Bacillus safensis and application thereof in preparation of flocculant

Abstract

This invention relates to the field of microbial technology, specifically to a strain of *Bacillus safranin* and its application in the preparation of flocculants. The strain is *Bacillus safranin*. (Bacillu s safensis)BS‑RCEES‑ 2025‑001 The strain was deposited at the China General Microbiological Culture Collection Center on December 30, 2025, with accession number [accession number missing]. CGMCCNo.37242。 The present invention is based on Bacillus sarfus. (Bacillus safensis)BS‑RCEES‑2025‑001 The strain secretes extracellular polysaccharides that have a strong flocculation effect, and the flocculation rate of the wastewater treated with the flocculant prepared from it is 80% to 94%.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a strain of Bacillus safranin and its application in the preparation of flocculants. Background Technology

[0002] Rapid industrialization and population growth have led to a rapid increase in wastewater containing a variety of pollutants, including suspended solids, heavy metals, dyes, and organic and inorganic contaminants. Improper treatment of such discharges poses a serious threat to aquatic environments, human health, and ecological security. Therefore, there is an urgent need to advance effective and sustainable wastewater treatment and pollutant removal technologies. Flocculation is widely used as a primary wastewater treatment technology due to its simplicity, economy, and significant efficiency in removing suspended and colloidal particles. Traditional flocculants used in wastewater treatment can be divided into inorganic flocculants (such as aluminum sulfate, ferric chloride, and polyaluminum chloride) and organic synthetic polymers (such as polyacrylamide and polyethyleneimine). Although these chemical flocculants have been widely adopted, they have several drawbacks, including high cost, potential toxicity, low biodegradability, and secondary pollution from residual flocculants and their degradation products.

[0003] In recent years, the development of environmentally friendly and sustainable flocculants as alternatives to traditional chemical flocculants has surged significantly. Microbial flocculants (MBFs), in particular, have attracted considerable attention due to their unique advantages, including high efficiency, selective action, low toxicity, and superior biodegradability. These environmentally friendly agents are essentially extracellular polymeric substances (EPS) produced by microorganisms during their growth and metabolism. These EPSs, composed of polysaccharides, proteins, nucleic acids, and lipids, possess a variety of functional groups (such as hydroxyl, carboxyl, amino, and phosphate groups), promoting interaction with pollutants through mechanisms such as charge neutralization, adsorption, and flocculation. In recent years, the application of MBFs in wastewater treatment and pollution remediation has gained considerable attention. Their application in treating a variety of wastewaters, including municipal sewage and industrial wastewater from industries such as textiles, dyeing, pulp and paper, tanneries, agricultural runoff, and landfill leachate, has demonstrated their versatility and capability. MBFs have been shown to effectively remove suspended solids, turbidity, chemical oxygen demand (COD), heavy metals, dyes, and other pollutants, often achieving removal efficiencies comparable to or even better than those of traditional chemical flocculants. Furthermore, the biodegradable and non-toxic properties of MBF reduce environmental responsibility related to residue and waste disposal, contributing to sustainable waste management practices. As an important functional microorganism, Bacillus possesses excellent environmental adaptability, strong tolerance, and the ability to produce various enzymes to decompose complex organic matter, making it an ideal candidate for developing highly efficient wastewater treatment agents. Summary of the Invention

[0004] The purpose of this invention is to provide a strain of *Bacillus safranin* and its application in the preparation of flocculants. (Bacillus safensis)BS-RCEES-2025-001It can secrete extracellular polysaccharides with highly efficient flocculation activity, and flocculants made from them have strong flocculation ability and stable performance.

[0005] To achieve the above objectives, the present invention adopts the following technical solution: The first aspect of this invention provides a strain of *Bacillus safranin*, which is *Bacillus safranin*. (Bacillus safensis)BS-RCEES-2025-001 The strain was deposited at the China General Microbiological Culture Collection Center on December 30, 2025, with accession number CGMCC No. 37242; the deposit address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0006] The Bacillus safortiformis provided by this invention (Bacillus safensis)BS-RCEES-2025-001 The strain was isolated and screened from the residual sludge of the Beijing Lunan Wastewater Treatment Plant. This strain can secrete extracellular polysaccharides with high flocculation activity, which can be used to prepare flocculants for wastewater flocculation treatment.

