Investigation of colistin resistance in carbapenem-resistant Klebsiella pneumoniae isolates by flow cytometry method

Abstract

Abstract Rapid detection of antimicrobial resistance is crucial for early initiation of appropriate therapy. The aim of this study was to investigate whether resistance to colistin, the last resort antibiotic, in carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates can be detected accurately and rapidly by flow cytometry (FCM) and whether this method is convenient for routine use in clinical laboratories. The VITEK 2 automated system was used to identify K. pneumoniae strains and to determine their resistance to carbapenems and other antibiotics except for colistin. The minimum inhibitory concentration (MIC) values for colistin in 85 CRKP strains were determined by the broth microdilution (BMD), which is the reference method. In addition, FCM was used, combined with DiBAC₄(3) “bis-(1,3-dibutylbarbituric acid) trimethine oxonol” fluorescent stain, which can detect the changes in the bacterial membrane potential caused by antibiotic treatment, to determine colistin susceptibility. The MIC₅₀ value of the strains, 80% of which were resistant to colistin by the BMD method, was 16 mg/L, and the MIC₉₀ value was 32 mg/L. When FCM was compared with the reference method, it was determined that the categorical agreement between both methods was 98.8%; the specificity, sensitivity, and positive and negative predictive values of FCM were 94%, 100%, 98.5%, and 100%, respectively. Colistin susceptibility results with FCM were obtained within an average of 2 hours. In conclusion, these findings suggest that FCM holds great promise as a rapid and reliable alternative method for detecting colistin resistance in CRKP strains.