Pyrosequencing for rapid detection of Mycobacterium tuberculosis second-line drugs and ethambutol resistance

• Pyrosequencing is useful for detecting drug resistance in tuberculosis isolates.
• The method agrees accurately with the phenotypic result for FLQ, KAN, AMK, and CAP.
• Concordance between pyrosequencing and the phenotypic method for EMB is moderate.
• LPA is in agreement with pyrosequencing regarding gyrA, rrs, and embB.
• Pyrosequencing results can be useful to make initial decisions about TB treatment.

The aim of this work was to study the diagnostic accuracy of pyrosequencing to detect resistance to fluoroquinolones, kanamycin, amikacin, capreomycin, and ethambutol (EMB) in Mycobacterium tuberculosis clinical strains. One hundred four clinical isolates previously characterized by BACTEC 460TB/MGIT 960 were included. Specific mutations were targeted in gyrA, rrs, eis promoter, and embB. When there was a discordant result between BACTEC and pyrosequencing, Genotype MTBDRsl (Hain Lifescience, Nehren, Germany) was performed. Sensitivity and specificity of pyrosequencing were 70.6% and 100%, respectively, for fluoroquinolones; 93.3% and 81.7%, respectively, for kanamycin; 94.1% and 95.9%, respectively, for amikacin; 90.0% and 100%, respectively, for capreomycin; and 64.8% and 87.8%, respectively, for EMB. This study shows that pyrosequencing may be a useful tool for making early decisions regarding second-line drugs and EMB resistance. However, for a correct management of patients with suspected extensively drug-resistant tuberculosis, susceptibility results obtained by molecular methods should be confirmed by a phenotypic method.