中国媒介生物学及控制杂志 ›› 2014, Vol. 25 ›› Issue (4): 289-293.DOI: 10.11853/j.issn.1003.4692.2014.04.001

• 论著 •    下一篇

海南省澄迈县斑点热群立克次体分离株多间隔序列分型

程雪琴1,金玉明2,老世军2,黄昌和2,黄芳2,贾鹏本2,张丽娟1   

  1. 1 中国疾病预防控制中心传染病预防控制所无形体室,北京102206;
    2 海南省疾病预防控制中心,海口570203
  • 收稿日期:2014-02-21 出版日期:2014-08-20 发布日期:2014-08-20
  • 通讯作者: 张丽娟,Email: zhanglijuan@icdc.cn
  • 作者简介:程雪琴,女,在读硕士,主要从事无形体及立克次体病研究,Email: cxq1cg@163.com;金玉明,男,研究员,主要从事传染病防控工作,Email: jym1030@126.com
  • 基金资助:
    “十二五”传染病重大专项课题(2008ZX10004-008,2012ZX10004215);国家基础研究项目(973计划)(2010CB530206)

Multispacer typing of spotted fever group rickettsiae isolated from Chengmai county of Hainan province, China

CHENG Xue-qin1, JIN Yu-ming2, LAO Shi-jun2, HUANG Chang-he2, HUANG Fang2, JIA Peng-ben2, ZHANG Li-juan1   

  1. 1 Department of Anaplasma,National Institute for Communicable Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing102206, China;
    2 Hainan Center for Disease Control and Prevention,Haikou570203,Hainan Province, China
  • Received:2014-02-21 Online:2014-08-20 Published:2014-08-20
  • Contact: ZHANG Li-juan, Email: zhanglijuan@icdc.cn
  • Supported by:
    Supported by the National Key Science and Technology Projects of China(No. 2008ZX10004-008,2012ZX10004215)and the National Basic Research Program of China(973 Program)(No. 2010CB530206)

摘要: 目的 采用多间隔序列分型(MST)对从海南省澄迈县发热患者及野鼠分离的62株斑点热群立克次体进行分型研究。方法 根据文献检索,应用9对成功用于斑点热群立克次体MST分型引物,对实验62株菌进行PCR扩增及测序。校正后序列进行Blast分析。同时,对序列进行Megalign比对分析单核苷酸多态性(SNPs)。将每株细菌成功测序的6段间隔序列按一定顺序拼接并构建进化树。结果 使用6对间隔序列引物扩增实验菌株,结果显示每对引物扩增序列均显示较大的变异性,共获得48个变异型,以h3为代表的主要变异型占21.7%(13/60),剩余的47个变异型为独立型,分别占1.6%。进化树分析结果显示48个变异型均归为一簇。同其他参考菌株比较发现,黑龙江立克次体与上述菌株关系密切。SNPs分析发现位点变异主要以T-C转换及A-G转换为主。结论 海南地区斑点热群立克次体分离株非编码间隔序列呈高度变异性。MST分型较传统的血清表型分型以及MLST分型更具有鉴别能力,对未来立克次体暴发流行可提供快速有效的流行株追踪。

关键词: 斑点热群立克次体, 多间隔序列分型, 海南省澄迈县

Abstract: Objective Spotted fever caused by spotted fever group rickettsiae (SFGR) is found throughout China. During 2007-2008, 28 human SFGR isolates and 34 rat SFGR isolates (including 15 isolates from Rattus fulvescens, 5 isolates from R. edwardsi, 7 isolates from Callosciurus erythraeus roberti, and 7 isolates from Dremomys rufigenis) were gathered by L929 cell culture. Previous research indicated that these isolates shared identical sequences of 16S rRNA,gltA, ompA, groEL, and 17×103 protein gene with each other, and were of the same serophenotype. This project aims to study the 62 SFGR isolates from fever patients and wild rodents in Chengmai county of Hainan province, China by multispacer typing (MST).Methods Nine pairs of primers which had been previously reported in MST of SFGR were used in amplification and sequencing of the 62 isolates in this study. Sequences were subjected to Blast analysis after data correction. Analysis of single nucleotide polymorphisms (SNPs) was performed with Megalign. Six spaced sequences that were successfully sequenced from each isolate were spliced and applied in the construction of phylogenetic tree. Results There were great variations among the 62 isolates. A total of 48 mutant genotypes were identified. The predominant genotype, h3, accounted for 21.7% (13/60) of all the isolates tested, and the remaining 47 genotypes were all unique, each accounting for 1.6% (1/60) of all isolates. Phylogenetic analysis showed that all these 48 genotypes were classified in the same cluster. Among all reference strains included in this study, R. heilongjiangensiswas genetically closest to these isolates. Thymine-to-cytosine transitions had the highest percentage (52/114, 45.6%) among all SNPs, followed by adenine-to-guanine transitions (37/114, 32.5%).Conclusion The non-coding spaced sequences of SFGR prevailing in Chengmai county of Hainan province are highly heterologous. The selected primers for MST of SFGR are capable of detecting rickettsial pathogens. MST can be used in the rapid and effective tracing of endemic strains in outbreak of rickettsiae.

Key words: Spotted fever group rickettsiae, Multispacer typing, Chengmai county of Hainan province

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