Saline-alkali-tolerant growth-promoting spiropolyspora pink and microbial fertilizer thereof

Abstract

The invention provides a salt-alkali-tolerant growth-promoting spiropolyspora pink strain, and the preservation number of the strain is CGMCC (China General Microbiological Culture Collection Center) No.42331. The invention also provides an application of the Pharmacosporium rosea strain and a microbial fertilizer containing the Pharmacosporium rosea strain. The spiropolyspora pink XRLZ-13-1 strain provided by the invention is derived from soil in Tibet region, has excellent environmental adaptability, and can stably grow under double stress of high salt (NaCl concentration of 5%) and high alkali (pH of 9.0). When the strain fermentation liquor is applied in a saline-alkali environment, the seed germination rate can be remarkably increased, the seed germination process is accelerated, tomato seedling growth and root development are promoted, biomass is increased, plant resistance is induced, and soil nutrient indexes are improved.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of microbial technology, and particularly relates to a pink spiral sporidaria strain, a microbial fertilizer containing the same and application. BACKGROUND

[0002] Current soil salinization and alkalization problems are becoming more and more prominent, which have seriously threatened global agricultural production and ecological environment, and even affected the living environment of human beings. According to statistics, more than 1.3 billion hectares of land (about 10% of the global soil area) are affected by salinization and alkalization, which has destroyed the soil structure, unbalanced the soil nutrients, and damaged the root function, leading to unbalanced osmotic pressure of plant cells and toxicity. Soil degradation and crop yield reduction caused by soil salinization and alkalization have become the main factors affecting the sustainable development of agriculture.

[0003] In order to alleviate the influence of salt and alkali stress on crop growth, people have tried and promoted various physical and agronomic measures. For example, improving irrigation methods to reduce salt accumulation, using mulch to reduce surface water evaporation, planting salt-tolerant crops, and adopting rotation mode, which can help to enhance the ecological stability of saline-alkali soil. At the same time, various chemical remediation methods are also widely used, such as applying gypsum and acid amendment to reduce soil pH value and improve soil structure, thereby helping crop growth. However, these methods are often high in cost, complicated in operation, poor in effect persistence, and may cause secondary environmental problems. In recent years, the use of salt-tolerant microorganisms, especially rhizosphere bacteria, to alleviate salt and alkali stress has attracted much attention. These beneficial microorganisms can alleviate the physiological damage of salt and alkali to plants through various mechanisms, and improve soil properties. They can secrete plant hormones, produce extracellular polysaccharides, increase nutrients, promote root growth, and regulate ion balance in plants, activate the defense system, and remove harmful active oxygen. In addition, rhizosphere microorganisms can also regulate soil acid-base balance, increase microbial diversity, and restore soil function.

[0004] Pink spiral sporidaria (formerly known as pink mucor) Clonostachys rosea ) is a facultative saprophytic fungus belonging to Ascomycota, Sordariomycetes, Xylariaceae, and is widely distributed in nature. Pink spiral sporidaria is also a kind of biocontrol fungus that can parasitize plant fungi, nematodes and other diseases. In addition, this kind of fungus can promote plant growth, induce plant resistance, and improve soil microenvironment, and is a kind of soil beneficial microorganism that is conducive to plant growth and environment friendly. It is an extremely attractive topic to use pink spiral sporidaria to alleviate soil salinization and alkalization problems. SUMMARY

[0005] In view of the problems existing in the prior art, the first object of the present application is to provide a salt-tolerant and alkali-tolerant growth-promoting pink spiral sporidaria strain; A second object of the present application is to propose the use of the said strain of Sporocytophaga rosea; A third object of the present application is to propose a microbial fertilizer containing the said strain of Sporocytophaga rosea.

[0006] The technical solution for achieving the above-mentioned objects of the present application is as follows: A salt-tolerant Sporocytophaga rosea strain, which is isolated from farmland soil in Lazi County, Lhasa, Tibet, and has a strain number of XRLZ-13-1 and a preservation number of CGMCC No. 42331.

[0007] We found that Sporocytophaga rosea can be isolated from nearly 50% of the sampling points in the investigation of microbial resources in the Tibet Autonomous Region. The Tibet region has high altitude, strong ultraviolet radiation, and serious soil salinization problem. It is more likely to obtain efficient or special beneficial microorganisms from such extreme habitats. Further utilization of Sporocytophaga rosea and other important microbial resources will help to improve agricultural production, protect food safety, protect the ecological environment, and meet the needs of sustainable and healthy agricultural development. Moreover, the strain grows fast, has strong sporulation ability, and is easy to cultivate on a large scale, which has great application potential.

[0008] The application of the said Sporocytophaga rosea strain in crop cultivation and saline-alkali land improvement.

[0009] The microbial fertilizer containing the said Sporocytophaga rosea strain is a fermentation broth obtained by liquid fermentation of the Sporocytophaga rosea XRLZ-13-1 strain, or a preparation obtained by mixing the fermentation broth with auxiliary materials and / or soil improvers; and the dosage form of the microbial fertilizer is a liquid agent, a granular agent or a powder agent.

