Streptomyces Qhu-M48 and application thereof in plant salt resistance

Abstract

The invention relates to the technical field of microorganisms, in particular to streptomyces Qhu-M48 and application thereof in plant salt resistance. The preservation number of the streptomyces Qhu-M48 is CGMCC (China General Microbiological Culture Collection Center) No.4.8023; tests prove that the strain can play an obvious role in promoting plant growth, and can also obviously promote plant growth under salt stress, so that the strain has an application prospect in the aspects of improving plant stress resistance and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, and in particular to Streptomyces Qhu-M48 and application thereof in plant salt resistance. Background Art

[0002] In natural environments, plant growth and development are often adversely affected by biotic and abiotic stresses, limiting crop yields. Climate change and agronomic practices such as improper irrigation and fertilization have led to increased soil salinization, causing significant losses to crop production, especially in arid and semi-arid regions. Most crops are highly susceptible to salinized soils. Plants grown under high salt concentrations have a higher intracellular Na + and Cl - The accumulation of salt can cause ion imbalance and induce hyperosmotic stress, leading to metabolic disorders and reduced photosynthesis. Their development is restricted by the osmotic effects of salt in the soil or the toxic effects of salt absorption by the plant. Although genetic improvement and molecular breeding can improve the resistance of these crops to salinization stress, these methods are often time-consuming and costly. Therefore, there is an urgent need to develop innovative strategies to effectively improve their salt tolerance and stress resistance.

[0003] Studies of plant-microbe interactions have shown that environmental stresses induce changes in the diversity of the plant microbiome, particularly the root-associated microbiome. This allows plants to selectively recruit specific beneficial microbes through root metabolites, which in turn promote nitrogen fixation, enhance nutrient uptake, improve soil properties, and enhance plant stress tolerance, thereby directly or indirectly improving plant growth and stress tolerance. This suggests that root-associated microbes are potential targets for improving plant tolerance to environmental stress. Plant growth-promoting rhizobacteria are bacteria that colonize the rhizosphere, or roots, and promote growth, nutrient uptake, and systemic stress resistance in their host plants. Increasing evidence suggests that exploiting the interactions between plant growth-promoting rhizobacteria and their hosts is an effective strategy for improving plant salt tolerance. As a cost-effective biotechnology tool, plant growth-promoting rhizobacteria have shown great potential for enhancing plant adaptability to adverse environmental conditions such as salinity. Streptomyces, among others, can produce a variety of bioactive secondary metabolites, including antimicrobial agents, phytohormones, and siderophores, establishing beneficial interactions with plants and endowing them with properties that promote growth and protect against environmental stress.

[0004] Therefore, developing new plant growth-promoting Streptomyces and applying them to forage cultivation in saline soils has become an important way to improve the quality of saline-alkali soil and increase the efficiency of agricultural and animal husbandry production. Summary of the Invention

[0005] The present invention aims to provide a Streptomyces sp. Qhu-M48, which can promote the growth of forage grass and, in addition, can reduce the impact of extreme environments on plants and increase the stress resistance of plants.

[0006] In a first aspect, the present invention provides a Streptomyces Qhu-M48 strain, with a deposit number of CGMCC No. 4.8023.

[0007] The strain Qhu-M48 was isolated from a soil sample of an alpine meadow on the Qinghai-Tibet Plateau.

[0008] In a second aspect, the present invention provides a use of Streptomyces Qhu-M48 in promoting plant growth.

[0009] In the present invention, the Streptomyces Qhu-M48 can promote seed germination and / or the growth of one or more of roots, stems, leaves, or flowers and fruits. For example, it can promote or inhibit root length, stem diameter, plant height, leaf width, leaf length, leaf number, leaf area, biomass, water content, chlorophyll content, yield, photosynthesis, and the like.

[0010] In a third aspect, the present invention provides use of Streptomyces Qhu-M48 in regulating plant stress resistance.

[0011] Plant stress resistance refers to certain traits that plants possess to resist adverse environments, such as cold resistance, drought resistance, saline-alkali resistance, and high temperature resistance.

[0012] In the present invention, the stress resistance includes but is not limited to saline-alkali resistance, cold resistance, drought resistance, and high temperature resistance.

[0013] In some specific embodiments of the present invention, the stress resistance is salt resistance.

