Detection of active pairs of XerC/D recognition sites mediating fusions and inversions in Acinetobacter baumannii plasmids carrying OXA-58 carbapenemase adaptive modules

CAMERANESI MM; MORAN BARRIO J; LIMANSKY AS; REPIZO G; VIALE AM

Frankfurt

Simposio; Acinetobacter 2019 - 12th International Symposium on the Biology of Acinetobacter; 2019

Institute of Molecular Biosciences Goethe-University Frankfurt

Acquired carbapenem-hydrolysing class-D β-lactamases (OXA-type) represent main factors of carbapenem resistance among multi-drug resistant (MDR) Acinetobacter baumannii (Aba) strains. Similarly to other OXA carbapenemases, OXA-58 genes (blaOXA-58) are embedded in plasmid-borne genetic structures which are flanked by a variable number of short 28-nucleotide motifs potentially recognized by the XerC/XerD site-specific recombination system (SSR) of the bacterial hosts (pXerC/D or pdif sites) [1]. It is poorly understood, however, how pXerC/D sites mediate mobilisation of the adaptive modules linked to them. We have recently demonstrated in Aba strains assigned to CC15 (Pasteur scheme) the existence of an active pair of pXerC/D sites promoting cointegrate formation between plasmids containing a blaOXA-58- and TnaphA6-resistance structure [1]. Here, we report the presence of additional pairs of pXerC/D sites mediating not only plasmid fusions and resolutions, but also the intra-molecular inversion of the adaptive module.The sequences of Ab825 plasmids were determined by 454 pyrosequencing and plasmid walking. A 28-nucleotide XerC/D consensus sequence was inferred from a local database of 215 Aba plasmids deposited in databases, and used to search potential sites in Ab825 plasmids. Cointegrate formation was detected by transformation of susceptible Acinetobacter strains, and the identification of sister pXerC/D pairs active in SSR by PCR and amplicon sequencing.Aba plasmids of less than 50 kbp were found to contain an average of 3-5 pXerC/D-like sites per molecule, larger plasmids were mostly devoid of these sites. Sequence analysis revealed the presence of three plasmids in Ab825: pAb825_27 (27 kbp), pAb825_12 (12 kbp), and pAb825_9 (9 kbp), each bearing several pXerC/D sites. A co-integrate (pAb825_36) was also detected resulting from the fusion of pAb825_27 and pAb825_9 mediated by an active pair of pXerC/D sites. Two additional active pairs were uncovered, one mediating the intra-molecular inversion of the blaOXA-58- and TnaphA6-containing structure and the other the fusion between pAb825_27 and pAb825_12. These observations shed light on the role of pXerC/D-mediated SSR in the evolutionary dynamics of Aba plasmids and the underlying mechanisms of dissemination of antimicrobial resistance and other adaptive structures among the Acinetobacter population.