A strain of *Primatellorum aestivum* T54, its inoculum and application

Abstract

This invention belongs to the field of microbial technology, specifically relating to a strain of *Primatellorum aspergillus* T54, its inoculum preparation, and its application. The *Primatellorum aspergillus* (… Priestia aryabhattai T54 was deposited on February 21, 2025, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33613. The *Priscilla argentea* T54 and its inoculum involved in this invention exhibit significant inhibitory effects against various plant pathogenic fungi and also promote plant growth.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a strain of Primatelellae T54, its inoculum, and its application. Background Technology

[0002] cowpea( Unguiculata vine (L.) Walp.) is one of the world's most economically valuable legumes, with a long history of cultivation. In Hainan, my country, cowpeas are an important winter vegetable crop grown and transported north. They are a major type of winter vegetable grown in the southern breeding area, and are harvested in large quantities from November to the end of May of the following year. More than 90% of them are sold outside the island, occupying an important share of the national market and playing a vital role in the development of the local agricultural economy.

[0003] However, the development of the cowpea industry is severely hampered by Fusarium wilt. Fusarium wilt is one of the most serious, difficult-to-control, and economically damaging diseases affecting cowpeas in Hainan. Its pathogen is *Fusarium* spp. (Deuteromycotina). Fusarium Fusarium oxysporum ( Fusarium oxysporum This disease is a typical soil-borne disease. It occurs in cowpea growing areas in tropical to subtropical regions worldwide, and in severe cases, it can lead to a yield reduction of more than 70%, posing a great threat to the yield and quality of cowpeas.

[0004] Currently, the control of cowpea wilt still relies heavily on chemical agents. However, the long-term use of chemical agents can easily lead to environmental pollution, increased pathogen resistance, and pesticide residues in agricultural products, which contradicts the development needs of efficient and green agriculture. Therefore, finding environmentally friendly alternatives to chemical agents has become an important research direction in the agricultural field. Among these, microbial agents, as an environmentally friendly and sustainable biological control method, have shown great potential in the control of cowpea wilt. However, existing technologies still suffer from insufficient targeting and unstable efficacy, necessitating further research and development of highly efficient microbial control technologies. Summary of the Invention

[0005] This invention provides a strain of *Primatellis argentea* and its application in the control of cowpea wilt, solving the problems of current cowpea wilt control mainly relying on chemical methods, which easily lead to residues and have limited control methods.

[0006] On the one hand, a strain of *Primatellorum auriculi* T54 was classified and named *Primatellorum auriculi*. Priestess aryabhattai The *Primatex auriculata* T54 was deposited at the China General Microbiological Culture Collection Center on February 21, 2025, with accession number CGMCC No. 33613.

[0007] On the other hand, there is the application of Aspergillus oryzae T54 in the inhibition of plant pathogenic fungi.

[0008] Preferably, the plant pathogenic fungus includes *Fusarium oxysporum*, the pathogen of cowpea wilt (…). Fusarium oxysporum f. sp. tracheal ), Fusarium graminearum, the causal agent of wheat scab ( Fusarium gramineae Fusarium oxysporum, the bacterium that causes banana wilt ( Fusarium oxysporum f.sp. from Cuba Coffee leaf blight Fusarium oxysporum (F. rubrum) Fusarium wilt Mango anthracnose (Colletotrichum gloeosporioides) Colletotrichum gloeosporioides Rice blast fungus ( Magnaporthe grisea Sisal leaf rot caused by Diplosporum cocovenenans ( Lasiodiplodia Theobroma cassava ), Coconut gray spot disease, *Pseudomonas pumilum* ( Pestalotiopsis microspora Corn leaf spot disease, Curvularia corniculatum ( Curvularia lunate Phytophthora blight of pepper ( Phytophthora capsicum ), and the pathogen of the leaf spot disease, *Cyclocarya* genus ( Diaporthe biconispora Dragon fruit canker disease, new dark-colored scabies ( Neocytalidium halved ) or coffee anthrax cahava spirochetes ( Colletotrichum kahawae ).

[0009] On the other hand, there is the application of Aspergillus oryzae T54 in plant growth promotion.

[0010] Preferably, the plant includes cowpea.

[0011] On the other hand, there is a microbial agent comprising *Primatecium auriculi* T54.

[0012] On the other hand, there is the application of microbial agents in inhibiting plant pathogenic fungi.