[0007] The second aspect of the present invention provides the above-mentioned Bacillus safortiformis. (Bacillus safensis)BS-RCEES-2025- 001 Application of bacterial strains in the preparation of flocculants.

[0008] A third aspect of this invention provides a method for preparing a flocculant, specifically comprising the following steps: S1, Bacillus sarfusca (Bacillus safensis)BS-RCEES-2025-001 The strain was inoculated into a solid culture medium and cultured at 25–30°C for 24–36 hours. Then, sterile water was added to prepare the inoculum solution. S2. The bacterial culture solution is inoculated into a liquid culture medium for fermentation to prepare a fermentation bacterial culture solution; the liquid culture medium has a C / P mass ratio of 1 to 10, a C / N mass ratio of 5 to 40, a KH2PO4 concentration of 2 to 3 g / L, a K2HPO4 concentration of 4 to 6 g / L, and a pH of 5 to 11. S3. Add a flocculant to the fermentation broth and continue fermentation to obtain the raw material broth; S4. Add anhydrous ethanol at 0-5°C to the raw material liquid and allow it to settle. Then, separate the solid and liquid phases and freeze-dry the resulting solid phase to obtain the flocculant.

[0009] This preparation method involves a two-step fermentation process to prepare the flocculant. The addition of ethanol and a flocculant carrier works together to maximize the sedimentation of extracellular polysaccharides, thereby increasing the yield. The flocculant added in step S3 causes the bacterial cells to aggregate on the carrier, promoting the sedimentation of extracellular polysaccharides. Furthermore, it reduces the amount of ethanol used, resulting in a flocculant with strong flocculation ability and stable performance.

[0010] Optionally, the solid culture medium is beef extract peptone medium.

[0011] Preferably, the beef extract peptone medium comprises: 3 g beef extract, 10 g peptone, 15 g NaCl, 15–20 g agar, 1000 mL water, and a pH of 7.0–7.2. This beef extract peptone medium is sterilized before use, specifically by sterilization at 121°C for 20 min.

[0012] More preferably, the volume ratio of the sterile water to the beef extract peptone culture medium in step S1 is 1.0 to 1.5:1.

[0013] Preferably, the liquid culture medium comprises: 18-22 g glucose, 4.5-5.5 g K₂HPO₄, 2-2.5 g KH₂PO₄, 0.09-0.11 g NaCl, 0.18-0.22 g (NH₄)₂SO₄, 0.45-0.55 g urea, 0.45-0.55 g yeast extract, 0.18-0.22 g MgSO₄, and 1000 mL water. This liquid culture medium also needs to be sterilized before use, specifically by sterilizing at pH 8.0 and 121°C for 30 min.

[0014] More preferably, the liquid culture medium is: 20 g glucose, 5 g K2HPO4, 2 g KH2PO4, 0.1 g NaCl, 0.2 g (NH4)2SO4, 0.5 g urea, 0.5 g yeast extract, 0.2 g MgSO4, 1000 mL water, pH 8.0, sterilized at 121℃ for 30 min.

[0015] More preferably, the inoculation concentration of the bacterial culture in S2 is 0.6-1.0 mL of bacterial culture / 100 mL of liquid culture medium, the fermentation temperature is 25-30℃, and the fermentation time is 1-3 days.

[0016] More preferably, the fermentation culture in S2 is carried out in a shaker with a rotation speed of 120-150 r / min.

[0017] Preferably, the flocculant in S3 is one or more of sand, silt, sea sand, diatomaceous earth, bentonite, activated carbon, biochar, chitosan, and shell powder, and the particle size of the flocculant is 48-75µm, and it is used after sterilization.

[0018] Preferably, the amount of flocculating carrier added in S3 is 5-10 g / 100 mL of fermentation liquid, and the culture time is 1-2 days.