[0010] Further, the fermentation broth is filtered or concentrated by centrifugation, and then mixed with the auxiliary materials and / or soil improvers to prepare the preparation; The soil improver is one or more of straw powder, biochar, humic acid, fly ash, organic fertilizer, gypsum, desulfurization gypsum, silicon fertilizer, wood ash, and potassium fulvic acid. The auxiliary material is one or more of diatomite, light calcium carbonate, talc powder, attapulgite, bentonite, kaolin, starch, dextrin, grass carbon, and rice husk powder.

[0011] In a preferred technical solution of the present application, the fermentation broth of the Sporocytophaga rosea strain is prepared by the following steps: 1) Inoculum preparation: inoculate the Sporocytophaga rosea strain on potato dextrose agar (PDA) medium, cultivate at 26-28℃ for 6-10 days, elute the spores to prepare a spore suspension, or cut the mycelium as inoculum; 2) Seed culture: inoculate the spore suspension or mycelium into potato dextrose broth (PDB) medium, and cultivate in dark at 25-28℃ for 48-84 hours to obtain seed liquid; 3) Liquid fermentation culture: prepare fermentation medium, inoculate the seed liquid after sterilization and cooling, and cultivate at 25-30℃ for 48-120 hours to obtain fermentation liquid.

[0012] More preferably, in step 3), the ingredients of the fermentation medium per liter include: sucrose 10-40 g, yeast extract powder 5-15 g, soybean meal powder 5-15 g, K2HPO4 0.5-2 g, MgSO4·7H2O 0.5-1.0 g, FeSO4·7H2O 0.05-0.1 g; and the pH value is 5.5-6.5.

[0013] The pH adjustment can use 1-20% NaOH solution, or 1-10% HCl solution, or other pH adjusters known in the art.

[0014] In step 3), the liquid fermentation culture conditions are: seed liquid inoculation amount 0.5-5%, rotation speed 150-250 r / min, culture temperature 25-28℃, culture time 3-4 days, The content of the obtained fermentation liquid is (0.5-5) × 10 8 Spores / mL.

[0015] The microbial fertilizer can be directly used as fermentation liquid, or can be added with auxiliary materials, or added with soil conditioners, or added with both auxiliary materials and soil conditioners. The following are the preferred addition ratios of auxiliary materials and soil conditioners.

[0016] After fermentation to obtain the fermentation liquid, the plate and frame filtration or centrifugal concentration is used to obtain mycelium with water content of 50-70%, auxiliary materials are added to the mycelium, or auxiliary materials are directly added to the fermentation liquid; after mixing and drying, crushing or granulation is performed; the addition amount of the auxiliary materials is 50-120% of the mycelium, or 80%-150% of the fermentation liquid.

[0017] The microbial fertilizer is added with soil conditioner, and the mass ratio of fermentation liquid to soil conditioner is 1:1-10, or the fermentation liquid is filtered or centrifugally concentrated to obtain mycelium with water content of 50-70%, and the mass ratio of mycelium to soil conditioner is 1:0.5-5.

[0018] Another preferred technical solution of the present application is that the soil conditioner is one of fly ash, humic acid, biochar, organic fertilizer, gypsum, desulfurization gypsum, silicon fertilizer, wood ash, potassium fulvate, and straw powder, and the bacterial content is 0.5-5 × 10 8The mass ratio of the fermentation broth of spores / mL and the soil conditioner is 1:1-5.

[0019] The present application has the advantages of: The pink spiral sporae strain XRLZ-13-1 provided by the present application is derived from the soil in Tibet region, has excellent environmental adaptability, and can stably grow under high-salt (NaCl concentration 5%) and high-alkali (pH 9.0) conditions. The application of the fermentation broth of the strain under saline-alkali environment can significantly improve the seed germination rate, accelerate the seed germination process, promote the growth and root of tomato seedlings, and increase the plant height and biomass.

[0020] In the pot experiment, the growth of tomato plants under saline-alkali stress is severely inhibited, the plants are short, the leaves show yellowing, curling, dry tips, and early senescence, and the root length and fresh weight significantly decrease. After applying the XRLZ-13-1 bacterial solution, the plant height and root length of tomato are increased by 21.6% and 113.6%, respectively. The root dry weight is increased by 2 times. The determination of the related enzymes and peroxidation metabolite related substances in the body shows that the activities of superoxide dismutase, peroxidase, and catalase are significantly improved, and the contents of malondialdehyde and hydrogen peroxide are significantly reduced, thereby effectively alleviating the oxidative damage caused by saline-alkali stress. At the same time, it is found that the pink spiral sporae can increase the microbial diversity of the root system soil and improve the abundance of beneficial microorganisms, thereby improving the soil health index.

