Detection of Mutations in 81-bpRifampin Resistance Determining Region (RRDR) of rpoB gene in Mycobacterium tuberculosis using GeneXpert MTB/RIF in Clinical Specimens from Quetta, Pakistan

GeneXpert MTBIRIF has revolutionized the tuberculosis diagnosis by simultaneous detection of Mycobacterium tuberculosis and resistance to RIF (rifampicin), a surrogate marker for multidrug-resistant TB in less than two hours. The RIF-resistance pattern in Balochistan, Pakistan, is not documented. This study was aimed to detect RIF-resistant TB and mutations in RNA polymerase beta (rpoB) gene of M. tuberculosis within 81-bp RRDR in Quetta, Pakistan using GeneXpert (R) MTB/RIF assay. In total, 2300 clinical specimens were collected from suspected TB patients at Fatima Jinnah General and Chest Hospital Quetta, Pakistan between January and August 2017. These specimens were analyzed by GeneXpert (R) MTB/RIF assay. The data was statistically analyzed using SPSS software. Out of 2300 clinical specimens, M. tuberculosis was positive in in 899 (39.1%) cases by GeneXpere MTB/RIF assay [positive respiratory cases 42.9% (871/2032) and non-respiratory 10.4% (28/268) with statistically significant difference (chi(2) = 104.5, p<0.001)]. Among 899 MTB positive cases, 46 (5.1%) were RIF-resistance caused by various rpoB gene mutations within 81-hp RRDR. Most of the RIF-resistant isolates were observed to harbor mutationsin Probe E 78.3% (n-36) whereas mutations in Probe A, B, D were observed 2.2% (n-1), 4.3% (n-2), and 6.5% (n-3), respectively. However, none of cases had RIF-resistance associated with Probe C. Out of 46 RRD cases, 21 (45.7 %) were males and 25 (54.3 %) were females. Additionally, Xpert (R) test showed higher detection rate than fluorescent microscopy (39.1% vs 31.2%, P<0.05) and detected MTB in 186 (11.8%) smear-negative specimens. Among 42 confirmed TB patients had MDR contact and eight patients were co-infected with HIV. In conclusion, 5.1% of the TB patients showed rifampicin resistance. The most frequent rpoB genetic mutations were observed in codons 531/533 (Probe F. 78.3%) whereas the least within the sequence 511 (Probe A, 2.2%).