Mycobacterium tuberculosis Whole Genome Sequences From Southern India Suggest Novel Resistance Mechanisms and the Need for Region-Specific Diagnostics

Manson, Abigail L.; Abeel, Thomas; Galagan, James E.; Sundaramurthi, Jagadish Chandrabose; Salazar, Alex N.; Gehrmann, Thies; Shanmugam, Siva Kumar; Palaniyandi, Kannan; Narayanan, Sujatha; Swaminathan, Soumya; Earl, Ashlee M.

doi:10.1093/cid/cix169

Cited by 65 publications

(66 citation statements)

References 45 publications

(59 reference statements)

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“…This scenario was already hypothesized by Torres and colleagues, who found Ͼ20 clinical katG mutations outside codon S315 by WGS and used site-directed mutagenesis to show that several could independently confer INH resistance to Mycobacterium smegmatis (34). There is evidence to suggest that the rise of noncanonical INH resistance mutations is already happening in India, where there is a high burden of drug-resistant TB (78). WGS or expanded targeted sequencing (conventional or NGS) could be used to detect such mutations, although it is important to consider the financial and infrastructural challenges at play in many high-burden regions of the world, any of which might limit the utility of shifting to NGS-based testing.…”

Section: Discussionmentioning

confidence: 72%

Validation of Novel Mycobacterium tuberculosis Isoniazid Resistance Mutations Not Detectable by Common Molecular Tests

Kandler

Mercante

Dalton

et al. 2018

Antimicrob Agents Chemother

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Resistance to the first-line antituberculosis (TB) drug isoniazid (INH) is widespread, and the mechanism of resistance is unknown in approximately 15% of INH-resistant (INH-R) strains. To improve molecular detection of INH-R TB, we used whole-genome sequencing (WGS) to analyze 52 phenotypically INH-R complex (MTBC) clinical isolates that lacked the common S315T or promoter mutations. Approximately 94% (49/52) of strains had mutations at known INH-associated loci that were likely to confer INH resistance. All such mutations would be detectable by sequencing more DNA adjacent to existing target regions. Use of WGS minimized the chances of missing infrequent INH resistance mutations outside commonly targeted hotspots. We used recombineering to generate 12 observed clinical mutations in the pansusceptible H37Rv reference strain and determined their impact on INH resistance. Our functional genetic experiments have confirmed the role of seven suspected INH resistance mutations and discovered five novel INH resistance mutations. All recombineered mutations conferred resistance to INH at a MIC of ≥0.25 μg/ml and should be added to the list of INH resistance determinants targeted by molecular diagnostic assays. We conclude that WGS is a useful tool for detecting uncommon INH resistance mutations that would otherwise be missed by current targeted molecular testing methods and suggest that its use (or use of expanded conventional or next-generation-based targeted sequencing) may provide earlier diagnosis of INH-R TB.

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Section: Discussionmentioning

confidence: 72%

Validation of Novel Mycobacterium tuberculosis Isoniazid Resistance Mutations Not Detectable by Common Molecular Tests

Kandler

Mercante

Dalton

et al. 2018

Antimicrob Agents Chemother

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“…22 Nearly 90% of INH resistance in India is caused by KatG mutations, associated with high-level resistance and poor treatment outcomes. 23 Development of INH resistance precedes the development of MDR-TB. 24 Initial INH resistance increases incidence rates of treatment failure and relapse compared with pan-sensitive strains (incidence rate ratio 10.9 and 1.8, respectively).…”

Section: Discussionmentioning

confidence: 99%

Progression from tuberculosis to multi drug resistance-TB in revised national tuberculosis control programme: perspectives from health system care givers

