KR20180033774A - Entomopathogenic fungus, Beauveria bassiana ANU1 and microbial agent for controlling Spodoptera exigua, Plutella xylostella, Helicoverpa assulta or Phaedon brassicae using it - Google Patents

Abstract

The present invention relates to a Beauveria bassiana ANU1 (Accession No. KCTC18468P) having insecticidal effects against Beet armyworm, diamondback moth, Oriental tobacco budworm, or Phaedon brassicae. The Beauveria bassiana ANU1 of the present invention shows excellent insecticidal ability against Beet armyworm, diamondback moth, Oriental tobacco budworm, or Phaedon brassicae, and thus can be useful for the control of the pests.

Description

{Entomopathogenic fungus, Beauveria bassiana ANU1 and microbial agent for controlling Sputoptera exigua, Plutella xylostella, Helicoverpa assulta or Phaedon using ANU1 and insect pathogenic fungus bovia barbiana ANU1 brassicae using it}

The present invention relates to a novel strain of Beauveria bassiana , a fungus with high insecticidal activity against pest bacterium, a Chinese cabbage moth, a tobacco moth and a narrow bell beetle, and a new strain of the bacterium Beauveria bassiana , Or a microbial agent for controlling narrow bosom beetles.

Insect pathogenic fungi (fungi) are composed of single cell type, or branched filaments, hyphae and mycelia of mats with these hyphae.

Insect pathogenic fungi penetrate through the body wall of host insects. The conidial spore (conidia) produced by asexual reproduction of insect pathogenic fungi is the most commonly used infective stage in microbial pesticides. That is, the mycelium of the insect pathogenic fungi attaches to the epidermis of the host insect, and the mycelium penetrates into the inside of the insect through the body wall of the host insect so that the mycelium penetrates into the inside of the host insect, After proliferation in body fluids, it invades into the tissues to block the immune function of host insects and secrete toxic substances to kill host insects.

The insect pathogenic fungus, Beauveria bassiana , belongs to Hyphomycetes, which plays a role in regulating the density of insects in nature by killing the host insects. It is harmless to humans, animals, and plants but has high virulence for insects (Lacey et al., 2001).

Insect pathogenic fungi have been reported to be suitable as biological agents for the biological control of heat-insect pests due to the insufficient resistance of the host's resistance mechanism, although the insecticidal range is limited due to the high degree of pathogenicity to the target insects (Lacey et al. , 2011).

Unlike insect pathogenic viruses or insect pathogenic bacteria, which are expressed by insect feeding, insect pathogenic fungi penetrate the epidermis physically through contact with insects, and kill insects through inhibition of host immune function and secretion of toxic substances (Charley 1997). In addition, the infecting insect pathogenic fungus uses nutrients present in the host's blood to proliferate in large quantities, leading to depletion of host nutrients and death. Spores formed from lethal host insects subsequently have pathogenic potential to infect further healthy individuals (Hajek et al., 1994).

Spodoptera exigua belongs to the Lepidoptera, which is a member of the family Lepidoptera, and is a member of the family Lepidoptera, which is a member of the genus Lepidoptera. It is a pest that affects the back. Plutella xylostella belongs to the family Lepidoptera, and is a pest that infects cruciferous vegetables such as radish, cabbage, cabbage, and rape. Helicoverpa assulta belongs to Lepidoptera, and is a pest that damages various crops such as tobacco leaf, pepper, tomato, cotton, corn, eggplant, bell pepper, pumpkin. Phaedon brassicae is a beetle belonging to the leaf-beetle family, and is a pest insecting cabbage or radish planted in houses. All of these pests are resistant to medicines and are difficult to control.

Korean Patent Publication No. 10-2016-0000537 Korean Patent Publication No. 10-2016-0084968

 Charnley, A. K. (1997) Entomopathogenic fungi and their role in pest control, The mycota Ⅳ Environmental and microbial relationships, Berlin, Springer. 185-201.  Hajek A. E. and R. J. St. Leger (1994) Interactions between fungal pathogens and insect hosts. Annu. Rev. Entomol. 39: 293-322.  Lacey, L. A., R. Frutos, H. K. Kaya and P. Vail (2001) Insect pathogens as biological control agents: Do they have a future? Biol. Control. 21: 230-248.  Bactericera cockerelli (Sulc) (Hemiptera: Triozidae) is an area of endemic for zebra chip disease, which is the most common form of zebrafish disease. potato Biol. Contr. 56, 271-278.

The present invention relates to the isolation and identification of a novel insect pathogenic fungus bovaria bacillus, ANU1, and to develop a microbial agent capable of controlling the fungus, the moth, the tobacco moth and the narrow beetle using this strain .

The present invention relates to the use of bovaria bacillus ANU1 ( Beauveria sp.), Which has an insecticidal effect against beetles, cabbage moths, tobacco moths and narrow chest beetles bassiana ANU1 (Accession No. KCTC18468P).