[0019] Preferably, the volume ratio of anhydrous ethanol to the raw material liquid in S4 is 2 to 4:1.

[0020] Preferably, the temperature for static sedimentation in S4 is 0–5°C and the time is at least 6 hours, more preferably at least 24 hours, in order to obtain more extracellular polysaccharides.

[0021] Preferably, the solid-liquid separation method described in S4 is centrifugation at a rate of 3000–5000 r / min.

[0022] Preferably, the freeze-drying temperature in S4 is -80°C, and the freeze-drying time is 3 to 4 hours.

[0023] A fourth aspect of the present invention also provides a flocculant prepared by the above-described preparation method.

[0024] The fifth aspect of the present invention provides the application of the above-mentioned flocculant in wastewater treatment.

[0025] The sixth aspect of the present invention provides a wastewater treatment method: adding the above-mentioned flocculant to the wastewater to be treated and mixing it at room temperature.

[0026] The beneficial effects of this invention are as follows: The present invention is based on Bacillus sarfus. (Bacillus safensis)BS-RCEES-2025-001 The strain secretes extracellular polysaccharides that have a strong flocculation effect, and the flocculation rate of the wastewater treated with the flocculant prepared from it is 80% to 94%. Attached Figure Description

[0027] Figure 1 The colony morphology of Bacillus sabinatus in Example 1 of this invention; Figure 2 The image shown is an electron microscope image of Bacillus sabinatus flocculant in Example 2 of this invention. Figure 3 This is a diagram illustrating the flocculation effect of the flocculant used in the test case of Bacillus sabolicii in this invention. Detailed Implementation

[0028] The present invention will now be described in detail with reference to specific embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that, unless otherwise specified, the embodiments and features described in the present invention can be combined with each other.

[0029] Unless otherwise specified, the chemical reagents used in the examples are all commercially available conventional reagents, and the technical means used in the examples are conventional means well known to those skilled in the art.

[0030] Microbial flocculants are novel, efficient, non-toxic, and inexpensive water treatment agents produced through fermentation using biotechnology. They possess bioflocculation properties, safety, and are inexpensive. Compared to chemical flocculants, they are widely available, environmentally safe, have good thermal stability, stable pH values, and can enhance the activity of microorganisms in the biochemical stage of water treatment. Currently, they have become a research hotspot in wastewater treatment both domestically and internationally. However, microbial flocculants have relatively low flocculation activity, requiring large dosages or long flocculation times when used alone.

[0031] This invention obtained a strain of *Bacillus safranin* from the residual sludge of a wastewater treatment plant in Beijing through separation and screening. (Bacillus safensis)BS-RCEES-2025-001 This strain can secrete extracellular polysaccharides with high flocculation activity, which can be used to prepare flocculants for wastewater flocculation treatment.

[0032] The embodiments of the invention also provide the use of this Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 Methods for preparing flocculants using bacterial strains: S1, Bacillus sarfusca (Bacillus safensis)BS-RCEES-2025-001 The strain was inoculated onto a solid culture medium and cultured, and then sterile water was added to prepare a bacterial culture solution.

[0033] S2. Inoculate the bacterial culture into a liquid culture medium and ferment to prepare a fermentation broth; the C / P mass ratio of the liquid culture medium is 1-10, the C / N mass ratio is 5-40, the K2HPO4 concentration is 2-3 g / L, the KH2PO4 concentration is 4-6 g / L, and the pH is 5-11. S3. Add flocculant to fermentation broth and continue fermentation to obtain raw material broth; S4. Add anhydrous ethanol at 0-5℃ to the raw material liquid and let it stand to settle. Then, separate the solid and liquid phases and freeze-dry the resulting solid phase to obtain the flocculant.

[0034] This invention also provides the use of this Bacillus safranin. (Bacillus safensis)BS-RCEES- 2025-001 A method for treating wastewater using flocculants made from the strain: Add the above-mentioned flocculant to the wastewater to be treated and mix at room temperature.

[0035] The present invention will be described below through specific embodiments.