[0014] In the present invention, Streptomyces Qhu-M48 can regulate the activity of antioxidant enzymes and the level of osmotic regulating substances to reduce the oxidative damage and osmotic regulation disorder caused by salt stress, thereby alleviating the salt stress damage suffered by plants.

[0015] In the present invention, when used, a single agent of Streptomyces Qhu-M48 or the agricultural product prepared therefrom is used to treat seeds, spray on leaves or irrigate roots.

[0016] In the present invention, the plants include plants for agriculture and animal husbandry.

[0017] In some embodiments of the present invention, the plant is selected from grasses of the Poaceae family, grasses of the Leguminosae family, grasses of the Asteraceae family, grasses of the Chenopodiaceae family, grasses of the Cyperaceae family, grasses of the Polygonaceae family, or grasses of the Cruciferae family.

[0018] In some embodiments of the present invention, Streptomyces Qhu-M48 is used to promote the growth of forage plants, particularly for plateau forage capacity.

[0019] In a fourth aspect, the present invention provides a microbial preparation containing the aforementioned Streptomyces Qhu-M48.

[0020] Beneficial effects of the present invention:

[0021] (1) The Streptomyces Qhu-M48 provided by the present invention can effectively promote plant seed germination and the growth of organs such as roots, stems, and leaves, increase biomass and photosynthesis efficiency, and can maintain normal growth and development of plants, especially under salt stress.

[0022] (2) The Streptomyces Qhu-M48 provided by the present invention significantly reduces the oxidative damage and osmotic imbalance caused by salt stress by regulating the activities of antioxidant enzymes (such as CAT, SOD, and POD) and the levels of osmotic regulating substances (such as proline and soluble sugars), thereby improving the salt tolerance of plants.

[0023] (3) Streptomyces Qhu-M48 has growth-promoting and stress-resistant effects on a variety of forage grasses such as Gramineae and Leguminosae. It has a wide range of applications and can be used for saline-alkali land improvement and agricultural and animal husbandry production.

[0024] The following abbreviations have the following meanings:

[0025] MDA is malondialdehyde;

[0026] SOD is superoxide dismutase;

[0027] POD is peroxidase;

[0028] CAT stands for catalase. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 : Phylogenetic tree of the 16S rRNA gene of strain Qhu-M48 constructed based on the neighbor-joining method;

[0030] Figure 2 : Qhu-M48 genome map;

[0031] Figure 3 : Morphological characteristics of strain Qhu-M48 grown on different culture media (A−G: ISP1−ISP7 medium; H: Gao's medium No. 1; I: NA medium; J: Czapek's medium);

[0032] Figure 4 : Microstructure of strain Qhu-M48 (A: scanning electron microscopy (10 000×); B: microscopic observation after carbolic acid staining (1 000×));

[0033] Figure 5 : Analysis of phospholipid composition of strain Qhu-M48 (A: ninhydrin staining; B: anisaldehyde staining; C: molybdophosphoric acid staining);

[0034] Figure 6 : Effects of Streptomyces Qhu-M48 on the phenotypes and physiology of oat seedlings under normal culture and salt stress (A: growth phenotype; B: total biomass; C: underground biomass; D: plant height; F: root length; G: relative water content; H: chlorophyll content; I: maximum photochemical efficiency of PSⅡ Fv / Fm);

[0035] Figure 7 : Effects of Qhu-M48 on the phenotypic and physiological parameters of Sainfoin under normal culture and salt stress (A: growth phenotype; B: total biomass; C: underground biomass; D: plant height; F: root length; G: relative water content; H: chlorophyll content; I: maximum photochemical efficiency of PSⅡ Fv / Fm);

[0036] Figure 8 : Effects of Streptomyces Qhu-M48 on the levels of peroxides, antioxidant enzymes, and osmotic regulation substances in Sainfoin seedlings (A: hydrogen peroxide content; B: malondialdehyde content; C: catalase activity; D: superoxide dismutase activity; E: peroxidase activity; E: proline content; F: soluble sugar content). DETAILED DESCRIPTION

[0037] In order to enable those skilled in the art to better understand the present invention, the present invention will be further described in detail below with reference to specific examples. It should be understood by those skilled in the art that this should not be construed as limiting the scope of the claims of the present invention. Unless otherwise defined, the technical and scientific terms used in the following examples have the same meanings as those generally understood by those skilled in the art to which the present invention belongs. It should also be noted that the reagents or instruments in the present invention, unless otherwise specified, are all conventional biochemical reagents or instruments and can be purchased commercially.

