CN110184221B

CN110184221B - Bacillus belgii and application thereof in prevention and treatment of phytophthora root rot of tomato

Info

Publication number: CN110184221B
Application number: CN201910486020.7A
Authority: CN
Inventors: 李宝聚; 谢学文; 孙雪莹; 李磊; 石延霞; 柴阿丽
Original assignee: Institute of Vegetables and Flowers Chinese Academy of Agricultural Sciences
Current assignee: Institute of Vegetables and Flowers Chinese Academy of Agricultural Sciences
Priority date: 2019-06-05
Filing date: 2019-06-05
Publication date: 2021-03-05
Anticipated expiration: 2039-06-05
Also published as: CN110184221A

Abstract

The invention discloses a Bacillus belgii strain and application thereof in preventing and treating phytophthora root rot of tomato. The invention provides Bacillus velezensis (Bacillus velezensis) ZF50, which has a preservation number of CGMCC No.16775 in China general microbiological culture Collection center. The invention provides a high-efficiency biocontrol strain for preventing and treating phytophthora root rot of tomatoes, and has great production significance for green prevention and control technology of tomato diseases and promotion of sustainable development of tomato industry.

Description

Bacillus belgii and application thereof in prevention and treatment of phytophthora root rot of tomato

Technical Field

The invention relates to the technical field of biological control of plant diseases, in particular to a Bacillus belgii strain and application thereof in control of phytophthora root rot of tomato.

Background

The Phytophthora root rot of tomatoes is a destructive fungal soil-borne disease caused by Phytophthora capsici Leonian (Phytophthora capsici Leonian), has the characteristics of rapid occurrence and strong destructiveness, has a short occurrence period, has the morbidity of 50 percent from morbidity to death of only 7 to 15 days, even is dead, is one of important diseases for tomato production in protected areas and open areas of many countries, and seriously restricts the development of the tomato industry. At present, few methods for preventing and treating phytophthora root rot of tomatoes are reported, and the selection of a disease-resistant variety is an economic and effective prevention and treatment measure, but the difficulty of disease-resistant breeding is high, while the traditional agricultural measures have limitations, chemical prevention is simple, convenient, quick and effective, but has poor degradability and is easy to cause environmental pollution, and the problem of drug resistance is highlighted day by singly using similar chemical agents such as metalaxyl and the like for a long time, so that a green prevention and treatment measure is urgently needed in production to ensure the sustainable development of the vegetable industry.

In recent years, biological control has attracted much attention because it is environmentally friendly and safe to humans and animals, and shows a good effect in disease control technologies. Proved by a large amount of time, the stability, the compatibility and the consistency of the Bacillus (Bacillus) are all superior to those of other biocontrol bacteria in the disease control research. At present, antagonistic bacillus is utilized in China to successfully develop various biocontrol bacillus for preventing and treating various diseases, and certain achievements are achieved.

Disclosure of Invention

In order to overcome the defects of the prior control technology, the invention aims to provide a Bacillus belgii strain and application thereof in controlling phytophthora root rot of tomato, and simultaneously has strong antagonistic effect on various pathogenic fungi such as multi-major corynebacterium sp.

In a first aspect, the invention claims a strain of bacillus belgii.

The Bacillus belgii required to be protected by the invention is specifically Bacillus belgii (Bacillus velezensis) ZF50, which is preserved in China general microbiological culture Collection center (CGMCC) in 11-26.2018 with the preservation number of CGMCC No. 16775.

In a second aspect, the presently claimed bacillus belgii is a microbial preparation.

The microbial preparation claimed in the present invention contains the Bacillus belgii (Bacillus velezensis) ZF 50.

Further, the biocontrol agent can be prepared by the Bacillus belief (Bacillus velezensis) ZF50 and an auxiliary agent or a substrate. When the biocontrol agent is in a liquid form, the effective content of the Bacillus belgii (Bacillus velezensis) ZF50 may be 1.0 × 10⁸cfu·mL^-1。

In a third aspect, the invention claims a bacterial suspension.

