v, at most about 0.3% v / v, at most about 0.4% v / v, at most about 0.5% v / v, at most about 0.6% v / v, at most about 0.7% v / v, at most about 0.8% v / v, at most about 0.9% v / v, at most about 1.0% v / v, at most about 1.5% v / v, at most about 2.0% v / v, at most about 2.5% v / v, at most about 3.0% v / v, at most about 3.5% v / v, at most about 4.0% v / v, at most about 4.5% v / v, at most about 5.0% v / v, at most about 5.5% v / v, at most about 6.0% v / v, at most about 6.5% v / v, at most about 7.0% v / v, at most about 7.5% v / v, at most about 10.0% v / v, at most about 12.5% v / v, or at most about 15.0% v / v.

[0282] In some cases, the PST WB provides a phosphorous source of rock phosphate to the working fluid of the bioreactor system 100. The PST WB can provide a first consortium to the bioreactor system 100 in order to populate a working fluid of a first reactor. In some embodiments, a ratio of floc: SPN may be 1 : 1, 1 :2, 1 :3, 1 :4, 1 :5, 1 :6, 1 :7, 1 :8, 1 :9, 1 : 10, 10: 1, 9: 1, 8: 1, 7: 1, 6:1, 5: 1, 4: 1, 3: 1, or 2: 1. The floc:SPN may be a combined mixture of floc (e.g.,biosolids) and supernatant from the clarifier. In some embodiments, the PST WB may be added into the first reactor 111. In some embodiments, the PST WB may be added into a second reactor 112, a third reactor 113, and / or a fourth reactor 114. In some embodiments, a concentration of range of 0.5-8% v / v of whole broth (WB) of a water-based Phosphate Solubilizing Technology (PwST), PwST WB (in a ratio of 1 : 1 floc: supernatant (SPN)), may also be added to a reactor on the span of the hydraulic retention time of the bioreactor system. In some embodiments, the PwST WB may be added into the first reactor 111. In some embodiments, the PwST WB may be added into a second reactor 112, a third reactor 113, and / or a fourth reactor 114. The PST WB and / or PwST WB may provide a phosphorous source to the bioreactor system. Addition of the PST WB and / or PwST WB may provide a first consortium to populate a working fluid of a first reactor 111 of the bioreactor system 100. A concentration of the PST WB or PwST WB may be based on a total concentration of components of a working fluid of a bioreactor system 100.

[0283] In some embodiments, the concentration range of PST WB or PwST WB input to the system may be at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.2% v / v, at least about 0.3% v / v, at least about 0.4% v / v, at least about 0.5% v / v, at least about 0.6% v / v, at least about 0.7% v / v, at least about 0.8% v / v, at least about 0.9% v / v, at least about 1.0% v / v, at least about 1.5% v / v, at least about 2.0% v / v, at least about 2.5% v / v, at least about 3.0% v / v, at least about 3.5% v / v, at least about 4.0% v / v, at least about 4.5% v / v, at least about 5.0% v / v, at least about 5.5% v / v, at least about 6.0% v / v, at least about 6.5% v / v, at least about 7.0% v / v, at least about 7.5% v / v, at least about 10.0% v / v, at least about 12.5% v / v, at least about 15.0% v / v. In some embodiments, the concentration range of PST WB or PwST WB input to the system may be at most about 0.05% v / v, at most about 0.1% v / v, at most about 0.2% v / v, at most about 0.3% v / v, at most about 0.4% v / v, at most about 0.5% v / v, at most about 0.6% v / v, at most about 0.7% v / v, at most about 0.8% v / v, at most about 0.9% v / v, at most about 1.0% v / v, at most about 1.5% v / v, at most about 2.0% v / v, at most about 2.5% v / v, at most about 3.0% v / v, at most about 3.5% v / v, at most about 4.0% v / v, at most about 4.5% v / v, at most about 5.0% v / v, at most about 5.5% v / v, at most about 6.0% v / v, at most about 6.5% v / v, at most about 7.0% v / v, at most about 7.5% v / v, at most about 10.0% v / v, at most about 12.5% v / v, at most about 15.0% v / v.

[0284] In some embodiments, the input composition, as described herein, may comprise calcium carbonate. The calcium carbonate may be added to the first reactor to at a concentration range of 0.02-0.2% w / v based on the span of the hydraulic retention time of the bioreactor system. In some embodiments, the calcium carbonate may be added to the first reactor to at a concentration range of a calcium carbonate in a working fluid of at least about 0.005%, at least about 0.075%,at least about 0.01%, at least about 0.015%, at least about 0.02%, at least about 0.025%, at least about 0.03%, at least about 0.05%, at least about 0.075%, at least about 0.1%, at least about 0.125%, at least about 0.15%, at least about 0.175%, at least about 0.2%, at least about 0.225%, at least about 0.25%, at least about 0.275%, at least about 0.3%, at least about 0.4%, at least about 0.5%, at least about 0.6%, at least about 0.75%, at least about 1.0%, at least about 1.5%, at least about 2.0%, at least about 3.0%, at least about 4.0%, or at least about 5.0% w / v. In some embodiments, the calcium carbonate may be added to the first reactor to at a concentration range of a calcium carbonate in a working fluid of at most about 0.005%, at most about 0.075%, at most about 0.01%, at most about 0.015%, at most about 0.02%, at most about 0.025%, at most about 0.03%, at most about 0.05%, at most about 0.075%, at most about 0.1%, at most about 0.125%, at most about 0.15%, at most about 0.175%, at most about 0.2%, at most about 0.225%, at most about 0.25%, at most about 0.275%, at most about 0.3%, at most about 0.4%, at most about 0.5%, at most about 0.6%, at most about 0.75%, at most about 1.0%, at most about 1.5%, at most about 2.0%, at most about 3.0%, at most about 4.0%, or at most about 5.0% w / v.