[0038] Strain deposit information:

[0039] Streptomyces Qhu-M48 was deposited in the General Microbiology Center of the China Culture Collection Administration on June 16, 2025. The strain deposit number is: CGMCC No. 4.8023; address: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing; the deposit name is: Streptomyces sp.

[0040] Example 1

[0041] 1 Experimental Methods

[0042] 1.1 Research Materials

[0043] Strain Qhu-M48 was isolated from a soil sample from an alpine meadow on the Qinghai-Tibet Plateau (N35°13.4752', E101°28.7727') and stored in 20% glycerol at -80°C at the College of Ecological and Environmental Engineering, Qinghai University. This strain has been deposited with the General Microbiology Center of the China General Culture Collection under the accession number CGMCC No. 4.8023.

[0044] Similar model strain Streptomyces exfoliatus NRRL B-2924 T The test soil was a 1:1:1 mixture of sand, peat, and vermiculite.

[0045] 1.2 Microbial morphological observation and identification

[0046] Seed liquid of the strain was streaked onto ISP1−ISP7, Gao's No. 1, NA, and Cha's medium and cultured at 28°C for 7 days for morphological analysis. Ultrastructure of the strain was observed under a scanning electron microscope after 10 days of culture on ISP2 medium using the insert method. Spore morphology was observed under a light microscope after staining with carbolic acid.

[0047] 1.3 Physiological and biochemical characteristics

[0048] To determine the strain's optimal temperature, pH, and salinity tolerance, 5% of the strain seed solution was inoculated into TSB medium and cultured at the appropriate temperature (5-40°C, with a 5°C gradient) at 180 rpm for 7 days. Cultures were also performed at 28°C and 180 rpm for 7 days at the appropriate pH values ​​(3.0-12.0) and sodium chloride concentrations (0-10%). To investigate the strain's carbon and nitrogen source utilization, 5% of the strain seed solution was inoculated into ISP9 carbon and nitrogen utilization medium and cultured at 28°C and 180 rpm for 7 days. The activities of catalase, urease, cellulose hydrolysis, H2S production, and starch hydrolysis were determined according to the "Rapid Identification and Systematic Classification of Actinomycetes." Further physical and chemical properties were tested according to the instructions of the API ZYM kit.

[0049] 1.4 Chemical classification characteristics analysis

[0050] A 5% seed solution of the strain was inoculated into TSB medium and then cultured at 28°C and 180 rpm for 7 days. Mycelia were then freeze-dried by centrifugation at 4°C and 6000 rpm for 10 min. Chemical taxonomic identification was performed primarily according to the "Rapid Identification and Systematic Classification of Actinomycetes." Whole-cell hydrolyzed sugar and amino acid composition analysis: 50 mg of freeze-dried cells were weighed and added to 100 μL of 0.5 mol / L HCl and 100 μL of 6 mol / L HCl, respectively, at 121°C. Thin-layer chromatography was performed on microcrystalline cellulose plates using a developing system of ethyl acetate:pyridine:glacial acetic acid:water (8:5:1:1.5, volume ratio) and methanol:pyridine:glacial acetic acid:water (5:0.5:0.125:2.5, volume ratio). Phospholipid composition analysis: 100 mg of lyophilized cells were ground and added to 15 mL of methanol in a boiling water bath for 10 min. After cooling, 10 mL of chloroform and an appropriate amount of 2% sodium chloride solution were added until the layers separated. The organic phase was concentrated at 37°C, dissolved in a 2:1 chloroform / methanol solution, and analyzed by two-phase thin-layer chromatography on GF254 silica gel plates (developing system: chloroform:methanol:water = 65:25:4; chloroform:acetic acid:methanol:water = 80:15:12:4, volume ratio). Chemical composition analysis of menaquinones: 100 mg of lyophilized cells were ground and added to 40 mL of a 2:1 chloroform / methanol solution. The mixture was shaken at 180 rpm overnight. The supernatant was dried under vacuum at 40°C and dissolved in 0.5 mL of acetone. The layer was developed on a GF254 silica gel plate with n-hexane:ether = 85:15 (volume ratio). The relative mobility R was measured by scraping under 254 nm UV light. f =0.8 dark brown band, after adding 0.5 mL of acetone, filtered through a 0.22 μm microporous filter and then analyzed by LC-MS / MS. In addition, the polar lipid composition was determined using a fully automated bacterial identification system.

