Whole-genome sequencing to establish relapse or re-infection with Mycobacterium tuberculosis: a retrospective observational study.
BACKGROUND
Recurrence of tuberculosis after treatment makes management difficult and is a key factor for determining treatment efficacy. Two processes can cause recurrence: relapse of the primary infection or re-infection with an exogenous strain. Although re-infection can and does occur, its importance to tuberculosis epidemiology and its biological basis is still debated. We used whole-genome sequencing-which is more accurate than conventional typing used to date-to assess the frequency of recurrence and to gain insight into the biological basis of re-infection.


METHODS
We assessed patients from the REMoxTB trial-a randomised controlled trial of tuberculosis treatment that enrolled previously untreated participants with Mycobacterium tuberculosis infection from Malaysia, South Africa, and Thailand. We did whole-genome sequencing and mycobacterial interspersed repetitive unit-variable number of tandem repeat (MIRU-VNTR) typing of pairs of isolates taken by sputum sampling: one from before treatment and another from either the end of failed treatment at 17 weeks or later or from a recurrent infection. We compared the number and location of SNPs between isolates collected at baseline and recurrence.


RESULTS
We assessed 47 pairs of isolates. Whole-genome sequencing identified 33 cases with little genetic distance (0-6 SNPs) between strains, deemed relapses, and three cases for which the genetic distance ranged from 1306 to 1419 SNPs, deemed re-infections. Six cases of relapse and six cases of mixed infection were classified differently by whole-genome sequencing and MIRU-VNTR. We detected five single positive isolates (positive culture followed by at least two negative cultures) without clinical evidence of disease.


CONCLUSIONS
Whole-genome sequencing enables the differentiation of relapse and re-infection cases with greater resolution than do genotyping methods used at present, such as MIRU-VNTR, and provides insights into the biology of recurrence. The additional clarity provided by whole-genome sequencing might have a role in defining endpoints for clinical trials.


BACKGROUND
Wellcome Trust, European Union, Medical Research Council, Global Alliance for TB Drug Development, European and Developing Country Clinical Trials Partnership.
Wellcome Trust Sanger Institute, Hinxton, UK.
Josephine M
JM
Bryant
Wellcome Trust Sanger Institute, Hinxton, UK.
Simon R
SR
Harris
Wellcome Trust Sanger Institute, Hinxton, UK.
Julian
J
Parkhill
Division of Pulmonology, University of Cape Town, Cape Town, South Africa.
Rodney
R
Dawson
DST/NRF Centre of Excellence for Biomedical Tuberculosis Research, MRC Centre for Molecular and Cellular Biology, Division of Molecular Biology and Human Genetics, Stellenbosch University, Tygerberg, South Africa.
Andreas H
AH
Diacon
DST/NRF Centre of Excellence for Biomedical Tuberculosis Research, MRC Centre for Molecular and Cellular Biology, Division of Molecular Biology and Human Genetics, Stellenbosch University, Tygerberg, South Africa.
Paul
P
van Helden
South African Medical Research Council and KwaZulu Research Institute for TB and HIV, Durban, South Africa.
Alex
A
Pym
Institute of Respiratory Medicine, Kuala Lumpur, Malaysia.
Aziah A
AA
Mahayiddin
Chest Disease Institute, Muang, Nothaburi, Thailand.
Charoen
C
Chuchottaworn
Clinical HIV Research Unit, Helen Joseph Hospital, Westdene, Johannesburg, South Africa.
Ian M
IM
Sanne
Madibeng Centre for Research, Brits, South Africa.
Cheryl
C
Louw
Radboud MD University Nijmegen Medical Centre/UCCZ Dekkerswald, Nijmegen, Netherlands.
Martin J
MJ
Boeree
Department of Infectious Diseases and Tropical Medicine, Klinikum, Ludwig-Maximilians-University, Munich, Germany; DZIF German Centre for Infection Research, Munich, Germany.
Michael
M
Hoelscher
Centre for Clinical Microbiology, Royal Free Campus, University College London, London, UK.
Timothy D
TD
McHugh
Centre for Clinical Microbiology, Royal Free Campus, University College London, London, UK.
Anna L C
AL
Bateson
Centre for Clinical Microbiology, Royal Free Campus, University College London, London, UK.
Robert D
RD
Hunt
Centre for Clinical Microbiology, Royal Free Campus, University College London, London, UK.
Solomon
S
Mwaigwisya
Centre for Clinical Microbiology, Royal Free Campus, University College London, London, UK.
Laura
L
Wright
School of Medicine, University of St Andrews, St Andrews, UK. Electronic address: shg3@st-andrews.ac.uk.
Stephen H
SH
Gillespie
Wellcome Trust Sanger Institute, Hinxton, UK.
Stephen D
SD
Bentley
D006801:Humans; D000995:Antitubercular Agents / Q000627:therapeutic use; D004269:DNA, Bacterial / Q000235:genetics; D005500:Follow-Up Studies; D055106:Genome-Wide Association Study / Q000379:methods; D005838:Genotype; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000302:isolation & purification; D012008:Recurrence; D012189:Retrospective Studies; D014376:Tuberculosis / Q000188:drug therapy / Q000235:genetics / Q000382:microbiology
D000995:Antitubercular Agents; D004269:DNA, Bacterial