中国感染控制杂志2017，Vol.16Issue(1)：1-5,5.DOI:10.3969/j.issn.1671-9638.2017.01.001
全基因测序分析耐甲氧西林金黄色葡萄球菌利奈唑胺耐药相关变异位点
Whole genome sequencing for analyzing mutation sites in linezolid-resistant methicillin-resistant Staphylococcus aureus
摘要
Abstract
Objective To understand genetic mutation sites in linezolid (LZD)-sensitive and inducible resistant strains of methicillin-resistant Staphylococcus aureus (MRSA) using whole-genome sequencing,and realize mutation sites of LZD-resistant gene.Methods MRSA-MS4 with explicit genotype and whole-genome sequences was induced by LZD of different concentration gradients,LZD-resistant strain MRSA-MS4-LZD100 was obtained,minimum inhibitory concentration(MIC) was detected,domain V of 23S rRNA and ribosomal proteins L3/L4 gene in MRSAMS4-LZD100 were amplified by polymerase chain reaction (PCR),the sequenced products obtained the corresponding mutation site in contrast with the wild-type strain;Illumina PE library was constructed through paired-end sequencing by Illumina HiSeq 2000 technique,and whole genome sequencing was completed based on bioinformatics.Results MRAS-MS4-LZD100 strain was induced after 32 passages,MIC of LZD was 96 μg/mL.Sequencing of PCR products indicated the genetic variations were G2447T mutation in multiple copies of domain V of 23S rRNA gene,and Gly113Val mutation in L3 protein respectively;the whole genome of MRSA-MS4-LZD100 contained 2 744 315 bp,annotation of the whole genome found a total of 2 509 genes,11 tRNA-encoding genes and 2 entire rRNA-encoding operons.The data were submitted to the PubMed,and the GeneBank accession number JXMJ00000000 was assigned;a total of 101 SNPs and 6 Small indels were found,16 of 101SNP mutations occurred in exon,of which the variant proteins with anmino acid sequence alterations included IstB ATP binding domain-containing protein,clumping factor A,IS1272 transposase and so on;3 of 6 Small indel mutations occurred in exon,of which the variant proteins with anmino acid sequence alterations included hypothetical protein,30S ribosomal protein S1,and clumping factor A.Conclusion LZD-resistant strain MRSA-MS4-LZD100 was successfully induced by LZD;beside 23S rRNA V domain and ribosomal L3 protein,the other mutant site exist in this resistant strain,which provide some direction for subsequent study of recessive LZD resistance mechanism.关键词
耐甲氧西林金黄色葡萄球菌/利奈唑胺/高通量测序/全基因组序列Key words
methicillin-resistant Staphylococcus aureus/linezolid/high-throughput sequencing/complete genome sequence分类
医药卫生引用本文复制引用
姚伟明,邓启文,余治健,陈重,蒲彰雅,王红燕,程航,李多云,郑金鑫,邓向斌,刘晓军..全基因测序分析耐甲氧西林金黄色葡萄球菌利奈唑胺耐药相关变异位点[J].中国感染控制杂志,2017,16(1):1-5,5.基金项目
深圳市重点学科资助项目(201506093) （201506093）
深圳市科技创新课题(No.JCYJ20150402152130173、No.JCYJ20150402152130167) （No.JCYJ20150402152130173、No.JCYJ20150402152130167）
深圳市重点学科经费资助 （）
深圳市南山区课题资助(No.2015019、2015022) （No.2015019、2015022）
