Paraburkholderia NRS-2-16, bacterial agent and application thereof

Abstract

The application belongs to the technical field of microorganisms, and particularly relates to Paraburkholderia fungorum NRS-2-16, a bacterial agent and application thereof. The Paraburkholderia fungorum of the application is preserved in the China General Microbiological Culture Collection Center, and has a preservation number of CGMCC No. 38750. The Paraburkholderia fungorum NRS-2-16 of the application has a significant inhibitory effect on cotton verticillium wilt (Lignyotum delavayi) and cotton boll blight (Phytophthora cinnamomi), and can be used for the prevention and treatment of cotton verticillium wilt and cotton boll blight.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to Burkholderia paraknegaceae NRS-2-16, its inoculum, and its application. Background Technology

[0002] Cotton is an important economic crop in my country, playing a pivotal role in the national economy. However, cotton production has long been seriously threatened by various diseases, among which Verticillium wilt and boll blight are the two most prominent and difficult to control diseases.

[0003] Verticillium wilt, or cotton wilt, is a devastating soil-borne vascular disease caused by Verticillium dahliae Kleb., widely distributed in major cotton-growing regions worldwide. The pathogen can form durable microsclerotia that can survive in the soil for over a decade. Upon encountering a suitable host, it invades through wounds at the root tip or lateral roots, penetrating the cortex and entering the vascular tissue. This obstructs the transport of water and nutrients, resulting in typical symptoms such as yellowing and wilting of leaves, interveinal chlorosis, and even the death of the entire plant. It is estimated that this disease causes billions of dollars in economic losses globally each year, making it one of the most difficult diseases to control in cotton production.

[0004] Boll blight, caused by *Phytophthora boehmeriae* Sawada, is a serious boll-stage disease that severely impacts cotton production, particularly prevalent and severe in cotton-growing areas of the Yellow River and Yangtze River basins in my country. This pathogen was first discovered and named on ramie leaves in Taiwan Province, my country, in 1927. Belonging to the genus *Phytophthora* within the phylum Oomycetes, its mycelium is slender, and its sporangia are mostly oval or obovate-pear-shaped. It can survive and spread in soil and diseased plant debris through various reproductive structures, including zoospores, chlamydospores, and oospores. Field surveys show that *Phytophthora boehmeriae* not only directly infects cotton bolls, causing boll blight, but the wounds it creates also significantly increase the probability of infection by other saprophytic pathogens, thus triggering a co-occurrence of cotton boll rot. Statistics show that boll blight accounts for over 90% of cotton boll rot cases across various regions, severely affecting cotton yield and fiber quality.

[0005] Currently, the control of both diseases primarily relies on chemical methods. Cotton Verticillium wilt control mainly uses chemical fumigants such as chloropicrin and methyl bromide, as well as some chemical fungicides; cotton boll blight control mainly relies on a few fungicides such as aluminum tris(ethyl phosphonate). However, long-term, large-scale application of chemical agents not only induces drug resistance in pathogens, leading to a gradual decline in control effectiveness, but also disrupts the balance of soil microbial communities, resulting in non-point source pollution and excessive pesticide residues in cotton products. More seriously, *Verticillium dahliae* has a wide host range and complex pathogenic mechanisms; while *Phytophthora ramie* exhibits homozygous mating characteristics, and sexual reproduction can lead to continuous variation in biological traits such as colony morphology, growth rate, and pathogenicity in asexual offspring, and it has already developed resistance to commonly used fungicides such as metalaxyl. At the same time, existing cotton germplasm resources generally lack highly resistant or immune varieties to these two diseases, conventional breeding methods are unlikely to cultivate effective resistant varieties in the short term, and genetic engineering for disease resistance has not yet achieved a commercial breakthrough. Therefore, the green control of cotton Verticillium wilt and cotton boll blight faces enormous challenges.

[0006] Against this backdrop, biocontrol using antagonistic microorganisms has gradually become an important research direction for the green control of the two diseases mentioned above due to its advantages such as good environmental compatibility, low resistance development, strong targeting, and no toxic residues. Existing literature reports that various microorganisms have inhibitory activity against *Verticillium wilt* or *Phytophthora ramieense*, mainly including *Bacillus* spp., *Trichoderma* spp., *Chaetomium globosum*, *Streptomyces* spp., and *Pythium oligandrum*. However, existing studies mostly focus on screening biocontrol bacteria for single diseases, and there are few reports of broad-spectrum biocontrol strains that can simultaneously exhibit highly efficient antagonistic activity against two different groups of pathogens (fungi and oomycetes) of both *Verticillium wilt* and *Phytophthora blight*.

