Biological control method for guava root-knot nematode

Abstract

The invention discloses a biological prevention and control method for guava root-knot nematodes, and belongs to the technical field of biological prevention and control of diseases and insect pests, and the biological prevention and control method for guava root-knot nematodes comprises the following steps: adding a complex microbial inoculant into excrement, the method comprises the following steps: fermenting a mixture of pseudomonas fluorescens ACCC 03936, pseudomonas fluorescens ACCC 01144, pseudomonas aeruginosa ACCC 60044 and proteusbacillus vulgaris ACCC 11766 to obtain a biological fermentation solution, applying the biological fermentation solution twice in a young psidium guajave shoot germination period and a nematode infection peak period, and performing conventional field management after the biological fermentation solution is applied. The method can effectively prevent and treat guava root-knot nematode infection, is ecological and environment-friendly, is simple and convenient, and can obviously improve the yield and quality of guavas.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological pest control, and particularly relates to a biological control method for root-knot nematode of Psidium guajava. BACKGROUND

[0002] Root-knot nematode (Meloidogyne spp.) is an important pathogenic organism that seriously threatens agricultural production and has a negative impact on most crops in the world. With the expansion of protected cultivation modes and the continuous increase of crop multiple cropping indexes, the occurrence of root-knot nematode disease is becoming more and more serious, and the current management measures cannot completely control the damage of root-knot nematode. Using chemical nematicides is still the most common short-term management strategy for controlling root-knot nematode at present.

[0003] Root-knot nematode is a major soil-borne disease of Psidium guajava, which invades the root system to form root knots, hinders water and nutrient absorption, causes tree vigor to weaken and yield to decrease, and can even cause plant death in severe cases. Chemical nematicides (such as abamectin) have problems of residual pollution, drug resistance and soil micro-ecological destruction.

[0004] Therefore, it is of great significance to develop a biological control method for effectively controlling root-knot nematode of Psidium guajava for green control of root-knot nematode disease of Psidium guajava. SUMMARY

[0005] To solve the above technical problems, the application provides a biological control method for root-knot nematode of Psidium guajava, which has great significance for green control of root-knot nematode disease of Psidium guajava.

[0006] To achieve the above purpose, the application provides a biological control method for root-knot nematode of Psidium guajava, comprising the following steps:

[0007] A compound microbial agent is added to the feces, the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766, and a biological fermentation liquor is obtained after fermentation;

[0008] The biological fermentation liquor is applied twice at the new shoot germination stage and the peak period of nematode infection of Psidium guajava, and then routine field management is performed.

[0009] The 2,4-diacetylphloroglucinol (2,4-DAPG) and pyoluteorin secreted by P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144 can directly destroy the body wall or nervous system of G. gossypioides. Both of the two P. fluorescens can secrete yellow-green fluorescent siderophores, and the ability of chelating iron ions is more than 100 times of ordinary microorganisms. The hatching of nematode eggs and the survival of juveniles are highly dependent on iron elements. The siderophores can significantly inhibit the reproduction of G. gossypioides by reducing the environmental iron concentration. P. aeruginosa ACCC 60044 has the ability to degrade aromatic compounds, and can decompose the attractant signal substances (such as phenylacetic acid) secreted by nematodes in the soil, reducing the aggregation of G. gossypioides to guava roots. Alginate lyase secreted by P. aeruginosa ACCC 60044 can also destroy the structure of nematode eggshell, which is complementary to the antibiotics produced by P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144. The optimum growth temperature of P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144 is 25-30℃, while P. aeruginosa ACCC 60044 still maintains metabolic activity at low temperature of 10-15℃. In the low temperature period of guava planting area (such as winter in South China), P. aeruginosa ACCC 60044 can continuously play a role in prevention and control, filling the gap of the decline of the activity of core strains. P. putida ACCC 11766 has strong protein-degrading ability, which can degrade macromolecular organic matter (such as cellulose and protein) in feces, and release more small molecular carbon sources (such as glucose and amino acids) for P. fluorescens and P. aeruginosa. Thiamine (vitamin B1) secreted by P. putida ACCC 11766 can also promote the growth of P. fluorescens and P. aeruginosa, and increase the number of viable bacteria in the biological fermentation broth. At the same time, the phenylalanine ammonia-lyase produced by P. putida ACCC 11766 can inhibit the growth of some pathogenic fungi (such as Fusarium), reducing the risk of complex infection of G. gossypioides and fungal diseases.