Kumar

Rana

Sundram

et al. 2019

Int J Community Med Public Health

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Background: The aims and objectives were to study the progression from tuberculosis to multi drug resistance-TB in revised national tuberculosis control programme: perspectives from health system care givers.Methods: The study was carried out in TB Sanatorium ITKI, Sadar Hospital Ranchi and RIMS Ranchi. The interview of various health personnel including SAHIYAs was taken using a semi-structured questionnaire based on programmatic management of multi drug resistant tuberculosis guidelines -2016.Results: Among Doctors knowledge level was good compared to other health personnel which had mean value 7.33 (±2.79), laboratory technician 3.45 (±2.64), STS 4.67 (±1.59), Sahiya 2.1 (±0.73). Regarding capacity enhancement level all health personnel needed refresher trainings in which doctors got 4.67 (±1.58), laboratory technician 3.45±2.64, STS 1.72±0.34, and Sahiya 0.5±0.52. Specially sahiya needs training regarding MDR-TB because they are the connecting link between health system and community. Regarding execution level, Doctors got 1.86 (±0.74), laboratory technician 1.64 (±0.56), STS 1.64 (±0.56) and Sahiya (ASHA) 2.2 (±0.44). Sahiya were better than other health personnel at execution level.Conclusions: Advocacy, communication, and social mobilization are important aspects of TB control, Policy makers and administrators should be sensitized for need of adequate and sustained funding for TB control to ensure quality capacity building. They need to provide continuous and quality training of staff at different levels and retention of trained staff and periodic reviews to identify gaps and take corrective steps.

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“…This scenario was already hypothesized by Torres and colleagues, who found >20 clinical katG mutations outside codon S315 by WGS of and used site-directed mutagenesis to show that several could independently confer INH resistance to Mycobacterium smegmatis (43). There is evidence to suggest that the rise of non-canonical INH resistance mutations is already happening in India, where there is a high burden of drug-resistant TB (65). WGS or expanded targeted sequencing could be used to detect such mutations.…”

Section: Discussionmentioning

confidence: 99%

Validation of novelMycobacterium tuberculosisisoniazid resistance mutations not detectable by common molecular tests

Kandler

Mercante

Dalton

et al. 2018

Preprint

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Resistance to the first-line anti-tuberculosis (TB) drug, isoniazid (INH), is widespread, and the mechanism of resistance is unknown in approximately 15% of INH-resistant (INH-R) strains. To improve molecular detection of INH-R TB, we used whole genome sequencing (WGS) to analyze 52 phenotypically INH-R Mycobacterium tuberculosis complex (MTBC) clinical isolates that lacked the common katG S315T or inhA promoter mutations. Approximately 94% (49/52) of strains had mutations at known INH-associated loci that were likely to confer INH resistance. All such mutations would be detectable by sequencing more DNA adjacent to existing target regions. Use of WGS minimized the chances of missing infrequent INH resistance mutations outside commonly targeted hotspots. We used recombineering to generate 12 observed clinical katG mutations in the pansusceptible H37Rv reference strain and determined their impact on INH resistance. Our functional genetic experiments have confirmed the role of seven suspected INH resistance mutations and discovered five novel INH resistance mutations. All recombineered katG mutations conferred resistance to INH at a minimum inhibitory concentration of ≥0.25 μg/mL and should be added to the list of INH resistance determinants targeted by molecular diagnostic assays. We conclude that WGS is a superior method for detection of INH-R MTBC compared to current targeted molecular testing methods and could provide earlier diagnosis of drug-resistant TB.

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Mycobacterium tuberculosis Whole Genome Sequences From Southern India Suggest Novel Resistance Mechanisms and the Need for Region-Specific Diagnostics

Abstract: Key pointsBy sequencing 223 M. tuberculosis strains from Southern India, we expanded the studied genetic diversity of lineages 1 and 3. We observed local transmission of strains; unexplained resistance; potential novel resistance mutations; and that isoniazid resistance was gained first.

Cited by 65 publications

References 45 publications

Validation of Novel Mycobacterium tuberculosis Isoniazid Resistance Mutations Not Detectable by Common Molecular Tests

Validation of Novel Mycobacterium tuberculosis Isoniazid Resistance Mutations Not Detectable by Common Molecular Tests

Progression from tuberculosis to multi drug resistance-TB in revised national tuberculosis control programme: perspectives from health system care givers

Validation of novelMycobacterium tuberculosisisoniazid resistance mutations not detectable by common molecular tests

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