CN113249250A

CN113249250A - Pseudomonas aeruginosa 9# and application thereof

Info

Publication number: CN113249250A
Application number: CN202110480700.5A
Authority: CN
Inventors: 林晓燕; 陈铭学; 曹赵云; 杨欢
Original assignee: China National Rice Research Institute
Current assignee: China National Rice Research Institute
Priority date: 2021-04-30
Filing date: 2021-04-30
Publication date: 2021-08-13
Anticipated expiration: 2041-04-30
Also published as: CN113249250B

Abstract

The invention discloses pseudomonas aeruginosa 9# and application thereof, and belongs to the technical field of microorganisms. The invention discloses a pseudomonas aeruginosa strain 9#, the preservation number of which is CCTCC NO: m2021178. The pseudomonas aeruginosa strain 9# disclosed by the invention can inhibit the growth of the ustilaginoidea virens strain YY7850, which is an example that the pseudomonas aeruginosa strain is found to have antagonism against ustilaginoidea virens pathogenic bacteria for the first time, and the strain has the potential of being developed into rice ustilaginoidea virens biocontrol bacteria due to the fact that the strain 9# has the capacity of inhibiting the ustilaginoidea virens pathogenic bacteria.

Description

Pseudomonas aeruginosa 9# and application thereof

Technical Field

The invention relates to the technical field of microorganisms, in particular to pseudomonas aeruginosa 9# and application thereof.

Background

False smut is a kind of ear fungal disease of rice caused by infection of Ustilaginodea virens (Cooke) Takahashi, which is a kind of green smut, and glume, glume and grain are infected in late stage of rice growth, commonly called "harvest disease". False smut is a worldwide disease of rice, occurring in many countries such as china, japan, usa, india, philippines, etc. In recent years, the number of diseases has been upgraded from secondary diseases on rice to one of the major diseases. A large amount of nutrients are consumed in the rice false smut germ breeding process, so that not only are the rice false smut grains damaged, but also the nutrition delivery of adjacent grains is influenced, the blight rate is increased, the setting rate and the thousand grain weight are reduced, and the yield is reduced. The false smut not only forms rice curly balls at the ear of rice to seriously affect the growth of grains and reduce the yield and quality of rice, but also has the toxic action on human and livestock by the false smut toxin generated by the pathogenic fungus of the false smut. At present, the extensive attention of the expert scholars to the ustilaginoidea virens producing bacteria and the ustilaginoidea virens toxin produced has been gradually raised.

In natural ecosystems, pathogenic bacteria may exist in a variety of "natural enemies" or "competitors," and thus microorganisms that kill or inhibit pathogenic bacteria are of potential biocontrol value. The antagonistic bacteria can be widely applied to the prevention and treatment of various plant diseases and insect pests. The microbial antagonistic preparation can directly generate a bacteriostatic effect through parasitism, antibiotic or competition and the like, and can also play an indirect protection effect through inducing the self resistance of plants. Biological control by extracting bacteriostatic substances with antagonistic activity on pathogenic bacteria or screening microorganisms with antagonistic activity on pathogenic bacteria is a novel measure with sustainable development potential. The biocontrol bacterium can be originated from the plant body and can also be originated from the related living environment.

Therefore, providing a pseudomonas aeruginosa strain 9# and application thereof is a problem to be solved urgently by the technical personnel in the field.

Disclosure of Invention

In view of the above, the invention provides pseudomonas aeruginosa 9# and application thereof, wherein the pseudomonas aeruginosa 9# is used for antagonizing ustilaginoidea virens and preventing ustilaginoidea virens.

In order to achieve the purpose, the invention adopts the following technical scheme:

pseudomonas aeruginosa (Pseudomonas aeruginosa)9#, with the preservation number of CCTCC NO: m2021178, having been preserved in China center for type culture Collection (CCTCC for short), with the address of university of Wuhan, in China, the preservation date of 2021, 02 months, and classified and named as Pseudomonas aeruginosa 9# and Pseudomonas aeruginosa 9 #.

Further, the pseudomonas aeruginosa 9# is applied to antagonism of ustilaginoidea virens.

Further, the pseudomonas aeruginosa 9# is applied to prevention and treatment of false smut.

Further, the pseudomonas aeruginosa 9# is applied to preparation of microbial agent for preventing and treating false smut.

According to the technical scheme, compared with the prior art, the invention discloses and provides the pseudomonas aeruginosa strain 9# and the application thereof, the pseudomonas aeruginosa strain 9# can inhibit the growth of the ustilaginoidea virens strain YY7850, which is an example that the pseudomonas aeruginosa strain is found to have antagonism against ustilaginoidea virens pathogenic bacteria for the first time, and the strain has the potential of being developed into rice ustilaginoidea virens biocontrol bacteria due to the fact that the strain 9# has the capacity of inhibiting the ustilaginoidea virens pathogenic bacteria.

In addition, the strain has strong tolerance to various heavy metals, such as Cd (2200mg/L), Zn (1800mg/L) and Pb (1200mg/L), wherein the strain has the strongest tolerance to Cd and is considered as a multi-metal-resistant bacterial strain; this strain is also sensitive to 13 different antibiotics. The strain is also a Cd adsorbing strain, and a simulated field test proves that the strain can obviously reduce the Cd accumulation in rice grains (Lin X Y, Mou R X, Cao Z Y, et al. Characterisation of calcium-resistant bacteria and the needle potential for reducing the accumulation of calcium in rice grains, 2016, 569. science of the Total Environment,2016, 570: 97-104.). Therefore, the strain is a multifunctional strain, and has great development and utilization values in the aspects of prevention and control of rice false smut and microbial remediation of rice field polluted soil.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the provided drawings without creative efforts.