[0010] Further, the mass ratio of the P. fluorescens ACCC 03936, P. fluorescens ACCC 01144, P. aeruginosa ACCC 60044 and P. putida ACCC 11766 is (1.0-1.5):(1.0-1.5):(0.8-1.2):(0.5-0.8).

[0011] Further, the mass ratio of the P. fluorescens ACCC 03936, P. fluorescens ACCC 01144, P. aeruginosa ACCC 60044 and P. putida ACCC 11766 is (1.0-1.2):(1.0-1.2):(0.8-1.0):(0.5-0.6).

[0012] Further, the complex bacterial agent accounts for 5-8% of the dry weight of the feces.

[0013] Further, the feces are cow feces or sheep feces, and the moisture content of the feces is adjusted to 55-65% and the carbon-nitrogen ratio (C / N) is (25-30):1 before fermentation.

[0014] Further, the fermentation conditions are as follows: temperature 28-32 DEG C, aeration amount 0.5-1.0 VVM (volume / volume / minute), fermentation time 5-7 days, and fermentation end point pH value 7.0-7.5.

[0015] Further, before the complex bacterial agent is added to the feces, 1-3% of molasses or 0.5-1.5% of humic acid is added as a fermentation accelerator, accounting for 1-3% of the dry weight of the feces.

[0016] Further, when the bio-fermentation liquid is applied, 0.1-0.3% of potassium silicate is synchronously added, accounting for 0.1-0.3% of the mass of the bio-fermentation liquid. After being absorbed by the root system, the potassium silicate is deposited on the cell wall to form a "siliconized layer" (SiO2·nH2O gel), which significantly enhances the mechanical strength of the cell wall: hinders the penetration of nematode stylets into the root epidermis; reduces the invasion success rate of second instar juveniles of root-knot nematodes. Silicon elements can also activate plant defense signaling pathways: promote the expression of key genes in the salicylic acid (SA) and jasmonic acid (JA) pathways, and promote the synthesis of resistance proteins; increase the secretion of chitinase, beta-1,3-glucanase and other nematode-inhibiting substances by the root system. Potassium silicate can also provide potassium nutrition, neutralize soil pH, promote the robustness of guava root systems, expand the colonization space of probiotics, improve the rhizosphere microecology, and inhibit nematode egg hatching.

[0017] Further, the guava new shoot germination period is from March to April, and the nematode infection peak period is from June to July.

[0018] Further, a circular ditch with a depth of 15-20 cm is opened along the drip line of the crown, and 80-100 mL of the bio-fermentation liquid is applied to each plant each time, and the soil is irrigated thoroughly after covering.

[0019] Compared with the prior art, the present application has the following advantages and technical effects:

[0020] 1) The present application first compounding Pseudomonas fluorescens double strains with Pseudomonas aeruginosa and Proteus vulgaris, breaking through the functional limitations of single bacterial species.

[0021] 2) The present application integrates four mechanisms of antibiotic killing, nutritional competition, environmental regulation and biodegradation, and realizes the integration of "prevention-treatment-repair".

[0022] 3) The present application is suitable for two-stage intervention of guava new shoots germination period and nematode infection peak period, filling the blank of the prior art for low temperature period protection.

[0023] 4) The method of the present application can effectively prevent and control the infection of guava root-knot nematode, is ecological and environmentally friendly, simple in method, and can significantly improve the yield and quality of guava. DETAILED DESCRIPTION

[0024] The various illustrative embodiments of the present application will now be described in detail below. This description is not to be considered limiting in scope, but rather as merely one explanation of certain aspects of the application, character, and embodiments.

[0025] It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the present application. Additionally, the use of the term "about" in relation to a value or a range of values is to be understood as encompassing each individual value or range of values falling within the range of the stated value or range of values. In any statement of a value or a range of values, the intermediate values and the smaller ranges within the range of the stated value or range of values are also included in the present application. The upper and lower limits of these smaller ranges can independently be included or excluded from the range.

[0026] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application, preferred methods and materials are described. All publications mentioned in this specification are herein incorporated by reference to disclose and describe the methods and / or materials in connection with which the publications are cited. The citation of any reference in this specification is not intended as an admission that the reference is prior art, but rather that the reference is part of the technical literature that is relevant to the disclosure.

