Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades: Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal

Keira Cohen, Thomas Abeel, Abigail Manson McGuire, Christopher A. Desjardins, Vanisha Munsamy, Terrance P. Shea, Bruce J. Walker, Nonkqubela Bantubani, Deepak V. Almeida, Lucia Alvarado, Sinéad B. Chapman, Nomonde R. Mvelase, Eamon Y. Duffy, Michael G. Fitzgerald, Pamla Govender, Sharvari Gujja, Susanna Hamilton, Clinton Howarth, Jeffrey D. Larimer, Kashmeel MaharajMatthew D. Pearson, Margaret E. Priest, Qiandong Zeng, Nesri Padayatchi, Jacques Grosset, Sarah K. Young, Jennifer Wortman, Koleka P. Mlisana, Max R. O'Donnell, Bruce W. Birren, William Ramses Bishai, Alexander S. Pym, Ashlee M. Earl

Research output: Contribution to journalArticle

Abstract

Background: The continued advance of antibiotic resistance threatens the treatment and control of many infectious diseases. This is exemplified by the largest global outbreak of extensively drug-resistant (XDR) tuberculosis (TB) identified in Tugela Ferry, KwaZulu-Natal, South Africa, in 2005 that continues today. It is unclear whether the emergence of XDR-TB in KwaZulu-Natal was due to recent inadequacies in TB control in conjunction with HIV or other factors. Understanding the origins of drug resistance in this fatal outbreak of XDR will inform the control and prevention of drug-resistant TB in other settings. In this study, we used whole genome sequencing and dating analysis to determine if XDR-TB had emerged recently or had ancient antecedents. Methods and Findings: We performed whole genome sequencing and drug susceptibility testing on 337 clinical isolates of Mycobacterium tuberculosis collected in KwaZulu-Natal from 2008 to 2013, in addition to three historical isolates, collected from patients in the same province and including an isolate from the 2005 Tugela Ferry XDR outbreak, a multidrug-resistant (MDR) isolate from 1994, and a pansusceptible isolate from 1995. We utilized an array of whole genome comparative techniques to assess the relatedness among strains, to establish the order of acquisition of drug resistance mutations, including the timing of acquisitions leading to XDR-TB in the LAM4 spoligotype, and to calculate the number of independent evolutionary emergences of MDR and XDR. Our sequencing and analysis revealed a 50-member clone of XDR M. tuberculosis that was highly related to the Tugela Ferry XDR outbreak strain. We estimated that mutations conferring isoniazid and streptomycin resistance in this clone were acquired 50 y prior to the Tugela Ferry outbreak (katG S315T [isoniazid]; gidB 130 bp deletion [streptomycin]; 1957 [95% highest posterior density (HPD): 1937–1971]), with the subsequent emergence of MDR and XDR occurring 20 y (rpoB L452P [rifampicin]; pncA 1 bp insertion [pyrazinamide]; 1984 [95% HPD: 1974–1992]) and 10 y (rpoB D435G [rifampicin]; rrs 1400 [kanamycin]; gyrA A90V [ofloxacin]; 1995 [95% HPD: 1988–1999]) prior to the outbreak, respectively. We observed frequent de novo evolution of MDR and XDR, with 56 and nine independent evolutionary events, respectively. Isoniazid resistance evolved before rifampicin resistance 46 times, whereas rifampicin resistance evolved prior to isoniazid only twice. We identified additional putative compensatory mutations to rifampicin in this dataset. One major limitation of this study is that the conclusions with respect to ordering and timing of acquisition of mutations may not represent universal patterns of drug resistance emergence in other areas of the globe. Conclusions: In the first whole genome-based analysis of the emergence of drug resistance among clinical isolates of M. tuberculosis, we show that the ancestral precursor of the LAM4 XDR outbreak strain in Tugela Ferry gained mutations to first-line drugs at the beginning of the antibiotic era. Subsequent accumulation of stepwise resistance mutations, occurring over decades and prior to the explosion of HIV in this region, yielded MDR and XDR, permitting the emergence of compensatory mutations. Our results suggest that drug-resistant strains circulating today reflect not only vulnerabilities of current TB control efforts but also those that date back 50 y. In drug-resistant TB, isoniazid resistance was overwhelmingly the initial resistance mutation to be acquired, which would not be detected by current rapid molecular diagnostics employed in South Africa that assess only rifampicin resistance.