[0051] 1.5 Molecular Biology Analysis

[0052] A 5% seed solution of the strain was cultured in TSB medium at 28°C and 180 rpm for 3 days. The cells were then harvested by centrifugation at 4°C and 6,000 rpm for 10 minutes, washed three times with sterile water, and resuspended in 0.5 mL of sterile water. DNA was extracted using a nucleic acid protein extractor at 6.5 M / s, 20 s fragmentation, and 20 s rest. This extraction was repeated three times, followed by centrifugation at 4°C and 12,000 rpm for 5 minutes. The supernatant was used as a PCR template. The 16S rRNA gene sequence was amplified using the universal bacterial primers 27F (5′-AGAGTTTGATCCTGGCTCAG-3′) and 1492R (5′-TACGGCTACCTTGTTACGACTT-3′). The PCR reaction system (25 μL) included 0.5 μL of each upstream and downstream primer (10 μmol / L), 12.5 μL of PCR high-fidelity polymerase (0.05 U / μL), 2 μL of template (56 ng / μL), and 9.5 μL of sterile ddH2O. PCR reaction conditions were: 95°C for 6 min, followed by 30 cycles of 95°C for 45 s, 56°C for 45 s, and 72°C for 90 s, and finally 72°C for 10 min. PCR products were verified by 1% agarose gel electrophoresis and then sequenced. The assembled sequences were aligned with the model strains in the EzBioCloud database. A phylogenetic tree was constructed using MEGA v7.0 software, and confidence intervals for the tree topology were tested using 1000 bootstraps. The genome of strain Qhu-M48 was sequenced by Beijing Biomike Biotechnology Co., Ltd. using the PacBio sequencing platform. Gene prediction was performed using Prodigal v2.6.3, and the genome circle map was created using Circos v0.66. To further analyze the genetic differences between the strains, digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) were calculated between Qhu-M48 and the genome of the most similar model strain. dDDH values ​​were calculated using the GGDC website (https: / / ggdc.dsmz.de / ), and ANI values ​​were obtained using pairwise BLAST using JSpecies.

[0053] 1.6 Study on the growth-promoting effect of the bacterial agent Qhu-M48 on Sainfoin and Avena sativa under salt stress

[0054] (1) Preparation of bacterial suspension

[0055] The strain Qhu-M48 was streaked onto TSA medium and cultured at 28°C for 96 h. A single colony was picked and cultured in TSB medium at 28°C and 180 rpm for 72 h. The cells were collected by centrifugation at 5000 rpm for 10 min and then resuspended in sterile deionized water. The bacterial suspension was adjusted to 1.0 × 10 8 cfu / mL for pot inoculation of Sainfoin.

[0056] (2) Potted plants and salt stress treatment

[0057] Full-grained oats and sainfoin seeds were selected and sterilized with 2% sodium hypochlorite for 5 minutes and 75% ethanol for 45 seconds. After rinsing five times with sterile water, the seeds were sown in circular pots (10 cm diameter, 9 cm height; soil matrix consisting of a 1:1:1 mixture of peat, vermiculite, and sand). Five sainfoin seeds and seven oats seeds were sown per pot. The pots were watered every three days under a 16-hour light (8000 lux) / 8-hour dark cycle at a temperature of 22-25°C. Four potted plant groups were used: normal culture (NC), normal culture plus inoculation with Qhu-M48 (NC+M48), salt stress (NaCl), and salt stress-induced inoculation with Qhu-M48 (NaCl+M48). After seed germination, oats and sainfoin were grown for 7 and 24 days, respectively. 30 mL of bacterial suspension was applied to the rhizosphere of the NC+M48 and NaCl+M48 groups, respectively. An equal volume of sterile deionized water was also applied to the NC and NaCl groups, respectively, and the plants were inoculated once every other day. After these two inoculations, 50 mL of a 200 mM NaCl solution was applied to the NaCl and NaCl+M48 groups, respectively, every other day. After 10 and 14 days of salt stress, the oats and sainfoin plants were photographed and observed for phenotype. Plant tissues were collected to measure relevant physiological and biochemical parameters.

