中华检验医学杂志
中華檢驗醫學雜誌
중화검험의학잡지
CHINESE JOURNAL OF LABORATORY MEDICINE
2010年
7期
594-598
,共5页
刘家云%徐修礼%孙惠平%龙铟%钱妙玲%张鹏亮%樊新%程晓东%马越云%苏明权%陈超扬%郝晓柯
劉傢雲%徐脩禮%孫惠平%龍銦%錢妙玲%張鵬亮%樊新%程曉東%馬越雲%囌明權%陳超颺%郝曉柯
류가운%서수례%손혜평%룡인%전묘령%장붕량%번신%정효동%마월운%소명권%진초양%학효가
分枝杆菌,结核%抗药性,细菌%聚合酶链反应%突变%序列分析,DNA
分枝桿菌,結覈%抗藥性,細菌%聚閤酶鏈反應%突變%序列分析,DNA
분지간균,결핵%항약성,세균%취합매련반응%돌변%서렬분석,DNA
Mycobacterium tuberculosis%Drug resistance,bacterial%Polymerase chain reaction%Mutation%Sequence analysis,DNA
目的 探讨结核分枝杆菌耐药表型与基因突变位点之间的相互关系.方法 采用序列特异性引物分别扩增92株结核分枝杆菌利福平耐药基因rpoB,异烟肼耐药基因katG、inhA、ahpC,链霉素耐药基因rrs、rpsL,乙胺丁醇耐药基因embB及喹诺酮耐药基因gyrA,SSCP筛选出突变序列,DNA测序分析突变性质.结果 59株利福平耐药株rpoB基因突变检出率94.9%(56/59),以Ser450Trp突变最多;90株异烟肼耐药株中,katG基因突变检出率38.9%(35/90),以Ser315Thr最多,3株检出inhA基因突变,ahpC基因无突变检出;34株喹诺酮耐药株中gyrA基因突变检出率82.4%(28/34),主要为Asp94Gly,其次为Ala90Val;31株链霉素耐药株中,15株检出rrs突变,最常见为A514C和A1041G,10株发生rpsL Lys88Arg突变,总的链霉素基因突变检出率为77.4%(24/31);31株乙胺丁醇耐药株中embB 基因突变检出率19.4%(6/31),主要为Met306Val.结论 耐药结核分枝杆菌耐药情况较为严重,以DNA测序为基础的基因突变分析能快速有效地检测结核分枝杆菌的rpoB、katG、gyrA、rrs、rpsL、embB 等耐药分子标识,显示了西安地区耐药性结核分枝杆菌的突变特点,为结核病的临床诊断和合理用药提供了实验依据.
目的 探討結覈分枝桿菌耐藥錶型與基因突變位點之間的相互關繫.方法 採用序列特異性引物分彆擴增92株結覈分枝桿菌利福平耐藥基因rpoB,異煙肼耐藥基因katG、inhA、ahpC,鏈黴素耐藥基因rrs、rpsL,乙胺丁醇耐藥基因embB及喹諾酮耐藥基因gyrA,SSCP篩選齣突變序列,DNA測序分析突變性質.結果 59株利福平耐藥株rpoB基因突變檢齣率94.9%(56/59),以Ser450Trp突變最多;90株異煙肼耐藥株中,katG基因突變檢齣率38.9%(35/90),以Ser315Thr最多,3株檢齣inhA基因突變,ahpC基因無突變檢齣;34株喹諾酮耐藥株中gyrA基因突變檢齣率82.4%(28/34),主要為Asp94Gly,其次為Ala90Val;31株鏈黴素耐藥株中,15株檢齣rrs突變,最常見為A514C和A1041G,10株髮生rpsL Lys88Arg突變,總的鏈黴素基因突變檢齣率為77.4%(24/31);31株乙胺丁醇耐藥株中embB 基因突變檢齣率19.4%(6/31),主要為Met306Val.結論 耐藥結覈分枝桿菌耐藥情況較為嚴重,以DNA測序為基礎的基因突變分析能快速有效地檢測結覈分枝桿菌的rpoB、katG、gyrA、rrs、rpsL、embB 等耐藥分子標識,顯示瞭西安地區耐藥性結覈分枝桿菌的突變特點,為結覈病的臨床診斷和閤理用藥提供瞭實驗依據.
목적 탐토결핵분지간균내약표형여기인돌변위점지간적상호관계.방법 채용서렬특이성인물분별확증92주결핵분지간균리복평내약기인rpoB,이연정내약기인katG、inhA、ahpC,련매소내약기인rrs、rpsL,을알정순내약기인embB급규낙동내약기인gyrA,SSCP사선출돌변서렬,DNA측서분석돌변성질.결과 59주리복평내약주rpoB기인돌변검출솔94.9%(56/59),이Ser450Trp돌변최다;90주이연정내약주중,katG기인돌변검출솔38.9%(35/90),이Ser315Thr최다,3주검출inhA기인돌변,ahpC기인무돌변검출;34주규낙동내약주중gyrA기인돌변검출솔82.4%(28/34),주요위Asp94Gly,기차위Ala90Val;31주련매소내약주중,15주검출rrs돌변,최상견위A514C화A1041G,10주발생rpsL Lys88Arg돌변,총적련매소기인돌변검출솔위77.4%(24/31);31주을알정순내약주중embB 기인돌변검출솔19.4%(6/31),주요위Met306Val.결론 내약결핵분지간균내약정황교위엄중,이DNA측서위기출적기인돌변분석능쾌속유효지검측결핵분지간균적rpoB、katG、gyrA、rrs、rpsL、embB 등내약분자표식,현시료서안지구내약성결핵분지간균적돌변특점,위결핵병적림상진단화합리용약제공료실험의거.
Objective To investigate the relationship between the phenotypes and the patterns of genetic mutations in the corresponding resistance genes (rpoB, katG, inhA, ahpC, rrs, rpsL, embB and gyrA) in resistant Mycobacterium tuberculosis (MTB) isolates. Methods Rifampicin-resistant gene (rpoB), isoniazid-resistant genes (katG, inhA, ahpC), streptomycin-resistant genes (rrs, rpsL), ethambutol-resistant gene (embB) and quinolinone-resistant gene (gyrA) were amplified by PCR with sequence-specific primers, then mutants screened by single-stranded conformation polymorphism (SSCP) were sequenced. Results rpoB mutation with predominant Ser450Trp pattern was 94. 9% (56/59) in 59 rifampicin-resistant isolates;katG mutation rate was 38. 9% (35/90) and the main pattern was Ser315Thr, but only 3 inhA mutants and no ahpC mutation were determined in 90 isoniazid-resistant isolates;gyrA mutation with main Asp94Gly then Ala90Val pattern was 82.4% (28/34) in 34 quinolinone-resistant isolates;the total mutation rate was 77.4% in 31 streptomycin-resistant isolates, of which 15 isolates mutated in rrs with main pattern A514C or A1041G, 10 isolates mutated in rpsL Lys88Arg;and embB mutation with main Met306Val accounted for 19.4% (6/31) in 31 ethambutol-resistant isolates. Conclusions The results showed that resistance of resistant MTB may be complicated, and DNA sequencing-based mutation analysis could efficiently detect the molecular makers such as rpoB, katG, gyrA, rrs, rpsL and embB in resistant MTB isolates. Meanwhile, it is notable that the rpoB mutation pattern in our isolates is different from previous report, further effort are needed to confirm the characteristics. The spectrum of potential resistance-related mutations in MTB clinical isolates may lay substantial foundation for the rapid molecular diagnosis and rational use of drug to MTB patients.