[0285] In some embodiments, the input composition, as described herein, may comprise plantbased materials (e.g., soluble plant-based materials). The plant-based material may be soy flour, corn flour, cereal flour, corn gluten, soy flour protein, soy protein hydrolysate, lentil flour, chickpea flour, green pea flour, yellow pea flour, white bean flour, or any combination thereof. The plant-based material (e.g., soy flour) may be added to the first reactor to at a concentration range of 0.2-3% w / v based on the span of the hydraulic retention time of the bioreactor system. In some embodiments, the plant-based material (e.g., soy flour) may be added to the first reactor at a concentration range of plant-based material in the working fluid of at least about 0.05%, at least about 0.075%, at least about 0.1%, at least about 0.125%, at least about 0.15%, at least about 0.175%, at least about 0.2%, at least about 0.25%, at least about 0.3%, at least about 0.5%, at least about 0.75%, at least about 1.0%, at least about 1.5%, at least about 2.0%, at least about 2.5%, at least about 3.0%, at least about 3.5%, at least about 4.0%, at least about 5.0%, at least about 6.0%, at least about 7.0%, at least about 8.0%, at least about 9.0%, or at least about 10.0% w / v. In some embodiments, the plant-based material (e.g., soy flour) may be added to the first reactor at a concentration range of plant-based material in the working fluid of at most about 0.05%, at most about 0.075%, at most about 0.1%, at most about 0.125%, at most about 0.15%, at most about 0.175%, at most about 0.2%, at most about 0.25%, at most about 0.3%, at most about 0.5%, at most about 0.75%, at most about 1.0%, at most about 1.5%, at most about 2.0%, at most about 2.5%, at most about 3.0%, at most about 3.5%, at most about 4.0%, at most about5.0%, at most about 6.0%, at most about 7.0%, at most about 8.0%, at most about 9.0%, or at most about 10.0% w / v.

[0286] In some cases, a bioreactor system 100 may be inoculated with an isolate or combination of isolates. In some cases, a bioreactor system 100 may not be reinoculated with an isolate or combination of isolates following a first inoculation. In some cases, a bioreactor system 100 may be reinoculated with an isolate or combination of isolates at least every 20 days, at least every 50 days, at least every 100 days, at least every 200 days, at least every 300 days, at least every 400 days, at least every 500 days, or more. In some cases, a bioreactor system 100 may be reinoculated with an isolate or combination of isolates at most every 500 days, at most every 400 days, at most every 300 days, at most every 200 days, at most every 100 days, at most every 50 days, at most every 20 days, or less. In some cases, a bioreactor system 100 may be inoculated with an isolate or combination of isolates on day 1 of a digestion process and reinoculated 1, 2, 3, 4, or 5 more times after day 1 of the digestion process.

[0287] The inoculum of a microbe may be an isolated microbe. An isolated microbe can comprise a microbe grown or enriched outside of a natural environment (e.g., in a culture medium or a streak plate method). In some embodiments, the inoculum of a microbe may comprise a mixture of two or more isolated microbes. The inoculum of a microbe may comprise a mixture of at least about 2, at least about 3, at least about 4, at least about 5, at least about 6, at least about 7, at least about 8, or more isolated microbes.

[0288] In some embodiments, the bioreactor system 100 may be inoculated with isolates MS4666, MS4689, MS3900, MS3907, MS4921, MS4687, MS2839, MS1835, or any combination thereof. A bioreactor system 100 may be inoculated with zinc solubilizing spore former. A bioreactor system 100 may be inoculated with phosphate solubilizing spore former. A bioreactor system 100 may be inoculated with nitrogen fixer. A bioreactor system 100 may be inoculated with zinc solubilizing rhizobacteria. A bioreactor system 100 may be inoculated with phosphate solubilizing rhizobacteria. A bioreactor system 100 may be inoculated with nitrogen fixing rhizobacteria. In some embodiments, the population of the microbe does not decrease by more than 60%, by more than 50%, by more than 45%, by more than 40%, by more than 35%, by more than 30%, by more than 25%, by more than 20%, by more than 15%, by more than 10%, or by more than 5% between a first and a second transferring of the isolate (e.g., microbe). In some embodiments, the population of the microbe is maintained by at least about 5%, at least 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, or at least about 75%.

[0289] In some embodiments, the incubation time, i.e., the amount of time that the working fluid is retained in the bioreactor system 100 before the product is removed, of the bioreactor system 100 may be 7 days. In some embodiments, the incubation time of the bioreactor system may be at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 25 days, at least about 30 days, or at least about 50 days. In some embodiments, the incubation time of the bioreactor system 100 may be at most about 50 days, at most about 30 days, at most about 25 days, at most about 20 days, at most about 19 days, at most about 18 days, at most about 17 days, at most about 16 days, at most about 15 days, at most about 14 days, at most about 13 days, at most about 12 days, at most about 11 days, at most about 10 days, at most about 9 days, at most about 8 days, at most about 7 days, at most about 6 days, at most about 5 days, at most about 4 days, at most about 3 days, at most about 2 days, or at most about 1 day.

[0290] In some embodiments, the incubation time of the bioreactor system may be about 1 day to about 30 days. In some embodiments, the incubation time of the bioreactor system may be about 1 day to about 3 days, about 1 day to about 5 days, about 1 day to about 8 days, about 1 day to about 10 days, about 1 day to about 12 days, about 1 day to about 14 days, about 1 day to about 16 days, about 1 day to about 18 days, about 1 day to about 20 days, about 1 day to about 25 days, about 1 day to about 30 days, about 3 days to about 5 days, about 3 days to about 8 days, about 3 days to about 10 days, about 3 days to about 12 days, about 3 days to about 14 days, about 3 days to about 16 days, about 3 days to about 18 days, about 3 days to about 20 days, about 3 days to about 25 days, about 3 days to about 30 days, about 5 days to about 8 days, about 5 days to about 10 days, about 5 days to about 12 days, about 5 days to about 14 days, about 5 days to about 16 days, about 5 days to about 18 days, about 5 days to about 20 days, about 5 days to about 25 days, about 5 days to about 30 days, about 8 days to about 10 days, about 8 days to about 12 days, about 8 days to about 14 days, about 8 days to about 16 days, about 8 days to about 18 days, about 8 days to about 20 days, about 8 days to about 25 days, about 8 days to about 30 days, about 10 days to about 12 days, about 10 days to about 14 days, about 10 days to about 16 days, about 10 days to about 18 days, about 10 days to about 20 days, about 10 days to about 25 days, about 10 days to about 30 days, about 12 days to about 14 days, about 12 days to about 16 days, about 12 days to about 18 days, about 12 days to about 20 days, about 12 days to about 25 days, about 12 days to about 30 days, about 14 days to about 16 days,about 14 days to about 18 days, about 14 days to about 20 days, about 14 days to about 25 days, about 14 days to about 30 days, about 16 days to about 18 days, about 16 days to about 20 days, about 16 days to about 25 days, about 16 days to about 30 days, about 18 days to about 20 days, about 18 days to about 25 days, about 18 days to about 30 days, about 20 days to about 25 days, about 20 days to about 30 days, or about 25 days to about 30 days.

[0291] In some embodiments, a bioreactor system may comprise two or more bioreactor systems 100 with each reactor producing a base product. Each reactor may operate individually and produce a base product.