Original languageEnglish (US)
Article numbere1001880
JournalPLoS Medicine
Volume12
Issue number9
DOIs
StatePublished - Sep 1 2015

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Extensively Drug-Resistant Tuberculosis
Mycobacterium tuberculosis
Rifampin
Disease Outbreaks
Genome
Isoniazid
Mutation
Tuberculosis
Drug Resistance
Multidrug-Resistant Tuberculosis
Streptomycin
South Africa
Clone Cells
HIV
Pharmaceutical Preparations
Pyrazinamide
Kanamycin
Ofloxacin
Molecular Pathology
Explosions

ASJC Scopus subject areas

  • Medicine(all)

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Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades : Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal. / Cohen, Keira; Abeel, Thomas; Manson McGuire, Abigail; Desjardins, Christopher A.; Munsamy, Vanisha; Shea, Terrance P.; Walker, Bruce J.; Bantubani, Nonkqubela; Almeida, Deepak V.; Alvarado, Lucia; Chapman, Sinéad B.; Mvelase, Nomonde R.; Duffy, Eamon Y.; Fitzgerald, Michael G.; Govender, Pamla; Gujja, Sharvari; Hamilton, Susanna; Howarth, Clinton; Larimer, Jeffrey D.; Maharaj, Kashmeel; Pearson, Matthew D.; Priest, Margaret E.; Zeng, Qiandong; Padayatchi, Nesri; Grosset, Jacques; Young, Sarah K.; Wortman, Jennifer; Mlisana, Koleka P.; O'Donnell, Max R.; Birren, Bruce W.; Bishai, William Ramses; Pym, Alexander S.; Earl, Ashlee M.

In: PLoS Medicine, Vol. 12, No. 9, e1001880, 01.09.2015.