[0013] Preferably, the plant pathogenic fungus includes *Fusarium oxysporum*, the pathogen of cowpea wilt (…). Fusarium oxysporum f. sp. tracheal ), Fusarium graminearum, the causal agent of wheat scab ( Fusarium gramineae Fusarium oxysporum, the bacterium that causes banana wilt ( Fusarium oxysporum f.sp. from Cuba Coffee leaf blight Fusarium oxysporum (F. rubrum) Fusarium wilt Mango anthracnose (Colletotrichum gloeosporioides) Colletotrichum gloeosporioides Rice blast fungus ( Magnaporthe grisea Sisal leaf rot caused by Diplosporum cocovenenans ( Lasiodiplodia Theobroma cassava ), Coconut gray spot disease, *Pseudomonas pumilum* ( Pestalotiopsis microsporaCorn leaf spot disease, Curvularia corniculatum ( Curvularia lunate Phytophthora blight of pepper ( Phytophthora capsicum ), and the pathogen of the leaf spot disease, *Cyclocarya* genus ( Diaporthe biconispora Dragon fruit canker disease, new dark-colored scabies ( Neocytalidium halved ) or coffee anthrax cahava spirochetes ( Colletotrichum kahawae ).

[0014] On the other hand, there is the application of microbial agents in promoting plant growth.

[0015] Preferably, the plant includes cowpea.

[0016] Beneficial effects

[0017] The present invention provides Priestia aryabhattai The T54 strain exhibits significant inhibitory effects against various plant pathogenic fungi. In the plate confrontation method experiment, it showed strong inhibition against *Fusarium oxysporum*, the pathogen causing cowpea wilt. Fusarium oxysporum f.sp. tracheal The inhibition rate reached 71.11%, and it was effective against rice blast fungus (*Strombus oryzae*). Magnaporthe grisea The inhibition rate of ) reached 64.69%, and it was effective against corn leaf spot disease Curvularia zei (Curvularia zei). Curvularia lunate The inhibition rate reached 61.65%, and it was effective against Diplosporum cocovenenans, the causal agent of sisal leaf rot. Lasiodiplodia theobromae The inhibition rate reached 59.50%, and it was effective against Fusarium graminearum, the causal agent of wheat scab. Fusarium gramineae The inhibition rate of ) reached 57.85%, and it was effective against coconut gray spot disease caused by *Pseudomonas aeruginosa* (…). Pestalotiopsis microspora The inhibition rate of ) reached 55.90%, and the inhibition rate of *Norophorus glomeratus* (a type of scab) against dragon fruit canker was also high. Neocytalidium halimidium The inhibition rate reached 55.67%, and it was effective against Fusarium oxysporum, the pathogen that causes wilt of bananas. Fusarium oxysporum f.sp. from Cuba The inhibition rate of ) reached 51.17%, and the inhibition rate of *Colletotrichum candida* (the causal agent of mango anthracnose) was also high. Colletotrichum gloeosporioides The inhibition rate reached 50.91%, and it was effective against Phytophthora blight of pepper (…). Phytophthora capsicum The inhibition rate of ) reached 50.97%, and the inhibition rate of Fusarium oxysporum (Fusarium oxysporum) against coffee leaf blight was also high. Fusarium wilt The inhibition rate of ) reached 49.42%, and the inhibition rate of coffee anthrax spores (Cachamus spp.) was also high. Colletotrichum kahawae The inhibition rate reached 43.82%, and the inhibition rate against the pathogen of leaf spot disease (Corylus spp.) was significantly higher. Diaporthe biconispora The inhibition rate reached 35.03%, demonstrating broad-spectrum and highly efficient antibacterial activity, which can effectively control plant diseases caused by various fungi.

[0018] In a pot experiment on the control of cowpea wilt, applying 1×10 7 CFU / mL, 1×108 CFU / mL, 1×10 9 CFU / mL Priestia aryabhattai After T54 bacterial suspension was administered, the disease index decreased by 27.08%, 31.25%, and 33.33% respectively compared with the Fusarium oxysporum control group, demonstrating a good disease control effect. It can significantly reduce the incidence of cowpea wilt and reduce yield loss caused by the disease.

