摘要
【目的】了解溶菌酶触发芽孢杆菌孢子萌发的异质性及其机制,为认识芽孢(Spore)萌发机制和杀灭芽孢提供参考。【方法】应用拉曼光谱和微分干涉差(DIC)显微镜成像技术高通量分析大量单个Bacillus subtilis(Bs)和B.megaterium(Bm)芽孢经溶菌酶触发的萌发动态。【结果】溶菌酶浓度和温度越高,芽孢萌发越快,孢内CaDPA开始快速释放时间(T_(lag))、快速释放所需时间(ΔT_(release))和芽孢皮层水解所需时间(ΔT_(lys))越短;低于20℃,Bs芽孢萌发的ΔT_(release)值是25℃时的4倍以上。SpoVA蛋白高表达菌株的ΔT_(release)值和普通菌株基本相同,而缺少皮层水解酶的菌株ΔT_(release)值高于普通菌株。95℃处理15min的孢子,其T_(lag)、ΔT_(release)和ΔT_(lys)值是对照的2倍以上。Bs芽孢萌发的异质性明显,不仅表现在芽孢间,也表现在菌株间。Bm芽孢对溶菌酶更敏感,芽孢间的异质性显著低于Bs。【结论】溶菌酶触发的芽孢萌发在物种、菌株和单细胞层面都存在显著的异质性;溶菌酶浓度和温度对芽孢萌发有重大影响;皮层水解酶也可能参与溶菌酶触发的芽孢萌发进程。
【Objective】The heterogeneous germination and its mechanism of Baillusspores trig-gered by lysozyme were studied in order to look insight into the mechanism of spore germination in general and provide new knowledge of inactivating bacterial spores.【Methods】The ki-netic of germinations of multiple individual decoated spores of Bacillus subtilis(Bs)and B.megaterium(Bm)triggered by lysozyme were followed by Raman spectroscopy and differentail interference contrast(DIC)microscopy.【Results】Higher concentrations of lysozyme and temperatures significantly speeded the germination of Bs decaoted spores and reduced the time T_(lag),at which spores began release of the great majority of spores' 1∶1 chelate of Ca^(2+) with dipicolinic acid(CaDPA),the period time of completing the release of CaDPA(named ΔT_(release))and the time period of hydrolysis of the spore's peptidoglycan cortex(named ΔT_(lys)).The ΔT_(release) values of germination below 20℃ were 4 times more than that at 25℃.The ΔT_(release) values of Bs spores with 4-fold-elevated level of SpoVA protein were basically similar to control strains,while spores lacking cortex-lytic enzymes exhibited notably highe ΔT_(release) values than wildtype spores.The values of T lag,ΔT_(release) and ΔT_(lys) of spores treated by wet heat at 95℃ for 15 min were two times more than the untreated spores.Heterogeneous germinations were observed not only among individual decoated Bs spores,but also among Bs strains.The decoated Bm spores were more sensitive to lysozyme and its germinations of individual spores were less heterogeneous.【Conclusion】The germinations of decoated B acillus sp ores triggered by lysozyme,on which the lysozyme concentration and temperature have a significant effect,are significantly heterogeneous at the levels of single spores and strains,as well as of species.The cortexlytic enzymes may be involved in the initiation of germination.
出处
《广西科学》
CAS
2016年第2期102-110,共9页
Guangxi Sciences
基金
国家自然科学基金项目(11264004,31460035)资助