Research output: Contribution to journalArticle

Cohen, K, Abeel, T, Manson McGuire, A, Desjardins, CA, Munsamy, V, Shea, TP, Walker, BJ, Bantubani, N, Almeida, DV, Alvarado, L, Chapman, SB, Mvelase, NR, Duffy, EY, Fitzgerald, MG, Govender, P, Gujja, S, Hamilton, S, Howarth, C, Larimer, JD, Maharaj, K, Pearson, MD, Priest, ME, Zeng, Q, Padayatchi, N, Grosset, J, Young, SK, Wortman, J, Mlisana, KP, O'Donnell, MR, Birren, BW, Bishai, WR, Pym, AS & Earl, AM 2015, 'Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades: Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal', PLoS Medicine, vol. 12, no. 9, e1001880. https://doi.org/10.1371/journal.pmed.1001880
Cohen, Keira ; Abeel, Thomas ; Manson McGuire, Abigail ; Desjardins, Christopher A. ; Munsamy, Vanisha ; Shea, Terrance P. ; Walker, Bruce J. ; Bantubani, Nonkqubela ; Almeida, Deepak V. ; Alvarado, Lucia ; Chapman, Sinéad B. ; Mvelase, Nomonde R. ; Duffy, Eamon Y. ; Fitzgerald, Michael G. ; Govender, Pamla ; Gujja, Sharvari ; Hamilton, Susanna ; Howarth, Clinton ; Larimer, Jeffrey D. ; Maharaj, Kashmeel ; Pearson, Matthew D. ; Priest, Margaret E. ; Zeng, Qiandong ; Padayatchi, Nesri ; Grosset, Jacques ; Young, Sarah K. ; Wortman, Jennifer ; Mlisana, Koleka P. ; O'Donnell, Max R. ; Birren, Bruce W. ; Bishai, William Ramses ; Pym, Alexander S. ; Earl, Ashlee M. / Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades : Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal. In: PLoS Medicine. 2015 ; Vol. 12, No. 9.
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title = "Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades: Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal",
abstract = "Background: The continued advance of antibiotic resistance threatens the treatment and control of many infectious diseases. This is exemplified by the largest global outbreak of extensively drug-resistant (XDR) tuberculosis (TB) identified in Tugela Ferry, KwaZulu-Natal, South Africa, in 2005 that continues today. It is unclear whether the emergence of XDR-TB in KwaZulu-Natal was due to recent inadequacies in TB control in conjunction with HIV or other factors. Understanding the origins of drug resistance in this fatal outbreak of XDR will inform the control and prevention of drug-resistant TB in other settings. In this study, we used whole genome sequencing and dating analysis to determine if XDR-TB had emerged recently or had ancient antecedents. Methods and Findings: We performed whole genome sequencing and drug susceptibility testing on 337 clinical isolates of Mycobacterium tuberculosis collected in KwaZulu-Natal from 2008 to 2013, in addition to three historical isolates, collected from patients in the same province and including an isolate from the 2005 Tugela Ferry XDR outbreak, a multidrug-resistant (MDR) isolate from 1994, and a pansusceptible isolate from 1995. We utilized an array of whole genome comparative techniques to assess the relatedness among strains, to establish the order of acquisition of drug resistance mutations, including the timing of acquisitions leading to XDR-TB in the LAM4 spoligotype, and to calculate the number of independent evolutionary emergences of MDR and XDR. Our sequencing and analysis revealed a 50-member clone of XDR M. tuberculosis that was highly related to the Tugela Ferry XDR outbreak strain. We estimated that mutations conferring isoniazid and streptomycin resistance in this clone were acquired 50 y prior to the Tugela Ferry outbreak (katG S315T [isoniazid]; gidB 130 bp deletion [streptomycin]; 1957 [95{\%} highest posterior density (HPD): 1937–1971]), with the subsequent emergence of MDR and XDR occurring 20 y (rpoB L452P [rifampicin]; pncA 1 bp insertion [pyrazinamide]; 1984 [95{\%} HPD: 1974–1992]) and 10 y (rpoB D435G [rifampicin]; rrs 1400 [kanamycin]; gyrA A90V [ofloxacin]; 1995 [95{\%} HPD: 1988–1999]) prior to the outbreak, respectively. We observed frequent de novo evolution of MDR and XDR, with 56 and nine independent evolutionary events, respectively. Isoniazid resistance evolved before rifampicin resistance 46 times, whereas rifampicin resistance evolved prior to isoniazid only twice. We identified additional putative compensatory mutations to rifampicin in this dataset. One major limitation of this study is that the conclusions with respect to ordering and timing of acquisition of mutations may not represent universal patterns of drug resistance emergence in other areas of the globe. Conclusions: In the first whole genome-based analysis of the emergence of drug resistance among clinical isolates of M. tuberculosis, we show that the ancestral precursor of the LAM4 XDR outbreak strain in Tugela Ferry gained mutations to first-line drugs at the beginning of the antibiotic era. Subsequent accumulation of stepwise resistance mutations, occurring over decades and prior to the explosion of HIV in this region, yielded MDR and XDR, permitting the emergence of compensatory mutations. Our results suggest that drug-resistant strains circulating today reflect not only vulnerabilities of current TB control efforts but also those that date back 50 y. In drug-resistant TB, isoniazid resistance was overwhelmingly the initial resistance mutation to be acquired, which would not be detected by current rapid molecular diagnostics employed in South Africa that assess only rifampicin resistance.",
author = "Keira Cohen and Thomas Abeel and {Manson McGuire}, Abigail and Desjardins, {Christopher A.} and Vanisha Munsamy and Shea, {Terrance P.} and Walker, {Bruce J.} and Nonkqubela Bantubani and Almeida, {Deepak V.} and Lucia Alvarado and Chapman, {Sin{\'e}ad B.} and Mvelase, {Nomonde R.} and Duffy, {Eamon Y.} and Fitzgerald, {Michael G.} and Pamla Govender and Sharvari Gujja and Susanna Hamilton and Clinton Howarth and Larimer, {Jeffrey D.} and Kashmeel Maharaj and Pearson, {Matthew D.} and Priest, {Margaret E.} and Qiandong Zeng and Nesri Padayatchi and Jacques Grosset and Young, {Sarah K.} and Jennifer Wortman and Mlisana, {Koleka P.} and O'Donnell, {Max R.} and Birren, {Bruce W.} and Bishai, {William Ramses} and Pym, {Alexander S.} and Earl, {Ashlee M.}",
year = "2015",
month = "9",
day = "1",
doi = "10.1371/journal.pmed.1001880",
language = "English (US)",
volume = "12",
journal = "Nature Clinical Practice Oncology",
issn = "1759-4774",
publisher = "Nature Publishing Group",
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TY - JOUR

T1 - Evolution of Extensively Drug-Resistant Tuberculosis over Four Decades

T2 - Whole Genome Sequencing and Dating Analysis of Mycobacterium tuberculosis Isolates from KwaZulu-Natal

AU - Cohen, Keira

AU - Abeel, Thomas

AU - Manson McGuire, Abigail

AU - Desjardins, Christopher A.