[0007] Therefore, there is a need to provide an improved technical solution that addresses the shortcomings of the existing technology. Summary of the Invention

[0008] The purpose of this invention is to provide Burkholderia paraknegrius NRS-2-16, its inoculant, and its application, in order to help solve or improve the problem that there are few strains in the prior art that can simultaneously achieve biological control of cotton Verticillium wilt and cotton boll blight.

[0009] To achieve the above objectives, the present invention provides the following technical solution: a Paraburkholderia fungorum NRS-2-16, wherein the Paraburkholderia fungorum NRS-2-16 is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 38750.

[0010] The present invention also provides a microbial agent, which adopts the following technical solution: a microbial agent containing Burkholderia paraknegrius NRS-2-16 as described above.

[0011] Preferably, the concentration of Burkholderia paraknegriensis NRS-2-16 in the bacterial agent is 1×10⁻⁶. 6 cfu / mL -1×10 8 cfu / mL.

[0012] The present invention also provides a method for preparing a bacterial agent, which adopts the following technical solution: the method for preparing the bacterial agent as described above includes the following steps: (1) culturing the *U. paraknechoica* NRS-2-16 in beef extract tryptone agar medium; (2) inoculating the *U. paraknechoica* NRS-2-16 obtained in step (1) into beef extract tryptone liquid medium and culturing it, adjusting the concentration of the *U. paraknechoica* NRS-2-16 to obtain the bacterial agent.

[0013] Preferably, in step (1), the culture temperature is 25-28℃ and the culture time is 48-72h; in step (2), the culture temperature is 25-28℃ and the culture time is 72-96h; the culture method is oscillation culture with a rotation speed of 160-200r / min.

[0014] The present invention also provides the application of *U. parakylcholine* NRS-2-16 or its agent as described above, which adopts the following technical solutions: the application of *U. parakylcholine* NRS-2-16 or its agent as described above in the control of cotton Verticillium wilt; and / or, the application of *U. parakylcholine* NRS-2-16 or its agent in the control of cotton boll blight.

[0015] Preferably, *U. parakk. Holdella* NRS-2-16 or the aforementioned agent is used to inhibit *Verticillium dahliae*; and / or, *U. parakk. Holdella* NRS-2-16 or the aforementioned agent is used to inhibit *Phytophthora ramie*.

[0016] Preferably, the procedure includes the following steps: during the growth of cotton, inoculating with Burkholderia paraknegrius NRS-2-16 or the aforementioned inoculum; the inoculum is inoculated into the soil where cotton is grown.

[0017] Beneficial effects: The *Paraburkholderia fungorum* NRS-2-16 strain of this invention is deposited at the China General Microbiological Culture Collection Center (No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing) on ​​May 19, 2026, with accession number CGMCC No. 38750. *Paraburkholderia fungorum* NRS-2-16 of this invention exhibits significant inhibitory effects against *Verticillium dahliae*, the causal agent of cotton wilt, and *Phytophthora ramie*, the causal agent of boll blight, and can be used for the control of these diseases.

[0018] Based on the *Burkholderia paraknegrius* NRS-2-16 of this invention, corresponding microbial agents or biopesticides can be developed to provide safe, efficient, and environmentally friendly microbial resources for the green integrated control of cotton Verticillium wilt and boll blight, reduce the use of chemical pesticides, ensure cotton yield and quality, and promote the sustainable development of the cotton industry.

[0019] The microbial agent of the present invention can be applied with water using a cotton film drip irrigation device, thereby achieving precise control of water, fertilizer and microorganisms under drip irrigation conditions. Attached Figure Description

[0020] The accompanying drawings, which form part of this application, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. Wherein: Figure 1 This is a colony morphology diagram of Burkholderia paraknegaceticus NRS-2-16, which is the present invention.

[0021] Figure 2 Phylogenetic tree of Burkholderia paraknegaceae NRS-2-16.

