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INVESTIGATIVE REPORT ActaDV ActaDV Advances in dermatology and venereology Acta Dermato-Venereologica
Molecular Characteristics of Mycobacterium tuberculosis Strains Isolated from Cutaneous Tuberculosis Patients in China
Haiqin JIANG 1, 2 #, Yali JIN 1, 2 #, Varalakshmi VISSA 1, 2, Liangfen ZHANG 3, Weijun LIU 1, 2, Lianhua QIN 4, Kanglin WAN 5, Xiaocui WU 5, Hongsheng WANG 1, 2, Weida LIU 1, 2 and Baoxi WANG 1, 2
1
Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College, 2 Jiangsu Key Laboratory of Molecular Biology for Skin Diseases and STIs, Nanjing, China, 3 Department of Pathology, University of Mississippi Medical Center, Jackson, USA,
4
Shanghai Key Laboratory of Tuberculosis, Shanghai Pulmonary Hospital, Medical School, Tongji University, Shanghai, and 5 State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China
#
These authors contributed equally to this work.
Cutaneous tuberculosis( CTB) is probably underreported due to difficulties in detection and diagnosis. To address this issue, genotypes of Mycobacterium tuberculosis strains isolated from 30 patients with CTB were mapped at multiple loci, namely, RD105 deletions, spacer oligonucleotides, and Mycobacterial Interspersed Repetitive Unit-Variable Number Tandem Repeats( MIRU-VNTRs). Fifty-eight strains of pulmonary tuberculosis( PTB) were mapped as experimental controls. Drug resistance-associated gene mutations were determined by amplicon sequencing of target regions within 7 genes. Beijing family isolates were the most prevalent strains in CTB and PTB. MIRU-VNTR typing separated the Beijing strains from the non-Beijing strains, and the majority of CTB could be separated from PTB counterparts. Drug resistance determining regions showed only one CTB strain expressing isomazid resistance. Thus, while the CTB strains belonged to the same phylogenetic lineages and sub-lineages as the PTB strains, they differed at the level of several MIRU-VNTRs and in the proportion of drug resistance.
Key words: tuberculosis; cutaneous; spoligotyping; MIRU- VNTR; RD deletion, drug resistance.
Accepted Nov 10, 2016; Epub ahead of print Nov 14, 2016 Acta Derm Venereol 2017; 97: 472 – 477.
Corr: Hongsheng Wang and Weida Liu, Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College, 12 Jiangwangmiao Road, Nanjing 210042, China. E-mail: whs33 @ vip. sina. com and wanghs @ ncstdlc. org
Tuberculosis( TB) is a major public health problem worldwide, especially in China. Extrapulmonary TB( EPTB) is well known in the history and literature of TB; however, there are few reports on the genetic, biochemical and drug sensitivity properties of EPTB( 1 – 3). Cutaneous TB( CTB) is a difficult to diagnose form of EPTB. One report estimated that CTB comprises approximately 1 – 2 % of all clinical forms of TB( 4, 5). Over the past 2 – 3 decades, CTB has been diagnosed and treated successfully at the Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College, Nanjing, China, and strains have been cultured from skin specimens( 6). The identification of CTB cases in a background of known high levels of pulmonary TB( PTB) and multidrug-resistant TB( MDR-TB) in mainland China prompted us to further characterize these archived CTB along with additional isolates derived from subsequently diagnosed cases.
Recent developments have led to rapid detection of Mycobacterium tuberculosis DNA or RNA in clinical specimens by in vitro nucleic acid amplification, enabling investigations into epidemiology, transmission and PTB outbreaks( 7). IS6110-based restriction fragment length polymorphism( RFLP) has been the gold standard in genotyping for more than a decade, but this technique is laborious and expensive, and requires a large amount of chromosomal DNA( 8). Spoligotyping is based on polymorphisms of the chromosomal direct repeat( DR) locus, which contains a variable number of short DRs interspersed with non-repetitive spacers( 9). This method is commonly used to differentiate M. tuberculosis complex strains. Mycobacterial Interspersed Repetitive Unit-Variable Number Tandem Repeat( MIRU-VNTR) analysis( 10, 11) is a rapid and simple genotyping method with a discriminatory power similar to that of RFLP( 12). MIRU-VNTR has been proposed by an international consortium as a standardized genotyping scheme, with 15- and 24-loci sets demonstrated to have adequate discriminatory power for tracing transmission and investigating the phylogenetics of TB( 13). Other markers, including single-nucleotide polymorphisms( SNPs) and deletions at regions of difference( RD) loci, identify deeper phylogenetic origins and their branches. Whole-genome sequencing, an emerging highly discriminatory, sensitive and affordable methodology, is yet to be universally adopted in clinics and research centres; therefore comparison of genotypes using a uniform set of markers is limited. At strain level, MIRU-VNTRs with their higher rates of change than RD and DR deletion markers, aid in breaking larger phylogenetic clusters, enabling discrimination levels suitable for tracing recent transmission links.
The Beijing family, a dominant M. tuberculosis genotype, which belongs to Lineage 2, is detected in PTB strains in China and the rest of Asia, and exhibits important pathogenic features that may be associated doi: 10.2340 / 00015555-2577 Acta Derm Venereol 2017; 97: 472 – 477
This is an open access article under the CC BY-NC license. www. medicaljournals. se / acta Journal Compilation © 2017 Acta Dermato-Venereologica.