[0058] (3) Physiological and biochemical index determination

[0059] Plant height and root length were measured using a tape measure for each treatment group. Total biomass, aboveground biomass, and root weight of oats were measured using a 1 / 10,000 balance. Three seedlings were randomly selected as a replicate, with three replicates for each indicator. For Sainfoin, four seedlings were randomly selected as a replicate and weighed using a 1 / 10,000 balance to determine total plant biomass. Aboveground and belowground biomass of individual plants were measured from three randomly selected seedlings, with three replicates for each indicator. 0.2 g of expanded leaf samples of the same plant height were oven-dried at 105°C for 15 minutes, dried at 80°C to constant weight, and weighed. Relative water content was calculated as follows: relative water content (%) = (FW - DW) / (TW - DW) × 100, where FW, TW, and DW represent fresh weight, saturated fresh weight, and dry weight, respectively. The maximum photochemical efficiency (Fv / Fm) of leaves from the same tissue site of plants under different treatments was measured using a chlorophyll fluorescence meter (FluorPen FP110). Chlorophyll content was measured using a chlorophyll content meter (Spad 502 Plus). Fresh leaf samples (0.1 g) of Sainfoin seedlings were collected and ground in liquid nitrogen. The activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), as well as the contents of soluble sugars, proline, malondialdehyde (MDA), and hydrogen peroxide (H2O2), were determined using biochemical kits (purchased from Suzhou Keming Biotechnology Co., Ltd.) according to the manufacturer's instructions. Three biological replicates were used for each experiment.

[0060] 2 Results Analysis

[0061] 2.1 Analysis of the taxonomic status of Qhu-M48 based on 16S rRNA gene sequences and phylogenetic relationships

[0062] The full-length sequence of 16S rRNA gene of Qhu-M48 was obtained by amplification and sequencing. The 16S rRNA gene sequence was compared with the EzBioCloud database and found that Qhu-M48 was similar to Streptomyces exfoliates NRRL B-2924. T 、S. venezuelae ATCC 10712 T and S. zaomyceticus NBRC 13348 T The similarities were high, 99.93%, 99.86% and 99.86% respectively. Based on the neighbor-joining method, a phylogenetic tree was constructed by combining the 16S rRNA genes of Qhu-M48 and its highly similar model strains ( Figure 1 ), found that Qhu-M48 formed an independent branch, and speculated that Qhu-M48 might be a new species of Streptomyces.

[0063] 2.2 Whole genome analysis of strain Qhu-M48

[0064] To further determine its taxonomic status, we performed whole-genome sequencing of Qhu-M48. Whole-genome sequencing using the PacBio sequencing platform yielded 128,047 clean reads totaling 1,116,576,418 base pairs. After assembly, the Qhu-M48 genome was 8.32 Mb in size with a GC content of 71.95%. Prodigal v2.6.3 software predicted 7,340 coding genes totaling 7,245,063 base pairs, representing 87.04% of the genome (Tables 1 and 2). Figure 2 ).

[0065] Table 1 Genomic characteristics of strain Qhu-M48

[0066]

[0067] Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) analysis revealed that strain Qhu-M48 was similar to similar strains S. exfoliates NRRL B-2924T and Streptomyces venezuelae ATCC 10712. T and Streptomyces zaomyceticus NBRC 13348 TThe dDDH and ANI values ​​of Qhu-M48 were lower than the dDDH and ANI thresholds of 70% and 95% for distinguishing new bacteria (TIEDJE JM. DNA-DNA hybridization values ​​and their relationship to whole-genome sequence similarities. International Journal of Systematic and Evolutionary Microbiology, 2007, 57(Pt 1):81-91; JAIN C, RODRIGUEZ-R LM, PHILLIPPY AM, KONSTANTINIDIS KT, ALURU S. High throughput ANI analysis of 90K prokaryotic genomes reveals clear species boundaries. Nature Communications, 2018, 9: 51-14.), further indicating that Qhu-M48 is a new species of Streptomyces and was named Streptomyces sp. Qhu-M48.

