Ashworth Maegan, Horan Kathleen L, Freeman Robert, Oren Eyal, Narita Masahiro, Cangelosi Gerard A
Seattle Biomedical Research Institute, 307 Westlake Avenue North, Suite 500, Seattle, WA 98109, USA.
J Clin Microbiol. 2008 Mar;46(3):856-62. doi: 10.1128/JCM.01146-07. Epub 2008 Jan 3.
Genotypic analysis of Mycobacterium tuberculosis isolates is increasingly applied in direct support of tuberculosis outbreak control activities. This is facilitated by PCR-based strain typing methods that enable the genotypic characterization of samples containing small numbers of M. tuberculosis cells. By using DNA extracted directly from primary diagnostic cultures, PCR-based methods were applied to a tuberculosis outbreak investigation and to surveillance in King County, Washington. In the outbreak investigation, five epidemiologically linked M. tuberculosis isolates had a unique pattern at mycobacterial interspersed repeating unit (MIRU) loci 10 and 23 when the pattern was compared to the patterns in a local MIRU locus database. In order to quickly identify new cases involving this strain (termed SBRI10), targeted genotyping at these two loci was performed with cultures from epidemiologically associated tuberculosis cases. Isolates with the characteristic genotypes at loci 10 and 23 were further analyzed by use of a 12-locus MIRU panel and by repetitive-unit-sequence-based PCR (rep-PCR). Between May 2004 and January 2005, 82 cases were screened, of which 14 were identified for further analysis and 13 were confirmed to be infected with SBRI10. Between September 2005 and August 2006, surveillance universal genotyping was performed by using the 12-locus MIRU panel with DNA from primary diagnostic enrichment cultures. A total of 161 samples were submitted for analysis, and 156 were successfully typed. Fifty-one cases formed 18 presumptive clusters by MIRU locus typing. Of these, 30 cases were confirmed to be members of 11 clusters by rep-PCR. Presumptive genotypic data were available rapidly, sometimes within 2 weeks of diagnosis. In this fashion, PCR-based genotyping provided data that can be used to prioritize disease control activities.
结核分枝杆菌分离株的基因分型越来越多地直接用于支持结核病暴发控制活动。基于聚合酶链反应（PCR）的菌株分型方法推动了这一应用，该方法能够对含有少量结核分枝杆菌细胞的样本进行基因分型。通过直接从初次诊断培养物中提取DNA，基于PCR的方法被应用于华盛顿州金县的一次结核病暴发调查和监测。在暴发调查中，将5株在流行病学上相关的结核分枝杆菌分离株的分枝杆菌散布重复单位（MIRU）位点10和23的图谱与当地MIRU位点数据库中的图谱进行比较时，发现它们具有独特的图谱。为了快速识别涉及该菌株（称为SBRI10）的新病例，对来自流行病学相关结核病病例的培养物在这两个位点进行了靶向基因分型。对在位点10和23具有特征性基因型的分离株，进一步使用12位点MIRU分型板和基于重复单位序列的PCR（rep-PCR）进行分析。在2004年5月至2005年1月期间，筛查了82例病例，其中14例被确定需进一步分析，13例被确认为感染了SBRI10。在2005年9月至2006年8月期间，使用12位点MIRU分型板对初次诊断富集培养物中的DNA进行监测通用基因分型。总共提交了161份样本进行分析，156份成功分型。通过MIRU位点分型，51例病例形成了18个推定聚类。其中，30例通过rep-PCR被确认为11个聚类的成员。推定的基因分型数据可迅速获得，有时在诊断后2周内即可获得。通过这种方式，基于PCR的基因分型提供的数据可用于确定疾病控制活动的优先次序。