[0022] Figure 3 The figure shows the experimental results of the antagonistic effect of Burkholderia paraknegaceae NRS-2-16 on Verticillium dahliae.

[0023] Figure 4 The figure shows the experimental results of the antagonistic effect of Burkholderia paraknegaceae NRS-2-16 against Phytophthora ramie.

[0024] Figure 5 Figure 1 shows the results of a pot experiment using an agent containing Burkholderia paraknegrius NRS-2-16 to control Verticillium wilt in cotton.

[0025] Figure 6 The image shows the gel electrophoresis results of 16S rDNA from Burkholderia paraknegrius NRS-2-16. Detailed Implementation

[0026] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention are within the scope of protection of the present invention.

[0027] The present invention will now be described in detail with reference to embodiments. It should be noted that, unless otherwise specified, the embodiments and features described in the embodiments of the present invention can be combined with each other.

[0028] This invention addresses the problem that there are few strains in the existing technology that can simultaneously achieve biological control of cotton Verticillium wilt and cotton boll blight, by providing a strain of *Paraburkholderia fungorum* NRS-2-16.

[0029] Paraburkholderia fungorum NRS-2-16, an embodiment of the present invention, is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 38750.

[0030] The *Burkholderia paraknockii* NRS-2-16 of this invention has a significant inhibitory effect on *Verticillium dahliae* (the causal agent of cotton wilt) and *Phytophthora dauricum* (the causal agent of boll blight), and can be used for the prevention and control of cotton wilt and boll blight.

[0031] Based on the *Burkholderia paraknegrius* NRS-2-16 of this invention, corresponding microbial agents or biopesticides can be developed to provide safe, efficient, and environmentally friendly microbial resources for the green integrated control of cotton Verticillium wilt and boll blight, reduce the use of chemical pesticides, ensure cotton yield and quality, and promote the sustainable development of the cotton industry.

[0032] The present invention also proposes a bacterial agent, wherein the bacterial agent of the present invention embodiment contains Burkholderia paraknegrius NRS-2-16 as described above.

[0033] In a preferred embodiment of the bacterial agent of the present invention, the concentration of Burkholderia paraknegriensis NRS-2-16 in the bacterial agent is 1 × 10⁻⁶. 6 cfu / mL -1×10 8 cfu / mL (e.g., 1×10⁻⁶) 6 cfu / mL, 5×10 6 cfu / mL, 1×10 7 cfu / mL, 5×10 7 cfu / mL or 1×10 8 (cfu / mL).

[0034] The present invention also proposes a method for preparing the bacterial agent as described above. The method for preparing the bacterial agent in the embodiment of the present invention includes the following steps: (1) culturing the *U. parakk. hold* NRS-2-16 in beef extract tryptone agar medium; (2) inoculating *U. parakk. hold* NRS-2-16 into beef extract tryptone liquid medium and culturing, adjusting the concentration of *U. parakk. hold* NRS-2-16 to obtain the bacterial agent.

[0035] In a preferred embodiment of the preparation method of the microbial agent of the present invention, in step (1), the culture temperature is 25-28℃ (e.g., 25℃, 26℃, 27℃ or 28℃), and the culture time is 48-72h (e.g., 48h, 54h, 60h, 66h or 72h); in step (2), the culture temperature is 25-28℃ (e.g., 25℃, 26℃, 27℃ or 28℃), and the culture time is 72-96h (e.g., 72h, 80h, 88h or 96h); the culture method is shaking culture, and the rotation speed is 160-200r / min (e.g., 160r / min, 170r / min, 180r / min, 190r / min or 200r / min).

[0036] This invention also proposes the application of *U. parakylcholine* NRS-2-16 or its inoculum as described above, specifically the application of *U. parakylcholine* NRS-2-16 or its inoculum in controlling cotton Verticillium wilt; and / or, the application of *U. parakylcholine* NRS-2-16 or its inoculum in controlling cotton boll blight. In a preferred embodiment of the application of the present invention, *U. parakk. Holder* NRS-2-16 or its agent is used to inhibit *Verticillium dahliae*; and / or, *U. parakk. Holder* NRS-2-16 or its agent is used to inhibit *Phytophthora ramie*.

[0037] In a preferred embodiment of the application of the present invention, the following steps are included: during the growth of cotton, inoculating with Burkholderia paraknegrius NRS-2-16 or a bacterial agent.

