一株降解黄曲霉毒素B1、拮抗黄曲霉的菌株鉴定及应用研究

Identification and application of a strain that degrades aflatoxin B1 and antagonizes Aspergillus flavus

目的筛选、鉴定一株降解黄曲霉毒素B1(aflatoxin B1,AFB1)及拮抗黄曲霉的菌株,并进行应用研究。方法以青岛市海洋、陆地、河流等不同地区的土壤、水质微生物为研究对象,利用平板划线法、AFB1降解率和黄曲霉对峙试验法筛选得到有效菌;通过形态学观察和同源性分析,鉴定有效菌的种类;分离该菌的上清液、悬菌液、胞内液并对各组分的AFB1降解率进行测定;提取有效菌的活性物质,对其提取物进行抑菌试验;主动侵染花生籽仁,验证有效菌对降解AFB1及抑制黄曲霉生长的效果。结果通过筛选,得到有效菌株LE160。综合形态学特征和16S rRNA基因分析结果,鉴定菌株LE160为毕节假单胞菌(Pseudomonas bijieensis),命名为毕节假单胞菌LE160。该菌株胞外上清液、菌悬液和胞内液对AFB1降解率分别为80.4%、16.3%、11.9%。该菌降解AFB1不是依赖细菌胞体的吸附作用,而是细菌代谢产生并分泌至胞外的活性物质主导的生物降解作用。菌株提取液对黄曲霉菌有明显抑制作用。花生应用试验结果表明该菌株的发酵液对降解AFB1及抑制黄曲霉生长有一定效果。结论菌株LE160对AFB1具有良好的降解效果,能有效拮抗黄曲霉的生长,具有应用的潜力。本研究结果为深入研究该菌的降解机制奠定了基础,也为花生等农产品防控黄曲霉侵染提供了新思路。

Objective To screen and identify a strain that can degrade aflatoxin B1(AFB1)and antagonize Aspergillus flavus,and study its application.Methods Based on the soil and water microorganisms in different areas such as sea,land and river in Qingdao,the effective strains were screened by streak plate method,degradation rate of AFB1 and Aspergillus flavus standoff test.The effective strains were identified by morphological observation and homology analysis.The supernate,bacterium suspension and intracellular fluid of the strain were isolated and the degradation rate of AFB1 in each fraction was determined.The active substance of effective strains were extracted and the antibacterial test were carried out.Aspergillus flavus actively infected peanut seeds to verify the effect of effective strains on the degradation of AFB1 and the inhibition of the growth of Aspergillus flavus.Results The effective strain LE160 was obtained through screening.Based on the morphological characteristics and 16S rRNA gene analysis,the strain LE160 was identified as Pseudomonas bijieensis,and named Pseudomonas bijieensis LE160.The degradation rates of AFB1 by the supernate,bacterium suspension and intracellular fluid of the strain were 80.4%,16.3%and 11.9%,respectively.The degradation of AFB1 by this bacterium was not dependent on the adsorption of bacterial cell body,but was dominated by the biodegradation of active substances produced by bacterial metabolism and secreted to the extracellular space.The strain extract had obvious inhibitory effect on Aspergillus flavus.The results of peanut application test showed that the fermentation broth of this strain had a certain effect on the degradation of AFB1 and the inhibition of the growth of Aspergillus flavus.Conclusion Strain LE160 has a good degradation effect on AFB1 and can effectively antagonize the growth of Aspergillus flavus.It has the potential of application.The results of this study lay the foundation for further study on the degradation mechanism of this strain,and also provides a new idea for the prevention and control of aflatoxin infection in peanut and other agricultural products.