Cuende J I, Jaime M L, Gómez M T, Del Campo F, Alba A, Pérez de Diego I J
Laboratorio de Biología Molecular, Hospital Provincial San Telmo, Palencia.
Med Clin (Barc). 1995 Feb 18;104(6):207-10.
Typing at species level of Mycobacterium is usually performed by microbiological and biochemical methods that require a long time and/or sufficient amount of bacteria. Molecular biology can avoid these problems using different techniques.
A colony growth of the following mycobacteria has been analyzed: M. tuberculosis, M. kansasii, M. avium, M. intracellulare, M. gordonae, M. phlei, M. aurum, M. fortuitum, M. flavescens, M. marinum, M. xenopi, M. nonchromogenicum, M. terrae and M. chelonei. Strains were grown in Löwenstein-Jensen medium. DNA was obtained by proteolytic digestion and fenol extraction. The 16S rRNA gen was amplified by polymerase chain reaction (PCR) and the amplification was digested by HaeIII, HpaII, RsaI and AluI restriction enzymes. Restriction fragment patterns were analyzed by agarose gel electrophoresis and UV transillumination.
The combination of the patterns obtained with HpaII and RsaI was sufficient to generate 13 different combined ones. The patterns of M. intracellulare and M. avium were the same.
PCR and restriction enzyme analysis is an useful method for typing at species level of clinical isolates of mycobacteria.
分枝杆菌的菌种分型通常采用微生物学和生物化学方法，这些方法需要较长时间且/或需要足够数量的细菌。分子生物学可以通过不同技术避免这些问题。
对以下分枝杆菌的菌落生长情况进行了分析：结核分枝杆菌、堪萨斯分枝杆菌、鸟分枝杆菌、胞内分枝杆菌、戈登分枝杆菌、草分枝杆菌、金分枝杆菌、偶然分枝杆菌、微黄分枝杆菌、海分枝杆菌、蟾分枝杆菌、非产色分枝杆菌、地分枝杆菌和龟分枝杆菌。菌株在罗氏培养基中生长。通过蛋白水解消化和酚提取获得DNA。16S rRNA基因通过聚合酶链反应（PCR）扩增，扩增产物用HaeIII、HpaII、RsaI和AluI限制性内切酶消化。通过琼脂糖凝胶电泳和紫外透射对限制性片段模式进行分析。
用HpaII和RsaI获得的模式组合足以产生13种不同的组合模式。胞内分枝杆菌和鸟分枝杆菌的模式相同。
PCR和限制性内切酶分析是对分枝杆菌临床分离株进行菌种分型的一种有用方法。