The content of Bacillus velezensis ZF50 bacterial suspension required by the invention is 1.0 multiplied by 10⁷～1.0×10⁹cfu/mL。

Further, the content of Bacillus velezensis ZF50 is 1.0 × 10⁸cfu/mL。

In a fourth aspect, the invention claims the use of said Bacillus belgii (Bacillus velezensis) ZF50 or said microbial preparation or said bacterial suspension in any of:

(A1) preventing and treating phytophthora root rot of tomatoes;

(A2) preparing a product for preventing and treating phytophthora root rot of tomatoes.

(B1) promoting the growth and development of plants;

(B2) preparing a product for promoting the growth and development of plants;

further, the plant is a vegetable;

further, the vegetable is tomato.

Wherein the promotion of plant growth and development is the promotion of plant height increase and/or fresh weight increase and/or root length increase and/or root weight increase.

In a fifth aspect, the invention claims the use of said Bacillus belgii (Bacillus velezensis) ZF50 or said microbial preparation or said bacterial suspension in any of:

(C1) inhibiting plant pathogenic fungi;

(C2) preparing a product for inhibiting phytopathogenic fungi;

(C3) preventing and controlling diseases caused by plant pathogenic fungi;

(C4) preparing products for preventing and treating diseases caused by plant pathogenic fungi.

Wherein the plant pathogenic fungi are Phytophthora capsici (Phytophthora Leonian), Corynespora cassicola (Corynespora cassicola), Fusarium oxysporum (Fusarium oxysporum), Rhizoctonia solani (Rhizoctonia solani), Colletotrichum (Colletotrichum capsicium), Staphylomyces aureus (Stemphylium) and/or Botrytis cinerea Pers (Botrytis cinerea).

In a sixth aspect, the invention claims the use of said Bacillus belgii (Bacillus velezensis) ZF50 or said microbial preparation or said bacterial suspension in any of:

(D1) producing IAA;

(D2) preparing a product for producing IAA.

Further, in the production of IAA, tryptophan (tryptophan can promote the production of IAA by Bacillus velezensis ZF 50) may be added.

In a seventh aspect, the invention claims a method for preventing and treating phytophthora root rot of tomato.

The method for preventing and treating the phytophthora root rot of the tomato comprises the following steps: applying the Bacillus velezensis ZF50 or the microbial preparation or the bacterial suspension to the rhizosphere soil of a tomato plant (e.g. irrigating the root in the plant rhizosphere soil).

Wherein the application amount of the Bacillus velezensis ZF50 during root irrigation can be 10 mL/strain, and 10mL means that the content of 10mL is 1.0 multiplied by 10⁸cfu·mL^-1The Bacillus velezensis ZF 50.

In an eighth aspect, the invention claims the use of said Bacillus belgii (Bacillus velezensis) ZF50 for the preparation of said microbial preparation or said bacterial suspension.

The invention has the beneficial effects that:

(1) a pot experiment is utilized to determine the control effect of Bacillus velezensis ZF50 on phytophthora rot of tomatoes and the growth promotion effect of tomato seedlings in different inoculation amounts, and the result shows that the Bacillus velezensis ZF50 can effectively control the phytophthora rot of tomatoes, the pot control effect on the phytophthora rot of tomatoes is the best when 10mL of Bacillus velezensis ZF is inoculated, and the disease index and the control effect are respectively 6.00 and 60.39%.

(2) Bacillus velezensis ZF50 has good growth promoting effect on tomato seedlings, the growth rate is 72.83%, the fresh weight growth rate is 37.26%, and the root length and root weight growth rate are 8.44% and 22.68% respectively. The field control effect of Bacillus velezensis ZF50 on phytophthora root rot of tomato is 87.5% through field tests, which shows that the strain has great development potential in the control of phytophthora root rot of tomato.

The invention provides an excellent biocontrol strain for preventing and treating phytophthora root rot of tomatoes, and has great production significance for green prevention and control technology of tomato diseases and promotion of sustainable development of tomato industry.

Deposit description

The strain name: bacillus belgii

Latin name: bacillus velezensis

According to the biological materials (strains): ZF50

The preservation organization: china general microbiological culture Collection center

The preservation organization is abbreviated as: CGMCC (China general microbiological culture Collection center)

Address: xilu No.1 Hospital No. 3 of Beijing market facing Yang district

The preservation date is as follows: 11/26/2018

Registration number of the preservation center: CGMCC No.16775

Drawings

FIG. 1 shows the inhibition effect of ZF50 strain on Phytophthora solani. Control on the left; the plate inhibition effect of the strain ZF50 on Phytophthora capsici is shown on the right.