[0021] It is also found that after applying the XRLZ-13-1 bacterial solution, the contents of available nitrogen and organic matter in the soil are increased by 14.1% and 37.3%, respectively, which shows a significant difference (P<0.05) compared with the saline-alkali soil. P < 0.05). The use of the pink spiral sporae compound soil conditioner can significantly improve the contents of available nitrogen, phosphorus, potassium, and organic matter in the soil, and also can reduce the soil pH value to a certain extent (P<0.05). It is shown that the pink spiral sporae microbial fertilizer can effectively improve the soil nutrition index and alleviate the saline-alkali stress. P

[0022] The pink spiral sporae strain and the microbial fertilizer provided by the present application not only can improve the tolerance of tomato to saline-alkali stress, but also can improve the nutrient absorption efficiency of crops and promote the growth of plants. It also has potential growth-promoting and stress-resistant effects on other crops (such as cucumber and lettuce). At the same time, the strain and the technology can also be used for saline-alkali land improvement and management, help to restore soil health, improve the ecological environment, and also help to alleviate the problem of insufficient arable land. The strain and the application method provided by the present application are safe, environmentally friendly, and pollution-free, which provides a new and sustainable solution for the ecological restoration of saline-alkali land and the healthy cultivation of crops, and has significant economic and ecological benefits. BRIEF DESCRIPTION OF DRAWINGS

[0023] Figure 1 ​The colony morphology of pink spiral sporulating fungus XRLZ-13-1 strain on PDA medium. A is the front of the colony; B is the back of the colony.

[0024] Figure 2 The ITS rDNA sequence homology tree of pink spiral sporulating fungus XRLZ-13-1 strain.

[0025] Figure 3 The growth diameters of different pink spiral sporulating fungus strains on PDA medium with added salt-alkali.

[0026] Figure 4 The effects of seed soaking of different pink spiral sporulating fungus strains on tomato seed germination under salt-alkali stress conditions.

[0027] Figure 5 The effects of pink spiral sporulating fungus strain fermentation liquor on tomato test tube seedling growth under salt-alkali solution hydroponics. From left to right in the figure are water treatment (no salt-alkali stress), salt-alkali solution control, pink spiral sporulating fungus XRLZ-13-1, XRDJ-1-2 and XRNM-1-1 strain treatments.

[0028] Figure 6 The effects of pink spiral sporulating fungus strain on tomato plant growth in salt-alkali soil. A is the plant growth state; B is the root development. From left to right in the figure are normal cultivated soil treatment (no salt-alkali stress), salt-alkali soil control, pink spiral sporulating fungus XRLZ-13-1, XRDJ-1-2 and XRNM-1-1 strain treatments.

[0029] Figure 7 The effects of pink spiral sporulating fungus XRLZ-13-1 fermentation liquor on the nutritional indicators in tomato leaves. A is the chlorophyll content; B is the nitrogen content. From left to right in the figure are cultivated soil (no stress state), salt-alkali soil control, salt-alkali soil + pink spiral sporulating fungus XRLZ-13-1 strain treatment.

[0030] Figure 8 The effects of pink spiral sporulating fungus XRLZ-13-1 fermentation liquor on the antioxidant enzyme activity and peroxidation products in tomato roots. A is the activity of superoxide dismutase (SOD); B is the activity of catalase (CAT); C is the content of malondialdehyde (MDA); D is the content of hydrogen peroxide (H2O2). From left to right in each figure are cultivated soil (no stress state), salt-alkali soil control, salt-alkali soil + pink spiral sporulating fungus XRLZ-13-1 strain treatment.

[0031] Figure 9The effects of fermentation broth of *Polyspora pulvinata* XRLZ-13-1 on the rhizosphere microbiota of tomato under salt-alkali stress. A represents bacterial composition (phylum level); B represents bacterial composition (genus level); C represents fungal composition (phylum level); D represents fungal composition (genus level).

[0032] Figure 10 This study illustrates the effects of different *Alternaria pinkis* compound microbial fertilizers on tomato growth under saline-alkali stress. From left to right in the figure, the fertilizers are: saline-alkali soil control, *Alternaria pinkis* XRLZ-13-1 fermentation broth, XRLZ-13-1 fermentation broth + fly ash (X+FA), XRLZ-13-1 fermentation broth + humic acid (X+HA), and XRLZ-13-1 fermentation broth + biochar (X+BC). Detailed Implementation

[0033] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention.

[0034] Unless otherwise specified, all test materials and instruments used in this instruction manual are commercially available.

[0035] Unless otherwise specified, all parts in the examples are by weight, and all percentages are by weight percentages.

[0036] The *Polyporus pulcherrimus* strain used in this invention was isolated from farmland soil in Resa Township, Lazi County, Shigatse Prefecture, Tibet Autonomous Region by our research team and is labeled as strain XRLZ-13-1. The strain has the CGMCC No. 42331 accession number, was deposited on December 1, 2025, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences. It is classified and named *Polyporus pulcherrimus*. Clonostachys rosea .

[0037] Example 1: Cultivation of Sclerotium sclerotia The preserved Sclerotium sclerotiorum (accession number ACCC 39161) was inoculated onto a PDA plate and incubated in a 28°C incubator for 7-10 days. The mycelial cakes were then punched out and inoculated into carrot agar, with 5-10 cakes per bottle. The plates were incubated at room temperature under light for 20 days. Once the agar was covered with a large number of sclerotia, the carrot agar was rinsed off the surface of the sclerotia with sterile water. The plates were then air-dried and stored in a cool place for later use.