[0036] The beef extract peptone culture medium used in the following examples has the following composition: 3 g beef extract, 10 g peptone, 5 g NaCl, 15 g agar, 1000 mL sterile water, pH 7; sterilized at 121℃ for 30 min.

[0037] Unless otherwise specified, the technical means used in the following examples are conventional means well known to those skilled in the art; the materials, reagents, culture media, etc. used in the following examples are all commercially available.

[0038] Example 1 This embodiment provides *Bacillus saliva*. (Bacillus safensis)BS-RCEES-2025-001 The process of screening and isolating strains.

[0039] 1. Bacillus safranin (Bacillus safensis)BS-RCEES-2025-001 The process of strain screening and isolation (1) Sampling: Take mud samples from the remaining sludge of a sewage treatment plant in Beijing, 5-10 cm below the surface, and put them into sterilized conical flasks for later use, or temporarily store them in a refrigerator at 4°C.

[0040] (2) Preparation of diluent (aseptic operation): Take 1 mL of sludge supernatant and pour it into an Erlenmeyer flask containing 99 mL of sterile water. Stir with a glass rod for 5 min to fully disperse the soil sample, thus obtaining a 10 mL diluent. -2 Soil suspension. Use a sterile pipette to aspirate 10... -2 1 mL of soil suspension was placed in a 9 mL sterile water test tube to make 10. -3 The diluted solution. Repeating this process, 10 solutions can be prepared sequentially. -3 -10 -7 The diluent. During operation, the tip of the pipette must not touch the liquid surface. Use a different pipette for each dilution. After each aspiration of the diluent, insert the pipette into the liquid surface and aspirate and spit three times. Each time, the liquid level drawn up should be higher than the previous one to reduce errors in dilution.

[0041] (3) Drawing lines on a flat surface: Take 10 -4 10 -5 10 -6 To prepare the dilution, use an inoculation loop to take one loopful of the dilution and streak the bacterial culture onto the surface of the petri dish. Invert the petri dish after streaking and incubate it in a 30°C biochemical incubator for 1–2 days.

[0042] (4) Selection and picking of colonies: After the colonies grow, select a culture dish with a colony count between 30 and 300. Select a single colony with a smooth and sticky surface and pick up a loopful with an inoculation loop into the slant culture medium.

[0043] (5) Colony purification: Incubate the slant culture in a 30℃ constant temperature incubator for 1-2 days. The microorganisms picked and cultured on the slant should be streaked multiple times until the characteristics of the colonies that grow on the plate after several streaks are similar and there are no heterogeneous colonies. Only then can the strain be considered to be relatively completely purified.

[0044] The plate separation medium and slant medium used in this separation process are both beef extract peptone medium.

[0045] Example 2 This embodiment provides a method using Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 The method for preparing flocculants using bacterial strains includes the following specific steps: (1) Bacillus sarfusae (Bacillus safensis)BS-RCEES-2025-001 The bacterial strain was inoculated onto beef extract peptone medium and cultured at 25°C for 1 day. Then, sterile water was added to prepare the bacterial culture solution. The volume ratio of sterile water to beef extract peptone medium was 1:1. (2) Inoculate the bacterial culture into the liquid culture medium at a concentration of 0.6 mL bacterial culture / 100 mL liquid culture medium, ferment at 30℃ for 1 day, and shake at 80 r / min to prepare the fermentation bacterial culture. The liquid culture medium consists of: 20 g glucose, 5 g K2HPO4, 2 g KH2PO4, 0.1 g NaCl, 0.2 g (NH4)2SO4, 0.5 g urea, 0.5 g yeast extract, 0.2 g MgSO4, 1000 mL water, pH 8.0, and sterilize at 121℃ for 30 min.

[0046] (3) Add flocculant to the fermentation liquid at a rate of 5 g / 100 mL. The flocculant is sand. After sieving through a 200-mesh sieve, it is sterilized and used. Continue fermentation for 1 day to obtain the raw material liquid. (4) Add 1℃ anhydrous ethanol to the raw material liquid. The volume ratio of anhydrous ethanol to the raw material liquid is 2:1. Then let it stand at 1℃ for 6 hours to settle. Centrifuge the solid at 3000 r / min to separate it. Then freeze it at -80℃ for 3 hours and vacuum dry it for 2 hours to obtain the flocculant.