[0027] Many modifications and variations of this application specification can be made in light of the above teachings without departing from the spirit or scope of the application. Other implementations of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application disclosed herein. The specification and examples given are exemplary only.

[0028] As used herein, the terms "comprises", "comprising", "includes", "including", "has", "having", and the like are open-ended terms that are intended to mean including, but not limited to.

[0029] The embodiment of the present application provides a biological control method for guava root-knot nematode, comprising the following steps:

[0030] The complex microbial agent is added into the feces, the complex microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766, and a biological fermentation liquor is obtained after fermentation;

[0031] The biological fermentation liquor is applied twice at the germination stage of new shoots of Psidium guajava and the peak period of nematode infection, and then routine field management is performed.

[0032] In the embodiment of the present application, the mass ratio of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 is (1.0-1.5):(1.0-1.5):(0.8-1.2):(0.5-0.8). Preferably, the mass ratio of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 is (1.0-1.2):(1.0-1.2):(0.8-1.0):(0.5-0.6).

[0033] The 2,4-diacetylphloroglucinol (2,4-DAPG) and pyoluteorin secreted by P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144 can directly destroy the body wall or nervous system of the root-knot nematode. Both of the two P. fluorescens can secrete yellow-green fluorescent siderophores, and the ability of chelating iron ions is more than 100 times of that of ordinary microorganisms. The hatching of nematode eggs and the survival of larvae are highly dependent on iron elements. The siderophores can significantly inhibit the reproduction of the root-knot nematode by reducing the environmental iron concentration. P. aeruginosa ACCC 60044 has the ability to degrade aromatic compounds, and can decompose the attractant signal substances (such as phenylacetic acid) secreted by the nematode in the soil, thereby reducing the aggregation of the root-knot nematode to the guava root system. The alginate lyase secreted by P. aeruginosa ACCC 60044 can also destroy the structure of the nematode egg shell, which is complementary to the antibiotics produced by P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144. The optimum growth temperature of P. fluorescens ACCC 03936 and P. fluorescens ACCC 01144 is 25-30℃, while P. aeruginosa ACCC 60044 still maintains metabolic activity at low temperature of 10-15℃. In the low temperature period of the guava planting area (such as the winter in South China), P. aeruginosa ACCC 60044 can continuously play a role in prevention and control, and fill the gap of the decline of the activity of the core strain. P. putida ACCC 11766 has strong protein-degrading ability, and can degrade macromolecular organic matter (such as cellulose and protein) in feces to release more small-molecule carbon sources (such as glucose and amino acids) for the use of P. fluorescens and P. aeruginosa. The thiamine (vitamin B1) secreted by P. putida ACCC 11766 can also promote the growth of P. fluorescens and P. aeruginosa, and increase the number of viable bacteria in the biological fermentation broth. At the same time, the phenylalanine ammonia-lyase produced by P. putida ACCC 11766 can inhibit the growth of some pathogenic fungi (such as Fusarium), thereby reducing the risk of complex infection of the root-knot nematode and fungal diseases.

[0034] In the embodiment of the present application, the compound microbial agent accounts for 5-8% of the dry weight of the feces.

[0035] In the embodiment of the present application, the feces are cow feces or sheep feces, and the moisture content of the feces is adjusted to 55%-65% and the carbon-nitrogen ratio (C / N) is (25-30):1 before fermentation.

[0036] In the embodiment of the present application, the fermentation conditions are as follows: temperature 28-32℃, aeration amount 0.5-1.0 VVM (volume / volume / minute), fermentation time 5-7 days, and pH value at the end of fermentation 7.0-7.5.

[0037] In the embodiments of the present application, before adding the complex microbial agent into the feces, 1-3% molasses or 0.5-1.5% humic acid is added as a fermentation promoter, accounting for 1-3% of the dry weight of the feces. The molasses can provide a transiently available carbon source for microorganisms, increase the biomass of the bacterial population, shorten the logarithmic growth phase of the bacterial population, and advance the enzyme / antibacterial substance synthesis phase. The reducing sugars (fructose, glucose) in the molasses can act as an electron donor, reduce the Eh (oxidation-reduction potential) of the fermentation broth to below -150 mV, promote the secretion of chitinase by facultative anaerobes (Proteus vulgaris), and improve the inhibition of Meloidogyne incognita. The quinone group (-C=O) in humic acid acts as an electron shuttle, which can activate the bacterial quorum sensing system, accelerate the formation of biofilm, induce Pseudomonas aeruginosa to express stress-resistant genes, and improve the survival rate in the field. The humic acid-bacterial complex is adsorbed on the soil particles to form a "microbial library", which can achieve a slow-release effect and prolong the control effect. At the same time, humic acid and the complex microbial agent can synergistically inhibit Meloidogyne incognita.