[0292] In some embodiments, water may act as a hydraulic source 140 and, when coupled to the first reactor 111, provides continuous flow of water to the first reactor 111 at an electrical conductivity of at least about 100 microsiemens / centimeter (pS / cm), In some embodiments, the water tank 141 when coupled to the first reactor 111 provides continuous flow of water to the first reactor 111 at an electrical conductivity of at least about 100 microsiemens / centimeter (pS / cm), at least about 150 pS / cm, at least about 200 pS / cm, at least about 250 pS / cm, at least about 300 pS / cm, at least about 350 pS / cm, at least about 400 pS / cm, at least about 450 pS / cm, at least about 500 pS / cm, at least about 550 pS / cm, or at least about 600 pS / cm.

[0293] In some embodiments, the water tank 141 when coupled to the first reactor 111 provides continuous flow of water to the first reactor 111 at an electrical conductivity of at most about 600 pS / cm, at most about 550 pS / cm, at most about 500 pS / cm, at most about 450 pS / cm, at most about 400 pS / cm, at most about 350 pS / cm, at most about 300 pS / cm, at most about 250 pS / cm, at most about 200 pS / cm, at most about 150 pS / cm, or at most about 100 pS / cm.

[0294] In some embodiments, the water tank 141 when coupled to the first reactor 111 provides continuous flow of water to the first reactor 111 at an electrical conductivity of about 50 pS / cm to about 800 pS / cm. In some embodiments, the water tank 141 when coupled to the first reactor 111 provides continuous flow of water to the first reactor 111 at an electrical conductivity of about 50 pS / cm to about 100 pS / cm, about 50 pS / cm to about 200 pS / cm, about 50 pS / cm to about 250 pS / cm, about 50 pS / cm to about 300 pS / cm, about 50 pS / cm to about 350 pS / cm, about 50 pS / cm to about 400 pS / cm, about 50 pS / cm to about 450 pS / cm, about 50 pS / cm to about 500 pS / cm, about 50 pS / cm to about 600 pS / cm, about 50 pS / cm to about 700 pS / cm, about 50 pS / cm to about 800 pS / cm, about 100 pS / cm to about 200 pS / cm, about 100 pS / cm to about 250 pS / cm, about 100 pS / cm to about 300 pS / cm, about 100 pS / cm to about 350 pS / cm, about 100 pS / cm to about 400 pS / cm, about 100 pS / cm to about 450 pS / cm, about 100 pS / cm to about 500 pS / cm, about 100 pS / cm to about 600 pS / cm, about 100 pS / cm to about 700 pS / cm, about 100 pS / cm to about 800 pS / cm, about 200 pS / cm to about 250 pS / cm, about 200pS / cm to about 300 pS / cm, about 200 pS / cm to about 350 pS / cm, about 200 pS / cm to about 400 pS / cm, about 200 pS / cm to about 450 pS / cm, about 200 pS / cm to about 500 pS / cm, about 200 pS / cm to about 600 pS / cm, about 200 pS / cm to about 700 pS / cm, about 200 pS / cm to about 800 pS / cm, about 250 pS / cm to about 300 pS / cm, about 250 pS / cm to about 350 pS / cm, about 250 pS / cm to about 400 pS / cm, about 250 pS / cm to about 450 pS / cm, about 250 pS / cm to about 500 pS / cm, about 250 pS / cm to about 600 pS / cm, about 250 pS / cm to about 700 pS / cm, about 250 pS / cm to about 800 pS / cm, about 300 pS / cm to about 350 pS / cm, about 300 pS / cm to about 400 pS / cm, about 300 pS / cm to about 450 pS / cm, about 300 pS / cm to about 500 pS / cm, about 300 pS / cm to about 600 pS / cm, about 300 pS / cm to about 700 pS / cm, about 300 pS / cm to about 800 pS / cm, about 350 pS / cm to about 400 pS / cm, about 350 pS / cm to about 450 pS / cm, about 350 pS / cm to about 500 pS / cm, about 350 pS / cm to about 600 pS / cm, about 350 pS / cm to about 700 pS / cm, about 350 pS / cm to about 800 pS / cm, about 400 pS / cm to about 450 pS / cm, about 400 pS / cm to about 500 pS / cm, about 400 pS / cm to about 600 pS / cm, about 400 pS / cm to about 700 pS / cm, about 400 pS / cm to about 800 pS / cm, about 450 pS / cm to about 500 pS / cm, about 450 pS / cm to about 600 pS / cm, about 450 pS / cm to about 700 pS / cm, about 450 pS / cm to about 800 pS / cm, about 500 pS / cm to about 600 pS / cm, about 500 pS / cm to about 700 pS / cm, about 500 pS / cm to about 800 pS / cm, about 600 pS / cm to about 700 pS / cm, about 600 pS / cm to about 800 pS / cm, or about 700 pS / cm to about 800 pS / cm.

[0295] In some embodiments, the hydraulic rate of flow through the bioreactor system may be 4.6 ml / min to maintain a retention time of 10 days and can be varied accordingly for a retention time from 7 to 21 days depending on production volume requirements. In some embodiments, the hydraulic rate of flow through the bioreactor system may be a rate of 15 ml / min or 11 ml / min. In some embodiments, the hydraulic rate of flow through the digestion system may be a rate of at least about 1 ml / min, 2 ml / min, 3 ml / min, 4 ml / min, 4.5 ml / min, 5 ml / min, 5.5 ml / min, 6 ml / min, 6.5 ml / min, 7 ml / min, 7.5 ml / min, 8 ml / min, 9 ml / min, 10 ml / min, 11 ml / min, 12 ml / min, 13 ml / min, 14 ml / min, 14.5 ml / min, 15 ml / min, 15.5 ml / min, 16 ml / min, 16.5 ml / min, 17 ml / min, 17.5 ml / min, 20 ml / min, 22.5 ml / min, 25 ml / min, or greater than about 25 ml / min. In some embodiments, the hydraulic rate of flow through the digestion system may be a rate of at most about 25 ml / min 22.5 ml / min, 20 ml / min, 17.5 ml / min, 17 ml / min, 16.5 ml / min, 16 ml / min, 15.5 ml / min, 15 ml / min, 14.5 ml / min, 14 ml / min, 13 ml / min, 12 ml / min, 11 ml / min, 10 ml / min, 9 ml / min, 8 ml / min, 7.5 ml / min, 7 ml / min, 6.5 ml / min, 6 ml / min, 5.5 ml / min, 5 ml / min, 4.5 ml / min, 4 ml / min, 3 ml / min, 2 ml / min, 1 ml / min, or less than about 1 ml / min.

