FUNCTIONAL CHARACTERIZATION OF aphA-6 GENE FROM A LOCAL Acinetobacter bereziniae CLINICAL ISOLATE

MORALES-FLORES, AI; LIMANSKY AS; MARCHIARO P; MORAN BARRIO J

Chapadmalal

Congreso; XVIII Congreso Argentino de Microbiología General; 2023

SAMIGE

Acinetobacter is an aerobic Gram-negative genus ubiquitous in nature. Some species are opportunistic human pathogens, being A. baumannii (Ab) the most commonly isolated from clinical specimens. However, other species have been frequently associated with nosocomial infections including A. nosocomialis, A. lwoffii, A. pittii and less frequently, A. bereziniae1. The carbapenem resistant A. bereziniae local isolate HPC229 was previously characterized in our group1,2. This strain carries pNDM229 plasmid harboring blaNDM-1 and aphA6 resistance genes, responsible of the resistance to carbapenem and to aminoglycosides (AG), respectively. Regarding AG, HPC229 showed resistance towards amikacin (AKNR) but sensitivity towards gentamicin (GENS). Upstream of aphA6, in pHPC229, there is an ISAba14 insertion sequence, while blaNDM-1 is inserted within the Tn125 transposon located downstream aphA61. The ISAba14-aphA6-Tn125 structure is widely distributed in carbapenem resistant Acinetobacter strains, and is mostly associated to AKNS due to a lack of promoter upstream aphA63. However, it has been reported that the recombinant production of aphA6 from its native promoter resulted in 4-fold increases in MIC against AKN in A. baumannii and E. coli strains4.The aim of this work is to characterize the functionality of aphA6 gene in the A. bereziniae local strain HPC229, considering it´s AKNR and GENS, and to evaluate the mobilization of this gene from HPC229 to different Acinetobacter strains by horizontal gene transfer (HGT). For this purpose, sensitive strains such as A. nosocomialis M2 and Ab ATCC 17978 were transformed employing the total HPC229 purified plasmids (pHPC229), further selected for AKNR.These assays resulted in transformants M2/pHPC229 and Ab17978/pHPC229 strains displaying AKNR, with a reduction (15 mm) in their AKN inhibition zone by disc diffusion method, and aphA6 gene was detected in the first one. Thus indicating not only the mobilization of this resistance gene, but also its functionality. In addition, and notably, we also observed GENR phenotype in these transformants indicating a broader substrate spectrum of the corresponding protein, APH(3´), in the new hosts. Altogether, our results show for the first time that HPC229 AG resistance is susceptible to dissemination by HGT, indicating the plasmid localization of the corresponding gene(s). In addition, this gene in A. bereziniae HPC229 does not confer resistance to gentamicin, while it does in A. nosocomialis and Ab17978, suggesting a differential display of the AG resistance in different hosts. Further characterization of M2/pHPC229 and Ab17978/pHPC229 plasmids will allow us to underscore the AG resistance regulation in clinical Acinetobacter strains. 1) Brovedan M et al. (2015) doi: 10.1128/AAC.00367-152) Brovedan M et al. (2019) doi.org/10.1371/journal.pone.02205843) Hu H et al. (2012) doi: 10.1128/AAC.06199-114) Yoon E et al. (2014) doi:10.1128/mBio.01972-14