The present invention also provides microorganism preparations for controlling Pak chilbai, Chinese cabbage moth, Tobacco moth or Narrow chestnut boll, containing bovaria bacillus or ANU1 or spores thereof as an active ingredient.

In addition, the present invention provides a method for controlling Phellodendron moth, Chinese cabbage moth, Tobacco moth or Narrow chest beetle characterized by treating a bacterium preparation containing bovaria bacillus or ANU1 or spores thereof as an active ingredient.

It was confirmed that the bovaria bacillus ANU1 of the present invention has a high level of pathological power against various species of lepidopteran insect pests such as Paralichthys olivaceus, Chinese cabbage moth, Tobacco moth, and Beetle pest. Therefore, the bovaria bacillus or ANU1 of the present invention can be used as a new microbial agent for controlling pest bugs, Chinese cabbage moths, tobacco moths and narrow chest beetles, which are difficult to control due to drug resistance in agricultural fields.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a nucleotide sequence of the β-tubulin gene of bovaria bacillus ANU1 of the present invention.
Fig. 2 shows a molecular system diagram using the β-tubulin gene of bovaria bacillus or ANU1 of the present invention.
Fig. 3 is a photograph of a spore and mycelium of bovaria bacillus or ANU1 of the present invention observed under a microscope.
FIG. 4 shows the results of examining the insecticidal activity at various temperatures on the larvae of the bovaria barbiana ANU1 of the present invention.
FIG. 5 shows the result of examining the insecticidal activity against the larvae of Biberia basiana or ANU1 according to the present invention at various humidities.
FIG. 6 shows the results of examining the insecticidal activity against the larvae of the bovaria bacillus and ANU1 of the present invention, the Chinese cabbage moth (Px), the tobacco moth Ha and the narrow chest beetle (Pb) larvae.

The boveriac basiana ANU1 of the present invention is an insect pathogenic fungus isolated from Andong, Kyungsangbuk-do. The larvae collected from the paddy field in Andong city, Gyeongbuk province were collected, and blood was collected, and then the pigs were placed on the PDA medium. After that, the cells were cultured for 5 days in a 28 ° C incubator and inoculated with PDB medium using the mycelium grown.

The β-tubulin gene was isolated from the isolated bovaria bacillus, and the nucleotide sequence of the β-tubulin gene was confirmed. As a result, it was found that there was a difference between the β-tubulin gene and the other bovaria barbiana (FIG. 1) (Fig. 2). The results are shown in Fig. 2 (b). Morphological analysis was also performed by observing the shape of the spores and mycelium of the isolated bobiera bacillus with a microscope (Fig. 3).

As a result, it was found that the insect pathogenic fungus of the present invention is a novel strain belonging to bobberry barbiana. Therefore, this insect pathogenic fungus was named as bovaria bacillus or ANU1, deposited with the Korea Research Institute of Bioscience and Biotechnology on May 25, 2016, and granted accession number KCTC18468P.

The bovaria bacillus ANU1 strain of the present invention grows well on PDB (potato dextrose broth) medium and grows well at a temperature range of 20 to 30 ° C. When culturing at less than 20 ° C, the growth rate of the strain is lowered due to the low temperature, and when the culture is performed at a temperature exceeding 30 ° C, the growth rate of the strain is lowered due to the high temperature.

The boveriac basin ANU1 of the present invention can be industrially used because it can be mass-produced when it is cultured under the above conditions.

The bovaria bacillus ANU1 of the present invention has been found to exhibit a very high insecticidal effect against the narrow-bore beetle, which is a pest of lepidoptera, parasitic moth, cabbage moth, tobacco moth, and beetle pest. Thus, ANU1 or its spores can be used as an active ingredient in microbial preparations for controlling the moth, moth, moth, or narrow bovine leaf beetle.

Since the microbial agent of the present invention is excellent in the survival period of the active fungi, the germination rate during the survival period, and the ability to maintain the insecticidal activity, it can be widely used as an insecticidal microbial pesticide, a foliar spraying agent,

The microbial formulation of the present invention can be applied to areas where there is a risk of receiving or receiving the harmful effects of parasitic moths, cabbage moths, tobacco moths or narrow breast beetles to control these pests. Be treated to contain ^{1 × 10 6 ~ 1 × 10} 8 spores per single unit dose when treatment (㎖) of the microorganism preparation of the present invention is preferred. The microorganism preparation of the present invention exhibits the best insecticidal effect when treated at a temperature of 20 to 30 ° C, more preferably 25 to 30 ° C, and a relative humidity of 30 to 60%, more preferably 40 to 60%.

Hereinafter, the present invention will be described in more detail with reference to specific examples. However, these examples are intended to further illustrate the present invention, and the scope of the present invention is not limited by these examples.

