鲍曼不动杆菌耐药持留菌的特征及Ⅱ型毒素-抗毒素系统的多样性

Characteristics of persister cells and the diversity of type Ⅱ toxin-antitoxin system in Acinetobacter baumannii

【目的】研究鲍曼不动杆菌持留菌的特征,分析鲍曼不动杆菌Ⅱ型毒素-抗毒素(TA)系统的多样性及分布,探讨TA系统与持留菌形成的潜在关联。【方法】采用分属于6大类中的6种不同抗生素分别筛选的方法分离并定量鲍曼不动杆菌的持留菌;应用PSI-BLAST及TBLASTN程序分析721株鲍曼不动杆菌中的Ⅱ型TA系统;通过聚合酶链式反应(PCR)方法检测44株临床多重耐药(MDR)的鲍曼不动杆菌中5个Ⅱ型TA系统的分布。【结果】我们发现不同抗生素处理得到的鲍曼不动杆菌持留菌的数量并不相同;对多数抗生素而言,浓度越高,得到的持留菌数量越少;处于生长平台期的菌株中相比对数期含有更多的持留菌;多粘菌素B及妥布霉素均具有杀灭部分持留菌的能力;鲍曼不动杆菌所有菌株中均含有Ⅱ型TA系统,HTH/GNAT类型在菌株中分布最为广泛;所有700余株鲍曼不动杆菌基因组中共含有15个潜在的Ⅱ型TA系统;三类普遍存在于已知基因组中的明确功能的Ⅱ型TA系统在临床多重耐药菌株中同样分布广泛,且Hig B/Hig A的表达量在标准株的持留菌中显著增高。【结论】鲍曼不动杆菌的持留菌水平与菌株的生长状态、抗生素的种类及浓度密切相关。Ⅱ型TA系统在鲍曼不动杆菌已知基因组和临床多重耐药菌株中普遍存在,其中,HTH/GNAT、GP49/Cro(Hig B/Hig A)及DUF497/COG3514三种类型的Ⅱ型TA系统可能在鲍曼不动杆菌持留菌的形成中扮演重要角色。

[Objective] To study the biological features of the Acinetobacter baumannii persisters and the correlation between Type II toxin-antitoxin systems（ TAs） and the persisters. [Methods] Different antibiotics were used to isolate the persisters; BLAST programs were adopted to analyze candidate TAs; PCR was used to detect the distribution of TAs.[Results]The number of persisters was different when treated by different antibiotics. For most antibiotics,the higher the concentration used,the lower the persister number was obtained. Persister numbers in exponential phase populations were lower than that in stationary phase. Polymyxin B and tobramycin could kill persisters. [Conclusion]The persistence level of A. baumannii is related to the growth state of the strains,the class and concentration of antibiotics.