Evidence of diversity among epidemiologically related carbapenemase- producing Acinetobacter baumannii strains belonging to international clonal lineage II

Carbapenem-resistant Acinetobacter baumannii strains belonging to international clonal lineage II (ICL-II) have become predominant in intensive care units (ICUs) throughout Italy. Between 2005 and 2009, the carbapenem-hydrolyzing class D β-lactamase (CHDL) bla _OXA-23 gene became more prevalent than bla _OXA-58 among epidemic ICL-II strains showing extensive genetic similarity. These findings posed the question of whether CHDL gene replacement occurred in the homogeneous ICL-II population or a new OXA-23 clone(s) emerged and spread in ICUs. In this study, the changes in the ICL-II A. baumannii population and CHDL gene carriage were investigated in 30 genetically related isolates collected during the bla _OXA-58-to- bla _OXA-23 transition period. Pulsotyping, randomly amplified polymorphic DNA (RAPD) analysis, and multilocus sequence typing (MLST) results were combined with multilocus variable-number tandem-repeat (VNTR) analysis (MLVA-8), siderotyping, and plasmid profiling to improve genotype-based discrimination between isolates. Pulsotyping, RAPD analysis, and MLST clustered isolates into a single type. MLVA-8 identified 19 types that clustered into three complexes. All OXA-23-producing isolates formed a single complex, while OXA-58 producers were split into two complexes. Southern blot analysis of the physical localization and genetic context of the CHDL genes showed that bla _OXA-58 was invariably located on plasmids, while bla _OXA-23 was present within Tn2006 on the chromosome or both the chromosome and plasmids. These data indicate that the apparently homogeneous population of CHDL-producing ICL-II strains was composed of several independent strains and that, between 2005 and 2009, distinct OXA-23 producers displaced the preexisting OXA-58 producers. Thus, MLVA-8 appears to be a suitable tool not only for investigating A. baumannii population structure but also for high-resolution epidemiological typing.