[0019] at the same time, Priestia aryabhattai The T54 strain exhibits a significant growth-promoting effect on cowpea plants, at 1×10⁻⁶. 8 After treatment with CFU / mL bacterial suspension, the plant height, fresh weight, and root length of cowpea plants increased by 47.57%, 54.37%, and 65.05%, respectively, compared with the control group; 1×10 9 After treatment with CFU / mL bacterial suspension, compared with the Fusarium oxysporum treatment group, plant height, leaf area and root length increased by 89.00%, 563.75% and 69.33% respectively, and compared with the water treatment group, plant height, leaf area and root length increased by 77.90%, 114.03% and 25.17% respectively. It can effectively promote the growth and development of cowpea plants and improve the growth status and biomass of plants.

[0020] Furthermore, as a natural microbial strain, this strain has good environmental compatibility and will not cause problems such as environmental pollution, pathogen resistance, and agricultural product residues caused by the use of chemical pesticides. It meets the needs of green agricultural development, has a wide range of application scenarios and potential commercial application value, and can be used as a microbial preparation for the prevention and control of various plant fungal diseases and the promotion of crop growth, providing strong technical support for sustainable agricultural development. Attached Figure Description

[0021] Figure 1 for Priestia aryabhattai Colony morphology of strain T54 on LB solid medium, where A is a single colony morphology and B is a streaked colony morphology on a plate.

[0022] Figure 2 for Priestia aryabhattai Physiological and biochemical reactions of strain T54, where A is... Priestia aryabhattai The ability of strain T54 to secrete siderophores, B being... Priestia aryabhattai Nitrogen fixation capacity of strain T54, C is Priestia aryabhattai Potassium solubilization ability of strain T54, D is Priestia aryabhattai Phosphate solubilization ability of strain T54, E is Priestia aryabhattai The ability of strain T54 to secrete auxin (IAA);

[0023] Figure 3 for Priestia aryabhattaiPhylogenetic tree of 16S rDNA of strain T54;

[0024] Figure 4 for Priestia aryabhattai T54 strain gyrB Genetic phylogenetic evolutionary tree;

[0025] Figure 5 for Priestia aryabhattai The inhibitory activity of strain T54 against the pathogen causing cowpea wilt was determined, where A was the control group for Fusarium oxysporum and B was the control group for Fusarium oxysporum. Priestia aryabhattai Diagram showing the confrontation between the antibacterial abilities of strain T54;

[0026] Figure 6 for Priestia aryabhattai The antibacterial spectrum of strain T54, where A represents *Dioscorea haematobium*, the causal agent of sisal leaf rot. Lasiodiplodia theobromae B is *Pseudomonas spp.*, a species of coconut gray spot disease. Pestalotiopsis microspora C represents corn leaf spot disease and corn bending ( Curvularia lunata D represents *Normacea glomerata*, a new dark-colored scab for dragon fruit canker. Neoscytalidium dimidiatum E represents the coffee anthrax spore *Caucasus kaharva* ( Colletotrichum kahawae F represents Fusarium oxysporum, the fungus that causes banana wilt. Fusarium oxysporum f.sp. cubense G represents *Aeroblastus collodion*, the pathogen causing mango anthracnose. Colletotrichum gloeosporioides H represents Fusarium graminearum, a fungus that causes wheat scab. Fusarium graminearum ), I is Phytophthora blight, the fungus that causes blight in peppers ( Phytophthora capsici J is the pathogen of the genus *Cyclocarya* in *Lactarius* leaf spot disease. Diaporthe biconispora K represents Fusarium tumefaciens, the blight pathogen of coffee leaves. Fusarium lateritium L represents the rice blast fungus ( ), where L is the pathogen causing rice blast. Magnaporthe grisea );

[0027] Figure 7 for Priestia aryabhattai The control effect and growth-promoting ability of strain T54 against cowpea wilt in potted plants were compared, with H2O serving as the water control group and FO as the positive control group, at a concentration of 1×10⁻⁶. 7 T54 is an inoculation solution of *Fusarium oxysporum* spore suspension and 1×10 7 T54 bacterial suspension at CFU / mL, 1×10 8 T54 is an inoculation solution of *Fusarium oxysporum* spore suspension and 1×10 8 T54 bacterial suspension at CFU / mL, 1×10 9 T54 is an inoculation solution of *Fusarium oxysporum* spore suspension and 1×10 9 T54 bacterial suspension at CFU / mL. Detailed Implementation

[0028] I. Experimental Materials

[0029] PDA medium: Boil 200 g potatoes for 15 min and keep the filtrate. Add 18 g glucose and 18 g agar to ddH2O to 1000 mL; sterilize at 121℃ for 20 min.