[0296] An amount of biosolids may be maintained in a bioreactor system. Maintenance of biosolids may support enrichment and / or growth of a population of the microbial strain, microbes of a microbial consortium, nutrients, or additional components of a bioreactor system as described herein. In some embodiments, as biosolids accumulate over time in the clarifier container 330, a range of at least about 20-25% biosolids v / v may be maintained in the bioreactor system. In some embodiments, additional floc may be harvested from the bioreactor system and removed. In some embodiments, at least about 5% biosolids v / v, at least about 10% biosolids v / v, at least about 15% biosolids v / v, at least about 16% biosolids v / v, at least about 17% biosolids v / v, at least about 18% biosolids v / v, at least about 19% biosolids v / v, at least about 20% biosolids v / v, at least about 21% biosolids v / v, at least about 22% biosolids v / v, at least about 23% biosolids v / v, at least about 24% biosolids v / v, at least about 25% biosolids v / v, at least about 26% biosolids v / v, at least about 27% biosolids v / v, at least about 28% biosolids v / v, at least about 29% biosolids v / v, at least about 30% biosolids v / v, at least about 35% biosolids v / v, at least about 40% biosolids v / v, at least about 45% biosolids v / v, or at least about 50% biosolids v / v may be maintained in the bioreactor system.

[0297] In some embodiments, at most about 50% biosolids v / v, at most about 45% biosolids v / v, at most about 40% biosolids v / v, at most about 35% biosolids v / v, at most about 30% biosolids v / v, at most about 29% biosolids v / v, at most about 28% biosolids v / v, at most about 27% biosolids v / v, at most about 26% biosolids v / v, at most about 25% biosolids v / v, at most about 24% biosolids v / v, at most about 23% biosolids v / v, at most about 22% biosolids v / v, at most about 21% biosolids v / v, at most about 20% biosolids v / v, at most about 19% biosolids v / v, at most about 18% biosolids v / v, at most about 17% biosolids v / v, at most about 16% biosolids v / v, at most about 15% biosolids v / v, at most about 14% biosolids v / v, at most about 13% biosolids v / v, at most about 12% biosolids v / v, at most about 11% biosolids v / v, at most about 10% biosolids v / v, or at most about 5% biosolids v / v may be maintained in the bioreactor system.

[0298] In some embodiments, about 0.1% biosolids v / v to about 60% biosolids v / v may be maintained in the bioreactor system. In some embodiments, about 0.1% biosolids v / v to about 1% biosolids v / v, about 0.1% biosolids v / v to about 5% biosolids v / v, about 0.1% biosolids v / v to about 10% biosolids v / v, abo...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A method of making a biostimulant composition having a plant growth promoting property, the method comprising:(a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the plant growth promoting property;(b) operating the bioreactor system for a duration of time by:(i) transferring into the first container an aqueous feedstock comprising a microbial consortium;(ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream;(iii) maintaining a concentration of the microbial strain throughout the duration of time in at least the first container at at least 80% of a concentration of the microbial strain at the beginning of the duration of time; and(iv) collecting at least a portion of the product outflow stream as the biostimulant composition, wherein the biostimulant composition has the plant growth promoting property; wherein the duration of time is at least 5 days; and wherein the microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the microbial strain in the first container.

2. The method of claim 1, wherein the maintaining of step (b)(iii) comprises maintaining the concentration of the microbial strain at at least 1x103CFU / ml.

3. The method of claim 1 or 2, wherein the maintaining of step (b)(iii) comprises providing nutrients in the first container that selectively promote growth of the microbial strain and sustain the established population.

4. The method of any one of claims 1-3, wherein the maintaining of step (b)(iii) comprises applying selective pressure in at least the first container.

5. The method of claim 4, wherein the selective pressure comprises conditions that reduce the growth rate of microbes present in the first container relative to the growth rate of the microbial strain in the first container.

6. The method of claim 5, wherein the selective pressure comprises conditions that shifts an ionic concentration of the working fluid.

7. The method of claim 6, wherein the microbial strain, microbes of the microbial consortium, or any combination thereof may be enriched following the shift in the ionic concentration of the working fluid.

8. The method of any one of claims 5-7, wherein the selective pressure increases an activity of microbes within the microbial consortium.

9. The method of claim 8, wherein the activity comprises solubilization of an inorganic substrate.

10. The method of claim 9, wherein the inorganic substrate comprises phosphate or zinc.

11. The method of any one of claims 1-10, wherein the biostimulant composition comprises an amount of the microbial strain.

12. The method of claim 11, wherein the amount of the microbial strain is effective to impart the plant growth promoting property to the biostimulant composition.

13. The method of claim 11, wherein the amount of the microbial strain is 1x102to 1x106CFU / ml.

14. The method of any one of claims 1-13, wherein the aqueous feedstock further comprises an organic substrate.

15. The method of any one of claims 1-14, wherein the organic substrate comprises manure, lignocellulosic material, wastewater biosolids, food waste, energy crops, yeast, guano, agricultural waste, algae, or any combination thereof.

16. The method of claim 15, wherein the biostimulant composition comprises digestion products produced by digestion of the organic substrate by the microbial strain and / or the microbial consortium.

17. The method of any one of claims 1-16, wherein the biostimulant composition comprises metabolites produced by the microbial strain and / or the microbial consortium.

18. The method of claim 17, wherein the metabolites and / or digestion products in the biostimulant composition are present in an amount effective to impart the plant growth promoting property to the biostimulant composition.

19. The method of any one of claims 1-18, wherein the biostimulant composition retains the plant growth promoting property after filter sterilization.

20. The method of any one of claims 1-19, wherein the plant growth promoting property comprises an ability to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof.

21. The method of any one of claims 1-20, wherein the plant growth promoting property comprises an ability to increase nitrogen use efficiency, zinc solubilization, or phosphate solubilization.

22. A method of making a biostimulant composition having a desired plant growth promoting property, the method comprising:(a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having the desired plant growth promoting property;(b) operating the bioreactor system for a duration of time by:(i) transferring into the first container an aqueous feedstock comprising a microbial consortium;(ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and(iii) collecting at least a portion of the product outflow stream as the biostimulant composition; wherein the established population of the microbial strain is not present in an aqueous feedstock or any other input in the bioreactor system during the duration of time at a concentration that is higher than 1% of the concentration of the established population of the microbial strain in the first container.

23. A method, comprising:(a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid,wherein a first container comprises an established population of a microbial strain having a plant growth promoting property;(b) operating the bioreactor system for a duration of time by:(i) transferring into the first container an aqueous feedstock comprising a microbial consortium;(ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and(iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the amount of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of a plant not administered the biostimulant composition.

24. The method of claim 22 or 23, wherein the nutrient uptake is measured by one or more of analysis of biomass, depletion method, or nutrient balance method, or by an assay as described in Example 4.

25. The method of any one of claims 22-24, wherein the nutrient uptake comprises macronutrients comprising nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof.

26. The method of any one of claims 22-24, wherein the nutrient uptake comprises micronutrients comprising boron, zinc, manganese, iron, copper, or a combination thereof.