[0038] In the embodiments of the present application, when the biological fermentation broth is applied, 0.1-0.3% potassium silicate is added synchronously, accounting for 0.1-0.3% of the mass of the biological fermentation broth. After being absorbed by the root system, the potassium silicate is deposited on the cell wall to form a "siliconized layer" (SiO2·nH2O gel), which significantly enhances the mechanical strength of the cell wall: hinders the penetration of nematode stylets into the root epidermis; and reduces the success rate of invasion of second instar larvae of root-knot nematodes. Silicon elements can also activate plant defense signaling pathways: promote the expression of key genes in the salicylic acid (SA) and jasmonic acid (JA) pathways, and promote the synthesis of resistance proteins; increase the secretion of chitinase, β-1,3-glucanase and other nematode-inhibiting substances by the root system. Potassium silicate can also provide potassium nutrition, neutralize soil pH, promote the robustness of Psidium guajava root system, expand the colonization space of probiotics, improve the rhizosphere microecology, and inhibit the hatching of nematode eggs. Moreover, potassium silicate has no growth inhibition on Pseudomonas fluorescens, Pseudomonas aeruginosa and Proteus vulgaris.

[0039] In the embodiments of the present application, the Psidium guajava new shoot germination period is from March to April, and the peak period of nematode infestation is from June to July.

[0040] In the embodiments of the present application, a circular ditch with a depth of 15-20 cm is opened along the drip line of the tree crown, and 80-100 mL of biological fermentation broth is applied to each plant each time, and the soil is irrigated thoroughly after covering.

[0041] VVM is the abbreviation of Air Volume per Culture Volume per Minute, its full meaning is the ratio of air volume per minute to the actual liquid volume of the tank, and it can also be understood as the amount of air required per cubic meter of fermentation liquid per minute under standard conditions. VVM reflects the proportion of air volume per unit time into the fermentation liquid and the volume of the fermentation liquid, which is an important parameter for measuring the intensity of oxygen supply in the fermentation process. For example: VVM = 1 indicates that the air volume per minute is equal to the volume of the fermentation liquid (such as 10L of fermentation liquid per minute 10L of air). VVM = 1.5 indicates that the air volume per minute is 1.5 times the volume of the fermentation liquid (such as 10L of fermentation liquid per minute 15L of air).

[0042] Unless otherwise specified, the room temperature in the present application is 25±2℃.

[0043] It should be noted that the part not described in detail in the present application is the conventional operation means in the art, and is not the focus of the present application. For example, the field management method of Psidium guajava can be managed according to the conventional management method in the art.

[0044] The strains used in the present application are obtained by purchase. As an example, Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 are purchased from China Agricultural Microbial Culture Collection Center; the purchased strains need to be activated, prepared seed liquid and prepared fermentation liquid before use. The activation, preparation of seed liquid and preparation of fermentation liquid are realized by using conventional technical means, and the effective viable bacteria content in the fermentation liquid of the four kinds of bacteria is (6-8) × 10 9 CFU / mL; four kinds of fermentation liquid are respectively prepared into four kinds of microbial agents (lyophilized powder) after spray drying.

[0045] The technical solutions of the present application are further illustrated by the following examples.

[0046] Example 1

[0047] A biological control method of Psidium guajava root-knot nematode, the steps are as follows:

[0048] The compound microbial agent is added in the cow dung, the compound microbial agent accounts for 6% of the dry weight of the cow dung, the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.2:1.2:1.0:0.6), the moisture content of the cow dung is adjusted to 60% before fermentation, the carbon-nitrogen ratio is 28:1, the fermentation is carried out under the condition that the temperature is 30 DEG C and the ventilation amount is 0.8 VVM for 6 days, the pH value at the end of fermentation is 7.2, and the biological fermentation liquor is obtained after the fermentation is completed;

[0049] The biological fermentation liquor is applied twice at the germination period (March-April) and the peak period of nematode infection (June-July) of the new shoots of Psidium guajava, the application process is that a circular ditch with a depth of 18 cm is opened along the drip line of the tree crown, 90 mL of the biological fermentation liquor is applied to each plant each time, the soil is covered and then water is poured, and routine field management is carried out after application.