[0038] Example 2: Isolation of *Alternaria pinki* strains from Tibet The crop rhizosphere soil was collected from six regions of Tibet (Ali region, Chamdo, Lhasa, Nyingchi, Naqu, and Shigatse), dried, passed through a 2 mm sieve, and 100 g of the soil sample was weighed and placed in a sterilized glass culture bottle. Twelve fresh sclerotia prepared in Example 1 were buried in each bottle (about 2 cm deep) to act as baits for attracting fungi with parasitic functions on other fungi. The culture bottles were placed in a constant temperature incubator and incubated at 28°C with a humidity of 60% to 70%. After 30 days, the sclerotia were removed from the soil sample, washed with tap water, disinfected with 1% NaClO for 30 seconds, washed with sterile water three times, excess water was removed, and then the sclerotia were placed in a sterilized culture dish with three layers of filter paper and incubated at 25°C with moisture. After 7 days, the surface of the sclerotia was observed for the presence of mycelium growth. The mycelium and spores that grew were streaked onto PDA medium, and single colonies were picked for purification. The isolated parasitic fungi were stored in a 4°C refrigerator.

[0039] From the 90 collected soil samples, 50 strains of H. roseogrisea were isolated, including 2 strains from the Ali region, 6 strains from Chamdo, 3 strains from Nyingchi, 4 strains from Naqu, and 35 strains from Shigatse. Figure 1 A colony morphology diagram of the XRLZ-13-1 strain on PDA medium is shown in FIG. 1, where A is the front of the colony and B is the back of the colony.

[0040] Example 3 Identification of the parasitic fungus XRLZ-13-1 The isolated parasitic fungi were identified by colony morphology, microscopic structure, and ITS sequencing analysis. The identification process for the XRLZ-13-1 strain is as follows: Morphological identification: The isolated XRLZ-13-1 strain was inoculated into PDA medium and incubated at 26°C for 7 days, and the colony morphology is shown in FIG. 1. Figure 1 The hyphae and conidia were picked and observed under a microscope for the characteristics of the conidial phialides and conidia. The hyphae were colorless and septate, the conidial phialides were broom-like branches, a large number of conidia were produced at the top of the phialides, the spores were oval or cylindrical with a diameter of 3-5 μm. Based on the morphological characteristics, the XRLZ-13-1 strain was initially identified as H. roseogrisea.

[0041] Molecular identification: Hyphae and conidia of strain XRLZ-13-1 were collected, and genomic DNA was extracted using the Ezup column-based fungal genomic DNA extraction kit. PCR amplification was performed using universal fungal primers ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3'). The amplification system consisted of 2 μL DNA template, 1 μL each of the 10 μM ITS primers, and 21 μL of 1.1×T3 Super PCR Mix enzyme. The amplification program was: 98℃ pre-denaturation for 3 min, 98℃ denaturation for 10 s, 55℃ annealing for 10 s, 72℃ extension for 10 s, for 35 cycles; followed by a 10 min extension at 72℃. After electrophoresis, the PCR products were sent to Beijing Qingke Biotechnology Co., Ltd. for sequencing. The obtained sequences were compared for homology using BLAST in the NCBI database, and a phylogenetic tree was constructed using the neighbor-joining method. The results showed that strain XRLZ-13-1 had the highest homology with *Polyspora pinkis*, and the homology comparison phylogenetic tree was as follows: Figure 2 As shown.

[0042] Example 4: Determination of the salt and alkali tolerance of *Polyspora pinkis* strains A salt- and alkali-stressed medium was prepared by adding 5% NaCl to PDA medium and adjusting the pH to 9.0 with 10% NaOH solution. The isolated *Polyspora pinkis* strain was inoculated onto PDA plates and incubated at 28°C for 5 days. Then, using a sterile punch, 5 mm diameter mycelial discs were taken from the edge of the colony and inoculated into the center of both a fresh, ordinary PDA plate (control) and a salt- and alkali-stressed PDA plate. The plates were incubated at 28°C for 7 days, and the colony diameter was measured. The relative growth rate (RGR) of each strain was calculated.

[0043] Relative growth rate = Colony diameter on saline-alkali medium / Colony diameter on PDA medium × 100%.

[0044] The results showed that the relative growth rates of different *Alternaria pinki* strains under salt-alkali stress ranged from 7.4% to 83.8%, exhibiting significant differences. P < 0.05 indicates rich genetic diversity within the *Alternaria pinki* species. Some strains exhibit high tolerance, with growth capacity under stress conditions exceeding or equaling half that under normal culture conditions (RGR > 50%). For example, the RGR value of strain XRLZ-13-1 is 83.8%, significantly higher than other tested strains ( P< 0.05), in addition, the RGR value of XCBS-2-1 strain was 55.5%, which also showed good salt-alkali tolerance. Most of the strains (accounting for 82%) showed moderate tolerance, with RGR values ranging from 20% to 50%. Some strains, such as XRRB-4-1 and XRSJ-2-3, showed high sensitivity to salt-alkali conditions, and their growth was severely inhibited by salt-alkali stress.

[0045] Figure 3 The growth and morphological differences of representative strains of P. roseum on PDA medium and salt-alkali medium are intuitively displayed. High-tolerance strains can still grow rapidly and form dense colonies under stress conditions, while sensitive strains grow slowly and have sparse mycelium. Through growth determination, 13 salt-alkali tolerant strains, including XRLZ-13-1, XCBS-2-1, XRXT-7-2 and XRDJ-1-2, were selected from 50 P. roseum strains for subsequent study on the effects on seed germination and hydroponic seedling growth under salt-alkali stress.