AU - Munsamy, Vanisha

AU - Shea, Terrance P.

AU - Walker, Bruce J.

AU - Bantubani, Nonkqubela

AU - Almeida, Deepak V.

AU - Alvarado, Lucia

AU - Chapman, Sinéad B.

AU - Mvelase, Nomonde R.

AU - Duffy, Eamon Y.

AU - Fitzgerald, Michael G.

AU - Govender, Pamla

AU - Gujja, Sharvari

AU - Hamilton, Susanna

AU - Howarth, Clinton

AU - Larimer, Jeffrey D.

AU - Maharaj, Kashmeel

AU - Pearson, Matthew D.

AU - Priest, Margaret E.

AU - Zeng, Qiandong

AU - Padayatchi, Nesri

AU - Grosset, Jacques

AU - Young, Sarah K.

AU - Wortman, Jennifer

AU - Mlisana, Koleka P.

AU - O'Donnell, Max R.

AU - Birren, Bruce W.

AU - Bishai, William Ramses

AU - Pym, Alexander S.

AU - Earl, Ashlee M.

PY - 2015/9/1

Y1 - 2015/9/1

N2 - Background: The continued advance of antibiotic resistance threatens the treatment and control of many infectious diseases. This is exemplified by the largest global outbreak of extensively drug-resistant (XDR) tuberculosis (TB) identified in Tugela Ferry, KwaZulu-Natal, South Africa, in 2005 that continues today. It is unclear whether the emergence of XDR-TB in KwaZulu-Natal was due to recent inadequacies in TB control in conjunction with HIV or other factors. Understanding the origins of drug resistance in this fatal outbreak of XDR will inform the control and prevention of drug-resistant TB in other settings. In this study, we used whole genome sequencing and dating analysis to determine if XDR-TB had emerged recently or had ancient antecedents. Methods and Findings: We performed whole genome sequencing and drug susceptibility testing on 337 clinical isolates of Mycobacterium tuberculosis collected in KwaZulu-Natal from 2008 to 2013, in addition to three historical isolates, collected from patients in the same province and including an isolate from the 2005 Tugela Ferry XDR outbreak, a multidrug-resistant (MDR) isolate from 1994, and a pansusceptible isolate from 1995. We utilized an array of whole genome comparative techniques to assess the relatedness among strains, to establish the order of acquisition of drug resistance mutations, including the timing of acquisitions leading to XDR-TB in the LAM4 spoligotype, and to calculate the number of independent evolutionary emergences of MDR and XDR. Our sequencing and analysis revealed a 50-member clone of XDR M. tuberculosis that was highly related to the Tugela Ferry XDR outbreak strain. We estimated that mutations conferring isoniazid and streptomycin resistance in this clone were acquired 50 y prior to the Tugela Ferry outbreak (katG S315T [isoniazid]; gidB 130 bp deletion [streptomycin]; 1957 [95% highest posterior density (HPD): 1937–1971]), with the subsequent emergence of MDR and XDR occurring 20 y (rpoB L452P [rifampicin]; pncA 1 bp insertion [pyrazinamide]; 1984 [95% HPD: 1974–1992]) and 10 y (rpoB D435G [rifampicin]; rrs 1400 [kanamycin]; gyrA A90V [ofloxacin]; 1995 [95% HPD: 1988–1999]) prior to the outbreak, respectively. We observed frequent de novo evolution of MDR and XDR, with 56 and nine independent evolutionary events, respectively. Isoniazid resistance evolved before rifampicin resistance 46 times, whereas rifampicin resistance evolved prior to isoniazid only twice. We identified additional putative compensatory mutations to rifampicin in this dataset. One major limitation of this study is that the conclusions with respect to ordering and timing of acquisition of mutations may not represent universal patterns of drug resistance emergence in other areas of the globe. Conclusions: In the first whole genome-based analysis of the emergence of drug resistance among clinical isolates of M. tuberculosis, we show that the ancestral precursor of the LAM4 XDR outbreak strain in Tugela Ferry gained mutations to first-line drugs at the beginning of the antibiotic era. Subsequent accumulation of stepwise resistance mutations, occurring over decades and prior to the explosion of HIV in this region, yielded MDR and XDR, permitting the emergence of compensatory mutations. Our results suggest that drug-resistant strains circulating today reflect not only vulnerabilities of current TB control efforts but also those that date back 50 y. In drug-resistant TB, isoniazid resistance was overwhelmingly the initial resistance mutation to be acquired, which would not be detected by current rapid molecular diagnostics employed in South Africa that assess only rifampicin resistance.