[0030] LB medium (Luria-Bertani medium): 10.0 g tryptone, 5.0 g yeast extract, 8.0 g sodium chloride (NaCl), 20 g agar, add ddH2O to 1000 mL; sterilize at 121℃ for 20 min.

[0031] NBRIP Inorganic Phosphorus Medium: 10.0 g glucose, 0.1 g ammonium sulfate, 0.25 g magnesium sulfate heptahydrate (MgSO4·7H2O), 0.2 g potassium chloride (KCl), 5.0 g magnesium chloride (MgCl2), 5.0 g calcium phosphate, add ddH2O to 1000 mL; sterilize at 121℃ for 20 min.

[0032] Alexandrite medium: 5.0 g sucrose, 2.0 g disodium hydrogen phosphate (Na2HPO4), 0.5 g magnesium sulfate heptahydrate (MgSO4·7H2O), 0.005 g ferric chloride (FeCl3), 0.1 g calcium carbonate (CaCO3), 2.0 g potassium feldspar, add ddH2O to 1000 mL; sterilize at 115℃ for 20 min.

[0033] Assumption nitrogen-free medium: dipotassium hydrogen phosphate (K2HPO4) 0.2 g, magnesium sulfate heptahydrate (MgSO4·7H2O) 0.2 g, calcium sulfate dihydrate (CaSO4·2H2O) 0.2 g, sodium chloride (NaCl) 0.2 g, calcium carbonate (CaCO3) 5.0 g, mannitol 10.0 g, add ddH2O to 1000 mL; sterilize at 121℃ for 20 min.

[0034] Chrome azurol (CAS) medium: The basal medium consists of 100.0 g glucose, 0.5 g magnesium sulfate heptahydrate (MgSO4·7H2O), 20.0 g tryptone, 0.5 g calcium chloride (CaCl2), and 10.0 g agar; the CAS assay solution consists of 0.06 g chrome azurol sulphonate (CAS), 0.0027 g ferric chloride (FeCl3), and 0.073 g hexadecy-1,3-methyl-ammonium bromide (HDTMA); 10× buffer PIPES 1 mol / L, pH 7.0; weigh 141 g of the medium and add ddH2O to bring the volume to 800 mL; sterilize at 115℃ for 20 min; cool the basal medium to 60℃ and slowly add 60 g of CAS solution. 100 mL of 10× buffer solution and 100 mL of 10× CAS detection solution, preheated to ℃.

[0035] YMB medium: 1.0 g yeast extract, 10.0 g mannitol, 0.5 g dipotassium hydrogen phosphate (K2HPO4), 0.2 g magnesium sulfate heptahydrate (MgSO4·7H2O), 0.1 g sodium chloride (NaCl), 1.0 g calcium carbonate (CaCO3), 2.0 g L-tryptophan, add ddH2O to 1000 mL; sterilize at 151℃ for 20 min.

[0036] Example 1: *Primatellis* ( Priestia aryabhattai The acquisition of T54.

[0037] Rhizosphere soil and tissue samples were collected from healthy cowpea plants in areas affected by cowpea wilt disease. Soil samples were separated using the dilution-spreading method: 10 g of soil sample was placed in a 250 mL Erlenmeyer flask, 90 mL of ddH2O was added, and the flask was placed in a shaker at 28 ℃ and 180 rpm for 30 min to mix thoroughly. The flask was then placed in an 80 ℃ water bath and allowed to stand for 10 min before being diluted to a final concentration of 10 g. -2 10 -3 10 -4 10 -5 Dilute the solution. Add 10-15 mL of preheated LB medium to a petri dish. After cooling, add 0.2 mL of the diluted solution to the plate and spread evenly with a glass spreader until dry. Repeat three times. After sealing, incubate at 28 ℃ under 14 h light-10 h dark conditions. Observe the colonies after 2 days. Select suspected colonies and streak them on LB plates for purification. Number and store the strains.

[0038] Using *Fusarium wilt*, the pathogen of cowpea wilt, as a control strain, the preserved strains were initially screened using the plate confrontation method. A 0.5 cm *Fusarium wilt* mycelium was inoculated in the center of a PDA solid medium plate. Four points 2.5 cm from the center were selected symmetrically in a cross pattern on each medium plate. The same numbered strain was then inoculated at each of the four points using a toothpick. Strains showing antagonistic effects were screened again using the same method and preserved. Colony diameter was measured using the cross-cross method. Inhibition rate / % = (control group colony diameter - treatment group colony diameter) / control group colony diameter × 100. The experiment was repeated three times, with three replicates for each treatment. Data were compiled using Excel, and SPSS (25) software was used for statistical analysis of data differences. Multiple comparisons were performed using Tukey's Method.

