PRELIMINARY MAPPING STUDY OF Mycobacterium tuberculosis' COMPLEX DNA BY Spoligotyping ON ISOLATED SPUTUM OF TUBERCULOSIS PATIENT FROM 10 PROVINCE CAPITAL CITY

Vivi Lisdawati, Ida Parwati, Pratiwi Sudarmono, T. Mirawati Sudiro, Ririn Ramadhany, Nelly Puspandari, Lutfah Rif’ati, Triyani S. .

Abstract


Mapping TB genotype of Mycobacterium tuberculosis (Mtb) is an important
study to identify their distribution or characteristic and also may lead to improve control
of the disease. This study conducted a preliminary mapping of the tubercle bacilli which
had been circulating in Indonesia. Cultures of DNA isolated from TB patients at urban
areas in 16 provinces in Indonesia, are chosen based on TB Case Detection Rate (CDR)
2006 from Indonesia Directorate General of Communicable Disease Control and
Environment Health (Ministry of Health), were analyzed by spoligotyping for strain
differentiation. In this first part, the analyzed result only came from urban areas in 10
provinces, i.e. Palembang, Bandar Lampung, Serang, Jakarta, Bandung, Surabaya,
Banjarmasin, Makassar, Pontianak and Ambon.
Sample were 269 DNA from 294 isolates collected from sputum of suspect TB patients
with sputum-smear positive (SS+) and age above 15 years old. All samples were obtained
from peripheral health laboratory in each province. The procedure collection is
accordance to Indonesia DOTs guidelines (AFB smears) and samples were transported
from those 10 areas to Bacteriology Laboratory at CBPRD. Sputum was taken for culture
in liquid media MGIT Bactec 960 and solid media Lowenstein Jensen. The DNAs from
positive liquid media MGIT Bactec 960 were isolated and analyzed by spoligotyping to
identify the spoligo pattern. The spoligotyping results converted into octal format within
Words & Excel spreadsheets and compared to International Spoligotype-database
(SpolDB4).
The previous study (Parwati et.al.) found some differences geographical distribution
between Beijing genotype strain of tubercle bacilli in West Indonesia compare to East
Indonesia, and the same pattern was also found in this study. Furthermore, the results in
this study showed the differences in spoligo pattern of Mtb complex at 10 urban areas in
West, Middle and East Indonesia. The percentage of Beijing strain family in the samples
from West Indonesia showed around 26.61% (31.48% in Sumatra, 28.83% in Java and
16.98% in Kalimantan); from the Middle Indonesia around 25.93%; and none were
found in samples from the East Indonesia. The SpolDB4 pattern also showed that the
majority of isolates belonged to major clades of M.tuberculosis, i.e. the East African-
Indian (EAI); Haarlem (H), Latin American-Mediterranean (LAM), the Central and
Middle Eastern Asian (CAS); U = undefined; T = T family; and the MANU/ Manu family.
We also found some isolates of Mycobacterium bovis. There were no significant
association showed between genotype families and gender, but strong significant
association found with ethnics and geography. Further confirmation of the results is still
Bul. Penelit. Kesehat, Vol. 38, No. 4, 2010: 169 - 185
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ongoing (κ value; RFLP and MIRU-VNTR). As conclusion, this study constituted a first
attempt to describe the preliminary mapping of genetic population structure of the
tubercle bacilli circulating in Indonesia.

Key words: preliminary mapping, Mtb complex, spoligotyping, Beijing genotype strain,
SpolDB4


References



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