AB - Background: The continued advance of antibiotic resistance threatens the treatment and control of many infectious diseases. This is exemplified by the largest global outbreak of extensively drug-resistant (XDR) tuberculosis (TB) identified in Tugela Ferry, KwaZulu-Natal, South Africa, in 2005 that continues today. It is unclear whether the emergence of XDR-TB in KwaZulu-Natal was due to recent inadequacies in TB control in conjunction with HIV or other factors. Understanding the origins of drug resistance in this fatal outbreak of XDR will inform the control and prevention of drug-resistant TB in other settings. In this study, we used whole genome sequencing and dating analysis to determine if XDR-TB had emerged recently or had ancient antecedents. Methods and Findings: We performed whole genome sequencing and drug susceptibility testing on 337 clinical isolates of Mycobacterium tuberculosis collected in KwaZulu-Natal from 2008 to 2013, in addition to three historical isolates, collected from patients in the same province and including an isolate from the 2005 Tugela Ferry XDR outbreak, a multidrug-resistant (MDR) isolate from 1994, and a pansusceptible isolate from 1995. We utilized an array of whole genome comparative techniques to assess the relatedness among strains, to establish the order of acquisition of drug resistance mutations, including the timing of acquisitions leading to XDR-TB in the LAM4 spoligotype, and to calculate the number of independent evolutionary emergences of MDR and XDR. Our sequencing and analysis revealed a 50-member clone of XDR M. tuberculosis that was highly related to the Tugela Ferry XDR outbreak strain. We estimated that mutations conferring isoniazid and streptomycin resistance in this clone were acquired 50 y prior to the Tugela Ferry outbreak (katG S315T [isoniazid]; gidB 130 bp deletion [streptomycin]; 1957 [95% highest posterior density (HPD): 1937–1971]), with the subsequent emergence of MDR and XDR occurring 20 y (rpoB L452P [rifampicin]; pncA 1 bp insertion [pyrazinamide]; 1984 [95% HPD: 1974–1992]) and 10 y (rpoB D435G [rifampicin]; rrs 1400 [kanamycin]; gyrA A90V [ofloxacin]; 1995 [95% HPD: 1988–1999]) prior to the outbreak, respectively. We observed frequent de novo evolution of MDR and XDR, with 56 and nine independent evolutionary events, respectively. Isoniazid resistance evolved before rifampicin resistance 46 times, whereas rifampicin resistance evolved prior to isoniazid only twice. We identified additional putative compensatory mutations to rifampicin in this dataset. One major limitation of this study is that the conclusions with respect to ordering and timing of acquisition of mutations may not represent universal patterns of drug resistance emergence in other areas of the globe. Conclusions: In the first whole genome-based analysis of the emergence of drug resistance among clinical isolates of M. tuberculosis, we show that the ancestral precursor of the LAM4 XDR outbreak strain in Tugela Ferry gained mutations to first-line drugs at the beginning of the antibiotic era. Subsequent accumulation of stepwise resistance mutations, occurring over decades and prior to the explosion of HIV in this region, yielded MDR and XDR, permitting the emergence of compensatory mutations. Our results suggest that drug-resistant strains circulating today reflect not only vulnerabilities of current TB control efforts but also those that date back 50 y. In drug-resistant TB, isoniazid resistance was overwhelmingly the initial resistance mutation to be acquired, which would not be detected by current rapid molecular diagnostics employed in South Africa that assess only rifampicin resistance.

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