Fig. 2 is a phylogenetic tree of ZF50 strain based on the 16S rDNA sequence.

Fig. 3 is a phylogenetic tree of ZF50 strain based on gyrB sequence.

Fig. 4 shows the bacteriostatic effect of ZF50 on 6 pathogenic fungi. A: stemphylium; b: corynespora casilicola; c: colletotrichum capsicii; d: botrytis cinerea; e: rhizoctonia solani; f: fusarium oxysporum;

FIG. 5 shows the results of the measurement of the yield of ZF50IAA strain.

Detailed Description

The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The various media referred to in the following examples are standard media commonly used in the art, for example:

LB liquid medium: 10.0g of peptone, 5.0g of yeast extract, 5.0g of sodium chloride and distilled water till the volume is 1000 mL.

Oat culture medium: 30g of oat, 20g of glucose, 17-20 g of agar powder and distilled water, wherein the volume of the distilled water is up to 1000 mL.

PDA culture medium: 200g of peeled potatoes, 20g of glucose, 20g of agar powder and distilled water which are made up to 1000 mL.

The following examples relate to the preparation of chemicals commonly used in the art, for example:

SolutionⅠ：10mmol^-1phosphoric acid.

SolutionⅡ：1mL 0.5mol·L^-1FeCl₃Dissolved in 50mL of 35% HClO₄。

Example 1 isolation, purification and characterization of Bacillus velezensis ZF50

1. Isolation of Bacillus

Weighing 10g of soil sample from the perennial continuous cropping soil of tomatoes collected from Shandong Shouguang, putting the soil sample into a triangular flask containing 100mL of sterile water, and placing the triangular flask on a shaking table at 27 ℃ and 200r/min to shake for 30min so as to fully and uniformly mix the soil sample. Treating 1mL of bacterial soil suspension in 80 deg.C water bath for 10min, and diluting to 10%^-1，10^-2，10^-3，10^-4The suspension of the soil (2) was dropped into the center of an LB plate by sucking 100. mu.L of each suspension of the gradient, and the suspension was spread with a spreader so that the plate was sufficiently absorbed, each gradient was repeated 3 times, and the plate was inverted and cultured in a 30 ℃ incubator for 2 d. Selecting bacterial colonies with different forms, carrying out plate streaking purification, numbering and storing, and carrying out co-separation to obtain 98 strains of bacillus.

2. Screening of Bacillus

And 5 mu L of biocontrol bacterium suspension is connected to the four corner points at the position 3 cm from the center of the dish by using the sterilization gun head, 3 times of repetition are carried out, a blank contrast only connected to the liquid LB culture medium is set, and the culture is carried out for 5 days at the temperature of 27 ℃. When the blank control is about to grow over the whole culture dish, the control growth amount (colony radius) and the treatment growth amount (growth radius after inoculation of bacteria) of the target bacteria are measured and expressed by the bacteriostasis rate.

Inhibition (%) - ((control colony diameter-treated colony diameter)/control colony diameter. times.100%)

The results show that the inhibition rate of the total 5 strains of biocontrol bacteria on phytophthora capsici is more than 55%, wherein the inhibition rate of the strain ZF50 is the best (figure 1), and the inhibition rate is 76% (table 1), so the strain ZF50 is selected as the phytophthora solani root rot antagonist.

TABLE 1 inhibition rate of five biocontrol bacteria on Phytophthora solani

Treatment of	Concentration (cfu. mL)^-1)	Colony diameter (mm)	Bacteriostatic ratio (%)
				ZF 23	1×10⁸	0.91±0.17	63.6±8a
ZF 26	1×10⁸	1.0±0.15	60±6a
				ZF 30	1×10⁸	1.0±0.25	60±4a
ZF
	50	1×10⁸	0.6±0.2	76±4.8a b
ZF 53					1×10⁸	0.93±0.1	62.8±4b
	LB control	1×10⁸	2.5	--

Note: different lower case letters indicate significant differences at the 0.05 level.

