大理地区HIV相关鸟结核分枝杆菌MLVA基因分型研究  

Study on MLVA genotyping of HIV-associated Mycobacterium avium in the Dali area

作　　者：翟凯新 刘丽婷 陈晓文 陈越 高婧华 吴利先 ZHAI Kai-xin;LIU Li-ting;CHEN Xiao-wen;CHEN Yue;GAO Jing-hua;WU Li-xian(Depart-ment of Microbiology and Immunology,Dali University,Dali,Yunnan,China 671000)

机构地区：[1]大理大学基础医学院微生物与免疫学教研室,云南大理671000

出　　处：《中国病原生物学杂志》2020年第8期929-933,共5页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.81760357)。

摘　　要：目的对云南大理地区HIV相关的12株非结核分枝杆菌(non-Tuberculous mycobacteria,NTM)进行鉴定,了解其菌种构成;探讨MLVA法在大理地区HIV相关鸟分枝杆菌(Mycobacterium avium,MAV)基因分型的应用,了解MAV基因型特征和耐药情况。方法对初步鉴定的NTM通过16SrRNA、hsp65和rpoB靶基因片段测序分析进行菌种鉴定;对鉴定后的HIV相关MAV进行MLVA基因分型。采用MLVA法选取15个数目可变串联重复序列(VNTR)特异位点,运用聚合酶链反应(PCR)和琼脂糖凝胶电泳技术检测MAV的DNA指纹图谱。运用Quantity One和BioNumerics(6.6)软件对结果进行数字化处理,采用UPGMA法和MST法进行聚类分析;通过比例法对HIV相关MAV进行6种二线抗结核药物的药敏试验。结果大理地区12株HIV相关NTM中,9株为MAV,1株为帕拉分枝杆菌,1株为戈登菌,1株为脓肿分枝杆菌。9株HIV相关MAV通过MLVA法进行基因分型,15个VNTR位点呈多态性,不同位点的HGDI值存在差异,其中MATR-14为0.75,MATR-14、MATR-9等13个位点均≥0.5。9株双感MAV经UPGMA法聚类分析分为3个基因群、8个基因型,用MST法聚类分析则分为3个簇;HIV相关MAV对丙硫异烟胺(TH1321)均敏感,阿米卡星(AK)耐药率为100%(9/9),左氧氟沙星(LFX)耐药率为77.8%(7/9),莫西沙星(MFX)耐药率为66.7%(6/9),卷曲霉素(CPM)耐药率为55.6%(5/9),利福布丁(RBU)耐药率为11.1%(1/9)。结论大理地区12株HIV相关NTM分为MAV、帕拉分枝杆菌和戈登菌,其中以MAV占多数。MATR-VNTR分析,不同MAV菌株同一位点的基因片段差异明显,有多态性。MATR-14、MATR-9等13个位点分辨力高,可作为大理地区HIV相关MAV的基因分型首选位点;MAV对丙硫异烟胺(TH1321)敏感,对阿米卡星耐药率高,而且出现多药耐药。Objective To identify 12 strains of non-tuberculous mycobacteria(NTM)associated with HIV in Dali,Yunnan,and to ascertain the composition of those strains in order to explore the use of MLVA in HIV-related genotyping of Mycobacterium aviumin Dali and to ascertain its genotype characteristics and drug resistance. Methods NTM were identified using 16 SrRNA,hsp65,and rpoB target gene sequencing.MLVA genotyping of HIV-associated M.avium was performed.The MLVA method selects 15 specific sites with a variable number of tandem repeats(VNTR)and uses a polymerase chain reaction(PCR)and agarose gel electrophoresis to detect the DNA fingerprint of M.avium.The results were then digitized using the software Quantity One and BioNumerics(6.6),and cluster analysis was performed using the UPGMA method and MST method.Six second-line antituberculosis drugs were tested against HIV-related M.avium using the proportional method. Results Among the 12 HIV-associated NTM strains in the Dali area,9 were M.avium,1 was Mycobacterium paraense,1 was Gordona,and 1 was M.abscessus.Nine strains of HIV-related M.avium were genotyped using MLVA.The 15 VNTR sites are polymorphic,and the HGDI values differed for different sites.The discriminatory power of MATR-14 was 0.75,and that of 13 loci such as MATR-14 and MATR-9 was higher than 0.5.Nine strains of double-sense M.avium were divided into 3 groups of genes and 8 genotypes according to UPGMA cluster analysis and three clusters according to MST cluster analysis.HIV-associated M.aviumis sensitive to prothionamide(TH1321),it has a resistance to amikacin(AK)of 100%(9/9),a resistance of 77.8%(7/9)to levofloxacin(LFX),a resistance of 66.7%(6/9)to moxifloxacin(MFX),a resistance of 55.6%(5/9)to capreomycin(CPM),and a resistance of 11.1%(1/9)to rifabutin(RBU). Conclusion The 12 HIV-associated NTM strains isolated in the Dali area were divided into M.avium,M.paraense,and Gordona,with M.avium constituting the majority.MATR-VNTR analysis of M.avium strains in the Dali area indicated that gene fragments of the

关 键 词：鸟分枝杆菌 菌种鉴定 非结核分枝杆菌 可变串联重复序列 药敏试验 

分 类 号：R378.911[医药卫生—病原生物学]