[0047] Example 3 This embodiment provides a method using Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 The method for preparing flocculants using bacterial strains includes the following specific steps: (1) Bacillus sarfusae (Bacillus safensis)BS-RCEES-2025-001 The bacterial strain was inoculated onto beef extract peptone medium and cultured at 25°C for 24 hours. Then, sterile water was added to prepare the bacterial culture solution. The volume ratio of sterile water to beef extract peptone medium was 1.25:1. (2) Inoculate the bacterial culture into the liquid culture medium at a concentration of 0.8 mL bacterial culture / 100 mL liquid culture medium, ferment at 30℃ for 2 days, and shake at 120 r / min to prepare the fermentation bacterial culture. The liquid culture medium consists of: 20 g glucose, 5 g K2HPO4, 2 g KH2PO4, 0.1 g NaCl, 0.2 g (NH4)2SO4, 0.5 g urea, 0.5 g yeast extract, 0.2 g MgSO4, 1000 mL water, pH 8.0, and sterilize at 121℃ for 30 min.

[0048] (3) Add flocculant to the fermentation liquid at a rate of 7.5 g / 100 mL. The flocculant is sand, which is sterilized after being sieved through a 200-mesh sieve. Continue fermentation for 1 day to obtain the raw material liquid. (4) Add 4℃ anhydrous ethanol to the raw material liquid. The volume ratio of anhydrous ethanol to the raw material liquid is 2:1. Then let it stand at 4℃ for 6 hours to settle. Centrifuge the solid at 4000 r / min to separate it. Then freeze it at -80℃ for 3 hours and vacuum dry it for 2 hours to obtain the flocculant.

[0049] Example 4 This embodiment provides a method using Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 The method for preparing flocculants using bacterial strains includes the following specific steps: (1) Bacillus sarfusae (Bacillus safensis)BS-RCEES-2025-001 The bacterial strain was inoculated onto beef extract peptone medium and cultured at 30°C for 36 hours. Then, sterile water was added to prepare the bacterial culture solution. The volume ratio of sterile water to beef extract peptone medium was 1.5:1. (2) Inoculate the bacterial culture into the liquid culture medium at a concentration of 1 mL bacterial culture / 100 mL liquid culture medium, ferment at 30℃ for 2 days, and shake at 150 r / min to prepare the fermentation bacterial culture. The liquid culture medium consists of: 20 g glucose, 5 g K2HPO4, 2 g KH2PO4, 0.1 g NaCl, 0.2 g (NH4)2SO4, 0.5 g urea, 0.5 g yeast extract, 0.2 g MgSO4, 1000 mL water, pH 8.0, and sterilize at 121℃ for 30 min.

[0050] (3) Add flocculant to the fermentation liquid at a rate of 10 g / 100 mL. The flocculant is sand. After sieving through a 200-mesh sieve, it is sterilized and used. Continue fermentation for 2 days to obtain the raw material liquid. (4) Add 5°C anhydrous ethanol to the raw material liquid. The volume ratio of anhydrous ethanol to the raw material liquid is 4:1. Then let it stand at 5°C for 10 hours to settle. Centrifuge the solid at 5000 r / min to separate it. Then freeze it at -80°C for 3 hours and vacuum dry it for 3 hours to obtain the flocculant.