[0046] Example 5 Fermentation culture of P. roseum XRLZ-13-1 The P. roseum XRLZ-13-1 strain was inoculated on PDA and cultured for 7 days, 5 mL sterile water was added to elute the spores, and a spore suspension was prepared. PDB medium (100 mL / 500 mL flask) was prepared, high-pressure wet heat sterilized, and when cooled to room temperature, the spore suspension was inoculated into the medium and placed on a shaker at 27°C for 72 h of dark culture to prepare a seed liquid.

[0047] The fermentation medium was prepared, containing 25 g of sucrose, 4 g of yeast extract powder, 6 g of soybean meal powder, 1.0 g of K2HPO4, 0.5 g of MgSO4·7H2O, and 0.05 g of FeSO4·7H2O per liter of medium, with a pH value of 6.0. The fermentation medium was divided into flasks and sterilized at 121°C for 30 min, then cooled to room temperature. The seed liquid was inoculated into the fermentation medium at a 3% inoculation amount, and cultured at 27°C with 180 r / min shaking for 4 days. The concentration of the strain fermentation liquid was 1.6×10 8 spores / mL.

[0048] Example 6 Effect of P. roseum on tomato seed germination under salt-alkali stress The salt-alkali tolerant P. roseum strains selected as described above were inoculated in the fermentation medium and cultured at 28°C with 180 r / min for 72 h to prepare the fermentation liquid. The concentration of the fermentation liquid was adjusted to 1×10 7Spores / mL. At the same time, a composite saline-alkali solution was prepared to simulate a saline-alkali stress environment. The saline-alkali solution contained NaCl, NaHCO3, Na2SO4, and Na2CO3, with a mass ratio of 1:9:9:1, and a total concentration of 50 mmol / L. Full and uniform tomato seeds were selected, surface-sterilized with 1% sodium hypochlorite solution for 5 min, and then washed with sterile distilled water for 3 times. The seeds were placed in 9 cm culture dishes lined with double filter paper, with 25 seeds per dish. 3 mL of the corresponding saline-alkali solution and 1 mL of the pink spiral polyangium fermentation broth were added to the culture dishes, and sterile water was used to replace the bacterial fermentation broth as a saline-alkali stress control. The culture dishes were placed in a 25°C constant temperature incubator in the dark, and the water loss was replenished quantitatively every day. The number of germinated seeds was observed and recorded every day, and the length of the radicle was taken as the criterion for germination. The germination rate was calculated and the ability of each strain to alleviate saline-alkali stress was evaluated by continuous measurement for 7 days. Each treatment was repeated 3 times.

[0049] The results are shown in Figure 4 After the addition of the saline-alkali solution, the germination of tomato seeds was strongly inhibited, while the addition of the pink spiral polyangium fermentation broth effectively alleviated this inhibition. Under non-stress conditions (water soaking), the germination rate of the seeds reached 100% after 7 days, while under saline-alkali stress (saline-alkali control), the germination rate of the seeds was only 28.7%. In the pink spiral polyangium broth treatment, the XRLZ-13-1 strain had the most outstanding effect, with a tomato seed germination rate of 78.0%, which was 2.1 times higher than under stress conditions, and the germination rate increased, showing a sustained and rapid germination trend from the 2nd day. Most of the tested pink spiral polyangium strains also showed significant alleviation of saline-alkali stress. For example, after treatment with strains XAPL-13-1, XRDJ-1-2, XRNM-1-1, and XRBL-1-1, the germination rate of tomato seeds was more than 60%, which was significantly higher than that of the stress control.

[0050] Example 7 Effect of pink spiral polyangium fermentation broth on the growth of hydroponic tomato test tube seedlings under saline-alkali stress Tomato seeds with full and uniform size were selected, and the surface was sterilized with 1% sodium hypochlorite solution for 5 min, and then washed with sterile distilled water for 3 times. The seeds were placed in a tissue culture plate for seedling culture, and when the tomato seedlings grew to two leaves with one heart, the seedlings with uniform growth were selected and placed in 15 mL test tubes. 9 mL of 1 / 8 MS culture solution (1.9 g of potassium nitrate, 1.65 g of ammonium nitrate, 0.17 g of potassium dihydrogen phosphate, 0.37 g of magnesium sulfate, 0.44 g of calcium chloride, 0.83 mg of potassium iodide, 6.2 mg of boric acid, 22.3 mg of manganese sulfate, 8.6 mg of zinc sulfate, 0.25 mg of sodium molybdate, 0.025 mg of copper sulfate, 0.5 mg of cobalt chloride, 0.5 mg of nicotinic acid, 37.3 mg of disodium ethylenediaminetetraacetate, 27.8 mg of ferrous sulfate, 100 mg of inositol, 2 mg of glycine, 0.1 mg of thiamine hydrochloride, and 0.5 mg of pyridoxal hydrochloride per liter of solution) and 1 mL of saline-alkali solution (NaCl, NaHCO3, Na2SO4, and Na2CO3, mass ratio of 1:9:9:1, total concentration of 500 mmol / L) were added to each test tube, and then 1 mL of the pink spiral sporangia fermentation liquid with a concentration of 10 7 Spores / mL prepared by the foregoing method were added, and the test tubes were cultured indoors under illumination, with the same amount of 1 / 8 MS culture solution being added as a control. Sterile distilled water was added daily to keep the liquid level in the test tubes consistent. After 7 days, the root length, fresh weight, dry weight, and chlorophyll content of the tomato were determined. There were 6 seedlings in each repeat, and each treatment was repeated 3 times.

