Mathuria Jitendra Prasad, Sharma Pragya, Prakash Pradyot, Samaria Jai Kumar, Katoch Vishwa Mohan, Anupurba Shampa
Department of Microbiology, Institute of Medical Sciences, Banaras Hindu University, Varanasi, India; Department of Medical Lab Technology, Paramedical Vigyaan Mahavidhyalaya, Uttar Pradesh University of Medical Sciences, Saifai, Etawah, India.
National Jalma Institute of Leprosy and Other Mycobacterial Diseases, Tajganj, Agra, India.
Int J Mycobacteriol. 2016 Dec;5 Suppl 1:S174-S175. doi: 10.1016/j.ijmyco.2016.11.015. Epub 2016 Nov 25.
OBJECTIVE/BACKGROUND: Molecular epidemiology methods are very useful for differentiating between strains, assessing their diversity, and measuring the prevalence of the most circulating strain in an area. Various molecular typing methods using different molecular markers have been utilized worldwide, such as restriction fragment length polymorphism (RFLP), spoligotyping, Mycobacterial Interspersed Repetitive Unit - Variable Number of Tandem Repeat (MIRU-VNTR), and Double repetitive element-PCR (DRE-PCR) typing, for simultaneous detection and epidemiologic typing of Mycobacterium tuberculosis. The present study is conducted to assess the genetic diversity of M. tuberculosis by IS6110-RFLP and spoligotyping in patients attending a tertiary care hospital in eastern Uttar Pradesh, North India.
A total of 83 representative isolates of M. tuberculosis were included in this study. These isolates were subjected to spoligotyping and IS6110-RFLP DNA fingerprinting techniques as described previously.
The spoligotype patterns were compared with SpolDB4.0; patterns of 64 out of 83 M. tuberculosis isolates were matched with the available data, while 19 isolates were found to be orphan, that is, absent in the SpolDB4.0 database. The majority of the M. tuberculosis strains (56.5%) belong to central Asian (32.5%), ill defined T (13.2%), and Beijing (10.8%) families. On IS6110-RFLP analysis, in 19.2% (16/83) of these isolates, IS6110 element was not found (0 copy number strains). Further, 15.6% (13/83) isolates were found to be low-copy-number strains having less than six copies of IS6110 element, and the remaining 65.0% (54/83) were multiple-copy-number strains with six or more copies of the element. On comparing the results of spoligotyping and IS-6110-RFLP, a total of 47 isolates were clustered by spoligotyping; out of these isolates, 40 were found to be unique by IS6110-RFLP.
Spoligotype analysis resulted in the grouping of a much larger number of isolates within apparently identical clusters compared with IS6110-RFLP typing, while IS6110-RFLP was not found to effectively distinguish between zero- and low-copy-number isolates. Therefore, we concluded that, in India, the use of both the techniques simultaneously for DNA fingerprinting of M. tuberculosis could be a better approach.
目的/背景：分子流行病学方法对于区分菌株、评估其多样性以及测定某地区最常见流行菌株的患病率非常有用。世界各地已采用了各种使用不同分子标记的分子分型方法，如限制性片段长度多态性（RFLP）、间隔寡核苷酸分型、分枝杆菌插入重复单位 - 可变数目串联重复序列（MIRU - VNTR）以及双重复元件PCR（DRE - PCR）分型，用于结核分枝杆菌的同时检测和流行病学分型。本研究旨在通过IS6110 - RFLP和间隔寡核苷酸分型评估印度北部北方邦东部一家三级护理医院患者中结核分枝杆菌的遗传多样性。
本研究共纳入83株具有代表性的结核分枝杆菌分离株。按照先前描述的方法，对这些分离株进行间隔寡核苷酸分型和IS6110 - RFLP DNA指纹图谱技术分析。
将间隔寡核苷酸分型模式与SpolDB4.0进行比较；83株结核分枝杆菌分离株中有64株的模式与现有数据匹配，而19株分离株被发现为孤儿株，即在SpolDB4.0数据库中不存在。大多数结核分枝杆菌菌株（56.5%）属于中亚家族（32.5%）、未明确的T家族（13.2%）和北京家族（10.8%）。在IS6110 - RFLP分析中，这些分离株中有19.2%（16/83）未发现IS6110元件（0拷贝数菌株）。此外，15.6%（13/83）的分离株被发现为低拷贝数菌株，其IS6110元件拷贝数少于6个，其余65.0%（54/83）为多拷贝数菌株，元件拷贝数为6个或更多。比较间隔寡核苷酸分型和IS - 6110 - RFLP的结果，共有47株分离株通过间隔寡核苷酸分型聚类；在这些分离株中，40株通过IS6110 - RFLP被发现是独特的。
与IS6110 - RFLP分型相比，间隔寡核苷酸分型分析导致在明显相同的聚类中分组的分离株数量更多，而IS6110 - RFLP未被发现能有效区分零拷贝数和低拷贝数分离株。因此，我们得出结论，在印度，同时使用这两种技术对结核分枝杆菌进行DNA指纹图谱分析可能是一种更好的方法。