[0038] Preferably, the microbial agent is inoculated into the soil where cotton is grown.

[0039] The following detailed description of the *Burkholderia paraknegrius* NRS-2-16, the inoculum agent, and its application, through specific embodiments, provides a detailed explanation of the present invention.

[0040] Unless otherwise specified, all reagents used in the following examples are commercially available; the main raw materials used in the following examples are from the following manufacturers: beef extract, tryptone, and PDA medium are manufactured by Beijing Coollab Technology Co., Ltd.; agar is manufactured by Beijing Solarbio Technology Co., Ltd.; sodium chloride is manufactured by Tianjin Damao Chemical Reagent Factory; and the Ezup column-based bacterial genomic DNA extraction kit and SanTaq Plus PCR amplification kit are manufactured by Sangon Biotech (Shanghai) Co., Ltd.

[0041] Example 1 Isolation, screening and identification of Burkholderia paraknegaceae NRS-2-16: The sample was isolated and screened from cotton rhizosphere soil samples in Weixian County, Hebei Province in 2025; the dilution-spreading method was used for isolation. The specific steps are as follows: (1) Weigh 10g of freshly collected soil and pour it into an Erlenmeyer flask containing 90mL of sterile water. Shake for 10min to obtain a suspension with a concentration of 1 / 10. Let it stand at room temperature for 30min, then take 1mL of the supernatant to prepare a suspension with a concentration of 10. -4 Or 10 -5 Prepare a diluted solution of the desired concentration.

[0042] (2) Sterilize the dissolved culture medium (the culture medium is beef extract tryptone agar medium, the components of which include: 10g beef extract, 5g tryptone, 5g sodium chloride, 18g agar and 1000ml distilled water, pH 7.0~7.2) and pour it into a sterilized petri dish; after the culture medium solidifies, take 20μL of the diluted solution obtained in step (1) and drop it onto the surface of the solidified culture medium, spread it evenly with a sterile spreader, and invert the petri dish in a 28℃ constant temperature incubator for 1~2 days.

[0043] (3) After the colonies grow, select single colonies that are round, yellow, flat, with relatively neat edges, and moist and smooth, and streak them on beef extract tryptone agar medium. Repeat the isolation and purification process three times. The final single colony (e.g.) Figure 1 As shown), it is named NRS-2-16.

[0044] (4) Physiological and biochemical identification and 16S rDNA sequencing were performed on the single colonies obtained from the above culture: The results of the physiological and biochemical identification are shown in Table 1 below: Table 1. Physiological and Biochemical Identification Results

[0045] The 16S rDNA of strain NRS-2-16 was amplified and sequenced: DNA extraction was performed using the Ezup column-based bacterial genomic DNA extraction kit, and 16S rDNA amplification was performed using the SanTaq Plus PCR amplification kit. Specifically: Table 2 PCR reaction system for 16S rRNA amplification

[0046] Table 3 PCR reaction conditions

[0047] The 16S rDNA of the amplified NRS-2-16 was subjected to gel electrophoresis and then sent to a sequencing company for sequencing. The results of gel electrophoresis are as follows Figure 6 As shown in the figure, a clear band appears at approximately 1.5 kb, indicating that the PCR successfully amplified the target 16S rDNA fragment of NRS-2-16.

[0048] The sequencing results showed that the 16S rDNA sequence length of strain NRS-2-16 was 1467 bp; the sequence was as follows:

[0049] The 16S rDNA sequencing results were compared with the sequences in GenBank, and the results showed that the gene sequence of strain NRS-2-16 was 99.79% homologous to that of Paraburkholderia fungorum strain PNG-April.

[0050] Based on the above identification results of strain NRS-2-16, strain NRS-2-16 was determined to belong to the fungus *Paraburkholderia fungorum*, and this strain was named *Paraburkholderia fungorum* NRS-2-16. For the phylogenetic tree of *Paraburkholderia fungorum* NRS-2-16 and related strains based on the 16S rRNA sequence, please refer to... Figure 2 .