[0051] Example 5 This embodiment provides a method using Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 The method for preparing flocculants using bacterial strains includes the following specific steps: (1) Bacillus sarfusae (Bacillus safensis)BS-RCEES-2025-001 The bacterial strain was inoculated onto beef extract peptone medium and cultured at 30°C for 36 hours. Then, sterile water was added to prepare the bacterial culture solution. The volume ratio of sterile water to beef extract peptone medium was 1.5:1. (2) Inoculate the bacterial culture into the liquid culture medium at a concentration of 1 mL bacterial culture / 100 mL liquid culture medium, ferment at 30℃ for 3 days, and shake at 150 r / min to prepare the fermentation bacterial culture. The liquid culture medium consists of: 18 g glucose, 4.5 g K2HPO4, 2.5 g KH2PO4, 0.09 g NaCl, 0.18 g (NH4)2SO4, 0.55 g urea, 0.45 g yeast extract, 0.22 g MgSO4, and 1000 mL water. Sterilize at 121℃ for 30 min.

[0052] (3) Add flocculant to the fermentation liquid at a rate of 10 g / 100 mL. The flocculant is sand. After sieving through a 200-mesh sieve, it is sterilized and used. Continue fermentation for 2 days to obtain the raw material liquid. (4) Add 5°C anhydrous ethanol to the raw material liquid. The volume ratio of anhydrous ethanol to the raw material liquid is 4:1. Then let it stand at 5°C for 10 hours to settle. Centrifuge the solid at 5000 r / min to separate it. Then freeze it at -80°C for 3 hours and vacuum dry it for 3 hours to obtain the flocculant.

[0053] Example 6 This embodiment provides a method using Bacillus salsa. (Bacillus safensis)BS-RCEES-2025- 001 The method for preparing flocculants using bacterial strains includes the following specific steps: (1) Bacillus sarfusae (Bacillus safensis)BS-RCEES-2025-001 The bacterial strain was inoculated onto beef extract peptone medium and cultured at 30°C for 36 hours. Then, sterile water was added to prepare the bacterial culture solution. The volume ratio of sterile water to beef extract peptone medium was 1.5:1. (2) Inoculate the bacterial culture into the liquid culture medium at a concentration of 1 mL bacterial culture / 100 mL liquid culture medium, ferment at 30℃ for 3 days, and shake at 150 r / min to prepare the fermentation bacterial culture. The liquid culture medium consists of: 22 g glucose, 5.5 g K2HPO4, 2 g KH2PO4, 0.11 g NaCl, 0.22 g (NH4)2SO4, 0.45 g urea, 0.55 g yeast extract, 0.18 g MgSO4, and 1000 mL water. Sterilize at 121℃ for 30 min.

[0054] (3) Add flocculant to the fermentation liquid at a rate of 10 g / 100 mL. The flocculant is sand. After sieving through a 200-mesh sieve, it is sterilized and used. Continue fermentation for 2 days to obtain the raw material liquid. (4) Add 5°C anhydrous ethanol to the raw material liquid. The volume ratio of anhydrous ethanol to the raw material liquid is 4:1. Then let it stand at 5°C for 10 hours to settle. Centrifuge the solid at 5000 r / min to separate it. Then freeze it at -80°C for 3 hours and vacuum dry it for 3 hours to obtain the flocculant.

[0055] Test Example This test example measures the performance of the flocculants prepared in Examples 2-4.

[0056] A 5 g / L kaolin suspension and a 10 g / L calcium chloride solution were prepared separately using distilled water. 100 mL of the kaolin suspension and 5 mL of the calcium chloride solution were mixed thoroughly to obtain a mixture. 5 mL of this mixture was collected in three 10 mL colorimetric tubes, labeled tube 1, tube 2, and tube 3 respectively. Then, 0.3 g of each of the flocculants prepared in Examples 2, 3, and 4 were added to tubes 1, 2, and 3 respectively. The mixtures were stirred at 300 r / min for 10 minutes, then at 50 r / min for 2 minutes, and then allowed to stand for 10 minutes. The absorbance (A) was measured at a wavelength of 550 nm. 测试样 The control sample was distilled water used to prepare the mixture (its absorbance was A). 对照组 The flocculation rate is calculated based on absorbance. The flocculation rate is the percentage of the absorbance of the test sample relative to the absorbance of the control sample, excluding the absorbance of the control sample.

[0057] .

[0058] Calculations show that the flocculation rates of Examples 2, 3, and 4 are 80.71%, 86.17%, and 94.32%, respectively.

