幽门螺杆菌形态变化超微结构分析

Ultrastructural analysis of Helicobacter pyloriin morphological transformation

目的 幽门螺杆菌(Helicobacter pylori,Hp)在不利的生长环境下会由螺旋杆状转变为球形体,简称为球形变,球形状态下的Hp通常是存活但不可培养的(VBNC)。当环境适宜时球形体又回复为螺旋杆状。目前对于Hp球形变及其回复的完整形态变化过程尚不清楚,转变机制亦未被完全揭示。本研究以两株Hp标准菌株(SS1和26695)为对象,从形态及超微结构层面揭示Hp的球形变及回复过程。方法 利用透射电子显微镜负染和超薄切片技术捕捉Hp在球形变及回复过程中不同时间点的形态和超微结构变化,并对大视野下球形体和螺旋体计数,分析对比球形转变及回复实验中Hp形态变化的速度。结果 Hp SS1与26695菌株形态相似,但SS1有更多鞭毛表达。当Hp发生形态转变时,螺旋杆状的Hp会缩短长度并折叠为“U型”,细胞内膜与外膜分离,周质空间加大;细菌内部结构发生重塑,进而缩紧成球形体,球形体的体积通常小于其螺旋杆状形态时的体积。Hp的两种标准菌株在次优生长状态下,SS1球形化的速度和比例均大于26695,回复为螺旋杆状的速度却慢于26695。结论 Hp的形态较为灵活,在球形转化时会出现“U”形的中间过度状态,不同菌株的球形变和最佳生长状态时间差异很大,在形态转换期间Hp内外膜分离,形态不规则,体外和体内试验中均应考虑菌株状态对结果可能造成的影响。

Objective Helicobacter pylori(Hp)is able to conduct morphological transformation from spiral to coccoid form under adverse growth conditions.This procedure is reversible,the coccoid Hp could resuscitate to spiral state in decent environment.However,there is few published data about the process of Hp morphological transformation,and its mechanism has not been fully understood.To reveal the morphological process of Hp coccoid conversion and its resuscitation via two Hp standard strains(SSl and 26695).Methods The ultrastructural changes of Hp in different time points during morphological transformation were captured by transmission electron microscope and ultrathin sectioning.Besides the spheroid and helicoid in each time points were counted under the large field of vision,and the speed of morphological changes was analyzed.Results Hp SSl showed similar morphology with 26695 despite more flagella expression.The spiral Hp initiated coccoid conversion by a"U-shaped"state,the inner membrane and outer membrane of cell wall separated and created a large periplasmic space;the internal structure of bacteria remolded,and then compressed into spheroid.The size of coccoid Hp is smaller than spiral shape.The speed of SSl transferring its shape to coccoid form was faster than 26695,while resuscitate to helicoid was slower than 26695.Conclusion Hp undergoes coccoid conversion via"Ushape"intermediate state.The culture time of optimal state or coccoid formation for different Hp strains vary greatly.The potential influence of Hp morphological state should be considered in both in vitro and in vivo experiments.