[0039] The morphology of strain T54 obtained through screening on LB solid medium is as follows: Figure 1 As shown, the colonies on the plate are pale yellow to yellowish-brown, with relatively smooth edges, opaque, and clearly spaced apart.

[0040] Example 2: *Primatellis* ( Priestia aryabhattai Physiological and biochemical characteristics of T54.

[0041] Determination of phosphorus solubilization ability: After activating strain T54, it was inoculated onto NBRIP inorganic phosphorus medium and placed in a constant temperature incubator at 28 ℃ for 7 days. The presence of transparent oil droplet-like halos around the colonies was observed. If present, it indicated that the strain had phosphorus solubilization ability. The size of the halo was measured.

[0042] Determination of potassium solubilizing ability: After activating strain T54, it was inoculated onto Alexandrite silicate medium and placed in a constant temperature incubator at 28 ℃ for 7 days. The presence of transparent oil droplet-like halos around the colonies was observed. If present, it indicated that the strain had potassium solubilizing ability. The size of the halo was measured.

[0043] Determination of nitrogen fixation function: After activation, strain T54 was inoculated onto Assumption nitrogen-free medium and placed in a constant temperature incubator at 28 ℃ for 7 days. Colony growth was observed. The ability to grow on Assumption nitrogen-free medium was considered to have nitrogen fixation function. The ammonia fixation capacity was judged based on the diameter of the colony growth.

[0044] Siderophore production capacity determination: After activation, strain T54 was inoculated onto chromium azurite (CAS) medium and placed in a constant temperature incubator at 28 ℃ for 7 days. The presence or absence of a yellow halo around the colony was observed, and the size of the halo was measured.

[0045] Auxin (IAA) secretion capacity: Strain T54 was inoculated into YMB medium and cultured at 28°C for 2 days. After centrifugation at 12,000 rpm for 2 min, 2 mL of the supernatant was transferred to a test tube, and an equal volume of Salkowski's reagent was added. The mixture was gently shaken and incubated in the dark for 30 min. A red color indicated the production of IAA. YMB culture medium with Salkowski's reagent served as a blank control (CK control group).

[0046] The results are as follows Figure 2 As shown, strain T54 possesses certain abilities in nitrogen fixation, phosphorus solubilization, potassium solubilization, and siderophore secretion. On CAS medium, a clear yellow halo with a diameter of 12.32 mm was observed in T54, indicating the strain's siderophore secretion ability. On Assumption medium without nitrogen, the secretion ring of T54 was smaller, indicating weaker nitrogen fixation ability. On Alexandrite silicate medium, the colony diameter of T54 was 7.22 × 7.86 mm, indicating a certain potassium solubilization ability. On NBRIP inorganic phosphorus bacteria medium, the secretion ring diameter of T54 was 24.21 mm, indicating the strain's ability to decompose organic phosphorus. In YMB medium, the color of the medium inoculated with T54 showed no significant change, indicating a weak IAA secretion ability.

[0047] Example 3: *Primatellis* ( Priestia aryabhattai Molecular identification of T54.

[0048] The T54 strain was inoculated into LB liquid medium and incubated at 180 rpm for 16 h. Using the bacterial DNA extraction kit from Tiangen Biotech (Beijing) Co., Ltd., the biocontrol bacterial DNA was extracted according to the instructions. The obtained bacterial DNA was used as a template for PCR amplification. The target gene was selected using universal primers for the conserved bacterial region ribosomal 16S rDNA gene and bacterial-specific primers for the helicase B subunit gene. gyrB The amplification primers were the universal 16S rDNA primers SEQ NO.1-27f (5′-AGAGTTTGATCCTGGCTCAG-3′) and SEQ NO.2-1492r (5′-TACGGCTACCTTGTTACGACTT-3′), and SEQ NO.3- primer. gyrB F (5′-GAAGTCATCATGACCGTTCTGCAYGCNGGNGGNAARTTYGA-3′) and SEQ NO.4- gyrB R (5′-AGCAGGGTACGGATGTGCGAGCCRTCNACRTCNGCRTCNGTCAT-3′).

