title: Directed mutagenesis of Mycobacterium smegmatis 16S rRNA to reconstruct the in vivo evolution of aminoglycoside resistance in Mycobacterium tuberculosis.
abstract: Drug resistance in Mycobacterium tuberculosis is a global problem, with major consequences for treatment and public health systems. As the emergence and spread of drug-resistant tuberculosis epidemics is largely influenced by the impact of the resistance mechanism on bacterial fitness, we wished to investigate whether compensatory evolution occurs in drug-resistant clinical isolates of M. tuberculosis. By combining information from molecular epidemiology studies of drug-resistant clinical M. tuberculosis isolates with genetic reconstructions and measurements of aminoglycoside susceptibility and fitness in Mycobacterium smegmatis, we have reconstructed a plausible pathway for how aminoglycoside resistance develops in clinical isolates of M. tuberculosis. Thus, we show by reconstruction experiments that base changes in the highly conserved A-site of 16S rRNA that: (i) cause aminoglycoside resistance, (ii) confer a high fitness cost and (iii) destabilize a stem-loop structure, are associated with a particular compensatory point mutation that restores rRNA secondary structure and bacterial fitness, while maintaining to a large extent the drug-resistant phenotype. The same types of resistance and associated mutations can be found in M. tuberculosis in clinical isolates, suggesting that compensatory evolution contributes to the spread of drug-resistant tuberculosis disease.
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Dmitri
authors.initials: D
authors.lastname: Shcherbakov
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Rashid
authors.initials: R
authors.lastname: Akbergenov
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Tanja
authors.initials: T
authors.lastname: Matt
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Peter
authors.initials: P
authors.lastname: Sander
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Dan I
authors.initials: DI
authors.lastname: Andersson
authors.affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Zürich, Schweiz.Nationales Zentrum für Mykobakterien, Universität Zürich, Zürich, Schweiz.Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Erik C
authors.initials: EC
authors.lastname: Böttger
mesh_terms: D000617:Aminoglycosides / Q000494:pharmacology*; D000995:Antitubercular Agents / Q000494:pharmacology*; D004252:DNA Mutational Analysis; D024881:Drug Resistance, Bacterial; D019143:Evolution, Molecular; D006801:Humans; D016296:Mutagenesis; D020102:Mycobacterium smegmatis / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D012641:Selection, Genetic; D014376:Tuberculosis / Q000382:microbiology
chemical_list: D000617:Aminoglycosides; D000995:Antitubercular Agents; D012336:RNA, Ribosomal, 16S