Genotyping Sekuens Nukleotida Gen 16S rRNA Mycobacterium tuberculosis Dari Pasien TB Paru dan Pengembangan Metode Multiplex PCR untuk Identifikasi Mycobacterium tuberculosis Dan MOTT

Indonesia merupakan negara dengan kasus tuberkulosis (TB) yang termasuk dalam high burden countries, dengan insiden dan prevalensi tinggi Domar 2 setelah India. Provinsi Jawa Timur tercatat 112 kasus TB /100.000 penduduk. Penegakan diagnostis yang cepat, tepat, dan akurat diperlukan untuk penentuan pengobatan tepat, cepat, dan adekuat memutus rantai penularan merupakan langkah strategi pengendalian pcnyakit TB. Metode diagnosis molekuler dengan target gen spesifik sangat diperlukan dalam upaya penegakan diagnosis infeksi Mycobacterium tuberculosis. Gen 165 rRNA merupakan gen conserved pada berbagai strain Mycobacterium tuberculosis, studi analisis genotyping dapat menentukan regio DNA conserved dan spesifik pada MTBC yang berbeda dengan MOTT, dapat menjadi gen spesifik untuk identifikasi dan diferensiasi dalam penegakan diagnosis infeksi penyakit TB paru

A 5‑year evaluation of chemoprophylactic treatment in elementary school children with subclinical leprosy

Subclinical leprosy is an infectious disease in which the immune system remains infected with Mycobacterium leprae (M. leprae). The progress of subclinical leprosy to clinical cases within 1 year of infection is 1.5%, with an increase to 6% in the following 4 years. Rifampicin is frequently used for prevention of leprosy, and clarithromycin has a bactericidal effect on M. leprae. Thus, the combination of both is expected to improve disease control in patients with subclinical leprosy. The aim of the present study was to evaluate the efficacy of a chemoprophylactic treatment involving rifampicin and clarithromycin against subclinical leprosy in elementary school children from endemic areas of East Java over a 5‑year period. The study was performed between 2011 and 2015. Samples were collected from 2,548 healthy elementary school children in Nguling (Pasuruan) and Raas (Sumenep), and analysed using ELISA for anti‑PGL (phenolic glycolipid)‑1 IgM antibodies. Children who were seropositive for anti‑PGL‑1 IgM antibodies received a chemoprophylactic regimen consisting of rifampicin (300 mg/day) and clarithromycin (250 mg/day) daily for the initial 10 days, followed by the same regimen every 2 weeks for 3 months. Clinical and serological evaluations were performed annually for 5 years. Amongst the 2,548 healthy elementary school children, 200 were seropositive. The anti‑PGL‑1 IgM antibody levels significantly decreased between 2011 and 2015 in Nguling (from 1,066.7 to 137.4 U/ml) and Raas (from 773.1 to 563.4 U/ml), the levels decreased every year. In addition, the proportion of patients with decreased anti‑PGL‑1 IgM antibody levels was consistently higher than patients with increased anti‑PGL‑1 IgM antibody levels in all periods, except during 2013‑2014, in Nguling and Raas. Chemoprophylactic treatment involving rifampicin and clarithromycin may thus be effective against subclinical leprosy amongst elementary school children

ANALISIS SEKUENS NUKLEOTIDA GEN 16S rRNA Mycobacterium tuberculosis Complex (MTBC) DARI SPUTUM PASIEN TB PARU

Tuberculosis is infectious disease which was caused by the member of pathogenic bacteria group Mycobacterium tuberculosis Complex (MTBC). This study was aimed to analyze nucleotide sequence of 16S rRNA gene from MTBC group isolated in pulmonary tuberculosis patient sputums. Sequencing results was compared to sequences of Mycobacterium tuberculosis strain H37Rv (wildtype) to determine species homology. MTBC species homology hopefully could be used as information for the benefit of pulmonary tuberculosis molecular epidemiology. This study used sputum samples of pulmonary TB patients from TB clinic, Dr. Soetomo General Hospital, Surabaya collected from September to November 2016, amounted up to 96 samples. Methods used includes smear BTA staining, microbe culture, rapid test, and molecular method, such as 16S rRNA gene amplification as used in this study to determine bacterial nucleotide sequences from pulmonary TB, in order to analyze bacterial homology to different virulent strain of Mycobacterium tuberculosis (strain H37Rv). The samples with positive PCR and positive culture results were proceeded to the next stage of sequencing. Sequences obtained from genotyping 16S rRNA gene in sputum of pulmonary TB had 70-76% homology to the MTBC strains gene from the Gene Bank. Percentage was caused by considerable variation in the regions suspected as the variable region. Analysis of Mycobacterium tuberculosis Complex using 16S rRNA gene target could not be used yet as basis for determining MTBC group but this gene could be used for identification or phylogenetic analysis and nucleotides variation in variable regio

MULTIPLEX PCR GEN 16S rRNA, rv0577, RD9, mtbk_20680, lineage 1-6 DAN PROFIL GENOTIPE GEN rpoB PADA IDENTIFIKASI SPESIES, STRAIN, DAN RESISTENSI RIFAMPISIN Mycobacterium tuberculosis ISOLAT PASIEN TUBERKULOSIS PARU DI PULAU JAWA

Objective: To classify the bacteria causing tuberculosis infection and also to identify gene mutations related with rifampicin resistance in Indonesia. Material and Methods: A total of 110 isolates of pulmonary tuberculosis patients from Java Island, Indonesia, were collected from July 2017- October 2018. All samples were examined by molecular methods using specific gene amplification to identify species, strains, and gene mutation in the Mycobacterium isolates, while VNTR and qPCR were also used as methods to confirm species and strains identification. Results: There were 60,9% Beijing strains and 39,1% non Beijing strains. In addition, identification using lineage analysis exists lineage 1: 0,9%; lineage 2: 57,3%; lineage 4: 37,3%; 3,6% mixed; 0,9% without SNPs; lineage 3, lineage 5, and lineage 6 were not found. The mixed strains and isolate without SNPs were continued to use VNTR and qPCR method for confirmation. Rifampicin resistance was confirmed by sequencing of rpoB gene (13,6%), and Asp435Phe mutations is firstly reported in Java Island. Conclusion: The Beijing strains and also the rpoB gene mutation in codon 450 were most commonly found in Java Island

Application of serial tests for Mycobacterium tuberculosis detection to active lung tuberculosis cases in Indonesia

Objective: Rapid detection and accurate diagnosis are very important in managing active tuberculosis because they provide an advantage in preventing further disease transmission. In accordance with the recommendation of the World Health Organization, the Indonesian Tuberculosis Control Program uses the acid fast bacilli (AFB) smear and Chest X-ray methods as the primary methods for detecting tuberculosis, especially in new cases of suspected tuberculosis. The genus Mycobacterium has many species, strains, and variants, and their natural differences may affect the clinical outcome of the diseases they induce. The purpose of this study was to assess different tuberculosis detection methods as part of serial tests and determine the best diagnostic approach for detecting active lung tuberculosis in Indonesia. Results: This study used clinical samples from tuberculosis patients and assessed them using a series of tests, aiming to increase the sensitivity of active tuberculosis detection. Some samples that yielded negative results in the AFB smear test were detected as positive for Mycobacterium tuberculosis using the nucleic acid amplification test, with a sensitivity of 83.1%. Additionally, nucleic acid amplification also detected positive results among samples assessed as M. tuberculosis-negative using the culture method, this method yielded the same results as the Gene Xpert test