[0049] The PCR reaction system consisted of: 12.5 μL of 2×Tap PCR MasterMix, 9.5 μL of ddH2O, 1 μL each of forward and reverse primers, and 1 μL of DNA. The 16S rDNA amplification program was as follows: pre-denaturation at 95 ℃ for 5 min, denaturation at 95 ℃ for 30 s, annealing at 55 ℃ for 45 s, extension at 72 ℃ for 1 min, and further extension at 72 ℃ for 10 min, for a total of 35 cycles. Gyr B The amplification program was as follows: pre-denaturation at 94 ℃ for 4 min, denaturation at 94 ℃ for 1 min, annealing at 57 ℃ for 1 min, extension at 72 ℃ for 80 s, and further extension at 72 ℃ for 10 min, for a total of 30 cycles.

[0050] PCR amplification products were detected by 1% agarose gel electrophoresis. The target fragment bands were observed and photographed using a gel imaging system. The PCR amplification products were then sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. Sequencing results were compared and analyzed for homology in the NCBI database, and highly similar gene sequences and their type strain sequences were downloaded.

[0051] For T54 16S rDNA and gyrB The gene fragment was amplified by PCR, yielding 1400 bp of 16S rDNA and... gyrB The target fragment was approximately 1200 bp. The PCR amplification product was sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. The obtained sequence was aligned using NCBI. 16S rDNA alignment results showed that T54 was associated with *Primatellorum auriculi*. Priestia aryabhattai B8W22 (NR_118442.1) has a similarity of 99.79%. gyrB The comparison results showed that T54 was related to *Primatellis*. P. aryabhattai The similarity between HJ.T3 (NZ_CP146499.1) and the target strain was 99.81%. Highly similar gene sequences and type strain sequences were downloaded, and the 16S rDNA of T54 was constructed using the maximum likelihood method with MEGA11 software. gyrB Phylogenetic tree, the results are as follows Figure 3 and Figure 4 As shown, T54 and *Primatellis* ( Priestia aryabhattai They clustered on the same branch. Therefore, T54 was identified as *Primatellis auriculata*. Priestia aryabhattai And the strain Priestia aryabhattaiT54 is deposited at the China General Microbiological Culture Collection Center (address: Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing) on ​​February 21, 2025, with accession number CGMCC No. 33613.

[0052] Example 4. *Priscilla argentea* ( Priestia aryabhattai T54 antibacterial ability determination.

[0053] The tested plant pathogenic fungi included *Fusarium oxysporum*, the pathogen of cowpea wilt (…). Fusarium oxysporum f.sp. tracheiphilum ), Fusarium graminearum, the causal agent of wheat scab ( Fusarium graminearum Fusarium oxysporum, the bacterium that causes banana wilt ( Fusarium oxysporum f.sp. cubense Coffee leaf blight Fusarium oxysporum (F. rubrum) Fusarium lateritium Mango anthracnose (Colletotrichum gloeosporioides) Colletotrichum gloeosporioides Rice blast fungus ( Magnaporthe grisea Sisal leaf rot caused by Diplosporum cocovenenans ( Lasiodiplodia theobromae ), Coconut gray spot disease, *Pseudomonas pumilum* ( Pestalotiopsis microspora Corn leaf spot disease, Curvularia corniculatum ( Curvularia lunata Phytophthora blight of pepper ( Phytophthora capsici ), and the pathogen of the leaf spot disease, *Cyclocarya* genus ( Diaporthe biconispora Dragon fruit canker disease, new dark-colored arthropoda ( Neoscytalidium dimidiatum Coffee anthrax disease, *Caucasus kaharva* ( Colletotrichum kahawae ).

[0054] Using the flat standoff method Priestia aryabhattai Determination of the antibacterial activity of strain T54. Priestia aryabhattai The T54 strain was cultured in LB liquid medium at 180 rpm and 28 ℃ for 16 h to obtain bacterial fermentation broth. Using the "cross-hatching method," sterile 5 mm filter paper discs were affixed to four points 2.5 cm apart at the center of PDA culture plates. Using a sterile 5 mm punch, mycelial discs of the same age were inoculated into the center of the PDA plate, targeting the tested pathogenic fungus. 1 μL of bacterial fermentation broth was added to the center of each of the four filter paper discs. Each treatment was repeated three times. The treated PDA plates were incubated at 28 ℃ for 4–5 days, during which colony growth was observed. When the colonies had covered approximately 3 / 4 of the culture plate, the antibacterial activity was measured, recorded, and photographed. The formula for calculating the antibacterial rate is as follows:

[0055] Inhibition rate / % = [(Diameter of pathogen growth in control group - Diameter of pathogen block) - (Diameter of pathogen growth in treatment group - Diameter of pathogen block)] / (Diameter of pathogen growth in control group - Diameter of pathogen block) × 100.

