中华结核和呼吸杂志
中華結覈和呼吸雜誌
중화결핵화호흡잡지
Chinese Journal of Tuberculosis and Respiratory Diseases
2012年
12期
907-910
,共4页
高诗会%赵英伟%胡忠义%王洁%陆俊梅%闫丽萍%郭琪%马慧%秦莲花
高詩會%趙英偉%鬍忠義%王潔%陸俊梅%閆麗萍%郭琪%馬慧%秦蓮花
고시회%조영위%호충의%왕길%륙준매%염려평%곽기%마혜%진연화
分枝杆菌属%核酸扩增技术%简单序列重复区间分型
分枝桿菌屬%覈痠擴增技術%簡單序列重複區間分型
분지간균속%핵산확증기술%간단서렬중복구간분형
Mycobacterium%Nucleic acid amplification techniques%Inter-simple sequences repeat analysis
目的 寻找新的鉴别诊断MTB复合群(MTC)高特异度和敏感的核酸靶标.方法 利用简单序列重复区间(ISSR)分型技术平台筛选MTC的特征片段,克隆测序获得特征序列并进行序列同源性分析,以该序列为基础设计MTC特征引物,并对211株分枝杆菌菌株(其中MTC107株、非结核分枝杆菌104株)进行鉴别检测.利用分枝杆菌属特征序列16s rRNA和MTC特征序列IS6110的鉴别结果,对MTC特征引物检测结果进行评估.结果 通过ISSR分型获得588 bp的MTC特征片段,该序列为MTC菌株的特征序列,位于MTB标准株H37Rv基因组的315947 ～316534位,以该序列为基础设计MTC鉴别引物MTCF/R,16s rRNA基因在211株分枝杆菌菌株中的扩增结果均为阳性,IS6110序列在107株MTC菌株中扩增阳性率为99％(106/107),在104株NTM菌株中阴性检出率为100％ (104/104),MTCF/R引物在MTC菌株中的扩增阳性率为100％(107/107),在非结核分枝杆菌菌株中的阴性检出率为100％ (104/104).结论 通过ISSR分型技术筛选出的MTC特征序列可作为新的核酸扩增靶标用于MTC和非结核分枝杆菌的鉴别诊断.
目的 尋找新的鑒彆診斷MTB複閤群(MTC)高特異度和敏感的覈痠靶標.方法 利用簡單序列重複區間(ISSR)分型技術平檯篩選MTC的特徵片段,剋隆測序穫得特徵序列併進行序列同源性分析,以該序列為基礎設計MTC特徵引物,併對211株分枝桿菌菌株(其中MTC107株、非結覈分枝桿菌104株)進行鑒彆檢測.利用分枝桿菌屬特徵序列16s rRNA和MTC特徵序列IS6110的鑒彆結果,對MTC特徵引物檢測結果進行評估.結果 通過ISSR分型穫得588 bp的MTC特徵片段,該序列為MTC菌株的特徵序列,位于MTB標準株H37Rv基因組的315947 ～316534位,以該序列為基礎設計MTC鑒彆引物MTCF/R,16s rRNA基因在211株分枝桿菌菌株中的擴增結果均為暘性,IS6110序列在107株MTC菌株中擴增暘性率為99％(106/107),在104株NTM菌株中陰性檢齣率為100％ (104/104),MTCF/R引物在MTC菌株中的擴增暘性率為100％(107/107),在非結覈分枝桿菌菌株中的陰性檢齣率為100％ (104/104).結論 通過ISSR分型技術篩選齣的MTC特徵序列可作為新的覈痠擴增靶標用于MTC和非結覈分枝桿菌的鑒彆診斷.
목적 심조신적감별진단MTB복합군(MTC)고특이도화민감적핵산파표.방법 이용간단서렬중복구간(ISSR)분형기술평태사선MTC적특정편단,극륭측서획득특정서렬병진행서렬동원성분석,이해서렬위기출설계MTC특정인물,병대211주분지간균균주(기중MTC107주、비결핵분지간균104주)진행감별검측.이용분지간균속특정서렬16s rRNA화MTC특정서렬IS6110적감별결과,대MTC특정인물검측결과진행평고.결과 통과ISSR분형획득588 bp적MTC특정편단,해서렬위MTC균주적특정서렬,위우MTB표준주H37Rv기인조적315947 ～316534위,이해서렬위기출설계MTC감별인물MTCF/R,16s rRNA기인재211주분지간균균주중적확증결과균위양성,IS6110서렬재107주MTC균주중확증양성솔위99％(106/107),재104주NTM균주중음성검출솔위100％ (104/104),MTCF/R인물재MTC균주중적확증양성솔위100％(107/107),재비결핵분지간균균주중적음성검출솔위100％ (104/104).결론 통과ISSR분형기술사선출적MTC특정서렬가작위신적핵산확증파표용우MTC화비결핵분지간균적감별진단.
Objective To identify and evaluate a new nucleic acid amplification (NAA) test target for specific detection of Mycobacterium tuberculosis (MTB) complex (MTC).Methods MTC-specific fragment was obtained by ISSR genotyping technology.Primer pairs were designed based on the sequences of MTC-specific fragment and tested in 211 mycobacterial strains including 107 MTC strains and 104 nontuberculous mycobacteria (NTM) strains.IS6110 element (specific identification of MTC strains) and 16s rRNA gene (specific identification of Mycobacterium) amplification were used as a control to evaluate the efficacy of the NAA test target in the detection of MTC strains.Results One MTC-specific fragment with the length of 588 bp,located in 315947-316534 of the genome from MTB reference strain H37 Rv,were obtained,cloned and sequenced.MTC-specific primer pairs MTCF/R were designed based on these sequences.All 211 mycobacterial strains accurately produced the genus-specific 16s rRNA amplicon.All MTC strains were positive in the MTCF/R PCR amplification while 99％ MTC strains (106/107) were positive in the amplification of IS6110 sequences.All NTM strains were negative in both IS6110 and MTCF/R PCR amplification.Conclusions The MTC-specific fragment developed in this study can be used as a new NAA test target to correctly distinguish MTC from NTM.