[0051] From Figure 5 It can be seen that the growth of tomato seedlings was seriously disturbed under saline-alkali stress, and the plants showed wilting phenomenon. After inoculation of pink spiral sporangia XRLZ-13-1, XRNM-1-1, and XRDJ-1-2, the growth of the seedlings was obviously improved. Among them, the dry weight of the aboveground part and the root of the tomato seedlings treated with XRLZ-13-1 fermentation liquid increased by 1.5 times and 6.5 times respectively compared with the stress control. After inoculation of XRLZ-13-1 and XRNM-1-1, the chlorophyll content of the leaves increased by 3.7 times and 3.5 times respectively compared with the control, and even increased by 19.4% and 15.7% respectively compared with the non-stress condition.

[0052] Example 8 Influence of pink spiral sporangia on tomato growth and antioxidant enzymes and peroxide products in saline-alkali soil Preparation of saline-alkali soil: The surface soil (0-20 cm depth) was collected from the Langfang experimental base of Chinese Academy of Agricultural Sciences, air-dried, passed through a 2 mm sieve, and mixed with 1 g of each of NaCl, Na2SO4, Na2CO3, and NaHCO3 (analytical grade) at a ratio of 0.4% per kg of soil to prepare the saline-alkali soil. The prepared soil sample was measured according to the national standard to determine the conductivity (EC) and pH value, with initial values of EC = 22.5 dS / m and pH = 8.51.

[0053] Greenhouse potting of tomatoes, determination of the response of tomato plants to saline-alkali stress after treatment with Pink Actinomycete XRLZ-13-1. Normal cultivated soil, saline-alkali soil, and saline-alkali soil with Pink Actinomycete liquid (2 mL / pot, fermentation broth prepared in Example 5, concentration of 1.6 x 10 8 Spores / mL, 300 g of soil per pot) were set up. After 10 days, the growth of tomatoes in different treatments was as shown in Figure 6 After treatment with Pink Actinomycete fermentation broth, tomato seedlings were not wilted, the stems were strong, and the leaves were dark green. In particular, after the addition of XRLZ-13-1 fermentation broth, the growth of tomatoes was not different from that in normal cultivated soil. The chlorophyll content and nitrogen content of the leaves were determined by a plant nutrition detector, and the results are shown in Figure 7 After treatment with the broth, the chlorophyll and nitrogen contents were increased by 64.1% and 63.2%, respectively, compared with the saline-alkali soil control.

[0054] Tomato roots grown for 10 days were taken, the soil clumps were washed away, and they were quickly frozen in liquid nitrogen and ground for use. The activities of superoxide dismutase (SOD) and catalase (CAT) and the contents of peroxidation products malondialdehyde (MDA) and hydrogen peroxide (H2O2) were determined using a kit according to the instructions. As shown in Figure 8 Saline-alkali stress (saline-alkali soil in the figure) led to an increase in the activities of antioxidant enzymes and accumulation of peroxidation products in the roots. Due to the activation of the plant antioxidant system by saline-alkali stress, the activities of SOD and CAT in the tomato were increased by 1-fold and 3.9-fold, respectively, compared with normal soil, and the contents of peroxidation products MDA and H2O2 were also significantly accumulated, with increases of 0.8-fold and 1.1-fold, respectively. After the application of XRLZ-13-1 microbial agent, the activities of antioxidant enzymes increased to varying degrees, among which the CAT activity had a significant difference, with an increase of 1.2-fold. The contents of MDA and H2O2 were reduced by 31.6% and 39.4%, respectively, compared with the saline-alkali control, close to those in normal soil.

[0055] Example 9 Effect on the rhizosphere microbial community of tomatoes Tomatoes were grown in a greenhouse (planting method as in Example 8). Soil samples were collected from both untreated (CK) and treated (XRLZ-13-1) soils. High-throughput sequencing was used to analyze the microbial community composition of the soil bacteria and fungi, and Mothurv 1.30.1 was used to calculate various α-diversity indices. The results showed that the application of XRLZ-13-1 had no significant effect on the α-diversity of the soil bacterial community, but significantly increased the abundance of the fungal community. The Sobs, Ace, and Chao indices increased from 455.7, 467.4, and 485.4 to 541.3, 562.4, and 558.9, respectively, with the increases in Ace and Chao indices reaching highly significant levels. P < 0.001). Microbial classification, sequencing, and gene function annotation cluster analysis of soils under different treatments showed that the microbial flora composition in saline-alkali soils changed after the application of XRLZ-13-1 bacterial solution. The pink spiral polycystic spp. agent increased the relative abundance of beneficial microbial groups, such as *Sphingomonas* and *Microbranchia* (e.g., ...). Figure 9 (As shown). Sphingomonas can produce catalase under saline-alkali conditions, and can play a role in nitrogen fixation and promote carbon cycling under high salinity conditions. Microbranchs help improve the bioavailability of soil phosphorus.

