embCAB sequence variation among ethambutol-resistant Mycobacterium tuberculosis isolates without embB306 mutation.
OBJECTIVE
Mechanisms of resistance to ethambutol in Mycobacterium tuberculosis remain inadequately described. Although there is mounting evidence that mutations of codon 306 in embB play a key role, a significant number of phenotypically ethambutol-resistant strains do not carry mutations in this codon. Here, other mutations in the embCAB operon are suggested to be involved in resistance development.


METHODS
The entire embCAB operon ( approximately 10 kb) was analysed in 34 phenotypically ethambutol-resistant M. tuberculosis strains without mutations in embB306 and in 12 ethambutol-susceptible strains. Furthermore, 106 control strains were investigated for the presence of particular mutations only.


RESULTS
Overall, 18 non-synonymous mutations in 15 distinct codons of the embCAB operon were identified in ethambutol-resistant strains but not in ethambutol-susceptible isolates. The majority occurred in the embB gene (10 distinct codons), in a 570 bp region also encompassing embB306. Mutations in embC and embA were found rarely and in most cases in combination with polymorphisms in embB. One synonymous mutation (embA 228 bp) and two non-synonymous mutations (embCVal981Leu and embCArg738Gln) were found in ethambutol-susceptible strains as well as resistant strains and were confirmed to represent phylogenetic markers for strains of the Beijing, Haarlem and Delhi/CAS genotypes, respectively.


CONCLUSIONS
Besides mutations in embB306, mutations in embB406 and embB497 were confirmed as hot spots for genomic variation in ethambutol-resistant clinical isolates. Of all resistant strains 70.6% carry a mutation in a relatively short region in embB, which therefore represents a promising target for inclusion in molecular assays for rapid detection of ethambutol resistance.
Plinke
Claudia
C
Molecular Mycobacteriology, Research Center Borstel, Borstel, Germany.
Cox
Helen S
HS
Zarkua
Nana
N
Karimovich
Hamraev A
HA
Braker
Kai
K
Diel
Roland
R
Rüsch-Gerdes
Sabine
S
Feuerriegel
Silke
S
Niemann
Stefan
S
D000995:Antitubercular Agents / Q000494:pharmacology*; D015373:Bacterial Typing Techniques; D016000:Cluster Analysis; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D024881:Drug Resistance, Bacterial; D004977:Ethambutol / Q000494:pharmacology*; D005838:Genotype; D006801:Humans; D008826:Microbial Sensitivity Tests; D020125:Mutation, Missense; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics; D009876:Operon; D010430:Pentosyltransferases / Q000235:genetics*; D017354:Point Mutation; D011110:Polymorphism, Genetic; D017422:Sequence Analysis, DNA
D000995:Antitubercular Agents; D004269:DNA, Bacterial; D004977:Ethambutol; C540269:EmbB protein, Mycobacterium tuberculosis; D010430:Pentosyltransferases