[0059] Example 7 This embodiment provides Bacillus salafulatus. (Bacillus safensis)BS-RCEES-2025-001Application of flocculants made from strains in wastewater treatment.

[0060] Take 100 mL of water from the pretreatment stage of a municipal wastewater treatment plant, with a pH of 7.39 and a turbidity of 10 NTU, and add Bacillus sabovellae. (Bacillus safensis)BS-RCEES-2025-001 0.3 g of flocculant prepared by the strain according to the method in Example 4 was rapidly stirred for 3 min at 23°C at a stirring rate of 160 r / min, then slowly stirred for 1 min at a stirring rate of 60 r / min, and then allowed to stand for 6 hours. Three batches were performed in parallel. The flocculation rate (calculated using the same method as the test example) of the three batches of flocculant treated the wastewater was found to be 80%–94%.

[0061] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely preferred examples and are not intended to limit the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of the present invention is defined by the appended claims and their equivalents.

Claims

1. A strain of *Bacillus safranin*, characterized in that, The bacterium is Bacillus safortiformis. (Bacillus safensis)BS- RCEES-2025-001 The strain has the accession number CGMCCNo.37242.

2. The *Bacillus sarfusca* as described in claim 1 (Bacillus safensis)BS-RCEES-2025-001 Application of strains in the preparation of microbial flocculants.

3. The method for preparing microbial flocculants as described in claim 2, characterized in that, Includes the following steps: S1, Bacillus sarfusca (Bacillus safensis)BS-RCEES-2025-001 The strain was inoculated into a solid culture medium and cultured at 25–30°C for 24–36 hours. Then, sterile water was added to prepare the inoculum solution. S2. The inoculum obtained in step S1 is inoculated into the fermentation medium at an inoculation concentration of 0.6-1.0 mL inoculum / 100 mL liquid medium. The culture is shaken and cultured for 1-3 days at a temperature of 25-30℃ and a rotation speed of 80-150 r / min to obtain the fermentation broth. S3. Add flocculant to fermentation broth and continue fermentation to obtain raw material broth; S4. Add ethanol to the raw material liquid and let it stand to settle. Then, centrifuge to separate the sediment and freeze-dry it to make a flocculant.

4. The method for preparing microbial flocculants as described in claim 3, characterized in that: In step S1, the solid culture medium is: 3 g beef extract, 10 g peptone, 5 g NaCl, 15-20 g agar, 1000 mL water, pH 7.0-7.2, sterilized at 121℃ for 20 min, and the volume ratio of sterile water to solid culture medium is 1.0-1.5:

1.

5. The method for preparing microbial flocculants as described in claim 3, characterized in that: In step S2, the liquid culture medium is: 20 g glucose, 5 g K2HPO4, 2 g KH2PO4, 0.1 g NaCl, 0.2 g (NH4)2SO4, 0.5 g urea, 0.5 g yeast extract, 0.2 g MgSO4, 1000 mL water, pH 8.0, sterilized at 121℃ for 30 min.

6. The method for preparing microbial flocculants as described in claim 3, characterized in that: In step S3, the amount of flocculant added is 5-10 g / 100 mL of fermentation liquid, and the culture is continued for 1-2 days.

7. The method for preparing microbial flocculants as described in claim 3, characterized in that: The flocculant is one or more of the following: sand, silt, sea sand, diatomaceous earth, bentonite, activated carbon, biochar, chitosan, and shell powder. The flocculant is sterilized after being sieved through a 200-300 mesh sieve.

8. The method for preparing microbial flocculants as described in claim 3, characterized in that: In step S4, the volume ratio of ethanol to raw material liquid is 2-4:1, the mixture is allowed to stand at 0-5°C for 6-10 hours, centrifuged at a speed of 3000-5000 r / min, frozen at -80°C, and freeze-dried for 3-4 hours.

9. A flocculant prepared by the method for preparing a microbial flocculant according to any one of claims 3-8.

10. The application of the flocculant as described in claim 9 in wastewater treatment.

Images