利用复苏可培养化VBNC(Viable but non-culturable bacteria)资源菌筛选了对毒死蜱有降解性能的菌种及其酶.应用最大或然数计数(MPN)法,添加复苏促进因子Rpf蛋白分子,获取可培养化VBNC菌种,确定菌种的系统进化属性,并用常规培养法筛选了高效毒死蜱降解菌及其酶,探索了适宜的酶降解优化条件.从分离到的可培养化VBNC菌株中筛选出一株降解毒死蜱效果达70%的菌株CHZYR63,经16S r RNA基因序列分析,初步鉴定为荧光假单胞菌(Pseudomonas fluorescens).继而,从该菌株提取出粗酶,其降解毒死蜱能力为50%.降解条件优化研究表明:降解酶促反应的最适温度为35℃,最适p H值为7;在最适条件下该降解酶在30 min内对毒死蜱(100 mg/L)的降解率可达60.26%.
VBNC(viable but non-culturable)是指处于"活的但非可培养"状态的微生物,此微生物体的细胞仍有代谢活性,但用常规方法无法分离培养。本文阐述了VBNC状态菌的形成机理、转变与种类、复苏、研究意义及其应用展望。并报道了我们在十余年间针对生态环境中处于VBNC状态菌的复苏、可培养化、系统进化关系及潜在功能等方面的一些研究成果,拟为微生物资源的开发与应用提供新的科学依据。
We aimed to investigate the composition and phylogenetic rela-tionships of the viable but non-culturable （VBNC） state bacteria in pharmaceutical wastewater. [Method] Soil filter was used for constructing bioreactor. Based on the resuscitation- and growth-promoting function of Resuscitation Promoting Factor （Rpf） for VBNC bacteria, VBNC bacteria were isolated by most probable number （MPN） method and dilution-plating method and 16S rRNA gene phylogenetic analysis was carried out. [Result] In MPN culture system, Rpf could promote the resuscitation and growth of some bacteria. There were VBNC advantage floras that sensitive to Rpf in pharmaceutical wastewater. The culturable VBNC bacteria in pharmaceutical wastewater consisted of high-GC gram-positive actinomycetes including genera Mi-crobacterium, Gordonia and Leucobacter, and gram-negative bacteria including gen-era Candidimonas, Xanthobacter and Aminobacter. Four strains （ZYM1, ZYM3, ZYZR4, ZYXR1） could be potential novel species. [Conclusion] This research re-vealed there were VBNC bacteria in pharmaceutical wastewater. These results could provide important ideas and methods for further studies on VBNC bacteria in the pharmaceutical wastewater, especial y the formation mechanism and recovery mech-anism of VBNC bacteria and the advanced degradation process improvement of pharmaceutical wastewater.
