摘要
目的:分析创口分离一株耐甲氧西林金黄色葡萄球菌(MRSA)的全基因组序列特点。方法:将深圳大学南山区人民医院一例创口脓液分离菌株采用BD Phoenix TM100自动细菌鉴定/药敏系统鉴定;用E-Test试验测定其利奈唑胺(LZD)最低抑菌浓度(MIC)值;扩增细菌提取细菌总DNA,应用多重PCR扩增及DNA测序方法行SCCmec-spaMLST分型;并采用Illumina Miseq结合第三代测序技术分别构建了Illumina Miseq PE文库(~500 bp)和Pac Bio文库(8~10 kb),普通PCR覆盖gap,对获得的测序数据进行质控后利用生物信息学分析手段完成该菌株的全基因组完成图绘制。结果:经鉴定此菌株为耐甲氧西林金黄色葡萄球菌(命名为MS4)。药敏结果显示该菌株对头孢西丁、氨苄西林、甲氧西林、青霉素P、阿莫西林/克拉维酸耐药,对利奈唑胺等其余抗菌素敏感。E-Test提示LZD MIC值为0.094 ug/m L。SCCmecspa-MLST分型结果显示该菌株属于III-t 3592-ST59型。MS4基因组包含2709797 bp,GC含量33.5%;通过注释后总共有2475个基因,11个t RNA编码基因,3个完整的r RNA基因编码操纵子,并包括mec A等耐药基因和γ-溶血素、溶血素III、烯醇酶、肠毒素等多种毒力因子,申请获得Genebank号CP009828。结论:完成了一株创口脓液来源MRSA的基因分型和全基因组完成图绘制,为MRSA基因组比对和系统进化发育分析提供了模板序列。
Objective To explore the complete genome sequence of a wound fester isolated methicillin-resistant staphylococcus aureus. Methods A strain separated from the wound secretions was subjected to BD Phoenix TM 100 Automatic bacteria identification/susceptibility system for determination of strain type and various drug MICs. LZD MIC was detected by E-Test approach. The bacteria DNA was extracted; SCCmec-spa-MLST types were made according to the target DNA fragment sequence amplified by multiple PCR using respective primers; Illumina Miseq PE library（ ~ 500bp） and Pac Bio library（8 ~ 10kb） were constructed using Illumina Miseq technique coupled with the third-generation sequencing, while the gaps were repaired by normal PCR; the complete genome sequence was finally gained after bioinformatic analysis for the original data. Results The strain was confirmed as methicillin-resistant staphylococcus aureus named as MS4, which was shown to resistant to Cefoxitin, ampicillin, methicillin, penicillin P, amoxicillin/clavulanic acid but remained sensitive to linezolid, and belonged to the III-t3592-ST59 type. The completed MS4 circular chromosome contained 2709797 bp with a GC content of 33.5 %. Annotation of the whole genome created a total of 2475 genes, 11 t RNA-encoding genes, three entire r RNA-encoding operons, one drug resistant gene mec A and various virulence factors such as hlg, hlg III, Eno, and other virulence factors（Genebank： CP〈009828）. Conclusion The genetic type and whole genomic sequencing of an MRSA strain isolated from the wound fester were completed, which facilitated the subsequent investigations of genome comparison and evolutionary development of MRSA through providing the DNA template.
出处
《深圳中西医结合杂志》
2017年第15期6-9,共4页
Shenzhen Journal of Integrated Traditional Chinese and Western Medicine
基金
深圳市科技创新委资助课题(JCYJ20150402152130173
JCYJ20150402152130167)
深圳市南山区资助课题(2015019
2015022)
深圳市重点学科资助课题(201506093)
深圳市重点学科经费资助课题