[0056] Example 10 Preparation of Pink Spiral Polyporus Microbial Fertilizer Biochar was selected as a soil conditioner and compounded with *Polyspora pinkis* to prepare microbial fertilizer. The fermentation broth of *Polyspora pinkis* XRLZ-13-1 obtained in Example 5 was placed in a low-speed centrifuge and centrifuged at 4000 r / min for 10 min. The supernatant was discarded, and the bacterial sludge was collected. The moisture content of the bacterial sludge was determined to be 58%. Diatomaceous earth and peat moss (diatomaceous earth to peat moss mass ratio 1:1) were added to the bacterial sludge at a 1:1 mass ratio and mixed thoroughly until the material was homogeneous. The resulting mixture was placed in a clean enamel dish and allowed to dry naturally in a cool, ventilated place until constant weight. It was then pulverized to prepare the microbial inoculant.

[0057] The microbial agent and biochar were then mixed at a mass ratio of 1:2 and stirred thoroughly to prepare a compound microbial fertilizer. The prepared sample was a dark brown powder with a loose texture and good adsorption properties, suitable for field trials.

[0058] Weigh 1.0 g of the prepared microbial fertilizer, add 9 mL of sterile water and shake. Perform serial dilutions, and spread 100 μL onto a PDA plate containing 50 mg / L penicillin and 50 mg / L streptomycin sulfate. Incubate at 28℃ for 3 days and count the components. Perform three replicates. The content of the active ingredient in the microbial fertilizer was determined to be 4.2 × 10⁻⁶. 7 CFU / g.

[0059] Example 11: Effects of Pink Spiral Polyporus Microbial Fertilizer on Tomato Growth The fermentation broth of S. goshikiensis XRLZ-13-1 was prepared according to the method described in Example 5, and the concentration was 1.4 x 10 8 Meanwhile, the saline-alkali soil was prepared according to the method described in Example 8, 300 g of soil was put in each pot, and the application effect of the S. goshikiensis microbial fertilizer on tomatoes was comprehensively evaluated by a pot experiment. A total of 5 treatments were set: (1) saline-alkali soil planting (CK); (2) adding 2 mL of the fermentation broth of S. goshikiensis XRLZ-13-1 (XRLZ); (3) the fermentation broth of the XRLZ-13-1 strain + fly ash (X+FA, mass ratio 1:3); (4) the fermentation broth of the XRLZ-13-1 strain + humic acid (X+HA, mass ratio 1:3); (5) the fermentation broth of the XRLZ-13-1 strain + biochar (X+BC, mass ratio 1:3).

[0060] Seedling was raised in a seedling tray, and when it grew to two leaves and one heart, it was transferred to a flowerpot of different treatment, with 1 plant per pot. An independent tray was placed at the bottom of each flowerpot, and the water seeping out of the tray was poured back into the pot, so as to ensure that the concentrations of various components in the soil of the pot were unchanged. The plant height, root length, fresh weight and dry weight of the tomato were determined 10 days after planting. There were 15 plants in each treatment, and 3 replicates.

[0061] The growth of tomatoes was as shown in Figure 10 There was a significant difference (P<0.05) in the effect of different treatments on the growth of tomatoes under saline-alkali stress. P Compared with the control (CK), the application of the fermentation broth of S. goshikiensis XRLZ-13-1 (XRLZ) greatly alleviated the inhibitory effect of saline-alkali stress on the growth of tomatoes, and the root length and biomass were increased by more than 40%, especially the aboveground dry weight was increased by 3.2 times. After applying the compound microbial fertilizer, especially after the treatment of S. goshikiensis + biochar (X+BC), the growth-promoting effect was further improved, and the aboveground and root biomass was increased by more than 50%, showing obvious synergistic effect, indicating that the application of the compound microbial fertilizer of S. goshikiensis and biochar could effectively improve the growth vigor and stress resistance of plants under saline-alkali stress, and improve the rhizosphere environment.

[0062] Example 12 Effect of S. goshikiensis microbial fertilizer on soil physical and chemical indexes The saline-alkali soil was prepared, and the S. goshikiensis microbial fertilizer prepared in Example 11 was used for a greenhouse pot experiment. A total of 3 treatments were set: (1) saline-alkali soil planting (CK); (2) the fermentation broth of S. goshikiensis XRLZ-13-1 treatment (XRLZ); (3) XRLZ-13-1 fermentation broth + biochar treatment (X+BC).

[0063] The rhizosphere soil samples of transplanted tomatoes were collected after 10 days and stored at -80℃. The physicochemical properties of the soil were determined according to the national standard. A certain amount of soil sample was weighed, diluted 5 times and 2.5 times with water, respectively, and then the soil conductivity and pH value were determined. The total nitrogen was determined by an elemental analyzer, the available nitrogen was determined by alkali hydrolysis diffusion method, the total phosphorus and available phosphorus were determined by molybdenum-antimony anti-colorimetric method, the total potassium and available potassium were determined by flame photometric method, and the soil organic matter was determined by potassium dichromate oxidation-titration method.

