Disc Diffusion Testing of Azithromycin Against Clinical Isolates of Typhoidal Salmonellae: A Diagnostic Conundrum

Introduction: Typhoid remains a major healthcare problem in low and middle-income countries. The emergence of extremely drug-resistant (XDR) typhoid strains from the Indian subcontinent has led to very limited therapeutic options. Azithromycin being the only oral option for XDR typhoid faces a threat of rapid resistance due to its overuse after the COVID-19 pandemic. Objective: To evaluate the reliability of azithromycin disc diffusion testing against clinical isolates of typhoidal salmonellae in comparison with E-test minimum inhibitory concentrations (MICs). Study design: This is a cross-sectional validation study. Place and duration of the study: The Department of Microbiology, Pakistan Navy Ship Shifa hospital, Karachi from June 1 to December 31, 2020. Methodology: Antimicrobial susceptibility was performed by Kirby Bauer disc diffusion method for 60 isolates including Salmonella enterica ser. Typhi and Paratyphi A using Clinical Laboratory Standard Institute (CLSI) guidelines. MICs by the E-test method were determined for Azithromycin only. Results: A significant proportion of the isolates (55%) had high azithromycin MIC in the wild-type distribution range (8-16 µg/ml). Ten (16.6%) isolates showed false resistance, i.e., zone diameter <13 mm by disc diffusion method when compared to E-test MIC results. Isolates with MICs close to breakpoint, i.e., 16 µg/ml were more likely to show discordant results. The sensitivity, specificity, negative predictive value, positive predictive value, and diagnostic accuracy of the disc diffusion method versus E-test were 100%, 83%, 100%, 9%, and 83%, respectively. Conclusions: Disc diffusion method as recommended by CLSI is not reliable for azithromycin susceptibility testing particularly for isolates with high MICs in the susceptible range. The E-test method may be a better alternative to disc diffusion provided appropriate training is done prior to its application.