FIG. 1 is a diagram showing the colony morphology of the strain 9# of the present invention;

FIG. 2 is a phylogenetic tree of strain # 9 of the present invention;

FIG. 3 is a diagram showing the bacteriostatic effect of the strain No. 9 of the present invention on Ustilago virens strain YY 7850;

wherein, A is a control; b is an antagonistic map.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

EXAMPLE 1 screening and identification of Strain 9#

Multiple bacterial strains obtained from soil of a test field of the national rice institute through enrichment culture, separation and purification are subjected to ustilaginoidea virens antagonistic experiment, and one bacterial strain is successfully screened through the antagonistic experiment and named as strain No. 9. The colony morphology is shown in FIG. 1. The strain forms large and flat colony after being cultured on common agar, is in a ground glass shape, has metallic luster and ginger flavor, can generate blue-green pigment, and is diffused in a culture medium.

The strain 9# was identified as a strain of Pseudomonas aeruginosa (Pseudomonas aeruginosa) by 16s sequencing and comparative analysis, and the phylogenetic tree is shown in fig. 2.

Example 2 antagonistic assay

(1) Culturing of bacterial strains

A. Culturing Ustilaginoidea virens strain YY 7850:

inoculating Ustilago virens strain YY7850 stored in the laboratory to a potato sucrose agar culture medium (PSA) flat plate, culturing at constant temperature of 28 ℃, inoculating to a potato sucrose liquid culture medium (PS) after bacterial colonies grow out, culturing at constant temperature of 150rpm and 28 ℃ for 7 days, and filtering with eight layers of gauze to obtain spore suspension for later use.

Potato sucrose agar medium (PSA): the culture medium is PSA agar culture medium prepared by Beijing Soilebao science and technology Limited, 47.0g of PSA agar culture medium is added per liter of water to prepare the PSA agar culture medium, the PSA agar culture medium is subpackaged, sterilized by high-pressure moist heat at 121 ℃ for 20 minutes, taken out and shaken evenly, and stored at 4 ℃ for later use.

Potato sucrose broth (PS): adding 1000mL of water into 200g of potatoes, boiling for 20-30 min, filtering with eight layers of gauze, adding 20g of cane sugar into the filtrate, fixing the volume to 1L, performing high-pressure moist heat sterilization at 121 ℃ for 20min, taking out, and cooling for later use.

B. Culture of antagonistic strain # 9:

inoculating the strain No. 9 to an LB inclined plane, culturing at the constant temperature of 37 ℃ for 1d, inoculating to an LB liquid culture medium after a bacterial colony grows out, and culturing at the constant temperature of 37 ℃ for 1d at 150rpm for later use.

LB inclined plane: the culture medium is LB agar culture medium produced by Biotechnology engineering (Shanghai) GmbH, preparing LB inclined plane according to the amount of 35.0g LB agar culture medium added per liter of water, subpackaging, sterilizing at 121 deg.C under high pressure and moist heat for 20min, placing the inclined plane while it is hot, cooling the inclined plane, and storing at 4 deg.C for use.

LB liquid medium: the culture medium is LB liquid culture medium produced by Biotechnology engineering (Shanghai) GmbH, preparing LB liquid culture medium according to the amount of 35.0g LB liquid culture medium added per liter of water, subpackaging, sterilizing under high pressure and moist heat at 121 ℃ for 20 minutes, taking out, and cooling for later use.

(2) Antagonistic test

Taking the Ustilaginoidea virens YY7850 spore suspension cultured in the step (1) (the number of spores is 1 multiplied by 10)⁶cfu/ml) of the strain, uniformly coating the mixture on a sterile PSA plate with the diameter of 9cm by using a sterilized coating rod, pre-culturing the mixture for 1 day at the constant temperature of 28 ℃, taking a sterilized filter paper sheet with the diameter of 6mm in the middle of the PSA plate, and adding 5 mul of antagonistic strain 9# bacterial liquid (1 multiplied by 10) cultured in the step (1)⁶cfu/ml) (plate coated with only Ustilaginoidea virens YY7850 spore suspension without antagonist 9# as control), and culturing at 28 deg.C until the control plate is full of growthThe colony is measured, the radius of the inhibition zone is measured, and the antagonism result is shown in figure 3. As can be seen from FIG. 3, the strain No. 9 has an obvious bacteriostatic effect on Ustilaginoidea virens strain YY7850, and the radius of a bacteriostatic zone is 3.3 cm. Therefore, the screened strain No. 9 has the capability of inhibiting pathogenic bacteria of the rice false smut and has the potential of developing the pathogenic bacteria of the rice false smut.

The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. Pseudomonas aeruginosa (Pseudomonas aeruginosa)9#, which is characterized in that the preservation number is CCTCC NO: m2021178.

2. The use of the pseudomonas aeruginosa # 9 of claim 1 for antagonizing ustilaginoidea virens.

3. The application of the pseudomonas aeruginosa 9# as claimed in claim 1 in preventing and treating false smut.

4. The application of the pseudomonas aeruginosa 9# as claimed in claim 1 in preparing a microbial agent for preventing and treating ustilaginoidea virens.