鲍曼不动杆菌原噬菌体phiOXA在耐药基因OXA-23传播中的作用研究

Drug-resistant A. baumannii has been a serious health problem, clarifying the mechanisms of drug resistance gene transmission is of great significance for controlling drug resistance level and reducing clinical hazards. Prophages mediate the transmission of drug-resistant genes through horizontal transfer, leading to changes in drug-resistant levels of host bacteria. Our previous studies have found that Acinetobacter baumannii prophage phiOXA carries the resistance gene OXA-23. The loss of phiOXA results in the decrease of resistance to Meropenem. However, whether phiOXA can induce the release and infection of other bacteria to transmit resistance genes is still unclear. This project aims to study the role of prophage phiOXA in the transmission of drug-resistant genes in Acinetobacter baumannii. Mitomycin C will be used to induce the production of phages and release of drug-resistant gene. Combining with commonly used disinfectants and antibiotics in the hospital, the induction efficiency is tested, different temperature and pH conditions are set, the stability of phages is tested, and the risk of drug-resistant gene release and persistence in the hospital environment is assessed. To determine the host spectrum of bacteriophages and the transduction efficiency of each host bacteria, analyze the stability of successive passages after bacteriophage transduction, evaluate the potential target, efficiency and stability of drug-resistant genes after transmission, and finally clarify the role of prophage phiOXA in the transmission of drug-resistant genes, This project will help to provide a new basis and ideas for the monitoring and control of drug resistance of Acinetobacter baumannii.

鲍曼不动杆菌耐药形势严峻，阐明耐药基因传播机制，对于控制耐药水平、降低临床危害意义重大。原噬菌体通过水平转移介导耐药基因传播，导致宿主菌耐药水平改变。我们前期研究发现，鲍曼不动杆菌原噬菌体phiOXA携带耐药基因OXA-23, phiOXA丢失导致对美罗培南耐药性降低，但phiOXA能否诱导释放、感染其它细菌从而传播耐药基因尚不清楚。本课题拟研究原噬菌体phiOXA在鲍曼不动杆菌耐药基因传播中的作用，采用丝裂霉素C诱导产生噬菌体、释放耐药基因；结合院内常用消毒剂和抗生素，分别检测诱导效率，设置不同温度和pH条件，测试噬菌体稳定性，评估院内环境中，耐药基因释放与持续存留的风险；测定噬菌体宿主谱和各宿主菌的转导效率，分析噬菌体转导后连续传代稳定性，评估耐药基因潜在传播对象、效率和传播后稳定性，最终阐明原噬菌体phiOXA在耐药基因传播中的作用，为鲍曼不动杆菌耐药的监测、防控提供新的依据和思路。

院内感染鲍曼不动杆菌耐药形势严峻，细菌基因组中原噬菌体可通过水平基因转移介导耐药基因传播，导致宿主菌耐药水平改变。本研究通过收集发生鲍曼不动杆菌院内感染的临床标本及病人基本信息，从中分离培养出多重耐药鲍曼不动杆菌，利用高通量全基因组重测序对多重耐药鲍曼不动杆菌基因组特征进行分析，通过phigaro软件对细菌基因组中原噬菌体结构进行预测鉴定。研究发现，在收集的临床感染标本中，61%来自患者痰液；MLST分型结果显示，96%的菌株属于ST2型，利用细菌全基因组序列构建进化树进行系统发育分析，将收集菌株进一步细分为4个不同的Clade分支。通过phigaro软件预测鉴定多重耐药鲍曼不动杆菌基因组上的原噬菌体，共发现原噬菌体518个，菌株携带原噬菌体数量为2-8个，平均每个鲍曼不动杆菌基因组中携带5个原噬菌体结构。其中长尾噬菌体 (Siphoviridae) 有261个，占比50.4%，肌尾噬菌体 (Myoviridae) 有86个，占比16.6%，未知噬菌体 (Unknow) 有143个，占比27.6%，短尾噬菌体 (Podoviridae) 有15个，占比2.9%。对预测的前噬菌体基因组数据进行分析，鲍曼不动杆菌前噬菌体的平均GC含量为39.85% (34.61-47.10%)，平均基因组大小为20.23kb (2.41-67.15kb)。通过上述研究，我们揭示了院内感染鲍曼不动杆菌在临床不同样本和科室中的分布情况，我们研究了院内感染鲍曼不动杆菌基因组中原噬菌体的分布情况和基因组特征，为监测预防鲍曼不动杆菌原噬菌体介导耐药基因传播扩散提供了理论依据。