Detection of Efflux Activity in Macrolide Resistant Streptococcus pyogenes Obtained from Patients with Throat Infections

Abstract

Background: Streptococcus pyogenes are bacteria which cause a wide variety of clinical manifestations including pharyngitis. Macrolides are alternative treatment options in cases of penicillin allergy or penicillin treatment failure. Due to a global rise in the emergence of macrolides resistance among these bacteria, the need for easy and rapid identification of isolates over-expressing efflux is paramount. This study detected the efflux-pump mediated resistance in macrolide resistant Streptococcus pyogenes using the ethidium-bromide (EtBr)–Agar cartwheel method

Methods: Overnight cultures of Streptococcus pyogenes isolates obtained from patients with throat infection were prepared. Agar plates containing EtBr of different concentrations were prepared. The minimum inhibitory concentration (MIC) of carbonyl cyanide m-chlorophenylhydrazine (CCCP) was determined.

Results: The isolates identified as isolates 190 (Reference strain), 22, 78, and 114 (iMLSB phenotypes) fluoresced at a concentration of 1.0 mg/l of EtBr; isolate 101 did not fluoresce even at a concentration of EtBr as high as 2.0 mg/l. The effect of ½ MIC of CCCP on the MIC of Erythromycin was tested and in the presence of CCCP at a concentration of 4μg/ml, the MIC of erythromycin was decreased by 2 to 4-fold.

Conclusion: The ethidium bromide-agar cartwheel method provided an easy and instrument-free means by which rapid evaluation of antibiotic resistance through the use of efflux pumps in bacterial isolates can be conducted.