< Example  1>

Bobiera Bassiana ANU1 The identification of

The larvae collected from the paddy field in Andong city, Gyeongbuk province were collected, and blood was collected, and then the pigs were placed on the PDA medium. After that, the cells were cultured for 5 days in a 28 ° C incubator and inoculated with PDB medium using the mycelium grown.

The genomic DNA of the isolated bovaria bacillus strains was isolated and subjected to β-tubulin gene analysis.

The β-tubulin gene analysis was performed by a domestic sequencing analysis company (Macrogen, Daejeon). The nucleotide sequence of the analyzed β-tubulin gene is shown in FIG.

Sequence analysis was performed on the separated β-tubulin gene using Neighbor-joining method (bootstrap 1500 value) using MEGA 6.0 program. The results are shown in Fig.

Morphological analysis was carried out by observing the morphology of spores and mycelium of the isolated bobiera bacillus strains with a microscope. The results are shown in FIG.

As a result of the molecular identification and the morphological analysis through the microscope, the isolated strain was confirmed to be a novel strain of bovaria barbiana and named as bovaria barbiana ANU1.

[ Experimental Example ]

Experimental insect breeding

Indoor artificial diets were used to breed the larvae of beetles, cabbage moths, tobacco moths, and narrow chest beetle larvae. The conditions of the incubation incubator were temperature 25 ± 1 ℃, light period 16: 8 h (L: D), and adults gave 10% sugar water.

Preparation of sample

Bovaria bacillus ANU1 was cultured in PDB (potato dextrose broth) medium and then homogenously stirred in sterilized distilled water. After counting the number of spores through the hemocyte counting unit, it was prepared at a concentration of 1 × 10 ⁷ conidia / ㎖ in artificial diets (or cabbage leaves).

< Experimental Example  1>

Bobiera Bassiana ANU1 of In the parrot parrot  By temperature Insecticide  Confirm

The antioxidant capacity of bovaria bacillus ANU1 was investigated as follows.

The bacteriocin ANU1 prepared at a concentration of 1 × 10 ⁷ conidia / ㎖ was treated at various temperature conditions (15 ° C, 20 ° C, 25 ° C and 30 ° C) The results are shown in Fig.

As can be seen from the graph in FIG. 4, the mortality rate of the parrot parrot began to increase from 2 days after the treatment, and continued to increase until 6 days after the treatment. The higher the temperature, the higher the mortality rate. The mortality rate was about 80% at 20 ℃ and nearly 100% at 25 ℃ and 30 ℃ after 6 days of treatment.

< Experimental Example  2>

Bobiera Bassiana ANU1 of In the parrot parrot  By humidity Insecticide  Confirm

The antimicrobial activity of bovaria bacillus ANU1 was examined as follows.

The bacteriocin ANU1 prepared at a concentration of 1 × 10 ⁷ conidia / ml was treated at various humidity conditions (30%, 40%, 50% and 60% relative humidity) The results are shown in Fig.

As can be seen from the graph of FIG. 5, the mortality rate of the parrot parrot began to increase from 2 days after the treatment, and continued to increase until 6 days after the treatment. The mortality rate was about 90% at relative humidity of 40%, 50% and 60%, and about 80% at relative humidity of 30% after 6 days of treatment. Of deaths were confirmed.

< Experimental Example  3>

Bobiera Bassiana ANU1 of Chestnut , Cabbage moth , Tobacco moths Narrow chest on leaf beetle  About Insecticide  Confirm

The bovaria bacillus ANU1 was treated at a concentration of 1 × 10 ⁷ conidia / ㎖ in the seaweed, cabbage moth (Px), tobacco moth (Ha) and narrow chest beetle (Pb) Respectively. The treatment conditions were a temperature of 25 캜 and a relative humidity of 40%. The results are shown in Fig.

As can be seen from the results in FIG. 6, the bovaria bacillus ANU1 of the present invention showed a high insecticidal power of 80% or more for both the parrot moth, Chinese cabbage moth, Tobacco moth and Narrow chest beetle. In particular, 100% insecticidal activity was shown for cabbage moths and tobacco moths.

Korea Biotechnology Research Institute KCTC18468P 20160525

Claims (3)

Bovaria bacillus ANU1 ( Beauveria), which has insecticidal effect against tobacco moth, cabbage moth, tobacco moth and narrow chest beetle bassiana ANU1) (Accession No. KCTC18468P). Microbiological preparations for controlling Phellodendron moth, Cabbage moth, Tobacco moth or Narrow chest beetle, containing bovaria bacillus ANU1 or spores thereof as active ingredients. Wherein the microbial agent containing bovaria bacillus ANU1 or spores thereof as an active ingredient is treated. Description: BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for controlling a bacterium, a Chinese cabbage moth, a tobacco moth or a narrow chest beetle.

Images