[0068] Table 2 Comparison of molecular taxonomic characteristics between strain Qhu-M48 and the most similar model strain

[0069]

[0070] 2.3 Morphological analysis of strain Qhu-M48

[0071] The strain Qhu-M48 can grow normally on ISP1-7 medium, Gao's No. 1, NA, and Cha's medium ( Figure 3 After 7 days of culture on ISP1-3 medium, Qhu-M48 produced white aerial hyphae, beige basal hyphae, a dry, wrinkled surface, neat colony edges, and gray spores. A few white spores were produced on Gao's No. 1 and Czapek's medium, but almost no spores were produced on other media. In addition, brown pigment accumulated on ISP1-3 medium. Cell ultrastructure was observed using a laser scanning electron microscope ( Figure 4 A) It was found that the mycelium of strain Qhu-M48 was cylindrical and filamentous, and after carbolic acid staining ( Figure 4 B) Optical microscopy revealed that the spores of the strain were straight and chain-like, which is consistent with the characteristics of Streptomyces.

[0072] 2.4 Physiological and biochemical characteristics of strain Qhu-M48

[0073] Tolerance experiments showed (Table 3) that strain Qhu-M48 can grow at temperatures between 10°C and 40°C, with an optimal growth temperature of 30°C. It can grow normally at NaCl concentrations of 0-3% (mass volume fraction), with a maximum NaCl tolerance of 3%. Furthermore, strain Qhu-M48 can grow between pH 5.0 and 11.0, with the best growth at pH 7.0. It is most similar to strain NRRL B-2924. T It also exhibits similar growth characteristics. Carbon source utilization experiments revealed that strain Qhu-M48 can utilize D-mannitol, rhamnose, D-galactose, inositol, fructose, sorbitol, ulose, mannose, and glucose, but cannot utilize D-arabinose, xylose, and ribose. Regarding nitrogen source utilization, the strain can utilize threonine, arginine, tyrosine, alanine, proline, aspartic acid, serine, glutamic acid, and glycine to maintain growth. Furthermore, Qhu-M48 exhibits starch hydrolysis and lipase activities, but does not produce H2S. API ZYM test found that the bacteria had the activities of alkaline phosphatase, lipase (C4), lipase (C8), lipase (C14), leucine arylamine, valine arylamine, cystine arylamine, chymosin, acid phosphatase, naphthol-AS-BI-phosphohydrolase, β-galactosidase, α-glucosidase, β-glucosidase and N-acetyl-glucosaminidase, while the activities of trypsin, α-galactosidase, β-uronidase, α-mannosidase and α-fucosidase were negative. The most similar model strain NRRL B-2924 was used to test the bacteria. T Comparison of physiological and biochemical characteristics revealed that the Qhu-M48 strain had certain differences in carbon source utilization. T It cannot utilize D-galactose and sorbitol, while Qhu-M48 can utilize them.

[0074] Table 3 Comparison of physiological and biochemical characteristics of strain Qhu-M48 and the most similar model strain

[0075]

[0076] +: positive; –: negative

[0077] 2.5 Chemotaxonomic characteristics of strain Qhu-M48

[0078] TLC analysis of the whole cell hydrolysate of strain Qhu-M48 revealed that the sugars in the whole cell hydrolysate were galactose and arabinose, and the amino acid was L, L-2,6-diaminopimelate (L, L-DAP). Based on LC-MS / MS analysis of the isopentenylquinone species of strain Qhu-M48, the molecular weight and retention time were determined to be MK-9 (H2), MK-9 (H4), and MK-9 (H10 ). Phospholipids were analyzed using biphasic thin layer chromatography ( Figure 5 ), mainly including phosphatidylethanolamine (PE) which is stained red by ninhydrin, blue by anisaldehyde and blue by molybdophosphate; phosphatidylinositol mannosides (PIM) which is stained green by anisaldehyde; phospholipids (PLS) which are stained blue by anisaldehyde and blue by molybdophosphate; phosphatidylmethylethanolamine (PME) which is stained blue by molybdophosphate; phosphatidylglycerol (DPG) which is stained blue by anisaldehyde and blue by molybdophosphate and an unknown phosphate (L1) which is stained blue by molybdophosphate. The polar lipid components of the strain are mainly Antieiso-C 15:0 (44.08 %)、Antieeso-C 17:0 (16.14 %)、iso-C 16:0 (12.23 %)、iso-C 15:0 (10.23 %), C 16:0 (5.16 %)、iso-C 17:0 (4.97 %)、iso-C 14:0 (3.17 %), etc.

