铜绿假单胞菌CRISPR/Cas系统的基因结构及其与耐药基因的关系  

Analysis of the gene structure of Pseudomonas aeruginosa CRISPR/Cas system and its relationship with antibiotic resistance-related genes

作　　者：刘兴雨[1] 毕春霞[2] 韩璐[1] 迟媛媛 王梦园 兰蕾 闫志勇[4] LIU Xing-yu;BI Chun-xia;CHI Yuan-yuan;Han Lu;WANG Meng-yuan;LAN Lei;YAN Zhi-yong(Department of Clinical Laboratory,the Affiliated Hospital of Qingdao University,Qingdao 266000,Shandong,China;Qingdao Municipal Hospital;Women and Children’s Hospital,Qingdao University;Medical College of Qingdao University)

机构地区：[1]青岛大学附属医院检验科,山东青岛266000 [2]青岛市市立医院 [3]青岛市妇女儿童医院 [4]青岛大学基础医学院

出　　处：《中国病原生物学杂志》2022年第12期1399-1403,共5页Journal of Pathogen Biology

摘　　要：目的研究铜绿假单胞菌CRISPR/Cas系统基因结构及其与耐药基因的关系。方法收集95株铜绿假单胞菌的全基因组序列信息,通过CRISPRs web server获取CRISPR系统的信息。通过ClustalW进行重复序列和cas基因的比对分析,采用MEGA7构建cas1和cas3的系统发育树,通过RNAfold预测重复序列RNA的二级结构。通过GenBank数据库对间隔序列进行BLAST比对,并进行同源性分析。通过基因注释查找耐药相关基因,并分析其与CRISPR系统之间的关系。结果95株铜绿假单胞菌中,共发现130个确定的CRISPR位点,分布于58个菌株中,其中47个具有结构完整的CRISPR系统,30个I-F型,7个I-C型,6个I-E型,I-F型可以和I-C型或I-E型共存于同一株菌中;CRISPR位点共有18种重复序列,具有一定的保守性,基本均可形成保守的哑铃状RNA二级结构。不同类型CRISPR系统的cas基因组成不同,但均有cas1和cas3。cas1和cas3比较保守,可作为分型的依据。不同位点中间隔序列的长度和数量不同,2132个间隔序列中526个与噬菌体序列同源,32个与质粒序列同源。耐药基因分析显示,含CRISPR系统的菌株IntI1和OXA-10的携带率高于不含该系统菌株的携带率。结论铜绿假单胞菌基因组中CRISPR系统主要为-F型,其次为-E和-F型;同种类型CRISPR系统中的cas1和cas3基因高度保守;与间隔序列同源的外源基因主要为噬菌体,少数是质粒。CRISPR系统的存在与某些抗生素耐药相关基因的携带率有关。Objective To explore the gene structure of CRISPR/Cas system in Pseudomonas aeruginosa and its relationship with drug resistance genes.Methods Collect the complete genome sequences of 95 P.aeruginosa strains from NCBI and obtain the information of the CRISPR system through the CRISPRs web server.The comparison and analysis of the repetitive sequence and the cas gene is completed by Clustal W in DNAstar,the phylogenetic tree of cas1 and cas3 is completed by MEGA7,and the secondary structure of the repetitive RNA is predicted by RNAfold.Blast the spacer sequence with the help of GenBank database to find the homologous gene sequence,and analyze its homologous sequence.Resistance-related genes is found by gene annotation.The relationship between CRISPR system and antibiotic resistance genes is tested by chi-square test,P<0.05 indicates that the difference is statistically significant Results Among 95 strains of P.aeruginosa,we found 130 identified CRISPR locus distributed in 58 strains.43 strains are able to find the cas gene,with a structurally complete CRISPR system,among these 30 are I-F type,and 7 are I-C type,6 are I-E type,others cannot be typed.I-F type can exist in the same strain with I-C type or I-E type at the same time.There are 18 types of repetitive sequences found in 130 CRISPR locus,which are conservative and can basically form a conservative dumbbell-shaped RNA secondary structure.Different types of CRISPR systems have different cas proteins,but they all have cas1 and cas3.The phylogenetic tree indicates that the branches of cas1 and cas3 in the three CRISPR systems are independent of each other.cas1 and cas3 are relatively conservative and can be used as a basis for typing.The length and number of spacer sequences in different sites are different.Among the spacer sequencer,526 are homologous to the phage sequence and 32 are homologous to the plasmid sequence.The analysis of drug resistance genes showes that the OXA-10 carrier rate of strains with CRISPR system(13.95%) was higher than that strains wi

关 键 词：铜绿假单胞菌 基因结构 CRISPR系统 耐药相关基因 

分 类 号：R378[医药卫生—病原生物学]