[0050] Example 2

[0051] A biological control method of Psidium guajava root-knot nematode, the steps are as follows:

[0052] The compound microbial agent is added in the sheep dung, the compound microbial agent accounts for 8% of the dry weight of the sheep dung, the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.0:1.0:0.8:0.5), the moisture content of the sheep dung is adjusted to 55% before fermentation, the carbon-nitrogen ratio is 25:1, the fermentation is carried out under the condition that the temperature is 28 DEG C and the ventilation amount is 1.0 VVM for 7 days, the pH value at the end of fermentation is 7.0, and the biological fermentation liquor is obtained after the fermentation is completed;

[0053] The biological fermentation liquor is applied twice at the germination period (March-April) and the peak period of nematode infection (June-July) of the new shoots of Psidium guajava, the application process is that a circular ditch with a depth of 15 cm is opened along the drip line of the tree crown, 80 mL of the biological fermentation liquor is applied to each plant each time, the soil is covered and then water is poured, and routine field management is carried out after application.

[0054] Example 3

[0055] A biological control method of Psidium guajava root-knot nematode, the steps are as follows:

[0056] The compound microbial agent is added in the cow dung, the compound microbial agent accounts for 5% of the dry weight of the cow dung, the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.5:1.5:1.2:0.8), the moisture content of the cow dung is adjusted to 65% before fermentation, the carbon-nitrogen ratio is 30:1, the fermentation is carried out under the condition that the temperature is 32 DEG C and the ventilation amount is 0.5 VVM for 5 days, the pH value at the end of fermentation is 7.5, and the biological fermentation liquor is obtained after the fermentation is completed;

[0057] The biological fermentation liquor is applied twice at the new shoot germination period (March-April) and the peak period of nematode infection (June-July) of Psidium guajava, the application process is that a circular ditch with a depth of 20 cm is opened along the drip line of the tree crown, 100 mL of the biological fermentation liquor is applied to each plant each time, the soil is covered and then water is poured, and routine field management is carried out after application.

[0058] Example 4

[0059] A biological control method of Psidium guajava root-knot nematode, the steps are as follows:

[0060] The compound microbial agent is added in the cow dung, the compound microbial agent accounts for 6% of the dry weight of the cow dung, 3% of the dry weight of the cow dung is added before the compound microbial agent is added, the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.2:1.2:1.0:0.6), the moisture content of the cow dung is adjusted to 60% before fermentation, the carbon-nitrogen ratio is 28:1, the fermentation is carried out under the condition that the temperature is 30 DEG C and the ventilation amount is 0.8 VVM for 6 days, the pH value at the end of fermentation is 7.2, and the biological fermentation liquor is obtained after the fermentation is completed;

[0061] The biological fermentation liquor is applied twice at the new shoot germination period (March-April) and the peak period of nematode infection (June-July) of Psidium guajava, the application process is that a circular ditch with a depth of 18 cm is opened along the drip line of the tree crown, 90 mL of the biological fermentation liquor is applied to each plant each time, the soil is covered and then water is poured, and routine field management is carried out after application.

[0062] Example 5

[0063] A biological control method of Psidium guajava root-knot nematode, the steps are as follows:

[0064] The compound microbial agent is added into the cow dung, and the compound microbial agent accounts for 6% of the dry weight of the cow dung. Before adding the compound microbial agent, 1.0% of humic acid in the dry weight of the cow dung is added. The compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.2:1.2:1.0:0.6). The moisture content of the cow dung is adjusted to 60% before fermentation. The carbon-nitrogen ratio is 28:1. The fermentation is carried out at a temperature of 30 DEG C and an aeration amount of 0.8 VVM for 6 days. The pH value at the end of fermentation is 7.2. The biological fermentation liquor is obtained after the fermentation is completed.

