Genetic and phenotypic characterization of drug-resistant Mycobacterium tuberculosis isolates in Hong Kong.
OBJECTIVE
To characterize 250 drug-resistant Mycobacterium tuberculosis (MTB) isolates in Hong Kong with respect to their drug susceptibility phenotypes to five common anti-tuberculosis drugs (ofloxacin, rifampicin, ethambutol, isoniazid and pyrazinamide) and the relationship between such phenotypes and the patterns of genetic mutations in the corresponding resistance genes (gyrA, rpoB, embB, katG, inhA, ahpC and pncA).


METHODS
The MIC values of the aforementioned anti-tuberculosis drugs were determined for each of the 250 drug-resistant MTB clinical isolates by the absolute concentration method. Genetic mutations in the corresponding resistance genes in these MTB isolates were identified by PCR-single-stranded conformation polymorphism/multiplex PCR amplimer conformation analysis (SSCP/MPAC), followed by DNA sequencing of the purified PCR products.


RESULTS
Resistance to four or five drugs was commonly observed in these MTB isolates; such phenotypes accounted for over 34% of the 250 isolates. The most frequently observed phenotypes were those involving both rifampicin and isoniazid, with or without additional resistance to the other drugs. A total of 102 novel mutations, which accounted for 80% of all mutation types detected in the 7 resistance genes, were recovered. Correlation between phenotypic and mutational data showed that genetic changes in the gyrA, rpoB and katG genes were more consistently associated with a significant resistance phenotype. Despite this, however, a considerable proportion of resistant MTB isolates were found to harbour no detectable mutations in the corresponding gene loci.


CONCLUSIONS
These findings expand the spectrum of potential resistance-related mutations in MTB clinical isolates and help consolidate the framework for the development of molecular methods for delineating the drug susceptibility profiles of MTB isolates in clinical laboratories.
Chan
Raphael C Y
RC
Department of Microbiology, The Chinese University of Hong Kong, The Prince of Wales Hospital, Shatin, New Territories, Hong Kong, China. chiuychan@cuhk.edu.hk
Hui
Mamie
M
Chan
Edward W C
EW
Au
T K
TK
Chin
Miu L
ML
Yip
Chun K
CK
AuYeang
Carrie K W
CK
Yeung
Christina Y L
CY
Kam
Kai M
KM
Yip
Peter C W
PC
Cheng
Augustine F B
AF
D000995:Antitubercular Agents / Q000494:pharmacology*; D005798:Genes, Bacterial; D006723:Hong Kong; D008826:Microbial Sensitivity Tests; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D010641:Phenotype; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D014376:Tuberculosis / Q000382:microbiology*
D000995:Antitubercular Agents