Screening,identification and antagonistic effect of antagonistic bacteria JTFM1001 against aflatoxin contamination in corn

Aflatoxin is a strong carcinogenic and toxic fungal toxin produced by Aspergillus flavus and other Aspergillus species,and can seriously threaten the health of consumers,thus becoming a global concern.Corn,as an important oil and economic crop,is highly susceptible contaminated by aflatoxin.In this study,antagonistic bacteria with strong inhibitory effect on aflatoxin were screened to provide support for the treatment aflatoxin contamination control in corn.Ten strains which have strong antagonistic effects against A.flavus were isolated from healthy corn from different corn producing areas in China.Among them,the antagonistic bacteria JTFM1001 through corn kernels in vivo and field experiment,the inhibition effect of aflatoxin contamination reached above 70%and 55%,respectively.And the strain was identified as Bacillus subtilis based on its morphological,physiological and biochemical characteristics and phylogenetic analysis of 16S rDNA.In addition,our data showed that it can colonize in the rhizosphere and survive for a long time,forming the dominant flora,with broad application prospect.Finally,we were surprised to find that the antibacterial metabolites secreted by the antagonistic bacteria was one of the mechanisms of its inhibition of A.flavus and aflatoxin.This will provide us with new ideas and perspectives on the effective prevention and control of aflatoxin contamination in corn and corn oil.

Isolation of Antagonistic Bacteria against Fusarium graminearum and Its Antagonistic Activity

[ Objective] To develop biological control resources of endophytic bacteria in wheat, the bacteria with antagonistic effect against Fusarium graminearum were screened and their antagonistic activity was studied. [ Method] The endophytic bacteria of wheat with antagonistic effect were screened using plate confrontation method; the antagonistic bacteria were identified through physiological and biochemical characteristics and 16S rDNA sequence analysis; the antagonistic spectrum of antagonistic bacteria was studied using plate confrontation method. [ Result] The antagonistic bacterium XY-3 was isolated from wheat leaves, and was identified as Bacillus amyloliquefaciens according to the physiological and biochemical characteristics and 16S rDNA sequence analysis. [ Conclusion] XY-3 had antagonistic effect against four kinds of common pathogenic fungi, presenting broad spectrum antagonistic activity against plant pathogenic fungi.

Antagonistic Effects of Sphingomonas and Pseudomonas aeruginosa on 4 Kinds of Pathogenic Bacteria of Ginseng

[Objectives]To explore effective biocontrol methods for diseases in the process of ginseng cultivation,and develop an efficient and environmentally friendly biocontrol agent.[Methods]In this study,2 strains were isolated from biogas slurry,and Cylindrocarpon destructans(XF),Fusarium solani(GF),Botrytis cinerea Pers(HM)and Alternaria panax Whetz(HB)were used as test materials.The strains were isolated and identified by dilution plate method,16S rDNA sequence identification method,confrontation culture method,filter paper method and ultraviolet spectrophotometer method,and the bacteriostatic activity and bacteriostatic rate were tested.[Results]Strain 15(Sphingomonas)and strain 19(Pseudomonas aeruginosa)were screened out through identification and analysis,and they grew stably within 8-10 d.The bacteriostatic rates of strain 15 against A.panax and B.cinerea were 47.37%and 43.40%,respectively,and the bacteriostatic rates of strain 19 against A.panax and B.cinerea were 62.30%and 63.27%,respectively.The bacteriostatic activity of the extract of strain 19 increased with the increase of OD_(600) value,and the bacteriostatic effect was optimal when the OD_(600) value was in the range of 0.8-1.0,up to 70%,so it had a strong biocontrol potential.[Conclusions]This experiment provides convenience for more effective inoculation,establishes a fast,simple and accurate method for the determination of the best bacteriostatic rate of P.aeruginosa culture solution to HM,and lays a foundation for large-scale culture of P.aeruginosa culture solution.Besides,it is expected to provide a theoretical basis for the efficient control of ginseng B.cinerea in field production,use it for the prevention and control of ginseng shoot diseases,and provide a reference for the efficient and diverse development of biocontrol agents for ginseng shoot diseases.

Gram-Positive Bacteria Associated with Rice in China and Their Antagonists Against the Pathogens of Sheath Blight and Bakanae Disease in Rice

It is necessary to understand the bacterial populations associated with rice so as to provide more information and natural resources for effective management of major diseases in rice. A survey on screening and identification of gram-positive bacteria was conducted during 1998 2004. Seven hundred and fifty-six rice samples were collected from Zhejiang, Jiangsu, Fujian and Yunnan Provinces, China. Over 1000 bacterial isolates were isolated and tested for colony morphology, pathogenicity, and some characteristics of bacteriology including Gram staining, fluorescent pigment on Kings medium B and microscopic observation for endospore. Together with five standard reference strains, 74 representative gram-positive bacterial isolates were confirmed by Biolog and gas chromatographic analysis of fatty acid methyl esters. Five bacterial species of Bacillus and other three genera were identified and isolates from Bacillus sublitis and Bacillus megaterium, exhibited the most effective inhibition against the pathogens of sheath blight and bakanae disease of rice. A few isolates from Bacillus pumilus and Bacillus megaterium showed weak virulent on rice together with some virulent isolates, risk should be considered when isolates from these species were screened for biocontrol agents.

