Effects of prophage regions in a plasmid carrying a carbapenemase gene on survival against antibiotic stress.

In this study, we investigated the evolutionary importance of cryptic prophage elements in a blaNDM-1 -bearing plasmid by exploring the effect of prophage regions on survival against antibiotic stress. While analyzing a plasmid harbouring an NDM-1-encoding gene in Klebsiella pneumoniae from South Korea, we found a prophage region within the plasmid. We constructed single-prophage knock-out mutants by gene replacement. The intact plasmid and plasmids with deleted prophages were conjugated into Escherichia coli DH5α. Growth rate and antibiotic susceptibility were determined, and survival rates of strains were evaluated in the presence of antibiotics such as imipenem, amikacin, gentamicin, cefotaxime, and piperacillin/tazobactam. A transcriptional response of sigma factor-coding genes (rpoS and rpoE) and reactive oxygen species (ROS)-related genes from different operons (soxS, fumC, oxyR, and katE) to a subinhibitory concentration of aminoglycodies were monitored by method of quantitative real-time PCR. The prophage region consists of four cryptic prophages 16,795 bp in size and 19 coding DNA sequences. An Escherichia coli transconjugant carrying the plasmid with intact prophages showed increased survival during treatment with various antibiotics including imipenem and amikacin; however, transconjugants carrying this plasmid with a single-prophage knockout did not. mRNA expression analyses revealed that sigma factor proteins (rpoB and rpoE) were highly upregulated by antibiotics. We propose that cryptic prophages in the antibiotic resistance plasmid may contribute to adaptation of the bacterial host to antibiotic stress. We are concerned that the combination of prophages with a drug resistance plasmid helps drug-resistant bacteria in a hostile environment and accelerates their dissemination.