3. Identification of Strain ZF50

The strain ZF50 was identified as follows:

(1) strain ZF50 culture character observation

The single colony of the purified strain ZF50 on an LB culture medium is milky white, round, wrinkled on the surface, sunken in the center of the colony and volcano-mouthed.

(2) Analysis of physiological and biochemical characteristics

With reference to the method of donxiu beads et al, gram stain, fructose, maltose, cellobiose, glucose, lactose, sucrose, NaCl, inositol, aspartic acid, etc. of the strain ZF50 were analyzed. The results are shown in Table 2.

TABLE 2 physio-biochemical characteristics of Strain ZF50

Item	Reaction of	Item	Reaction of
				Gram stain	+	Lactose	-
Glycerol	-	Sucrose	+
				Fructose	+	4％NaCl	+
Maltose	-	8％NaCl	+
				Cellobiose	w	Inositol	-
Glucose	+	Aspartic acid	-

The results of physiological and biochemical analysis show that: the strain ZF50 is gram-positive, fructose, glucose, sucrose and NaCl are positive, cellobiose is weak positive, other indexes are negative, and the strain is primarily identified to belong to Bacillus belief.

(3)16s rDNA sequence analysis and phylogenetic tree establishment

The DNA of the strain ZF50 is extracted by using a bacterial DNA extraction kit purchased from Tiangen Biochemical technology Co., Ltd, and PCR amplification is carried out by using the extracted DNA as a template, wherein the amplification primer adopts a bacterial universal primer. The 16rDNA primer information is F27: 5'-TACGGTACCTTGTTACGACT-3', R1492: 5'-CTGAGCCAGGATCA AACT-3' are provided. The primers were synthesized by Beijing Bomaide Biotechnology GmbH, and the reaction system (50. mu.L) was: 2 XTaq PCR Master Mix 25. mu.L, upstream and downstream primers 1.25. mu.L each, DNA template 2.5. mu.L, ddH₂O make up to 50. mu.L. Reaction procedure: pre-denaturation at 94 ℃ for 5 min; denaturation at 94 ℃ for 30s, annealing at 60 ℃ for 30s, and extension at 72 ℃ for 30s for 35 cycles; extension was supplemented at 72 ℃ for 10 min. The phylogenetic tree was constructed using the MEGA5.0 adjacency (Neighbor-Joining, NJ).

A partial effective sequence (SEQ ID No.1) of a strain ZF 5016S rDNA gene and NCBI accession number MH978894 are obtained through PCR amplification, the sequence is compared with a known sequence in GenBank, multiple comparison is carried out by MEGA5.0 software, a strain ZF50 strain 16S rDNA gene sequence phylogenetic tree is constructed by adopting an adjacent method, and the result shows that the genetic relationship between the strain ZF50 and a strain BH21 (accession number KT889364) is nearest (figure 2), the sequence similarity is 99 percent, and only 1 base difference exists.

(4) gyrB gene sequence analysis and identification and construction of phylogenetic tree thereof

The DNA of ZF50 is extracted by adopting a bacterial DNA extraction kit of Tiangen Biochemical technology Co., Ltd, and the extracted DNA is taken as a template. The gyrB primer information is UP 1F: 5 '-GAAGTCATCATGACCGTTCTGCAYGCNGG NGGNAARTTYGA-3'; UP 2R: 5 '-AGCAGGGTACGGATGTGCGAGCCRTCNACR TCN GCRTCNGTCAT-3'; the reaction system (50. mu.L) was: 2 XTaq PCR Master Mix 25. mu.L, upstream and downstream primers 1.25. mu.L each, DNA template 2.5. mu.L, ddH₂O make up to 50. mu.L. Reaction procedure: pre-denaturation at 94 ℃ for 5 min; denaturation at 94 ℃ for 30s, annealing at 60 ℃30s, extension at 72 ℃ for 30s, for 35 cycles; extension was supplemented at 72 ℃ for 10 min. The sequencing work was done by Beijing Bomaide Biotech, and Blast sequence homology alignment was performed via Genbank as a result of sequencing.

