A Rapid Antimicrobial Susceptibility Test for Klebsiella pneumoniae Using a Broth Micro-Dilution Combined with MALDI TOF MS.

Abstract

BackgroundMatrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) is a novel method that can be used to identify pathogens and has potential applications in the detection of drug-resistant bacteria.PurposeTo evaluate the ability of a MALDI-TOF MS-based broth micro-dilution method in detecting the minimum inhibitory concentration (MIC) values of Klebsiella pneumoniae to ceftriaxone and imipenem.Materials and MethodsSixty strains of K. pneumoniae with different levels of resistance to carbapenems and cephalosporins were randomly collected. The 0.5 McFarland (Mc) concentration of the bacterial suspension was inoculated in cation-adjusted Mueller-Hinton broth (CAMHB) with a final cell turbidity of 5×105 CFU/mL. The broth was incubated with serial concentrations of antibiotics. After centrifuging the bacterial suspensions, the lysed cells were analyzed by MALDI-TOF MS to identify the growth-promoting or inhibitory effects on K. pneumoniae. The molecular mechanisms of resistance were investigated by PCR and DNA sequencing analysis.ResultsThe expression of known resistance genes (blaKPC, blaFOX, blaDHA, blaCTX-M and blaTEM) was detected in the 30 carbapenems-resistant strains. The agreement between the MIC values derived from the MALDI-TOF MS analysis and from the broth micro-dilution method was 61.7% for ceftriaxone and 71.7% for imipenem. According to the Clinical and Laboratory Standards Institute (CLSI) breakpoint of resistance to ceftriaxone and imipenem, the 60 isolates were accurately classified as resistant or susceptible isolates with 100% sensitivity and 100% specificity.ConclusionThe transmission and infection of multidrug-resistant bacteria could be better managed and treated with the rapid identification of strains and antimicrobial susceptibility. A MALDI-TOF MS-based susceptibility test could be used to identify resistance of K. pneumoniae within a short time-frame. This approach could potentially be used as a supplementary antimicrobial susceptibility test that could be investigated on more bacterial species combined with different antibiotics.