Screening of Antagonistic Bacillus spp. Strains against Phytophthora infestans (Mont.) de Bary and Isolation of lipopeptide Substances

[ Objective] The paper was to screen the antagonistic Bacillus spp. strains against Phytophthora infestans （Mont.） de Bary and analyze its lipepeptide substances. [ Methods] Using flat isolation and culture method, the bacteria that could produce Bacillus spp. were isolated from rhizosphere soil, and carried out primary and secondary screening by confrontation culture. The lipopeptide substances of Y-3 strain with better antifungal effect were carried out I-IPLC analysis. [Result] Nine strains of bacteria had certain antagonistic effects against P. infestans in primary screening, Y-1 and Y-3 strains had significant inhibition effect against the growth of P. infestans in secondary screening, with inhibition rate higher than 80% ; the crude extract of lipepeptide produced by Y-3 strain had better antagonistic effect against P. infestans, with inhibition rate higher than 90%. HPLC analysis showed that the crude extract of lipepeptide mainly contained 8 components. [ Conclusion] The lipopeptide substances produced by Y-3 strain had significant inhibition effect, which had potential application value for biological control of potato late blight.

Fire Blight Disease Caused by Erwinia amylovora on Rosaceae Plants in Kyrgyzstan and Biological Agents to Control This Disease

Erwinia amylovora species were isolated from the blossoms, exudates, infected fruits, leaves and bent branches of diseased apple, pear and hawthorn trees, selected in the Chy, Osh and Jalal Abad regions. Biochemical and pathogenicity tests, alongside PCR analyses, were conducted to identify the local isolates of Erwinia amylovora. The alternative antagonistic microorganisms which combat bacterium E. amylovora were tested within in vitro and in vivo conditions. The results revealed the ability of Streptomyces antagonistic bacteria to decrease fire blight severity on pear and apple trees during the first stage of the fire blight disease in leaf tissues. Streptomyces strain C1-4 suppressed E. amylovora disease symptoms in the leaf tissues and excised apple and pear shoots. The incidence of fire blight on leaves was reduced by about 70% with two applications of bacterial antagonists. Further studies at different locations in Kyrgyzstan, using large scale application, would allow for stronger recommendations to be made, including studies and recommendations on their ability to prevent disease and to use them as main components in an integrated pest management program.

Preliminary Study on Production Conditions and Action Mechanism of Antifuingal Protein from Strain HJX1

Antagonistic bacterium HJXI is a Bacillus subtilis strain isolated from field rhizasphere soil where banana wih was serious, with certain control efficacy on banana Fusarium wilt. The crude extract obtained from its fermentation broth after ammonium sulfate precipitation has an inhibitory effect on various pathogenic fungi. The culture conditions for this strain were optimized in this study with Fusarium oxysporum f. sp. ettbense as an indicator, and the results showed that the fermentation broth obtained from culture in LB liquid medium under 30 ℃ at 170 r/min for48 h had the strongest inhibition against F. oxysparum f. 8p. cubense. It was observed that the inhibition was mainly reflected by causing breakage and distortion of myeelia, followed by appearance of vesicles and swelling, ablation, and leakage of protoplasm. This study provides a scientific basis for the study on the action mechanism of crude protein from bacterial strain HJX1.

Studies on Possible Modes of Action and Tolerance to Environmental Stress Conditions of Different Biocontrol Agents of Foliar Diseases in Maize

The present study evaluates the possible modes of action of antagonistic bacteria and their tolerance to UV radiation, temperature and osmotic stress. The partial 16S-23S rRNA gene sequencing of eight antagonistic bacteria had a high match with three bacterial genera: Curtobacterium, Pantoea and Bacillus. In this study, the three Bacillus isolates showed the most relevant production of enzymes, volatile organic compounds and antibiosis against Exserohilum turcicum. Respect to UV radiation and temperature Pantoea and Bacillus isolates were more tolerant;whereas the eight isolates were tolerant to osmotic stress in varying degree. The three Bacillus isolates have the greatest potential as biocontrol agents for foliar diseases in maize. The antagonistic action could be explained through different modes of action such as enzymes, volatile organic compounds and/or direct antibiosis by other secondary metabolites. Bacillus isolates tolerance to environmental stresses including UV radiation, temperature and osmotic stress is relevant for survival and persistence on the leaf surface. This work provides new information about the mode of action of antagonistic bacteria with proven efficacy against maize leaf pathogens. In addition, it provides information about the tolerance of antagonistic bacteria against different stress conditions. The data of the present study could contribute to the development of a successful foliar biofungicide.