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目的对2011年湖南51株非结核分枝杆菌(NTM)临床分离株进行菌种鉴定。方法从湖南省结核病防治所接收到分枝杆菌临床分离株后,通过罗氏培养基(L-J)、对硝基苯甲酸(p-nitrobenzoic acid,PNB)和噻吩-2-羧酸肼(2-thiophene carboxylic acid hydrazide,TCH)鉴别培养基和多位点PCR方法鉴别出非结核分枝杆菌后,对其进行基于rpoB基因的限制性酶切片段长度多态性(PRA)和基因测序方法的鉴定。结果从2011年湖南省51份临床分离菌株中培养出51份抗酸杆菌,通过初步鉴别(即L-J、PNB/TCH鉴别培养基培养)鉴别出NTM有51株,经rpoB-PRA及进一步的测序鉴别出50株(98.04%)NTM,分别为胞内分枝杆菌(Mycobacterium intracellμlare)22株(43.14%),鸟分枝杆菌(M.avium)13株(25.49%),脓肿分枝杆菌(M.abscessus)6株(11.76%),偶发分枝杆菌(M.fortuitum),戈登分枝杆菌(M.gordonae),新产色分枝杆菌(M.neoaurum)各2株(均占3.92%),堪萨斯分枝杆菌(M.kansasii),马赛分枝杆菌(M.massiliense),福西亚分枝杆菌(M.phocaicum)各1株(分别占1.96%);以及鼻疽诺卡菌(Nocardia farcinica)1株(1.96%)。结论抗酸染色阳性、PNB/TCH鉴定为非结核分枝杆菌临床分离株中包含不同种类的非结核分枝杆菌和诺卡菌。湖南非结核分枝杆菌病的病原至少有9种。使用分子生物学方法能够快速、准确地对其鉴别。
Objective To identify 51 strains of non-tuberculosis (NTM) clinical isolates in Hunan in 2011. Methods After receiving mycobacteria clinical isolates from Hunan Tuberculosis Prevention and Control Institute, the mycobacterium tuberculosis isolates were isolated and purified by Roche medium (LJ), p-nitrobenzoic acid (PNB) and 2-thiophene (TCH) identification medium and multi-locus PCR method to identify non-tuberculous mycobacteria, the rpoB gene-based restriction fragment length polymorphism (PRA) and the gene sequencing method were identified. Results Fifty-one strains of acid-fast bacilli were isolated from 51 clinical isolates in Hunan Province in 2011. Totally 51 strains of NTM were identified by preliminary identification (ie, LJ, PNB / TCH differential medium culture). The rpoB-PRA and further sequencing Fifty isolates (98.04%) of NTM were identified, including 22 strains of Mycobacterium intracelllare (43.14%), 13 strains of M. avium (25.49%) and 5 strains of Mycobacterium abscess (2.76%), M. fortuitum, M. Gordonae and M.neaurum each accounted for 3.92% ), M.kansasii, M.massiliense and M.phocaicum (1.96% respectively); and Nocardia farcinica) (strain 1.96%). Conclusions Acid-fast staining was positive and PNB / TCH was identified as non-MTB clinical isolates containing different types of non-TB mycobacteria and Nocardia. There are at least 9 pathogens in Hunan’s non-tuberculous mycosis disease. It can be quickly and accurately identified using molecular biology methods.