[0065] The biological fermentation liquor is applied twice at the germination period (March-April) and the peak period of nematode infection (June-July) of Psidium guajava. The application process is as follows: a circular ditch with a depth of 18 cm is opened along the drip line of the tree crown. 90 mL of the biological fermentation liquor is applied to each plant each time. After covering the soil, the water is poured thoroughly. After the application, the routine field management is carried out.

[0066] Example 6

[0067] A biological control method of Psidium guajava root-knot nematode is provided, and the steps are as follows:

[0068] The compound microbial agent is added into the cow dung, and the compound microbial agent accounts for 6% of the dry weight of the cow dung. The compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766 (mass ratio is 1.2:1.2:1.0:0.6). The moisture content of the cow dung is adjusted to 60% before fermentation. The carbon-nitrogen ratio is 28:1. The fermentation is carried out at a temperature of 30 DEG C and an aeration amount of 0.8 VVM for 6 days. The pH value at the end of fermentation is 7.2. The biological fermentation liquor is obtained after the fermentation is completed.

[0069] The biological fermentation liquor is applied twice at the germination period (March-April) and the peak period of nematode infection (June-July) of Psidium guajava. The application process is as follows: a circular ditch with a depth of 18 cm is opened along the drip line of the tree crown. 90 mL of the biological fermentation liquor is applied to each plant each time. 0.2% of potassium silicate in the mass of the biological fermentation liquor is synchronously added. After covering the soil, the water is poured thoroughly. After the application, the routine field management is carried out.

[0070] Comparative Example 1

[0071] A biological control method of Psidium guajava root-knot nematode is provided, which is the same as example 1, except that the compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936 and Pseudomonas fluorescens ACCC 01144, and the mass ratio of the two is 1:1.

[0072] Comparative Example 2

[0073] A biological control method of Meloidogyne incognita, same as example 1, the only difference is that the complex microbial agent is a mixture of Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766, and the mass ratio of the two is 1:1.

[0074] Experimental example 1 indoor trapping experiment

[0075] The test crop of this experimental example is guava, and the test soil is sterilized at a high temperature of 80°C in advance, and no root-knot nematode is found to survive after inspection. The guava seedlings are transplanted into the cultivation box for use.

[0076] The test nematode is the second instar larva of Meloidogyne incognita, and before the experiment starts, Meloidogyne incognita is inoculated into the soil at a ratio of 100 per 1000 grams of soil. The temperature in the cultivation box is controlled at a temperature suitable for the growth of Meloidogyne incognita (28°C), and then healthy guava seedlings are transplanted into the cultivation box. After the crop is planted for 5 days, sufficient water is poured into the experimental group and the blank control group at one time, respectively, and the water in the experimental group is added with the biological fermentation liquid prepared in examples 1-6 and comparative examples 1-2 of the present application (the application amount is 2 mL / kg of soil), and the blank control group is a treatment group without adding any drug preparation. One day after application, the experimental group and the control group are cultivated in a conventional manner, respectively, and after 3 days, the nematode population density in the soil in each cultivation box is sampled and checked, and the results are shown in Table 1.

[0077] Table 1

[0078] Meloidogyne incognita density (nematodes / kg soil) Reduction rate (%) Example 1 15±21.8 85 Example 2 18±2.1 82 Example 3 22±2.2 78 Example 4 8±1.2 92 Example 5 10±1.5 90 Example 6 6±0.9 94 Comparative Example 1 45±3.0 55 Comparative Example 2 50±3.2 50 Blank control group 98±3.5 2 (natural attenuation)

[0079] Note:

[0080] As can be seen from Table 1, the nematode density of the blank control group is close to the initial value (98 per kg), indicating that the natural mortality rate of the nematode is very low without intervention. The density of the base complex microbial agent in examples 1-3 ranges from 15 to 22 per kg, and the reduction rate is 78%-85%, due to the synergistic effect of the four bacteria (Pseudomonas fluorescens secretes DAPG / PCA to destroy the nematode body wall, and Proteus secretes chitinase to dissolve the eggshell). The nematode density of the optimized group of examples 4-6 is further reduced to 6-10 per kg, which may be because example 4 adds molasses, which can provide readily available carbon sources to promote the synthesis of nematode-inhibiting substances by the bacterial community. Example 5 adds humic acid, which can complex trace elements and activate the stress response genes of the bacterial community, prolonging the soil colonization period. Example 6 adds potassium silicate, which can form a siliconized layer on the roots, physically blocking the invasion of nematodes. Comparative example 1 only contains Pseudomonas fluorescens, and lacks chitinase, so the egg hatching inhibition is insufficient; comparative example 2 lacks Pseudomonas fluorescens, and the production of bacteriostatic substances (DAPG / HCN) is low.