[0079] 2.6 Streptomyces Qhu-M48 promotes growth of oat seedlings and alleviates salt stress

[0080] In order to further evaluate whether Streptomyces Qhu-M48 can promote plant growth and improve plant salt stress tolerance, the present invention observed the growth-promoting effects of Streptomyces Qhu-M48 on different plants, oats and sainfoin, under normal and salt stress conditions.

[0081] Under normal culture conditions, after the oat seedlings were inoculated with Qhu-M48, the total biomass and aboveground biomass of the seedlings increased significantly by 10.0% and 14.6% compared with the normal control (NC). Figure 6 AD). Under salt stress conditions, the growth of oats was inhibited, and the total biomass, aboveground biomass, and underground biomass decreased significantly compared with the normal group (NC). However, the salt-stressed seedlings inoculated with fungicides (NaCl+M48) increased by 43.4%, 39.3%, and 70.5%, respectively, compared with the salt stress group (NaCl), and the total biomass and aboveground biomass returned to the level of the normal group ( Figure 6AD). The plant height of the seedlings inoculated with the microbial agent (NC+M48) increased by 8.2% compared with the NC, while that of the seedlings in the NaCl group decreased by 29.8%. However, compared with the NaCl group, the plant height was significantly increased by 35.9% after inoculation with the Qhu-M48 microbial agent, which was not significantly different from the NC group ( Figure 6 E). In addition, salt stress inhibited the elongation of seedling roots, while the bacterial agent Qhu-M48 significantly promoted root development under salt stress, increasing root length by 32.2% ( Figure 6 F). Although the bacterial agent Qhu-M48 had no effect on the relative water content and photosynthesis-related indicators of seedlings under normal conditions, it could significantly increase the relative water content, chlorophyll content and the maximum photochemical efficiency of PSⅡ Fv / Fm of seedlings under salt stress ( Figure 6 GI).

[0082] It can be seen from this that Streptomyces Qhu-M48 can not only promote the growth of oats, but also alleviate the adverse effects of salt stress on the growth of oat seedlings.

[0083] 2.7 Physiological and biochemical mechanisms of Streptomyces Qhu-M48 in improving salt tolerance of Sainfoin

[0084] The present invention further evaluated the growth-promoting and salt stress-alleviating effects of Streptomyces Qhu-M48 on the leguminous forage grass Sainfoin. The results showed that the growth of Sainfoin seedlings was inhibited under salt stress ( Figure 7 A), and the total biomass, aboveground biomass, underground biomass, plant height and root length phenotypes were significantly reduced compared with the normal group, while the fungus Qhu-M48 reversed the inhibition of salt stress on seedling growth ( Figure 7 BG). Compared with the eye stress group, the fungicide Qhu-M48 increased the total biomass, aboveground biomass, belowground biomass, plant height and root length of Sainfoin seedlings under salt stress by 21.7%, 23.3%, 17.7%, 29.4% and 6.7%, respectively ( Figure 7 BG). In addition, the relative water content and photosynthetic efficiency of seedlings decreased under salt stress, but the bacterial agent Qhu-M48 also reversed the downward trend to a certain extent, increasing the relative water content, chlorophyll content and Fv / Fm by 24.9%, 15.9% and 6.7%, respectively ( Figure 7 HJ). This shows that Streptomyces Qhu-M48 can effectively alleviate the salt stress inhibition of Sainfoin.