27. A method, comprising:(a) providing a bioreactor system comprising two or more containers arranged in a series, each of the two or more containers comprising a volume of a working fluid, wherein a first container comprises an established population of a microbial strain having a plant growth promoting property;(b) operating the bioreactor system for a duration of time by:(i) transferring into the first container an aqueous feedstock comprising a microbial consortium;(ii) transferring a portion of the working fluid out of each of the two or more containers into either a subsequent container of the bioreactor system or a product outflow stream; and(iii) collecting at least a portion of the product outflow stream as a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain; wherein the biostimulant composition is configured to promote a plant growth property of a plant administered the biostimulant composition as compared to that of a plant not administered the biostimulant composition.

28. The method of claim 27, wherein the biostimulant composition is configured to promote uptake of a nutrient by a plant, increase the bioavailability of a nutrient in soil, recruit microbes having the plant growth promoting property to the root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof.

29. The method of claim 28, wherein the total biomass of the plant administered the biostimulant composition is increased by at least 5% as compared to the plant not administered the biostimulant composition.

30. The method of claim 29, wherein the total biomass of the plant administered the biostimulant composition is increased by at least 10% as compared to the plant not administered the biostimulant composition.

31. The method of claim 27 or 28, wherein an average leaf area of the plant administered the biostimulant composition is increased by at least 10% compared the plant not administered the biostimulant composition.

32. The method of claim 31, wherein an average leaf area of the plant administered the biostimulant composition is increased by at least 25% compared the plant not administered the biostimulant composition.

33. The method of any one of claims 27-32, wherein the established population of the microbial strain comprises a zinc solubilizing microbial strain.

34. The method of claim 33, wherein administering the biostimulant composition to the plant increases an average leaf area by at least 25% as compared to an average leaf area of a plant not administered the biostimulant composition.

35. The method of claim 34, wherein administering the biostimulant composition to the plant increases an average leaf area by at least 10% compared to an average leaf area of a plant not administered the biostimulant composition.

36. The method of claim 34, wherein administering the biostimulant composition to a plant increases a nitrogen content, a phosphorous content, a potassium content, or any combination thereof by at least 20 ug compared to that of a plant not administered the biostimulant composition.

37. The method of any one of claims 27-32, wherein the established population of the microbial strain comprises a phosphate solubilizing microbial strain.

38. The method of claim 37, wherein administering the biostimulant composition to one or more plants increases a crop yield by at least 5% as compared to a crop yield of one or more plants not administered the biostimulant composition.

39. The method of claim 37 or 28, wherein administering the biostimulant composition to one or more plants increases a dry biomass by at least 25% compared a dry biomass of one or more plants not administered the biostimulant composition.

40. The method of any one of claims 27-32, wherein the established population of the microbial strain comprises a nitrogen use efficiency -promoting microbial strain.

41. The method of claim 40, wherein administering the biostimulant composition to the plant increases an average leaf area by at least 50% compared to an average leaf area of a plant not administered the biostimulant composition.

42. The method of claim 40 or 41, wherein administering the biostimulant composition to the plant increases a dry shoot weight, a dry root weight, a stem diameter, or any combination thereof by at least 10% compared to that of a plant not administered the biostimulant composition.

43. The method of any one of claims 40-42, wherein administering the biostimulant composition to the plant increases a leaf chlorophyll content by at least 10% compared to that of a plant not administered the biostimulant composition.

44. The method of any one of claims 22-43, wherein the established population of the strain in (b) is incubated for a duration of time.

45. The method of claim 44, wherein the duration of time is at least 5 days.

46. The method of any one of claims 22-45, further comprising in (b), operating the bioreactor system for the duration of time by: (iii) maintaining a concentration of the microbial strain throughout the duration of time in at least the first container at at least 80% of a concentration of the microbial strain at the beginning of the duration of time.

47. The method of claim 46, wherein the maintaining of step (b)(iii) comprises maintaining the concentration of the microbial strain at at least 1x103CFU / ml.

48. The method of claim 46 or 47, wherein the maintaining of step (b)(iii) comprises providing nutrients in the first container that selectively promote growth of the microbial strain and sustain the established population.

49. The method of any one of claims 46-48, wherein the maintaining of step (b)(iii) comprises applying selective pressure in at least the first container.

50. The method of claim 49, wherein the selective pressure comprises conditions that reduce the growth rate of microbes present in the first container relative to the growth rate of the microbial strain in the first container.

51. The method of claim 49 or 50, wherein the selective pressure comprises conditions that shifts an ionic concentration of the working fluid.

52. The method of claim 51, wherein the microbial strain, microbes of the microbial consortium, or any combination thereof may be enriched following the shift in the ionic concentration of the working fluid.

53. The method of any one of claims 1, 22, and 27, wherein the desired plant growth promoting property comprises an ability to promote uptake of a nutrient by one or more plants, increase bioavailability of a nutrient in soil, recruit microbes having the desired plant growth promoting property to root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof.

54. The method of claim 53, wherein the nutrient comprises macronutrients.

55. The method of claim 54, wherein the macronutrients comprises nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof.

56. The method of any one of claims 53-55, wherein the nutrient comprises micronutrients.

57. The method of claim 56, wherein the micronutrients comprises boron, zinc, manganese, iron, copper, or any combination thereof.

58. The method of any one of claims 1-57, wherein the microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time.

59. The method of any one of claims 1-58, wherein the microbial strain is present in the aqueous feedstock or any other input into the bioreactor system during the duration of time.

60. The method of any one of claims 1-59, further comprising, before (a), adding an inoculum of the microbial strain to the bioreactor system, wherein the inoculum of the microbial strain produces an initial population of the microbial strain of at least 0.5x104CFU / ml in at least one container.

61. The method of claim 60, wherein, before adding the inoculum of the microbial strain, the concentration of the microbial strain is less than 1x102CFU / ml.

62. The method of any one of claims 1-61, wherein the aqueous feedstock further comprises an organic material at least partially digestible by microbes present in at least one of the containers.

63. The method of claim 62, wherein, before the transferring of step (b)(i), the organic material had been partially digested by microbes endogenous to the organic material.

64. The method of claim 62 or 63, further comprising digesting the organic material in two or more serially connected containers before the transferring of step (b)(i).

65. The method of any one of claims 62-64, wherein the organic material comprises manure and / or material produced by microbial digestion of manure.

66. The method of any one of claims 1-65, wherein the aqueous feedstock further comprises an inorganic material.

67. The method of claim 66, wherein the inorganic material comprises rock phosphate particles.

68. The method of claim 67, wherein, prior to the transferring of step (b)(i), the rock phosphate particles had been partially digested by microbes present in the aqueous feedstock.

69. The method of claim 67 or 68, further comprising partially digesting the rock phosphate particles in two or more serially connected containers before the transferring of step (b)(i).

