黄曲霉毒素B1降解菌的分离鉴定及其降解特性

【目的】黄曲霉毒素是一类强毒、致癌的真菌次级代谢产物。本文旨在筛选出能高效降解黄曲霉毒素B1(AFB1)的细菌。【方法】以AFB1结构类似物香豆素为惟一碳源进行AFB1降解菌株初筛,得到的活性菌株的培养液分别与AFB1标准品(2.5μg/mL)共同作用,以AFB1降解率为指标进行复筛。对降解活性最好的菌株通过形态、生理生化特性以及16S rRNA序列分析进行初步鉴定;并对细胞浓度、pH、温度、金属离子等对菌株降解活性的影响进行考察。【结果】初筛获得了10株在香豆素培养基上生长良好的细菌,复筛发现这些菌均具有良好的AFB1降解活性,其中从金毛羚牛粪便中筛选出的菌株F4降解活性最好,去除AFB1能力达到90.03%。根据F4菌株16S rRNA序列同源性分析,结合形态、生理生化特性,初步确定菌株F4为施氏假单胞菌(Pseudomonas stutzeri)。F4的降解活性与细胞浓度呈正相关。当pH 7.0,35℃,菌细胞作用72 h后毒素降解率达到82.91%。Mg2+可增强F4的降解活性,降解率提高7.68%,而Cu2+可抑制其降解活性,降解率降低51.1%。【结论】筛选到能高效降解AFB1的施氏假单胞菌(Pseudomonas stutzeri)F4,F4降解毒素的活性物质主要存在于菌体细胞,其作用受到温度、pH等的影响,可能是一种胞内酶。

Isolation and characterisation of an aflatoxin B1-degrading bacterium

Objective] Aflatoxins are toxic and carcinogenic fungal metabolites.The aim of this research was to isolate aflatoxin B1(AFB1)-degrading bacteria.Methods] Samples from various sources were screened by using coumarin as the sole carbon source.The strains that could grow in the medium with coumarin carbon source were detected for their degradation of AFB1 ability by addition of AFB1(2.5 μg/mL) to the cultured broth.Among the positive strains,F4 strain with the highest activities was identified according to its morphological,physiological and biochemical properties together with phylogenetic analysis of its 16S rRNA sequence.The effects of degradation conditions such as bacterial cell concentration,pH,temperature,metal ions on the degrading AFB1 were investigated.Results]Ten isolates showed good AFB1 reduction activity and they could grow well on coumarin carbon source.Strain F4,obtained from Budorcas taxicolor feces could reduce AFB1 by 90.03% after incubation in the liquid medium at 37℃ for 72 h.F4 was preliminary identified to be Pseudomonas stutzeri with morphology,physiological and biochemical properties and 16S rRNA gene sequence.The active degrading component existed in the cell of F4,and the degrading activity was interrelated with cell concentration,temperature,pH and metal ions.Conclusion] An AFB1-degrading strain F4 was isolated from animal feces and identified as Pseudomonas stutzeri.The activive component of AFB1 degradation was mainly in the cell of F4.