The effective sequence (SEQ ID No.2) of the strain ZF50gyrB gene part is obtained through PCR amplification and the NCBI accession number is MK238275, the sequence is used for being compared with the known sequence in GenBank, MEGA5.0 software is used, the strain ZF50gyrB gene sequence phylogenetic tree is constructed by adopting an adjacent method, the phylogenetic tree of the strain ZF50 can be obtained, and the result shows that the strain ZF50 and the Bellis bacillus strain JT3-1 (accession number is CP032506) are gathered in the same branch, the sequence similarity is 100%, and no base difference exists (figure 3).

By combining the morphological, physiological and biochemical characteristics and the genome sequence determination result, the strain ZF50 is finally determined to be Bacillus velezensis (Bacillus velezensis).

The strain ZF50 has been deposited in China general microbiological culture Collection center (CGMCC). Address: western road No.1, north chen west road, north kyo, chaoyang, institute of microbiology, china academy of sciences, zip code 100101. The preservation date is 2018, 11 and 26. The preservation number is CGMCC No. 16775. The strain name: bacillus belgii; latin name: bacillus velezensis; according to the biological materials (strains): ZF 50.

Example 2 determination of the bacteriostatic Effect of Bacillus velezensis ZF50 on six pathogenic fungi

Inoculating pathogenic fungi onto a PDA flat plate for 24h, and inoculating 5 mu L of Bacillus velezensis (ZF 50) suspension (1.0 x 10) with a sterilizing gun head at four corner points 3 cm away from the fungus cake⁸cfu·mL^-1) In 3 replicates, a blank of liquid LB medium alone was inoculated and incubated at 27 ℃ for 5 days. When the blank control is about to grow over the whole culture dish, the control growth amount (colony diameter) and the treatment growth amount (growth diameter after inoculation of bacteria) of the target bacteria are measured and expressed by the bacteriostasis rate.

The results show (fig. 4 and table 3) that Bacillus belief (Bacillus velezensis) ZF50 has a good bacteriostatic effect on 6 tested pathogenic bacteria, wherein the bacteriostatic effect on colletotrichum is the highest, the bacteriostatic rate is 80.73%, is significantly higher than that of other 5 strains of pathogenic bacteria, and the bacteriostatic rate on botrytis cinerea is the lowest, and is 53.88%. The above results indicate that Bacillus velezensis ZF50 has a broad spectrum of bacteriostatic action.

TABLE 3 bacteriostatic effect of Bacillus velezensis ZF50 on 6 pathogenic fungi

Treatment of	Bacteriostatic ratio (%)	Treatment of	Bacteriostatic ratio (%)
				Fusarium oxysporum	54.44±1.11a	Exotidium torvum	71.05±3.94c
Staphylococcus aureus	79.44±0.66a	Rhizoctonia solani	65.73±1.16d
				Colletotrichum	80.73±0.85b	Botrytis cinerea	53.88±0.56d

Example 3 controlling Effect of Bacillus velezensis ZF50 on Phytophthora tomato

1. Potted plant control effect of strain ZF50 on phytophthora root rot of tomato

Preparation of phytophthora capsici: activating phytophthora capsici on an oat culture medium, and inoculating the bacterial disease on pumpkin for propagation for later use; preparing biocontrol bacillus: activating a strain ZF50 on an LB plate, selecting a single colony by using an inoculating needle, inoculating the single colony in an LB nutrient solution, culturing at 37 ℃ at 200r/min for 12-16 h, and preparing into a product of 1 multiplied by 10⁸cfu·mL^-1Biocontrol bacteria suspension; preparing biocontrol bacillus cultivation soil: 1000g of matrix soil is respectively packed in a plug (5 × 5cm), 1mL, 5mL and 10mL of Bacillus belgii ZF50 bacterial suspension (with the concentration of 1 × 10) is respectively added into each plug hole⁸cfu·mL^-1). Preparing pyraclostrobin cultivation soil: 1000g of matrix soil are separately loaded in a hole tray (5 multiplied by 5cm), 10mL of the matrix soil with the concentration of 0.25 mg/kg is added into each hole^-1The pyraclostrobin liquor.

And sowing the hastened tomato seeds in each substrate soil added with the good biocontrol bacteria, and taking the treatment without adding the Bacillus belgii ZF50 bacterial suspension as a control. Transplanting to a nutrition pot mixed with phytophthora capsici when the plant grows to 3 true leaves (the content of phytophthora capsici is 2 multiplied by 10)⁶cfu·mL^-1) And (5) performing normal-temperature management, and beginning to investigate the disease condition of the plants after the plants are subjected to clear water control.