[0051] Example 2 Confrontation culture assay to determine the inhibitory effect of the fungus *Paraburkholderia fungorum* NRS-2-16 on cotton verticillium wilt and cotton boll blight pathogens: Take a 6mm diameter dish containing Verticillium dahliae (the causal agent of cotton wilt) and Phytophthora bovis (the causal agent of cotton boll rot). Place the Verticillium dahliae (Verticillium dahliae) and Phytophthora bovis (Phytophthora ramie) dishes in the center of a petri dish containing PDA. At a distance of 2.5cm from the dish on all sides, use an inoculation loop containing strain NRS-2-16 to make a circular mark. Incubate Verticillium dahliae at 28℃ for 3 days and measure the colony diameter. Incubate Phytophthora ramie for 7 days and measure the colony diameter.

[0052] The experimental results show that strain NRS-2-16 can strongly inhibit the growth of cotton Verticillium wilt and cotton boll blight.

[0053] Strain NRS-2-16 strongly inhibited the growth of Verticillium dahliae (the causal agent of cotton verticillium wilt) and Phytophthora dauricum (the causal agent of cotton boll rot), with inhibition rates reaching 92.7% and 92.7%, respectively. Figure 3 The left figure shows the experimental results of the CK group (uninoculated with strain NRS-2-16); the right figure shows the experimental results of the treatment group (inoculated with strain NRS-2-16) and 65.4% ( Figure 4 The left figure shows the experimental results of the CK group without inoculation with strain NRS-2-16; the right figure shows the experimental results of the treatment group inoculated with strain NRS-2-16. Among them, the inhibition rate (%) = colony diameter (CK) - colony diameter (treatment group) / colony diameter (CK).

[0054] Example 3 Pot experiments were conducted to determine the control effect of strain NRS-2-16 against cotton Verticillium wilt: Preparation of NRS-2-16 bacterial agent: NRS-2-16 was transferred to fresh beef extract tryptone agar medium and cultured at 28℃ for 2 days. Then, it was inoculated into a 500mL Erlenmeyer flask containing 100mL of beef extract tryptone liquid medium (components: 10g beef extract, 5g tryptone, 5g sodium chloride, and 1000mL distilled water, pH=7.0~7.2). The culture was incubated at 28℃ on a rotary shaker at 180r / min for 72–96 hours to obtain the bacterial suspension. The suspension was diluted with distilled water and, under a microscope, adjusted to show 2–3 spores (1.0 × 10⁻¹⁰) in the smallest square of a hemocytometer. 7 The cfu / mL concentration is preferably used to obtain the NRS-2-16 bacterial agent of this embodiment (prepare fresh before use).

[0055] Preparation of Verticillium dahliae solution: Verticillium dahliae present at 4℃ was transferred to fresh PDA medium, sealed with sealing film, and then incubated upside down in a dark incubator at 25℃ for 7 days. The grown colonies were then divided into approximately 1cm blocks using an inoculation needle. These blocks were placed in PDA medium and incubated on a shaker (25℃, 130rpm). After filtering the solution through gauze to remove mycelia, a spore suspension was obtained. The suspension was diluted with distilled water and, under a microscope, adjusted to 2-3 spores (1.0 × 10⁻⁶) in the smallest square of a hemocytometer. 7 The concentration of cfu / mL should be ideal, and the solution should be prepared fresh each time it is needed.

[0056] Wash the 8cm×8cm×6cm flowerpots with clean water, then wipe them with 70% alcohol by volume and let them dry. Fill each flowerpot with 100g of nutrient soil and sow 6 cotton seeds per pot at a depth of 2cm-3cm. Place the seedling pots in a 28cm×44cm plastic tray, with 15 flowerpots in each tray. Add 1L of tap water to each tray to allow the soil to absorb water naturally. Place the plastic trays in an artificial climate chamber for cultivation. The cultivation conditions are: 16 hours of daytime, temperature 24℃, humidity 50%, light 60%; 8 hours of nighttime, temperature 20℃, humidity 70%, light 0%.

[0057] During the seedling stage, water with the same amount of tap water every 4 days to promote germination. Tender shoots will emerge in about 3 days. The cotton seedlings will grow to one leaf and one heart in about 15 days. N+V group: When cotton seedlings reach the stage of one leaf and one bud, inoculate with NRS-2-16 inoculant (directly inoculated into the soil), 20 mL (1.0 × 10⁻⁶) per pot. 7 CFU / mL). After approximately 20 days, when the cotton seedlings have grown to two leaves and a central bud, inoculate them (directly into the soil) with a *Verticillium dahliae* solution (1.0 × 10⁻⁶ CFU / mL). 720 mL (cfu / mL). Twenty days after inoculation with Verticillium dahliae, the incidence of Verticillium wilt in cotton seedlings was observed, and the disease index was investigated.

