Abstract
Carbapenem resistance presents severe threat to the treatment of multidrug resistant Pseudomonas aeruginosa infections. The study was undertaken to investigate the role of efflux pumps in conferring meropenem resistance and effect of single dose exposure of meropenem on transcription level of mexA gene in clinical isolates of P. aeruginosa from a tertiary referral hospital of India. Further, in this investigation an effort was made to assess whether different components of MexAB-OprM operon expresses in the same manner and the extent of contributions of those components in meropenem resistance in its natural host (P. aeruginosa) and in a heterologous host (E. coli). Out of 83 meropenem nonsusceptible isolates, 22 isolates were found to possess efflux pump activity phenotypically. Modified hodge test and multiplex PCR confirmed the absence of carbapenemase genes in those isolates. All of them were of multidrug resistant phenotype and were resistant to all the carbepenem drug tested. MexAB-OprM efflux pump was found to be overexpressed in all the study isolates. It could be observed that single dose exposure meropenem could give rise to trivial increase in transcription of mexA gene. Different constructs of MexAB-OprM (mexR-mexA-mexB-OprM; mexA-mexB-OprM; mexA-mexB) could be expressed in both its natural (P. aeruginosa PAO1) and heterologous host (E. coli JM107) but transcription level of mexA gene varied in both the hosts before and after single dose exposure of meropenem. Different components of the operon failed to enhance meropenem resistance in E. coli JM107 and P. aeruginosa PAO1. This study could prove that MexAB-OprM efflux pump can significantly contribute to meropenem resistance in hospital isolates of P. aeruginosa where an acquired resistant mechanism is absent. Thus, equal importance should be given for diagnosis of intrinsic resistance mechanism so as to minimize treatment failure. As meropenem could not enhance mexA transcriptions significantly, there might be a possibility that the increase in expression of efflux pump genes does not mediated by single antibiotic but rather by a combination of antipseudomonal drugs which are used during treatments. Early detection of efflux genes will help in selection of proper therapeutic options.
Highlights
Pseudomonas aeruginosa pose severe threat in treatment of nosocomial infections because of its intrinsic and acquired resistance towards a wide range of antimicrobial agents
Antimicrobial susceptibility testing and Minimum Inhibitory Concentration determination All the isolates with phenotypically predicted efflux pump activity were of multidrug resistant phenotype
As the test isolates with high Minimum inhibitory concentration (MIC) value were devoid of carbapenemase activity, so the expression of MexAB-OprM efflux pump was investigated in the isolates from each MIC
Summary
Pseudomonas aeruginosa pose severe threat in treatment of nosocomial infections because of its intrinsic and acquired resistance towards a wide range of antimicrobial agents. The mutation leads to loss of DNA binding capacity of the mexR repressor protein to the mexR-mexA intergenic region and thereby losing its ability to repress MexAB-OprM operon. This results in hyperexpression of MexAB-OprM in nalB mutants [4,5]. NalD and nalC type of mutants have been identified which occurs in response to mutation in nalC and nalD gene respectively Both type of the mutants lead to upregulation of MexAB-OprM [6]
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