Scarless removal of large resistance island AbaR results in antibiotic susceptibility and increased natural transformability in Acinetobacter baumannii

Abstract

With a great diversity in gene composition including multiple putative antibiotic-resistance genes, AbaR islands are potential contributors to multi-drug resistance in Acinetobacter baumannii. However, the effective contribution of AbaR to antibiotic resistance and bacterial physiology remains elusive. To address this, we exploited natural transformation to accurately remove AbaR islands and restore the integrity of their insertion site. We performed this genetic modification in two recent A. baumannii clinical strains: the strain AB5075 and the nosocomial strain AYE which carry AbaR11 and AbaR1 islands of 19.7 kbp and 86.2 kbp, respectively. Antibiotic susceptibilities were then compared between the parental strains and their AbaR-cured derivatives. As anticipated by the predicted function of the ORF of this island, the antibiotic resistance profiles were identical between the wild-type and the AbaR11cured strain. In contrast, AbaR1 carries 25 ORFs with a predicted resistance to several classes of antibiotics and the AbaR1-cured derivative showed restored susceptibility to multiple classes of antibiotics. Moreover, curing of AbaRs restored high levels of natural transformability. Indeed, most AbaR islands are inserted into the comM gene involved in natural transformation. Our data indicate that AbaR insertion effectively inactivates comM and that the restored comM is functional. Curing of AbaR consistently resulted in highly transformable, and therefore, easily genetically tractable strains.

Dissemination of novel Tn7 family transposons carrying genes for synthesis and uptake of fimsbactin siderophores among Acinetobacter baumannii isolates

“…Granted that Tn7 is known to transpose very efficiently into its preferred target site, the most surprising aspect of the analysis was the finding that the Tn6171 had been acquired both by targeted transposition and by replacement of the target region, presumably already carrying the transposon, with the corresponding sequence from another source. A. baumannii is known to exhibit high levels of competence [35][36][37] and extensive replacement of specific chromosomal segments in members of a specific clonal complex is increasingly reported. Multiple instances of introduction of an IS upstream of the ampC gene leading to cephalosporin resistance have been recorded [38][39][40].…”

Pilus Production in Acinetobacter baumannii Is Growth Phase Dependent and Essential for Natural Transformation

“…The lack of transformation defects in the absence of comM was in line with the minor impact observed for similar mutants in diverse V. cholerae strains (61,62), given that the current transformation assay was based on genomic DNA as transforming material that contained a resistance marker for transformant scoring. In contrast, there was a significant decrease of the transformability in comM mutants of V. cholerae and A. baumannii when PCR-amplified fragments served as transforming material, which led to the exchange of single nucleotides or short stretches of DNA (27,63). PilU, on the other hand, is a secondary retraction ATPase (PilT is the primary enzyme) that is present in many Gram-negative bacteria including P. aeruginosa in which it is essential for TFP-mediated twitching motility (64).…”

Interbacterial transfer of carbapenem resistance and large antibiotic resistance islands by natural transformation in pathogenic Acinetobacter

“…We thus tested whether this island could be acquired by the M2 strain. In a previous work, we showed that the AbaR1 island conferred resistance to aminoglycosides, tetracycline and to some beta-lactams (37). We thus used the tetracycline resistance as a marker for the transfer of AbaR1 from the A. baumannii strain AYE (donor strain) to the A. nosocomialis strain M2 (recipient strain).…”