[0058] CK group: When cotton seedlings reach the stage of one leaf and one bud, inoculate with sterile water (directly into the soil), 20 mL per pot. After about 20 days, when the cotton seedlings reach the stage of two leaves and one bud, inoculate with 20 mL of sterile water (directly into the soil). After 20 days, observe the incidence of Verticillium wilt in the cotton seedlings and investigate the disease index.

[0059] Group V: When cotton seedlings reach the stage of one leaf and one bud, inoculate with sterile water (directly into the soil), 20 mL per pot. After approximately 20 days, when the seedlings reach the stage of two leaves and one bud, inoculate (directly into the soil) with a *Verticillium dahliae* solution (1.0 × 10⁻⁶). 7 20 mL (cfu / mL). After 20 days, observe the incidence of Verticillium wilt in cotton seedlings and investigate the disease index.

[0060] The method for investigating the disease index is as follows: Grade 0: The plant is healthy, with well-developed stems and leaves and no disease symptoms; Grade 1: 1-2 cotyledons of the plant are yellow, and a few true leaves are withered, yellow, and curled; Grade 2: One true leaf is infected, turning yellow and wilting, with dry, curled, or brown mottled edges; Level 3: Two or more true leaves are affected, turning yellow and wilting, with dry and curled leaf edges or the leaves falling off, leaving only the heart leaves; Level 4: All leaves have fallen off or the plant has died; Calculation formula: Disease index = Σ (number of diseased plants at each level × corresponding disease level) / total number of plants surveyed × highest disease level; The control effect of the inoculant in this embodiment on cotton Verticillium wilt is as follows: Figure 4 ( Figure 4 The following table shows the test results for the CK group, V group, and N+V group, arranged from left to right: Table 4. Control efficacy of NRS-2-16 inoculant against Verticillium wilt.

[0061] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A strain of *Paraburkholderia fungorum* NRS-2-16, characterized in that, The Paraburkholderia fungorum NRS-2-16 strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 38750.

2. A microbial agent, characterized in that, The bacterial agent contains Burkholderia paraknegaceticus NRS-2-16 as described in claim 1.

3. The microbial agent as described in claim 2, characterized in that, The concentration of Burkholderia paraknegriensis NRS-2-16 in the bacterial agent is 1×10⁻⁶. 6 cfu / mL -1×10 8 cfu / mL.

4. The method for preparing the microbial agent as described in claim 2, characterized in that, Includes the following steps: (1) Burkholderia paraknegrius NRS-2-16 was cultured on beef extract tryptone agar medium; (2) Inoculate the Burkholderia paraknee NRS-2-16 obtained in step (1) into beef extract tryptone liquid medium and adjust the concentration of the Burkholderia paraknee NRS-2-16 to obtain the bacterial agent.

5. The method for preparing the microbial agent as described in claim 4, characterized in that, In step (1), the culture temperature is 25-28℃ and the culture time is 48-72h; In step (2), the culture temperature is 25-28℃ and the culture time is 72-96h; the culture method is shaking culture with a rotation speed of 160-200r / min.

6. The application of Burkholderia paraknegriensis NRS-2-16 as described in claim 1 or the inoculant as described in any one of claims 2-3 in the control of Verticillium wilt in cotton; and / or, The application of the aforementioned Burkholderia paraknegrius NRS-2-16 or the aforementioned inoculum in the prevention and control of boll blight.

7. The application as described in claim 6, characterized in that, Burkholderia paraknegaceae NRS-2-16 or the aforementioned agent is used to inhibit Verticillium dahliae; and / or, Burkholderia paraknegaceae NRS-2-16 or the aforementioned agent is used to inhibit Phytophthora ramie.

8. The application as described in claim 6, characterized in that, The process includes the following steps: during the growth of cotton, inoculating it with Burkholderia paraknockhausensis NRS-2-16 or the aforementioned inoculum; The microbial agent is inoculated into the soil where cotton is grown.

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