论文部分内容阅读
目的筛选结核病(tuberculosis,TB)药效评价用结核分枝杆菌(Mycobacterium tuberculosis,MTB)国家参考菌株。方法取169株MTB临床分离株,采用结核杆菌生长指示管(mycobacterial growth indicator tube,MGIT)960液体培养基,对4种一线抗TB药物[链霉素(streptomycin,STR)、异烟肼(isoniazid,INH)、利福平(rifampin,RFP)和乙胺丁醇(ethambutol,EMB)]进行药敏试验,并对耐INH和/或耐RFP的菌株的相关耐药基因位点进行测序,确认突变位点;用间隔区寡核苷酸分型法(Spoligotyping)对所有菌株进行基因分型,多位点可变数量串联重复序列分型方法(mutiple loci VNTR analysis,MLVA)对北京家族MTB临床分离株进行基因分型。结果 169株MTB临床分离株中耐STR 118株,INH 143株,RFP 129株,EMB 95株,耐多药(至少对NIH和RFP耐药)120株,单耐INH 5株,单耐RFP7株,四种抗TB药均敏感共11株。143株INH耐药菌株中,kat G基因突变菌株占76.9%,inh A基因突变菌株占27.3%,aph C基因突变菌株占25.2%,至少2个基因发生突变的菌株有31株;129株RFP耐药菌株中,rpo B基因531位点发生突变,占67.4%,526位发生突变,占17.8%,516位点发生突变,占16.3%,至少2个位点发生突变的菌株有10株。169株MTB临床分离株分为北京家族和非北京家族,北京家族111株;非北京家族58株,主要有CAS家族2株,EAI家族7株,H家族11株,LAM家族6株,MANU家族6株,T家族7株,U家族1株,X3家族3株,还有15株在数据库中未表现的新基因型。111株北京家族MTB临床分离株分为4大群,其中Ⅱ群分布最广,占68.5%,代表性最好。筛选出耐多药菌株27株,敏感菌株5株,单耐异烟肼2株,单耐利福平3株。结论成功筛选出敏感菌株、单耐药菌株、MDR菌株作为候选菌株。
Objective To screen out the national reference strain of Mycobacterium tuberculosis (MTB) for evaluating the efficacy of tuberculosis (TB). Methods A total of 169 clinical isolates of MTB were obtained from four kinds of MTB mycobacterial growth indicator tubes (MGIT) 960 liquid medium. Four kinds of anti-TB drugs (streptomycin, isoniazid, , INH), rifampin (RFP) and ethambutol (EMB)], and the related resistance gene loci of INH and / or RFP-resistant strains were sequenced and confirmed Mutation sites. All strains were genotyped by Spoligotyping and MLVA was performed on Beijing family MTB clinics with multi-locus number-variable tandem repeat typing (MLVA) Isolates were genotyped. Results Among the 169 MTB clinical isolates, STR 118, INH 143, RFP 129, EMB 95, multidrug resistance (at least to NIH and RFP resistance) 120, INH 5 and RFP 7 Four anti-TB drugs were sensitive to a total of 11 strains. Among 143 strains of INH-resistant strains, 76.9% of them were kat G gene mutation, 27.3% of inh A gene mutation strains, 25.2% of aph C gene mutation strains and 31 of at least 2 gene mutation strains. 129 RFP Among the resistant strains, the rpo B gene was mutated at 531 loci, accounting for 67.4%, 526 mutated, accounting for 17.8%, 516 loci representing 16.3%, and 10 loci mutated at least 2 sites. 169 MTB clinical isolates were divided into Beijing family and non-Beijing family, Beijing family 111; Non Beijing family 58, mainly CAS family 2, EAI family 7, H family 11, LAM family 6, MANU family 6 strains, 7 T families, 1 U family, 3 X3 families and 15 new genotypes that were not shown in the database. 111 clinical isolates of Beijing family MTB were divided into 4 groups, of which group Ⅱ was the most widely distributed, accounting for 68.5% of the total, with the best representation. 27 strains of multidrug-resistant strains, 5 strains of sensitive strains, 2 strains of isoniazid alone and 3 strains of rifampicin were screened out. Conclusion The successful screening of sensitive strains, single drug-resistant strains, MDR strains as candidate strains.