Genetic Basis Of The Dissemination Of OXA-23and OXA-24/40-like Carbapenemases Among Acinetobacter Spp.

Acinetobacter has become an infectious agent of importance to hospitals worldwide, of which A. calcoaceticus-A. baumannii complex plays a dominant role. Acinetobacter spp. can cause pneumonia, skin and soft tissue infections, wound infections, urinary tract infections, meningitis and bloodstream infections. In recent years, carbapenem resistance among Acinetobacter spp. has been rising and it poses a great challenge to clinicians. Production of OXA-type carbapenemases is the most common reason for the carbapenem resistance in Acinetobacter spp. In this study, the genetic basis of dissemination of OXA-23and OXA-24/40-like carbapenemases in Acinetobacter spp. was studied.OXA-23is the most common one of OXA-type carbapenemases in Acinetobacter spp. Sixty A. baumannii and three A. nosocomialis isolates representing different genetic backgrounds were collected from28hospitals in18provinces for this study. Through PCR mapping the genetic surrounding of blaXOA-23gene was detected. S1/ApaI-PFGE-Southern blot was used to reveal the genetic location of the blaOXA-23gene. Meropenem and imipenem MICs were determined by the broth dilution method. The relationship between carbapenem susceptibilities and the genetic locations or copies of blaOXA-23gene was studied. In addition, transferability of the blaXOA-23gene was identified by electroporation and conjugation. Typical plasmid was extracted and sequenced through high-throughput sequencing.A total of35A. baumannii isolates had chromosome-carrying blaXOA-23gene while25A. baumannii and three A. nosocomialis isolates carried the blaOXA-23gene on plasmids, of which three A. baumannii isolates also had one copy of chromosome-encoded blaoxA-23gene. The MIC levels of carbapenems of these isolates were not significantly associated with the genomic locations or the copy numbers of blaOXA-23-In this study only two types of plasmids were detected:A ca.78kb plasmid, designated as pAZJ221, was found in twenty-three A. baumannii and three A. nosocomialis isolates while a novel ca.50kb plasmid carried by only two other A. baumannii isolates. Transformation of both plasmids succeeded but only pAZJ221was conjugative. Plasmid pAZJ221was sequenced completely and found to carry no other previously known resistance genes except blaXOA-23.Two main domains were identified in pAZJ221:Tn2009and conjugation elements. The blaXOA23genetic environments were correlated with Tn2009, Tn2008and Tn2006in57,5and one isolates, respectively.Overall, these observations suggest that chromosome-carrying blaXOA-23have a higher frequency than plasmid-encoded blaoxA-23and clonal spread of OXA-23producing bacteria should be concerned. Plasmid pAZJ221and Tn2009have effectively contributed to the wide dissemination of blaoxA-23in Acinetobacter spp. in China and horizontal gene transfer may play an important role in the dissemination of blaXOA-23gene.The study about the genetic mechanism of dissemination of OXA-24/40-like carbapenemases was conducted later. In our former study elven OXA-24/40-like carbapenemase-producing isolates were detected from2880Acinetobacter sp. isolates. Of the eleven isolates only one carried the blaXOA24/4o-like gene on chromosome and the others had plasmid-encoded blaOXA24/40-like genes. PFGE clustered all the eleven isolates into ten different clones and only two A. baumannii isolates were of the same clone. Through PCR walking, plasmid extraction, sequencing and analysis, it was found that blaOXA24/40-like genes were flanked by XerC/XerD-like binding sites. By comparison of the replicons, two A. baumannii and one A. pitti isolates carried the same type of plasmid while the plasmids extracted from the other six isolates were grouped into different types.Some of the plasmids could encode plasmid mobilization protein and VapBC type toxin/antitoxin system. Through sequence analysis of plasmid pA2503, plasmid pMCCU3and pMMD were found to share similar regions with pA2503. Both of pMCCU and pMMD were isolated from the same French hospital at the same period.So far blaOXA24/40-like harboring Acinetobacter spp. has not yet appeared pandemic in China. Horizontal spread mediated by plasmids plays a key role in the dissemination of blaOXA24/40-like gene while clonal spread is not common. Plasmid mobilization proteins and toxin/antitoxin system may facilitate the dissemination or enhance the stability of blaOXA24/40-like gene carrying plasmids. XerC/XerD homologous recombination system may be essential to the mobilization of blaXOA24/40-like gene.