70. The method of any one of claims 1-69, wherein the microbial consortium comprises at least 1x105CFU / ml.

71. The method of any one of claims 1-70, wherein the microbial consortium comprises microbes derived from manure and from rock phosphate particles.

72. The method of any one of claims 1-71, wherein the operating of step (b) further comprises producing microbial metabolites that have the plant growth promoting property.

73. The method of any one of claims 1-72, wherein the transferring of step (b)(i), the transferring of step (b)(ii), and the collecting of step (b) are performed continuously throughout the duration of time.

74. The method of any one of claims 1-73, wherein the transferring of step (b), the transferring of step (b), and the collecting of step (b) are performed periodically throughout the duration of time.

75. The method of any one of claims 1-74, further comprising adding one or more carbon sources to at least one container of the bioreactor system.

76. The method of claim 75, wherein the one or more carbon sources are comprised in the aqueous feedstock.

77. The method of any one of claims 1-76, wherein the bioreactor system comprises a clarifier container comprising a clarifier working fluid.

78. The method of claims 77, further comprising separating a supernatant portion of the clarifier working fluid from a floc portion of the clarifier working fluid within the clarifier container.

79. The method of claim 78, wherein the separating comprises gravity separation.

80. The method of claim 78 or 79, further comprising folding the floc portion of the clarifier working fluid.

81. The method of claim 80, wherein the folding further comprises releasing a population of the microbial strain into the supernatant portion without introducing floc solids into the supernatant portion.

82. The method of claim 80 or 81, wherein the folding is performed by folding wipers in a bottom portion of the clarifier container.

83. The method of any one of claims 78-82, wherein the operating further comprises transferring the floc portion from the clarifier container to an earlier container in the bioreactor system.

84. The method of any one of claims 78-83, wherein the product outflow stream comprises the supernatant portion of the clarifier working fluid.

85. The method of any one of claims 1-84, wherein the method further comprises producing at least 1x104CFU / ml of the microbial strain in the product outflow stream.

86. The method of any one of claims 1-85, wherein the bioreactor system comprises the first container comprising a volume of a first working fluid, a second container comprising a volume of a second working fluid, and a third container comprising a volume of a third working fluid.

87. The method of claim 86, wherein at least one of the first working fluid, the second working fluid, or the third working fluid comprises a pH buffering system.

88. The method of claim 87, further comprising maintaining the pH of at least one of the first working fluid, the second working fluid, or the third working fluid between 6 and 9 throughout the duration of time.

89. The method of any one of claims 86-88, wherein the first container comprises an outlet port fluidly connected to an inlet port of the second container and the second container comprises an outlet port fluidly connected to an input port of the third container.

90. The method of any one of claims 86-89, wherein the third container comprises an outlet port fluidly connected to a clarifier container.

91. The method of any one of claims 86-90, further comprising maintaining the volume of each of the first working fluid, the second working fluid, and the third working fluid constant throughout the duration of time.

92. The method of any one of claims 1-91, wherein step (b) comprises operating the bioreactor system in a hydraulically balanced manner.

93. The method of any one of claims 1-92, wherein the transferring of step (b), the transferring of step (b, and the collecting of step (b) are driven by gravity.

94. The method of any one of claims 1-93, wherein the operating comprises maintaining a flow rate that results in a hydraulic retention time of at least 5 days.

95. The method of any one of claims 1-94, wherein the operating comprises maintaining the product outflow stream at a flow rate of at least 100 gallons per day.

96. The method of any one of claims 1-95, wherein the volume of working fluid in each of the two or more containers is at least 100 gallons.

97. The method of any one of claims 1-96, wherein at least one of the two or more containers is a fluidized bed reactor.

98. The method of any one of claims 1-97, wherein at least one of the two or more containers is a packed bed reactor.

99. The method of any one of claims 1-98, further comprising maintaining at least one of the two or more containers under aerobic conditions.

100. The method of any one of claims 1-99, further comprising maintaining at least one of the two or more containers under microaerobic conditions.

101. The method of any one of claims 1-100, wherein the bioreactor system is operated continuously for at least 90 days.

102. The method of any one of claims 1-101, wherein the aqueous feedstock does not comprise the microbial strain at a concentration higher than 10 CFU / ml.

103. The method of any one of claims 1-102, wherein the microbial strain is not added to the bioreactor system during the duration of time at a concentration that is higher than 10 CFU / ml.

104. The method of any one of claims 1-103, wherein the bioreactor system comprises at least one container placed in the series before the first container.

105. The method of any one of claims 1-104, wherein the volume of working fluid of one of the two or more containers comprises the microbial consortium, wherein each microbial consortium is distinct from all of the microbial consortia in other working fluids.

106. The method of any one of claims 1-105, further comprising producing a population of sporulated bacteria in the product outflow stream.

107. The method of any one of claims 1-106, further comprising producing a population of the microbial strain in the product outflow stream that is sporulated.

108. The method of claim 107, wherein the population of the microbial strain that is sporulated comprises at least 1x102CFU / ml.

109. The method of any one of claims 1-108, further comprising adding an additional population of the microbial strain to the biostimulant product.

110. The method of any one of claims 1-109, wherein the plant growth promoting property does not comprise nitrogen use efficiency, zinc solubilization, or phosphate solubilization.

111. The method of any one of claims 1-110, wherein the microbial strain is a nitrogen use efficiency-promoting microbial strain.

112. The method of claim 111, wherein the nitrogen use efficiency-promoting microbial strain is one that performs nitrogen fixation, promotes nitrogen fixation in the tissues of plants, recruits nitrogen fixers to the root zones or other tissues of plants, or increases organic nitrogen content and / or mineralization of organic nitrogen in soil.

113. The method of claim 111 or 112, wherein the nitrogen use efficiency-promoting microbial strain is positive for a nifH gene.

114. The method of any one of claims 111-113, wherein the nitrogen use efficiency-promoting microbial strain is one that promotes plant growth in a nitrogen-poor growth medium.

115. The method of claim 114, wherein the nitrogen-poor growth medium comprises nitrate at less than 10 ppm.

116. The method of any one of claims 111-115, wherein the nitrogen use efficiency-promoting microbial strain is of the genus Kosakonia, Klebsiella, Rahnella, Kluyvera, Enterobacter, Achromobacter, Microbacterium, Gluconobacter, Methylobacterium, Pseudomonas, Pantoea, Azospirillum, Azocarus, Herbaspirillum, Burkholderia, Cyanobacteria, Bacillus, Paenibacillus, or any combination thereof.

117. The method of any one of claims 111-116, wherein the nitrogen use efficiency-promoting microbial strain is of the species Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enterobacter spp., Achromobacter marplatensis, Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Paenibacillus borealis, Bacillus megaterium (Priestia megaterium), or Paenibacillus sonchi.