[0056] The results are shown in Table 1. Figure 5 and Figure 6 As shown, T54 exhibits good inhibitory activity against *Fusarium oxysporum*, the pathogen causing cowpea wilt, with an inhibition rate of 71.11%. T54 also demonstrates good inhibitory activity against other tested pathogenic fungi: 57.85% against *Fusarium graminearum* (wheat scab), 51.17% against *Fusarium oxysporum* (banana wilt), 49.42% against *Fusarium oxysporum* (coffee leaf blight), 50.91% against *Colletotrichum gloeosporioides* (mango anthracnose), and [missing data - likely related to rice blast disease]. The percentages of pathogens against virulence were 64.69% for *Phytophthora infestans*, 59.50% for *Diplosporium cocovenenans* (causing leaf rot in sisal), 55.90% for *Polytrichum gloeosporioides* (causing gray spot disease in coconut), 61.65% for *Curvularia zeylans* (causing leaf spot disease in maize), 50.97% for *Phytophthora infestans* (causing pepper blight), 35.03% for *Cytotrichum spp.* (causing leaf spot disease in heart-leaf spot), 55.67% for *Neococcus faecium* (causing dragon fruit canker), and 43.82% for *Colletotrichum cahavarensis* (causing coffee anthracnose).

[0057] Table 1. *Priscilla argentea* ( Priestia aryabhattai T54 antibacterial ability

[0058]

[0059] Example 5. *Priscilla argentea* ( Priestia aryabhattai The potted plant control efficacy and growth-promoting ability of T54 against cowpea wilt disease.

[0060] For indoor potted cowpea seedlings at the 2-leaf, 1-heart stage, the pathogen *Fusarium oxysporum* was prepared at a concentration of 1×10⁻⁶. 7 CFU / mL spore suspension. After rinsing the soil off the roots of seedlings, five fibrous roots were cut to injure the roots. The roots were then soaked in the pathogen spore suspension for 15 minutes before transplanting into flowerpots. Five treatment groups were set up: H2O (water control group, no root injury treatment), FO (positive control group, inoculated with Fusarium oxysporum spore suspension), FO+1×10 7 T54 (inoculated with Fusarium oxysporum spore suspension and 1×10 7 (T54 bacterial suspension at CFU / mL), FO+1×10 8 T54 (inoculated with Fusarium oxysporum spore suspension and 1×10 8 (T54 bacterial suspension at CFU / mL), FO+1×10 9 T54 (inoculated with Fusarium oxysporum spore suspension and 1×10 9T54 bacterial suspension (CFU / mL) was applied to different treatment groups every 5 days. After 20 days, the disease index was evaluated and the control effect was determined. At the same time, the morphological indicators of cowpea plants (plant height, leaf area, fresh weight, root length) were measured to evaluate the growth-promoting effect of biocontrol bacteria on cowpea plants.

[0061] The disease index evaluation for cowpea wilt is based on the following standards: Grade 0: No discoloration of the root and stem vascular bundles, no yellowing or wilting of leaves; Grade 1: Discoloration of less than 1 / 4 of the root and stem vascular bundles, some leaves discolor, no significant change in plant growth; Grade 2: Discoloration of less than 1 / 2 of the root and stem vascular bundles, leaves discolor and wither, plant growth is inhibited; Grade 3: Discoloration of nearly 3 / 4 of the root and stem vascular bundles, significant inhibition of the entire plant's growth, leaves yellowing, wilting, and falling off from the bottom up; Grade 4: Complete discoloration of the root and stem vascular bundles, the entire plant withers and dies. The disease index evaluation and control effect determination are calculated using the following formula:

[0062] Disease index = [Σ(number of plants with disease at each disease level × number of disease levels) / total number of potted plants × genus with the highest disease index] × 100.

[0063] Prevention and control effect / % = [Disease index of control group - Disease index of treatment group] / Disease index of control group × 100.

