Investigation of colistin-resistant non-human Acinetobacter seifertii isolates

The Acinetobacter calcoaceticus-baumannii (ACB) complex comprises multiple nosocomial infection-causing Acinetobacter species, the best-known being Acinetobacter baumannii. Once mainly considered opportunistic, Acinetobacter species are now being isolated from agricultural and food sources. We obtained and sequenced four Acinetobacter isolates from tank milk in Bogor, Indonesia. Using Average Nucleotide Identity and digital DNA:DNA hybridization, they were identified as Acinetobacter seifertii, a member of the ACB complex. Interestingly, all four isolates were resistant to colistin, a membrane-interacting antimicrobial polycationic peptide considered a drug of last resort. No known mechanism of colistin resistance was identified through analysis with the Comprehensive Antibiotic Resistance Database, and no known genetic substitutions that confer colistin resistance were found. Lipidomic analysis revealed lyso-phosphatidylethanolamine and an acyl-homoserine lactone quorum sensing molecule in significant abundance compared to colistin-sensitive A. baumannii ATCC 17978-VU. A membrane permeabilization assay with the four isolates showed muted interactions with colistin, and virulence-associated phenotypes did not vary greatly from the reference strain. Finally, the A. seifertii isolates were equally or less virulent than an A. seifertii reference strain in an insect infection model. Together, this suggests a potential unidentified mechanism of colistin resistance in A. seifertii or the discovery of tolerant isolates. Exploring non-human antibiotic-resistant bacterial species helps fill the gap in our understanding of the emergence of new pathogens.IMPORTANCEInvestigating drug-resistant bacterial species found outside of hospital settings is critical to understand and combat antibiotic resistance. Microbiological connections between humans, animals, and the environment can inform on how to use antimicrobial compounds appropriately, which in turn may curb the rise of bacterial antimicrobial resistance and failure of more antibiotics. Acinetobacter species are a group of soil- and water-dwelling microbes that are increasingly the cause of serious opportunistic infections, the best-known species being the multidrug-resistant pathogen A. baumannii. In this study, we discovered A. seifertii in tank milk in Indonesia that was resistant to a last-resort antibiotic, but the known genetic and cellular mechanisms of this resistance were not detected. Study and surveillance of such bacterial isolates will help us uncover and better understand bacterial antibiotic avoidance mechanisms and how pathogens emerge and move between different niches.