118. The method of any one of claims 111-117, wherein the nitrogen use efficiency -promoting microbial strain is the strain deposited under ATCC Accession No. PTA-127654 (MS3907), the strain deposited under ATCC Accession No. PTA-127653 (MS3900), the strain deposited under ATCC Accession No. PTA-127655 (MS4921), or the strain deposited under ATCC Accession No. PTA-127652 (MS2748).

119. The method of any one of claims 111-118, wherein the nitrogen use efficiency-promoting microbial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration of greater than 100 CFU / ml.

120. The method of any one of claims 1-119, wherein the microbial strain is a zinc solubilizing bacterial strain.

121. The method of claim 120, wherein the zinc-solubilizing bacterial strain is of the genusBacillus.

122. The method of claim 120 or 121, wherein the zinc-solubilizing bacterial strain is of the species Bacillus safensis or Bacillus megaterium.

123. The method of any one of claims 120-122, wherein the zinc-solubilizing bacterial strain is one of the following:(a) a Bacillus safensis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 1;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 4; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 7;(b) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 2;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 5; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 8; or(c) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 3;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 6; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 9.

124. The method of any one of claims 120-123, wherein the zinc-solubilizing bacterial strain is the Bacillus safensis strain deposited under ATCC Accession No. PTA-127681, the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127683, or the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127682.

125. The method of any one of claims 1-124, wherein the microbial strain is a phosphate solubilizing bacterial strain.

126. The method of claim 125, wherein the phosphate solubilizing bacterial strain is of the genus Bacillus.

127. The method of claim 125 or 126, wherein the phosphate solubilizing bacterial strain is of the species Bacillus amyloliquefaciens or Bacillus licheniformis .

128. The method of any one of claims 125-127, wherein the phosphate solubilizing bacterial strain is one of the following:(a) a Bacillus amyloliquefaciens strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 10;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 12; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 14; or(b) a Bacillus licheniformis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 11;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 13; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 15.

129. The method of any one of claims 125-128, wherein the phosphate solubilizing bacterial strain is the Bacillus amyloliquefaciens strain deposited under ATCC Accession No. PTA-127657 or the Bacillus licheniformis strain deposited under ATCC Accession No. PTA-127656.

130. The method of any one of claims 125-129, wherein the phosphate solubilizing bacterial strain is not present in the aqueous feedstock or any other input into the bioreactor system during the duration of time at a concentration of greater than 100 CFU / ml.

131. A biostimulant composition made by the method of any one of claims 1-130.

132. A method of promoting plant growth comprising contacting a plant, seed, or plant growth medium with the biostimulant composition of claim 131.

133. A bioreactor system comprising:(a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property, wherein a concentration of the microbial strain in the first working fluid is at least 100 times higher than a concentration of the microbial strain in the aqueous feedstock stream or in any other input into the bioreactor system;(b) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and(c) a product outflow stream in fluid communication with the product outflow stream port.

134. A bioreactor system comprising:(a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property;(b) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and(c) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition,wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain is configured to increase nutrient uptake of a plant administered the biostimulant composition as compared to a nutrient uptake of the plant not administered the biostimulant composition.

135. A bioreactor system comprising:(a) a stream of an aqueous feedstock in fluid communication with a first container comprising a volume of a first working fluid, wherein the aqueous feedstock comprises a microbial consortium, wherein the first working fluid comprises a population of a microbial strain having a desired plant growth promoting property;(b) one or more additional containers arranged in a series that comprises the first container, wherein each of the one or more additional containers comprises a volume of a working fluid and is in fluid communication with at least one other container in the series, and wherein at least one of the one or more additional containers comprises a product outflow stream port; and(c) a product outflow stream in fluid communication with the product outflow stream port, wherein the product outflow stream comprises a biostimulant composition, wherein the biostimulant composition comprises an amount of the microbial strain, wherein the amount of the microbial strain is configured to promote a plant growth promoting property of a plant administered the biostimulant composition as compared to the plant not administered the biostimulant composition.

136. The bioreactor system of claim 133 or 135, wherein the desired plant growth promoting property comprises an ability to promote uptake of a nutrient by one or more plants, increase bioavailability of a nutrient in soil, recruit microbes having the desired plant growth promoting property to root zones or other tissues of plants, stimulate plant growth, increase yield of a crop, increase shoot biomass, increase root biomass, increase deaminase activity, increase acid production, increase leaf area, increase chlorophyll content, increase heat tolerance, increase cold tolerance, increase drought tolerance, increase salt tolerance, increase total biomass, or any combination thereof.

137. The bioreactor system of claim 136, wherein the nutrient comprises macronutrients.

138. The bioreactor system of claim 137, wherein the macronutrients comprises nitrogen, sulfur, phosphorous, potassium, magnesium, calcium, or a combination thereof.

139. The bioreactor system of claim 136, wherein the nutrient comprises micronutrients.

140. The bioreactor system of claim 139, wherein the micronutrients comprises boron, zinc, manganese, iron, copper, or any combination thereof.

141. The bioreactor system of any one of claims 133-140, wherein the bioreactor system is a continuous flow bioreactor system and the stream of the aqueous feedstock is a continuous stream.

142. The bioreactor system of any one of claims 133-141, wherein each of the volume of the working fluid is constant.

143. The bioreactor system of any one of claims 133-142, wherein each of the first container and the one or more additional containers comprises a concentration of the microbial strain that remains at least 1x104CFU / ml during operation of the bioreactor system.

144. The bioreactor system of any one of claims 133-143, wherein the aqueous feedstock and any other input into the bioreactor system does not comprise the population of the microbial strain or does not comprise a concentration of the microbial strain at level higher than 100 CFU / ml.

145. The bioreactor system of any one of claims 133-144, wherein the microbial consortium comprises at least 1x104CFU / ml.

146. The bioreactor system of any one of claims 133-145, wherein the aqueous feedstock further comprises an organic material digestible by microbes present in the container.

147. The bioreactor system of claim 146, wherein the organic material comprises manure or material derived from manure.

148. The bioreactor system of any one of claims 133-147, wherein the aqueous feedstock further comprises rock phosphate particles.

149. The bioreactor system of claim 148, wherein the microbial consortium comprises microbes derived from manure and rock phosphate particles.

150. The bioreactor system of any one of claims 133-149, wherein the container comprising the product outflow stream port is a clarifier container configured to separate a portion of a working fluid in the clarifier container into a supernatant portion and a floc portion.

151. The bioreactor system of claim 150, wherein the clarifier container comprises one or more floc folding flights configured to agitate settled floc in the clarifier container without resuspending solids in the floc portion into the supernatant portion.

152. The bioreactor system of claim 150 or 151, further comprising a floc return stream that flows from the clarifier to an earlier container in the series.

