【摘要】 【目的】对细胞培养体系中出现的杆状污染物进行分离和鉴定,并探讨如何清除该污染物。【方法】采用固体培养基平板划线法分离细菌株,通过荧光染色和透射电镜对其进行形态学观察;结合16S rRNA基因序列分析,进行菌株鉴定;用生长状态良好的细胞上清复苏冻存的已经污染的细胞,检测细胞复苏的存活率。【结果】该污染物经形态学和16S rRNA基因序列鉴定为苯基杆菌。形态学观察表明它有一个二态生命周期:即游动期和附着期。大多数情况下该菌可以与宿主细胞共生,常规抗生素均不能彻底清除该细菌。采用生长状态良好的细胞上清复苏冻存细胞可以明显提高了细胞的存活率。【结论】本实验报导了苯基杆菌的二态生命周期,同时我们发现用细胞上清复苏冻存细胞可以显著提高细胞的存活率。更多还原

【Abstract】 [Objective] To identify and characterize an unknown microorganism causing contamination in several mammalian cell cultures. [Methods] This bacterium was identified by 16 S rRNA sequencing and studied by DAPI and Di OC6( 3) staining,Gram staining,acid-fast staining,and electron microscopy. The isolated bacterium was also used to infect host cells to observe antibiotic effectiveness and its relationship with host cells. [Results]The 16 S rRNA sequence analysis shows that this rod-shaped microorganism belongs to the family Caulobacteraceae,class Alphaproteobacteria,and was most closely related to Phenylobacterium zucineum HLK1 Tstrain. The bacterium collected in the "swimming"stage was Gram staining negative,but Gram staining positive in the "sessile " stage. Under the electron microscope both flagellated and non-flagellated types were found. So far,no antibiotics were effective to inhibit this microorganism. The contamination with this bacterium frequently led to failed resuscitation of thawed cells. We found that the cells resuscitated with the used culture supernatants were increased in number by 3- 4 folds as compared to those resuscitated with freshly prepared media. [Conclusion] Phenylobacterium may have a dimorphic life cycle including a swimming stage and a sessile stalked stage.更多还原