Bedaquiline and clofazimine resistance in Mycobacterium tuberculosis: an in-vitro and in-silico data analysis
Creators: name: Sonnenkalb, Lindsay
name: Carter, Joshua James
name: Spitaleri, Andrea
name: Iqbal, Zamin
name: Hunt, Martin
name: Malone, Kerri Marie
name: Utpatel, Christian
name: Cirillo, Daniela Maria
name: Rodrigues, Camilla
name: Nilgiriwala, Kayzad Soli
name: Fowler, Philip William
name: Merker, Matthias
name: Niemann, Stefan
name: Consortium, The Comprehensive Resistance Prediction for Tuberculosis: an International
name: Barilar, I
name: Battaglia, S
name: Borroni, E
name: Brandao, AP
name: Brankin, A
name: Cabibbe, AM
name: Carter, J
name: Claxton, P
name: Clifton, DA
name: Cohen, T
name: Coronel, J
name: Crook, DW
name: Dreyer, V
name: Earle, SG
name: Escuyer, V
name: Ferrazoli, L
name: Fowler, PW
name: Gao, G Fu
name: Gardy, J
name: Gharbia, S
name: Ghisi, KT
name: Ghodousi, A
name: Cruz, AL Gibertoni
name: Grandjean, L
name: Grazian, C
name: Groenheit, R
name: Guthrie, JL
name: He, W
name: Hoffmann, H
name: Hoosdally, SJ
name: Ismail, NA
name: Jarrett, L
name: Joseph, L
name: Jou, R
name: Kambli, P
name: Khot, R
name: Knaggs, J
name: Koch, A
name: Kohlerschmidt, D
name: Kouchaki, S
name: Lachapelle, AS
name: Lalvani, A
name: Lapierre, S Grandjean
name: Laurenson, IF
name: Letcher, B
name: Lin, WH
name: Liu, C
name: Liu, D
name: Malone, KM
name: Mandal, A
name: Mansjö, M
name: Matias, D
name: Meintjes, G
name: de Freitas Mendes, F
name: Mihalic, M
name: Millard, J
name: Miotto, P
name: Mistry, N
name: Moore, D
name: Musser, KA
name: Ngcamu, D
name: Hoang, NN
name: Nimmo, C
name: Okozi, N
name: Oliveira, RS
name: Omar, SV
name: Paton, N
name: Peto, TE
name: Pinhata, JM Watanabe
name: Plesnik, S
name: Puyen, ZM
name: Rabodoarivelo, MS
name: Rakotosamimanana, N
name: Rancoita, PM
name: Rathod, P
name: Rodger, G
name: Rodwell, TC
name: Roohi, E
name: Santos-Lazaro, D
name: Shah, S
name: Kohl, TA
name: Smith, G
name: Solano, W
name: Supply, P
name: Surve, U
name: Tahseen, S
name: Thuong, NTT
Publisher: name: The Francis Crick Institute
Year: 2023
Subjects: subject: Model organisms
subject: Infectious Disease
subject: Human Biology &amp; Physiology
subject: Immunology
subject: FOS: Clinical medicine

description: Background: Bedaquiline is a core drug for the treatment of multidrug-resistant tuberculosis; however, the understanding of resistance mechanisms is poor, which is hampering rapid molecular diagnostics. Some bedaquiline-resistant mutants are also cross-resistant to clofazimine. To decipher bedaquiline and clofazimine resistance determinants, we combined experimental evolution, protein modelling, genome sequencing, and phenotypic data. Methods: For this in-vitro and in-silico data analysis, we used a novel in-vitro evolutionary model using subinhibitory drug concentrations to select bedaquiline-resistant and clofazimine-resistant mutants. We determined bedaquiline and clofazimine minimum inhibitory concentrations and did Illumina and PacBio sequencing to characterise selected mutants and establish a mutation catalogue. This catalogue also includes phenotypic and genotypic data of a global collection of more than 14 000 clinical Mycobacterium tuberculosis complex isolates, and publicly available data. We investigated variants implicated in bedaquiline resistance by protein modelling and dynamic simulations. Findings: We discerned 265 genomic variants implicated in bedaquiline resistance, with 250 (94%) variants affecting the transcriptional repressor (Rv0678) of the MmpS5–MmpL5 efflux system. We identified 40 new variants in vitro, and a new bedaquiline resistance mechanism caused by a large-scale genomic rearrangement. Additionally, we identified in vitro 15 (7%) of 208 mutations found in clinical bedaquiline-resistant isolates. From our in-vitro work, we detected 14 (16%) of 88 mutations so far identified as being associated with clofazimine resistance and also seen in clinically resistant strains, and catalogued 35 new mutations. Structural modelling of Rv0678 showed four major mechanisms of bedaquiline resistance: impaired DNA binding, reduction in protein stability, disruption of protein dimerisation, and alteration in affinity for its fatty acid ligand. Interpretation: Our findings advance the understanding of drug resistance mechanisms in M tuberculosis complex strains. We have established an extended mutation catalogue, comprising variants implicated in resistance and susceptibility to bedaquiline and clofazimine. Our data emphasise that genotypic testing can delineate clinical isolates with borderline phenotypes, which is essential for the design of effective treatments. Funding: Leibniz ScienceCampus Evolutionary Medicine of the Lung, Deutsche Forschungsgemeinschaft, Research Training Group 2501 TransEvo, Rhodes Trust, Stanford University Medical Scientist Training Program, National Institute for Health and Care Research Oxford Biomedical Research Centre, Oxford University Hospitals NHS Foundation Trust, Bill &amp; Melinda Gates Foundation, Wellcome Trust, and Marie Skłodowska-Curie Actions.