[0081] Experimental example 2 field experiment

[0082] Taking the bio-fermentation liquor prepared by the examples 1, 5-6 as an example, the guavas farmland seriously damaged by root-knot nematode (the average content of root-knot nematode in soil is 92 per kg soil) is selected for experiment, at the beginning of the experiment, the bio-fermentation liquor prepared by the examples 1, 5-6 is applied in soil at one time (along the drip line of the crown, the annular ditch with the depth of 18 cm, 90 mL of bio-fermentation liquor is applied to each plant, and the soil is covered after watering, and the routine farmland management can be carried out after the application); the control group is watered with the same amount of water, and no any medicine is added, and the subsequent normal watering culture can be carried out. The nematode population density in the soil of the experimental group farmland and the control group farmland is sampled and checked after 3 days of the experiment, three sampling checking points are extracted from the experimental group and the control group farmland respectively, the nematode population density in the soil is checked respectively, and the average value is calculated, and the result is shown in table 2.

[0083] Table 2

[0084] Nematode density (nematodes / kg soil) Significance of difference from control group Example 1 19.3±2.8 ** Example 5 12.6±1.9 *** Example 6 7.8±1.9 *** Control group 87.4±4.2 -

[0085] Note: “**” represents P<0.01 (very significant difference); “*** represents” P<0.001 (highly significant difference).

[0086] The above is only the preferred specific embodiment of the present application, but the protection scope of the present application is not limited to this, any person skilled in the art can easily think of the changes or replacements within the technical range disclosed by the present application, which should be covered in the protection scope of the present application.

Claims

1. A biological control method for guava root-knot nematodes, characterized in that, Includes the following steps: A compound microbial agent is added to the feces. The compound microbial agent is a mixture of Pseudomonas fluorescens ACCC 03936, Pseudomonas fluorescens ACCC 01144, Pseudomonas aeruginosa ACCC 60044 and Proteus vulgaris ACCC 11766. After fermentation, a bio-fermentation broth is obtained. The bio-fermentation liquid is applied twice, once during the guava shoot emergence period and once during the peak nematode infestation period. After application, conventional field management can be carried out.

2. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, The mass ratio of *Pseudomonas fluorescens* ACCC 03936, *Pseudomonas fluorescens* ACCC 01144, *Pseudomonas aeruginosa* ACCC 60044, and *Proteus vulgaris* ACCC 11766 was (1.0-1.5):(1.0-1.5):(0.8-1.2):(0.5-0.8).

3. The biological control method for guava root-knot nematodes according to claim 2, characterized in that, The mass ratio of *Pseudomonas fluorescens* ACCC 03936, *Pseudomonas fluorescens* ACCC 01144, *Pseudomonas aeruginosa* ACCC 60044, and *Proteus vulgaris* ACCC 11766 was (1.0-1.2):(1.0-1.2):(0.8-1.0):(0.5-0.6).

4. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, The compound microbial agent accounts for 5-8% of the dry weight of the feces.

5. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, The excrement is cow dung or sheep dung, and the moisture content of the excrement is adjusted to 55%-65% before fermentation, with a carbon-to-nitrogen ratio of (25-30):

1.

6. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, The fermentation conditions are: temperature 28-32℃, aeration rate 0.5-1.0 VVM, fermentation time 5-7 days, and final pH value of 7.0-7.

5.

7. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, Before adding the compound microbial agent to the feces, molasses or humic acid, accounting for 1%-3% of the dry weight of the feces, is also added.

8. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, When applying the bio-fermentation broth, potassium silicate at a mass of 0.1%-0.3% is added simultaneously.

9. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, The guava shoots sprout in March-April, and the nematode infestation peaks in June-July.

10. The biological control method for guava root-knot nematodes according to claim 1, characterized in that, Dig a circular trench 15-20cm deep along the drip line of the tree canopy, apply 80-100mL of the bio-fermentation liquid to each tree each time, cover with soil and water thoroughly.