Rv1458c: a new diagnostic marker for identification of Mycobacterium tuberculosis complex in a novel duplex PCR assay

Shrivastava, Kamal; Garima, Kushal; Narang, Asmita; Bhattacharyya, Kausik; Vishnoi, Ekta; Singh, Roshan Kumar; Chaudhry, Anil; Prasad, Rajendra; Bose, Mridula; Varma-Basil, Mandira

doi:10.1099/jmm.0.000440

Cited by 11 publications

(6 citation statements)

References 26 publications

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“…The identification of genes that do not show any SNV in a large number of global clinical isolates (invariant regions), would be potential diagnostic markers for identification of M. tuberculosis complex [ 30 – 32 ]. We found several invariant genes in 1553 global clinical isolate including VPCI591.…”

Section: Discussionmentioning

confidence: 99%

“…We found several invariant genes in 1553 global clinical isolate including VPCI591. It has been shown that the invariant region in 190 bp region of Rv1458c and partial regions of Rv0440 can be used as diagnostic markers for the identification of M. tuberculosis complex (MTBC) [ 30 ]. The major group of invariant genes in our study includes, toxin antitoxin pathway genes like Rv0661c , Rv2760c , Rv0664 , and Rv0581 , and transposase of IS elements (Insertion elements) like IS6110 .…”

Section: Discussionmentioning

confidence: 99%

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Correlation of drug resistance with single nucleotide variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate of M. tuberculosis

et al. 2020

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Background Genome sequencing and genetic polymorphism analysis of clinical isolates of M. tuberculosis is carried out to gain further insight into molecular pathogenesis and host-pathogen interaction. Therefore the functional evaluation of the effect of single nucleotide variation (SNV) is essential. At the same time, the identification of invariant sequences unique to M. tuberculosis contributes to infection detection by sensitive methods. In the present study, genome analysis is accompanied by evaluation of the functional implication of the SNVs in a MDR clinical isolate VPCI591. Result By sequencing and comparative analysis of VPCI591 genome with 1553 global clinical isolates of M. tuberculosis (GMTV and tbVar databases), we identified 141 unique strain specific SNVs. A novel intergenic variation in VPCI591 in the putative promoter/regulatory region mapping between embC ( Rv3793 ) and embA ( Rv3794 ) genes was found to enhance the expression of embAB , which correlates with the high resistance of the VPCI591 to ethambutol. Similarly, the unique combination of three genic SNVs in RNA polymerase β gene ( rpoB ) in VPCI591 was evaluated for its effect on rifampicin resistance through molecular docking analysis. The comparative genomics also showed that along with variations, there are genes that remain invariant. 173 such genes were identified in our analysis. Conclusion The genetic variation in M. tuberculosis clinical isolate VPCI591 is found in almost all functional classes of genes. We have shown that SNV in rpoB gene mapping outside the drug binding site along with two SNVs in the binding site can contribute to quantitative change in MIC for rifampicin. Our results show the collective effect of SNVs on the structure of the protein, impacting the interaction between the target protein and the drug molecule in rpoB as an example. The study shows that intergenic variations bring about quantitative changes in transcription in embAB and in turn can lead to drug resistance.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Correlation of drug resistance with single nucleotide variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate of M. tuberculosis

et al. 2020

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show abstract

“…The identification of genes that do not show any SNV in a large number of global clinical isolates (invariant regions), would be potential diagnostic markers for identification of M.tuberculosis complex (51-53). We found several invariant genes in 1553 global clinical isolate including VPCI591.…”

Section: Discussionmentioning

confidence: 99%

“…We found several invariant genes in 1553 global clinical isolate including VPCI591. It has been shown that the invariant region in 190bp region of Rv1458c and partial regions of Rv0440 can be used as diagnostic markers for identification of M.tuberculosis complex (MTBC)(51). The major group of invariant genes in our study includes, toxin antitoxin pathway genes like Rv0661c, Rv2760c, Rv0664 , and Rv0581 , and transposase of IS elements (Insertion elements) like IS6110 .…”

Section: Discussionmentioning

confidence: 99%

Correlation of drug resistance with Single Nucleotide Variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate ofM.tuberculosis

Bhattacharyya

Nemaysh²,

Joon

et al. 2020

Preprint

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BackgroundGenome sequencing and genetic polymorphism analysis of clinical isolates of M.tuberculosis is carried out to gain further insight into molecular pathogenesis and host-pathogen interaction. Therefore the functional evaluation of the effect of single nucleotide variation (SNV) is essential. At the same time, the identification of invariant sequences unique to M.tuberculosis contributes to infection detection by sensitive methods. In the present study, genome analysis is accompanied by evaluation of the functional implication of the SNVs in a MDR clinical isolate VPCI591.ResultBy sequencing and comparative analysis of VPCI591 genome with 1553 global clinical isolates of M.tuberculosis, we identified 143 unique strain specific SNVs. A novel intergenic variation in VPCI591 in the putative promoter/regulatory region mapping between embC (Rv3793) and embA (Rv3794) genes was found to enhance the expression of embAB, which correlates with the high resistance of the VPCI591 to ethambutol. Similarly, the unique combination of three genic SNVs in RNA polymerase β gene (rpoB) in VPCI591 was evaluated for its effect on rifampicin resistance through molecular docking analysis. The comparative genomics also showed that along with variations, there are genes that remain invariant. 173 such genes were identified in our analysis.ConclusionWe have demonstrated that SNVs are not always benign and can also have functional effect. We show that variations bring about quantitative changes in transcription. Our results show the collective effect of SNVs on the structure of protein, impacting the interaction between the target protein and the drug molecule, with rpoB as an example.

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“…Moreover, sensitivity of LAM assay differs significantly between varying health settings. [3][4][5][6] In sub-Saharan Africa, 32% of the global TB burden is found in patients co-infected with HIV/AIDS, whereas in South-East Asia this figure…”

Section: Introductionmentioning

confidence: 99%

The MPB64 immunochromatography assay: an analysis of doubtful results

et al. 2020

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Culture remains the gold standard for tuberculosis (TB) diagnosis, and the mycobacteria growth indicator tube (MGIT), endorsed by the World Health Organization (WHO), is widely used. Further identification of a positive culture is done with the help of an immunochromatography assay, which often shows faint bands that are difficult to interpret. We analysed 125 BACTEC MGIT culture positive results, of which 11/16 (68.7%) of the doubtful assays, analysed by MGIT™ TBc Identification test (TBcId), were positive for Mycobacterium tuberculosis complex (MTBC), the remaining being non-tuberculous mycobacteria as determined by an in-house duplex polymerase chain reaction and line probe assay. Guidelines on faint or doubtful bands in immunochromatography assays are important so as not to overlook true-positive cases of TB.

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Rv1458c: a new diagnostic marker for identification of Mycobacterium tuberculosis complex in a novel duplex PCR assay

Abstract: The duplex PCR assay could be developed for use as a screening test to identify MTBC in clinical specimens in peripheral laboratories with limited resources.

Cited by 11 publications

References 26 publications

Correlation of drug resistance with single nucleotide variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate of M. tuberculosis

Correlation of drug resistance with single nucleotide variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate of M. tuberculosis

Correlation of drug resistance with Single Nucleotide Variations through genome analysis and experimental validation in a multi-drug resistant clinical isolate ofM.tuberculosis

The MPB64 immunochromatography assay: an analysis of doubtful results

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