[0064] The results of the determination of the physicochemical indexes of the soil sample are shown in Table 1. After applying the XRLZ-13-1 bacterial liquid, the contents of the available nitrogen and the organic matter in the soil were increased by 14.1% and 37.3% respectively compared with the saline-alkali soil control, which showed a significant difference (P<0.05). P < 0.05). The microbial fertilizer prepared by compounding the XRLZ-13-1 strain and the biochar also increased the contents of the total phosphorus, the available phosphorus and the available potassium, played a synergistic effect, and significantly reduced the soil pH, which indicated that the application of the pink spiral polysporal microbial fertilizer could effectively improve the soil nutrition indexes and relieve the saline-alkali stress.

[0065] Table 1 Influence of pink spiral polysporal microbial fertilizer on the physicochemical indexes of soil

[0066] Note: TN: total nitrogen; TP: total phosphorus; TK: total potassium; AN: available nitrogen; AP: available phosphorus; AK: available potassium; SOM: soil organic matter.

[0067] Although the present application is described above through examples, those skilled in the art should understand that, under the premise of not deviating from the spirit and essence of the present application, the improvements and modifications made to the present application shall all belong to the protection scope of the present application.

Claims

1. A halophilic growth-promoting strain of Pink Trichothecium, characterized by, CGMCC No. 42331.

2. The pink spiral polyangium strain of claim 1 is applied in saline-alkali soil crop cultivation and saline-alkali soil improvement.

3. The microbial fertilizer comprising the strain of pink spirogyra as claimed in claim 1, characterized in that, The microbial fertilizer is a fermentation broth of the pink spiral polyangium strain obtained by liquid fermentation, or is obtained by mixing the fermentation broth with auxiliary materials and / or soil improvers and then preparing a preparation; the dosage form of the microbial fertilizer is a liquid agent, a granular agent or a powder agent.

4. The microbial fertilizer according to claim 3, wherein the strain of Sporomusa pinkii is characterized in that, The fermentation broth is filtered or concentrated by centrifugation, and then auxiliary materials and / or soil improvers are added to prepare a preparation; The soil improver is one or more of straw powder, biochar, humic acid, fly ash, organic fertilizer, gypsum, desulfurization gypsum, silicon fertilizer, wood ash, potassium fulvic acid; The auxiliary material is one or more of diatomite, light calcium carbonate, talc powder, attapulgite, bentonite, kaolin, starch, dextrin, grass carbon, rice husk powder.

5. The microbial fertilizer according to claim 3, characterized in that, The fermentation broth of the pink spiral polyangium strain is prepared by the following steps: 1) Inoculum preparation: the pink spiral polyangium strain is inoculated on potato dextrose agar medium, and cultured at 26-28°C for 6-10 days, and spores are prepared into a spore suspension by eluting the spores; or a mycelial block is cut as an inoculum; 2) Seed culture: the spore suspension or mycelial block is inoculated in PDB medium, and cultured at 25-28°C in the dark with shaking for 48-84 h to obtain a seed liquid; 3) Liquid fermentation culture: a fermentation medium is prepared, sterilized and cooled, and then the seed liquid is inoculated, and cultured at 25-30°C for 48-120 h to obtain a fermentation broth.

6. The microbial fertilizer according to claim 5, characterized in that, In step 3), the components of the fermentation medium per liter include: sucrose 10-40 g, yeast extract powder 5-15 g, soybean meal powder 5-15 g, K2HPO4 0.5-2 g, MgSO4·7H2O 0.5-1.0 g, FeSO4·7H2O 0.05-0.1 g; the pH value is 5.5-6.

5.

7. The microbial fertilizer according to claim 5, characterized in that, In step 3), the liquid fermentation culture conditions are: seed liquid inoculation amount 0.5-5%, rotation speed 150-250 r / min, culture temperature 25-28°C, culture time 3-4 days.

8. The microbial fertilizer of the strain of S. gossypii according to any one of claims 3 to 7, characterized in that, After the fermentation broth is obtained, the plate and frame filtration or centrifugal concentration is carried out to obtain a mycelium with a water content of 50-70%, auxiliary materials are added to the mycelium, or the auxiliary materials are directly added to the fermentation broth; after mixing and drying, the mixture is crushed or granulated; the addition amount of the auxiliary materials is 50-120% of the mass percentage of the mycelium, or 80-150% of the mass percentage of the fermentation broth.

9. The microbial fertilizer of the strain of S. gossypii according to any one of claims 3 to 7, characterized in that, The soil improver is added to the microbial fertilizer, and the mass ratio of the addition is 1:1-10 of the fermentation broth to the soil improver, or the fermentation broth is filtered or concentrated by centrifugation to obtain a mycelium with a water content of 50-70%, and the mass ratio of the mycelium to the soil improver is 1:0.5-5.

10. The microbial fertilizer according to claim 9, characterized in that, The soil conditioner is one of fly ash, humic acid, biochar, organic fertilizer, gypsum, desulfurization gypsum, silicon fertilizer, wood ash, potassium fulvic acid and straw powder, and the bacterial content is 0.5-5×10 8 The mass ratio of the fermentation liquor containing spores of 0.5-5×10

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