Viana-Niero C, de Haas P E, van Soolingen D, Leão S C
Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo - Escola Paulista de Medicina (UNIFESP-EPM), Rua Botucatu, 862 3° andar, 04023-062, São Paulo, Brazil.
Diagnostic Laboratory of Infectious Diseases and Perinatal Screening, National Institute of Public Health and the Environment (RIVM), PO Box 1, 3720 BA, The Netherlands.
Microbiology (Reading). 2004 Apr;150(Pt 4):967-978. doi: 10.1099/mic.0.26778-0.
The Mycobacterium tuberculosis genome contains four highly related genes which present significant similarity to Pseudomonas aeruginosa genes encoding phospholipase C enzymes. Three of these genes, plcA, plcB and plcC, are organized in tandem (locus plcABC). The fourth gene, plcD, is located in a different region. This study investigates variations in plcABC and plcD genes in clinical isolates of M. tuberculosis, Mycobacterium africanum and 'Mycobacterium canettii'. Genetic polymorphisms were examined by PCR, Southern blot hybridization, sequence analysis and RT-PCR. Seven M. tuberculosis isolates contain insertions of IS6110 elements within plcA, plcC or plcD. In 19 of 25 M. tuberculosis isolates examined, genomic deletions were identified, resulting in loss of parts of genes or complete genes from the plcABC and/or plcD loci. Partial plcD deletion was observed in one M. africanum isolate. In each case, deletions were associated with the presence of a copy of the IS6110 element and in all occurrences IS6110 was transposed in the same orientation. A mechanism of deletion resulting from homologous recombination of two copies of IS6110 was recognized in a group of genetically related M. tuberculosis isolates. Five M. tuberculosis isolates presented major polymorphisms in the plcABC and plcD regions, along with loss of expression competence that affected all four plc genes. Phospholipase C is a well-known bacterial virulence factor. The precise role of phospholipase C in the pathogenicity of M. tuberculosis is unknown, but considering the potential importance that the plc genes may have in the virulence of the tubercle bacillus, the study of isolates cultured from patients with active tuberculosis bearing genetic variations affecting these genes may provide insights into the significance of phospholipase C enzymes for tuberculosis pathogenicity.
结核分枝杆菌基因组包含四个高度相关的基因，它们与铜绿假单胞菌中编码磷脂酶C的基因具有显著相似性。其中三个基因，即plcA、plcB和plcC，串联排列（基因座plcABC）。第四个基因plcD位于不同区域。本研究调查了结核分枝杆菌、非洲分枝杆菌和“卡内蒂分枝杆菌”临床分离株中plcABC和plcD基因的变异情况。通过聚合酶链反应（PCR）、Southern印迹杂交、序列分析和逆转录聚合酶链反应（RT-PCR）检测基因多态性。七株结核分枝杆菌分离株在plcA、plcC或plcD基因内存在IS6110元件插入。在检测的25株结核分枝杆菌分离株中，有19株鉴定出基因组缺失，导致plcABC和/或plcD基因座部分基因或完整基因缺失。在一株非洲分枝杆菌分离株中观察到plcD部分缺失。在每种情况下，缺失都与IS6110元件的一个拷贝的存在相关，并且在所有情况下IS6110都以相同方向转座。在一组遗传相关的结核分枝杆菌分离株中识别出由两个IS6110拷贝同源重组导致的缺失机制。五株结核分枝杆菌分离株在plcABC和plcD区域呈现主要多态性，同时表达能力丧失，影响了所有四个plc基因。磷脂酶C是一种著名的细菌毒力因子。磷脂酶C在结核分枝杆菌致病性中的精确作用尚不清楚，但考虑到plc基因可能在结核杆菌毒力中具有的潜在重要性，对患有活动性结核病且携带影响这些基因的遗传变异的患者培养的分离株进行研究，可能会深入了解磷脂酶C酶对结核病致病性的意义。
