不动杆菌CRISPR系统基因结构的分析及其与耐药性关系的研究  

Analysis of the genetic structure of the CRISPR system in Acinetobacter and its relationship to drug resistance

作　　者：付恒霞 毕春霞[2] 张蒙蒙[1] 秦江楠 罗玮[2] 王斌[1] 闫志勇[1] FU Heng-xia;BI Chun-xia;ZHANG Meng-meng;QIN Jiang-nan;LUO Wei;WANG Bin;YAN Zhi-yong(Department of Microbiology,Medical College of Oingdao University,Qingdao,Shandong,China 266071;Qingdao Municipal Hospital;401st Hospital of the PLA)

机构地区：[1]青岛大学基础医学院病原生物学系,山东青岛266071 [2]青岛市市立医院 [3]中国人民解放军第四〇一医院

出　　处：《中国病原生物学杂志》2018年第10期1073-1077,1083,共6页Journal of Pathogen Biology

基　　金：山东省优秀中青年科学家科研奖励项目(No.BS2011SW0005).

摘　　要：目的研究不动杆菌CRISPR-Cas系统的基因结构并探讨其与耐药性的关系。方法收集CRISPR db数据库所有不动杆菌菌株序列信息,利用CRISPRfinder软件分析CRISPR基因座;通过MEGA软件对cas基因序列进行对比及系统进化分析;收集自GenBank中的菌株基因注释,查找相关耐药基因信息,分析其与CRISPR基因座之间的关系。结果 80株不动杆菌包含11个种,21.3%的菌株含有确定CRISPR基因座,26.5%的菌株只含可疑基因座。基因座位于染色体或质粒上。大部分不动杆菌的CRISPR系统属于I-F型,并且根据cas1、cas3、csy2、csy3、cas6/csy4基因序列及csy1基因的存在均可分为I-Fa和I-Fb两个亚型。不动杆菌CRISPR基因座中有14种不同的重复序列,长度为24~31bp,平均重复次数为53,且不同亚型间的重复序列不同,而同一亚型内保守性较高。无CRISPR系统的菌株中A类和D类β-内酰胺酶、乙酰转移酶、核苷酸转移酶及16SrRNA甲基化酶等耐药基因的携带率均显著高于有CRISPR系统菌株。结论不动杆菌CRISPR系统属于I-Fa和I-Fb型,cas3、csy2、csy3、csy4、cas1基因序列均可单独作为不动杆菌属CRISPR系统分亚型的依据;CRISPR基因座的存在可降低某些耐药基因的水平转移,而基因组中相对较低的CRISPR系统携带率可能是导致不动杆菌耐药率日趋增高及更易出现多重耐药的原因之一。Objective To study the genetic structure of the CRISPR-Cas system and to explore its relationship to the drug resistance of Acinetobacter. Methods Positions of the CRISPR-cas loci were retrieved from the CRISPR database. CRISPR loci were identified in strains using CRISPRFinder. All cas loci were detected via annotation of the complete ge- nome sequence of Acinetobacter downloaded from PubMed, and the extracted cas gene sequences were imported into the software MEGA7. 0 for multiple sequence alignment and phylogenetic analysis. Information on the drug resistance of Acinetobacter strains with detailed annotations was retrieved from the GenBank database, and its relationship to the pres- ence or absence of CRISPR was statistically analyzed. Results Eighty strains belonged to 11 species of Acinetobacter. Of those strains, 17 contained confirmed CRISPR loci （21.3%） and 21 contained only possible loci （26.5%）. CRISPR loci can be located on a chromosome or plasmid. Most of the CRISPR Cas systems in the genus were of type I-F and can be divided into two subtypes, I-Fa and I-Fb, based on the sequence of cas1, cas3, csy2, csy3, and cas6/csy4 and the presence of the csyl gene. A total of 14 unique repeat sequences in confirmed CRISPR arrays of Acinetobacter were analyzed. The average number of direct repeats was 53, and these sequences were typically 24-31 bp in length. The repeat sequences are highly conserved within the same subtype and differ among different subtypes. Drug resistance genes such as class A and D β-lactamase, acetyltransferases, nucleotidyltransferases, and 16s rRNA methylase were detected at a significantly higher rate in strains with no CRISPR system than in strains with a CRISPR system （including a possible system）. Conclusion CRISPR arrays of Acinetobacter are mainly found in the I-Fa and I-Fb subtypes. In addition to cas1, cas3, csy2, csy3, and csy4 gene sequences can all be used to classify strains in the genus Acinetobacter. A CRISPR system （including a possible system） can prevent

关 键 词：CRISPR-Cas系统 不动杆菌 耐药基因 基因结构 

分 类 号：R378.4[医药卫生—病原生物学]