The disease investigation adopts tomato root rot grading standard: level 0: no disease; level 1: slight water-stain-like lesions appear on the stems; and 2, stage: the lesion spots on the stem are expanded but not exceed the plant height 1/4, and the plant does not wilt; and 3, level: the diseased part exceeds the whole plant 1/4, the diseased part extends downwards to the root part and does not exceed the plant height 3/4, the stem base is sunken and wilted; 4, level: the diseased part exceeds the whole plant or spreads to the whole plant, including roots and petioles, the stem base is seriously overflowed, the leaves are withered, and the disease is dead.

Disease index ═ Σ (number of diseased plants at each stage × corresponding stage)/(total number of investigated plants × maximum stage) × 100;

the preventing and treating effect (%) is (blank control disease index-treatment disease index)/blank control disease index x 100.

The results show (table 4) that inoculation with 10mL of ZF50 bacterial suspension gave the best control, with disease index and control of 6 and 60.39%, respectively. The control effect of inoculating 5mL of ZF50 bacterial suspension is the worst, and the disease index and the control effect are 14% and 34.17% respectively.

TABLE 4 potted control of Phytophthora solani by Strain ZF50

Treatment of	Concentration (cfu/mL)	Index of disease condition	Control effect (%)
				ZF50 1mL	1×10⁸	10	47±7.52a
ZF50 5mL	1×10⁸	14	34.17±3.29b
				ZF50 10mL	1×10⁸	6	60.39±11.37c
CK	--	21.25	--

2. Strain ZF50 field control effect on phytophthora root rot of tomato

The experiment was carried out with a total of 3 treatments, strain ZF50 (1.0X 10) suspension⁸cfu·mL^-1. ) Root irrigation treatment, 25% pyraclostrobin OD 0.25 mg-kg^-1Root irrigation treatment and clear water control, 4 replicates per treatment, 6 tomato seedlings per replicate. Transplanting the seedlings into a nursery after 3-4 true leaves of the seedlings are planted, irrigating the roots with 10 mL/seed of the prepared ZF50 bacterial suspension, and beginning to investigate the disease condition of the plants after the control is ill.

The disease investigation method is the same as above.

The results show (table 5) that the disease index and control effect of strain ZF50 were 20.83% and 68.75%, respectively, and the disease index and control effect of the control agent were 18 and 73%, respectively.

The results indicate that the field control effect of the strain ZF50 is higher than that of the control medicament.

The Bacillus belgii ZF50 is a biocontrol strain with strong inhibition activity on phytophthora root rot of tomato.

TABLE 5 field control of Phytophthora tomato by Strain ZF50

Treatment of	Concentration of	Index of disease condition	Control effect (%)
				ZF50 10mL	1×10⁸Cfu·mL^-1	20.83	68.75a
Pyraclostrobin (Kresoxim-methyl)	0.25mg·kg^-1	18	73b
				CK	--	66.66	--

Example 4 application of Bacillus velezensis ZF50 in tomato growth promotion

When the tomato seedlings have 2 leaves and 1 heart, transplanting the tomato seedlings into a nutrition pot (5cm multiplied by 5cm), and inoculating 15mL of Bacillus velezensis ZF50 bacterial suspension (1 multiplied by 10)⁸Cfu·mL^-1) And (3) repeating the treatment for 3 times, repeating 10 seedlings for each time, wherein the monitoring period is two weeks, and then counting biological indexes of the overground part plant height, the overground part fresh quality, the root length, the root weight and the like of the seedlings to analyze the growth promoting effect of the strains.

The results show that: the tomato seedlings inoculated with the ZF50 bacterial suspension had a plant height growth rate of 72.83%, a fresh weight growth rate of 37.26%, and root length and root weight growth rates of 8.44% and 22.68%, respectively (Table 6).