153. The bioreactor system of any one of claims 150-152, wherein the product outflow stream comprises the supernatant portion.

154. The bioreactor system of claim 153, wherein the product outflow stream comprises at least 1x104CFU / ml of the microbial strain.

155. The bioreactor system of claim 153 or 154, wherein the product outflow stream comprises at least 1x102CFU / ml of a sporulated form of the microbial strain.

156. The bioreactor system of any one of claims 153-155, wherein the product outflow stream comprises a total dry weight of 0.2 to 2.5 mg / ml.

157. The bioreactor system of any one of claims 153-156, wherein the product outflow stream has a chemical oxygen demand between 80 to 1200 mg / L.

158. The bioreactor system of any one of claims 153-157, wherein the product outflow stream has an electrical conductivity between 0.1 and 3.0 mS / cm.

159. The bioreactor system of any one of claims 133-158, wherein at least the first container comprises a mixer configured to aerate the first working fluid.

160. The bioreactor system of any one of claims 133-159, wherein the first working fluid and / or the working fluid in at least one of the one or more additional containers comprises aerobic conditions.

161. The bioreactor system of any one of claims 133-160, wherein the microbial strain is a nitrogen use efficiency-promoting microbial strain.

162. The bioreactor system of claim 161, wherein the nitrogen use efficiency-promoting microbial strain is one that performs nitrogen fixation, promotes nitrogen fixation in the tissues of plants, recruits nitrogen fixers to the root zones or other tissues of plants, or increases organic nitrogen content and / or mineralization of organic nitrogen in soil.

163. The bioreactor system of claim 161 or 162, wherein the nitrogen use efficiency -promoting microbial strain is positive for a nifH gene.

164. The bioreactor system of any one of claims 161-163, wherein the nitrogen use efficiencypromoting microbial strain is one that promotes plant growth in a nitrogen-poor growth medium.

165. The bioreactor system of claim 164, wherein the nitrogen-poor growth medium comprises nitrate at less than 10 ppm.

166. The bioreactor system of any one of claims 161-165, wherein the nitrogen use efficiencypromoting microbial strain is of the genus Kosakonia, Klebsiella, Rahnella, Kluyvera, Enterobacter, Achromobacter, Microbacterium, Gluconobacter, Methylobacterium, Pseudomonas, Pantoea, Azospirillum, Azocarus, Herbaspirillum, Burkholderia, Cyanobacteria, Bacillus, Paenibacillus, or any combination thereof.

167. The bioreactor system of any one of claims 161-166, wherein the nitrogen use efficiencypromoting microbial strain is of the species Kosakonia sacchari, Klebsiella variicola, Rahnella aquatilis, Kluyvera intermedia, Kosakonia pseusosacchari, Enter obacter spp., Achromobacter marplatensis, Azopirillum lipoferum, Microbacterium murale, Gluconobacter diazotrophicus, Methylobacterium symbioticum, Paenibacillus borealis, Bacillus megaterium (Priestia megaterium), or Paenibacillus sonchi.

168. The bioreactor system of any one of claims 161-167, wherein the nitrogen use efficiencypromoting microbial strain is the strain deposited under ATCC Accession No. PTA- 127654 (MS3907), the strain deposited under ATCC Accession No. PTA-127653 (MS3900), the strain deposited under ATCC Accession No. PTA-127655 (MS4921), or the strain deposited under ATCC Accession No. PTA-127652 (MS2748).

169. The bioreactor system of any one of claims 133-168, wherein the microbial strain is a phosphate solubilizing bacterial strain.

170. The bioreactor system of claim 169, wherein the phosphate solubilizing bacterial strain is of the genus Bacillus.

171. The bioreactor system of claim 169 or 170, wherein the phosphate solubilizing bacterial strain is of the species Bacillus amyloliquefaciens or Bacillus licheniformis.

172. The bioreactor system of any one of claims 169-171, wherein the phosphate solubilizing bacterial strain is one of the following:(a) a Bacillus amyloliquefaciens strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 10;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 12; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 14; or(b) a Bacillus licheniformis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 11;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 13; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 15.

173. The bioreactor system of any one of claims 169-172, wherein the phosphate solubilizing bacterial strain is the Bacillus amyloliquefaciens strain deposited under ATCC Accession No. PTA-127657 or the Bacillus licheniformis strain deposited under ATCC Accession No. PTA-127656.

174. The bioreactor system of any one of claims 133-173, wherein the microbial strain is a zinc solubilizing bacterial strain.

175. The bioreactor system of claim 174, wherein the zinc-solubilizing bacterial strain is of the genus Bacillus.

176. The bioreactor system of claim 174 or 175, wherein the zinc-solubilizing bacterial strain is of the species Bacillus safensis or Bacillus megaterium.

177. The bioreactor system of any one of claims 174-176, wherein the zinc-solubilizing bacterial strain is one of the following:(a) a Bacillus safensis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 1;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 4; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 7;(b) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 2;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 5; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 8; or(c) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 3;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 6; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 9.

178. The bioreactor system of any one of claims 174-177, wherein the zinc-solubilizing bacterial strain is the Bacillus safensis strain deposited under ATCC Accession No. PTA-127681, the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127683, or the Bacillus megaterium strain deposited under ATCC Accession No. PTA-127682.

179. A biostimulant composition made by the system of any one of claims 133-178.

180. A method of promoting plant growth comprising contacting a plant, seed, or plant growth medium with the biostimulant composition of claim 179.

181. The method of any one of claims 37-39, wherein administering the biostimulant composition to a plant increases an average leaf area by at least 10% compared to an average leaf area of a plant not administered the biostimulant composition.

182. The method of any one of claims 111, 112, 114, and 116-117, wherein the nitrogen use efficiency-promoting microbial strain is one of the following:(a) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 16;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 20; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 24;(b) a Paenibacillus borealis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 17;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 21; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 25;(c) a Paenibacillus sonchi strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 18;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 22; and(iii)a rpoB gene sequence at least 95% identical to SEQ ID NO: 26; or(d) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 19;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 23; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 27.

183. The bioreactor system of any one of claims 161, 162, 164, and 166-167, wherein the nitrogen use efficiency-promoting microbial strain is one of the following:(a) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 16;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 20; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 24;(b) a Paenibacillus borealis strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 17;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 21; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 25;(c) a Paenibacillus sonchi strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 18;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 22; and(iii)a rpoB gene sequence at least 95% identical to SEQ ID NO: 26; or(d) a Bacillus megaterium strain having one or more of the following:(i) a 16S rRNA gene sequence at least 95% identical to SEQ ID NO: 19;(ii) a gyrB gene sequence at least 95% identical to SEQ ID NO: 23; and(iii) a rpoB gene sequence at least 95% identical to SEQ ID NO: 27.

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