基于全基因组序列的耐碳青霉烯鲍曼不动杆菌的耐药与毒力研究

Whole-genome sequencing analysis on drug resistance and virulence genes of carbapenem-resistant Acinetobacter baumannii

目的明确耐碳青霉烯类鲍曼不动杆菌(CRAB)临床主要流行株CC92克隆复合群(CC92)基因组中耐药基因和毒力基因的分布,并探讨CRAB耐药和毒力的分子特征。方法本研究共纳入2019年1~7月自首都医科大学附属北京地坛医院检验科分离的33株CRAB分离株,行细菌全基因组测序研究。利用CLC Genomics Workbench v10.0软件对全基因组测序数据进行序列的质量修整后,开展序列拼接、组装与注释,并进行耐药基因与毒力基因研究,分析CC92克隆复合群CRAB基因组中携带的耐药基因和毒力基因的分布。结果基因组测序数据分析表明,33株CRAB菌株中30株为CC92型克隆复合群(包括ST195、ST208、ST369和ST540),2株为ST229型,1株为国际上未报道新序列型别。以上CRAB中包括大量的耐药基因,包括大环内酯类、四环素和链霉素耐药基因在内的8~18个耐药基因,以及包括编码与侵袭和黏附等毒力功能相关的12种毒力基因。同时,本研究发现CC92克隆复合群,除携带碳青霉烯类耐药基因以外,还携带了其他非CC92克隆群不具备的耐药基因(如blaOXA-66和blaTEM-1D)和毒力基因(如bauA和bap基因)。此外,在CC92克隆复合群中可能存在大环内脂类耐药基因[msr(E)和mph(E)]、氨基糖苷类的耐药基因(armA)、链霉素耐药基因(strA、strB)与四环素耐药基因[tet(B)]的重组。结论CC92克隆复合群的CRAB基因组存在大量耐药基因和毒力基因,且存在耐药基因重组现象,迫切需要对CRAB进一步开展全基因组水平监测,为临床病原诊断和治疗提供必要的依据。

Objective To definite the distribution and molecular characteristics of drug resistance genes and virulence genes in clonal complex 92(CC92)which was a major epidemic strain of carbapenemresistant Acinetobacter baumannii(CRAB).Methods Total of 33 strains of CRAB from different clinical samples during January to July,2019 in Beijing Ditan Hospital,Capital Medical University were analyzed by whole genome sequencing analysis.The quality of the whole genome sequencing data were modified by CLC Genomics Workbench v10.0 software,then the sequence splicing,assembly and annotation were carried out,and the drug resistance genes and virulence genes were studied,and the distribution of the drug resistance genes and virulence genes carried by CRAB of the genome of the clone complex group were analyzed.Results The analysis of whole genome sequence of CRAB showed that among 33 strains of CRAB,30 strains belonged to CC92,including ST195,ST208,ST369 and ST540.Two strains belonged to ST229,and the other one was identified as a new sequence type which had not been reported internationally.The analysis of the drug resistance genes showed the genome of these CRAB strains carried 8-18 drug resistance genes,covering carbapenem,macrolides,tetracycline,streptomycin resistance genes,etc and 12 kinds of virulence genes,which encoded adherence-related protein and invasion-related protein,etc.Meanwhile,the CC92 clone complex carried not only carbapenem resistance genes,but also other drug resistance genes(such as blaOXA-66 and blaTEM-1D)and virulence genes(such as bauA and bap)which cannot be found in non-CC92 clone complex.In addition,potential genetic integrations in macrolides[msr(E)and mph(E)],aminoglycoside(armA),streptomycin(strA,strB)and tetracycline[tet(B)]drug-resistance genes in CC92 genome were found.Conclusions CC92 carried a large number of drug-resistance genes and virulence genes,exhibited gene integration phenomenon which requires further investigation.An urgent need for clinical CRAB whole genomic surveillance should be carried out in order to provide the necessary basis for the diagnosis and treatment of clinical pathogens.