TABLE 6 growth promoting Effect of Strain ZF50 on tomato seedlings

Note: the different lower case letter representations were significantly different at the 0.05 level

Example 5 determination of IAA production of Bacillus velezensis ZF50

Bacillus velezensis ZF50 suspension was inoculated into a 2mL centrifuge tube containing 1.5mL DF + (Tryptophan-containing DF medium) medium at a final concentration of 1X 10 ZF50⁸Cfu·mL^-1. Culturing at 28 ℃ for 7 days, centrifuging at 12000rpm for 5min, taking 1mL of supernatant, adding 50 μ L of Solution I and 2mL of Solution II into a test tube, mixing uniformly, reacting at room temperature for 25min, detecting absorbance at 530mm wavelength by using a spectrophotometer, and calculating the IAA yield by using a standard curve according to the obtained OD value.

The results show (fig. 5): the strain ZF50 has the capability of producing IAA, tryptophan can induce the strain ZF50 to produce IAA, the strain ZF50 has the largest IAA production amount on DF + culture medium, and the secretion amount can reach 9.976 mug.mL^-1。

The present invention is not limited to the above-described embodiments, and any variations, modifications, and substitutions which may occur to those skilled in the art may be made without departing from the spirit of the invention.

Sequence listing

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<120> Bacillus belgii and application thereof in prevention and treatment of phytophthora root rot of tomato

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Claims

1. Bacillus velezensis (Bacillus velezensis) ZF50, wherein the preservation number of the Bacillus velezensis ZF in China general microbiological culture Collection center is CGMCC No. 16775.

2. A microbial preparation characterized by: the microbial preparation contains Bacillus belgii (Bacillus velezensis) ZF50 as claimed in claim 1.

3. A bacterial suspension characterized by: the bacterial suspension is the bacterial suspension of Bacillus velezensis ZF50 as claimed in claim 1, wherein the content of Bacillus velezensis ZF50 is 1.0 x 10⁷~1.0×10⁹Cfu/mL。

4. A bacterial suspension according to claim 3, wherein: the content of Bacillus velezensis ZF50 is 1 × 10⁸Cfu·mL^-1。

5. Use of a Bacillus belief (Bacillus velezensis) ZF50 as claimed in claim 1 or a microbial preparation as claimed in claim 2 or a bacterial suspension as claimed in claim 3 or 4 in any of the following:

(A1) preventing and treating phytophthora root rot of tomatoes;

6. Use of a Bacillus belief (Bacillus velezensis) ZF50 as claimed in claim 1 or a microbial preparation as claimed in claim 2 or a bacterial suspension as claimed in claim 3 or 4 in any of the following:

(B1) promoting the growth and development of plants;

(B2) preparing the product for promoting the growth and development of plants.

7. Use according to claim 6, characterized in that: the plant is vegetable.

8. Use according to claim 7, characterized in that: the vegetable is tomato.

9. Use of a Bacillus belief (Bacillus velezensis) ZF50 as claimed in claim 1 or a microbial preparation as claimed in claim 2 or a bacterial suspension as claimed in claim 3 or 4 in any of the following:

(C1) inhibiting plant pathogenic fungi;

(C2) preparing a product for inhibiting phytopathogenic fungi;

(C3) preventing and controlling diseases caused by plant pathogenic fungi;

10. Use according to claim 9, characterized in that: the plant pathogenic fungi are Phytophthora capsici (Phytophthora capsici Leonians), Phytophthora polyspora (Corynespora cassicola), Fusarium oxysporum (Fusarium oxysporum), Rhizoctonia solani (Rhizoctonia solani), Colletotrichum (Colletotrichum capsicii), Staphylomyces aureus (Stemphylium) and/or Botrytis cinerea Pers.

11. Use of a Bacillus belief (Bacillus velezensis) ZF50 as claimed in claim 1 or a microbial preparation as claimed in claim 2 or a bacterial suspension as claimed in claim 3 or 4 in any of the following:

(D1) producing IAA;

(D2) preparing a product for producing IAA.

12. A method for preventing and controlling phytophthora root rot of tomatoes comprises the following steps: applying Bacillus subtilis (Bacillus velezensis) ZF50 according to claim 1 or the microbial preparation according to claim 2 or the bacterial suspension according to claim 3 or 4 to the rhizosphere soil of tomato plants.

13. Use of Bacillus belief (Bacillus velezensis) ZF50 according to claim 1 for the preparation of a microbial preparation according to claim 2 or a suspension according to claim 3 or 4.