[0064] The results of the pot experiment showed that the water control group did not develop the disease, while the control group with the pathogen *Fusarium oxysporum* developed the disease, with a disease index of 85.83 (Table 2). Figure 7 The plants in the T54 bacterial suspension treatment group did not show obvious disease symptoms on the outside, but some brownish-red areas were still visible when the vascular bundles were cut open. The higher the concentration of the bacterial suspension and the lower the disease index, the better the control effect. Apply 1×10 7 CFU / mL, 1×10 8 CFU / mL, 1×10 9 The T54 bacterial suspension at CFU / mL showed a 47.57%, 54.37%, and 65.05% decrease in disease index compared to the Fusarium oxysporum control group, respectively. Furthermore, as shown in Table 3 and... Figure 7 As shown, 1×10 7 CFU / mL, 1×10 8 CFU / mL, 1×10 9 All three concentrations of T54 bacterial suspension (CFU / mL) exhibited a certain growth-promoting effect on cowpea plants. Specifically, 1×10⁻⁶ CFU / mL suspension... 9 Cowpea plants treated with CFU / mL bacterial suspension showed increased plant height, leaf area, and root length by 89.00%, 563.75%, and 69.33%, respectively, compared to the FO treatment group; and increased plant height, leaf area, and root length by 77.90%, 114.03%, and 25.17%, respectively, compared to the H2O treatment group.

[0065] Table 2. *Priscilla argentea* ( Priestia aryabhattai Potted plant control efficacy of T54 against cowpea wilt disease

[0066]

[0067] Table 3. Morphological indicators of cowpea plants under different treatments

[0068]

[0069] Note: Different lowercase letters in the same column indicate that the differences between treatments are significant at the 0.05 level.

Claims

1. A strain of *Primatellis* ( Priestia aryabhattai T54, characterized in that, The *Primate* T54 strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33613.

2. The application of *Priscilla argentea* T54 as described in claim 1 in inhibiting plant pathogenic fungi, characterized in that... The plant pathogenic fungi include *Fusarium oxysporum*, which causes cowpea wilt. Fusarium oxysporum f. sp. tracheiphilum Fusarium tumefaciens, which causes coffee leaf blight Fusarium lateritium Colletotrichum gloeosporioides, which causes mango anthracnose. Colletotrichum gloeosporioides The pathogen that causes rice blast Magnaporthe grisea The fungus *Diplosporium cocovenenans*, which causes sisal leaf rot. Lasiodiplodia theobromae *Plasmodium spp.*, which causes coconut gray spot disease Pestalotiopsis microspora Curvularia corniculatum, which causes corn leaf spot disease Curvularia lunata Phytophthora blight, the fungus that causes blight in peppers Phytophthora capsici Pathogens that cause leaf spot disease Diaporthe biconispora A new type of dark-colored arthropod spore that causes dragon fruit canker. Neoscytalidium dimidiatum Or bacteria that cause coffee anthrax Colletotrichum kahawae .

3. The application of *Primatecium argentea* T54 as described in claim 1 in the prevention and control of cowpea wilt and the promotion of cowpea growth.

4. A microbial agent, characterized in that, The bacterial agent includes *Primaterella auriculata* T54 as described in claim 1.

5. The application of the fungal agent according to claim 4 in inhibiting plant pathogenic fungi, characterized in that, The plant pathogenic fungi include *Fusarium oxysporum*, which causes cowpea wilt. Fusarium oxysporum f. sp. tracheiphilum Fusarium tumefaciens, which causes coffee leaf blight Fusarium lateritium Colletotrichum gloeosporioides, which causes mango anthracnose. Colletotrichum gloeosporioides The pathogen that causes rice blast Magnaporthe grisea The fungus *Diplosporium cocovenenans*, which causes sisal leaf rot. Lasiodiplodia theobromae *Plasmodium spp.*, which causes coconut gray spot disease Pestalotiopsis microspora Curvularia corniculatum, which causes corn leaf spot disease Curvularia lunata Phytophthora blight, the fungus that causes blight in peppers Phytophthora capsici Pathogens that cause leaf spot disease Diaporthe biconispora A new type of dark-colored arthropod spore that causes dragon fruit canker. Neoscytalidium dimidiatum Or bacteria that cause coffee anthrax Colletotrichum kahawae .

6. The application of the microbial agent according to claim 4 in the prevention and control of cowpea wilt and the promotion of cowpea growth.

Images