[0085] To further explore the biochemical mechanism of Streptomyces Qhu-M48 in improving the salt stress tolerance of red bean grass, the present invention measured the peroxide level, antioxidant enzyme activity and osmotic regulation substance content. The results showed that salt stress caused the accumulation of peroxides such as hydrogen peroxide (H2O2) and malondialdehyde (MDA). Figure 8AB), which may cause severe oxidative stress in seedlings. Although the levels of H2O2 and MDA in the seedlings of the NaCl+Qhu-M48 group were still significantly higher than those in the normal culture NC group, the contents of the two substances were significantly reduced by 20.2% and 30.5% compared with the NaCl treatment group, indicating that the bacterial agent Qhu-M48 significantly reduced the degree of oxidation caused by salt stress ( Figure 8 A, B). Salt stress induced the accumulation of peroxides, which further activated the activity of antioxidant enzymes in red clover. Under normal culture conditions, the activities of catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD) were low. However, under salt stress, the activities of CAT, SOD, and POD were significantly increased compared with the CK group, which may be related to the ability of antioxidant enzymes to scavenge reactive oxygen species. Compared with the salt stress treatment group, the bacterial agent Qhu-M48 further increased the CAT and POD activities of seedlings under salt stress by 45.5% and 65.2%, respectively. The activity of SOD was not significantly different from that of the NaCl treatment group ( Figure 8 CE), it is speculated that the bacterial agent Qhu-M48 reduces the peroxide content in sainfoin seedlings under salt stress by increasing the activity of plant antioxidant enzymes, thereby alleviating oxidative stress. Osmotic regulating substances such as proline and soluble polysaccharides play an important role in regulating the cellular osmotic pressure of plants under salt stress. Compared with the CK group, the bacterial agent Qhu-M48 significantly increased the proline and soluble sugar contents in sainfoin seedlings under normal culture and salt stress. The proline and soluble sugar levels in the salt stress inoculated bacterial agent Qhu-M48 group also increased to a certain extent compared with the salt stress group, but there was no significant difference with the salt stress treatment group ( Figure 8 F, G).

[0086] The above studies showed that Streptomyces Qhu-M48 improved the resistance of sainfoin to salt stress by increasing the activities of antioxidant enzymes CAT, SOD and POD, the content of osmotic regulating substances proline and soluble polysaccharides, and reducing the levels of peroxides H2O2 and MDA.

[0087] In summary, this study determined the taxonomic status of a new Streptomyces species isolated from an alpine meadow soil sample on the Qinghai-Tibet Plateau through morphological, physiological and biochemical characteristics, chemotaxonomic characteristics, 16S rRNA phylogenetic relationships, and genomic analysis. The species was named Streptomyces sp. Qhu-M48. Further salt tolerance evaluation of the grasses (Avena sativa) and the leguminous grass (Sainfoin) inoculated with Qhu-M48 under salt stress revealed that the inoculant Qhu-M48 alleviated salt-induced oxidative damage and osmotic disturbances by improving physiological parameters such as root development, plant height, relative water content, and photosynthesis, as well as regulating antioxidant enzyme activity and osmotic regulatory substance levels. This research has practical value for efficient forage cultivation in saline-alkali lands and provides a bacterial strain resource for the development of salt-tolerant agents.

[0088] Those skilled in the art will appreciate that the embodiments of the present invention described above and shown in the accompanying drawings are intended to be illustrative only and are not intended to limit the present invention. The objectives of the present invention have been fully and effectively achieved. The functional and structural principles of the present invention have been demonstrated and illustrated in the embodiments. Any variations or modifications may be made to the embodiments of the present invention without departing from the principles described.

Claims

1. Streptomyces Qhu-M48 strain, characterized in that The deposit number is: CGMCC No. 4.8023.

2. Use of the Streptomyces Qhu-M48 according to claim 1 in promoting plant growth.

3. The use according to claim 2, characterized in that The Streptomyces Qhu-M48 can promote seed germination and / or the growth of one or more of roots, stems, leaves or flowers and fruits.

4. Use of the Streptomyces Qhu-M48 according to claim 1 in regulating plant stress resistance.

5. The use according to claim 4, characterized in that The stress resistance is at least one of salt resistance, cold resistance, drought resistance and high temperature resistance.

6. The use according to claim 5, characterized in that The stress resistance is salt resistance.

7. The use according to claim 2 or 4, characterized in that When in use, a single agent of Streptomyces Qhu-M48 or a prepared agricultural product is used to treat seeds, spray leaves or irrigate roots.

8. The use according to any one of claims 2 to 7, characterized in that: The plants include plants used for agriculture and animal husbandry.

9. The use according to claim 8, characterized in that The plant is selected from grasses of the Poaceae family, grasses of the Leguminosae family, grasses of the Asteraceae family, grasses of the Chenopodiaceae family, grasses of the Cyperaceae family, grasses of the Polygonaceae family or grasses of the Cruciferae family.

10. A microbial preparation, characterized in that Contains the Streptomyces Qhu-M48 according to claim 1.

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