Abstract
Members of the genus Acinetobacter have been the focus recent attention due to both their clinical significance and application to molecular biology. The soil commensal bacterium Acinetobacter baylyi ADP1 has been proposed as a model system for molecular and genetic studies, whereas in a clinical environment, Acinetobacter spp. are of increasing importance due to their propensity to cause serious and intractable systemic infections. Clinically, a major factor in the success of Acinetobacter spp. as opportunistic pathogens can be attributed to their ability to rapidly evolve resistance to common antimicrobial compounds. Whole genome sequencing of clinical and environmental Acinetobacter spp. isolates has revealed the presence of numerous multidrug transporters within the core and accessory genomes, suggesting that efflux is an important host defense response in this genus. In this work, we used the drug-susceptible organism A. baylyi ADP1 as a model for studies into the evolution of efflux mediated resistance in genus Acinetobacter, due to the high level of conservation of efflux determinants across four diverse Acinetobacter strains, including clinical isolates. A single exposure of therapeutic concentrations of chloramphenicol to populations of A. baylyi ADP1 cells produced five individual colonies displaying multidrug resistance. The major facilitator superfamily pump craA was upregulated in one mutant strain, whereas the resistance nodulation division pump adeJ was upregulated in the remaining four. Within the adeJ upregulated population, two different levels of adeJ mRNA transcription were observed, suggesting at least three separate mutations were selected after single-step exposure to chloramphenicol. In the craA upregulated strain, a T to G substitution 12 nt upstream of the craA translation initiation codon was observed. Subsequent mRNA stability analyses using this strain revealed that the half-life of mutant craA mRNA was significantly greater than that of wild-type craA mRNA.
Highlights
Acinetobacter spp. are ubiquitous environmental organisms, and are readily isolated from soil, water, human skin, food items, and sewage effluent
Four diverse Acinetobacter spp. genomes were used for TransAAP multidrug transporter prediction: the soil commensal strain A. baylyi ADP1, clinical strains A. baumannii American Type Culture Collection (ATCC) 17978 and A. baumannii AYE, and the human louse isolate A. baumannii SDF
The non-pathogenic soil-dwelling Acinetobacter species A. baylyi ADP1 has been proposed as a novel model organism for metabolic and genetic studies, since, amongst other traits, A. baylyi ADP1 is highly recombinogenic and transformable
Summary
Acinetobacter spp. are ubiquitous environmental organisms, and are readily isolated from soil, water, human skin, food items, and sewage effluent. The non-pathogenic soil-dwelling Acinetobacter species, A. baylyi ADP1, has been proposed as an alternative model for molecular, genetic, and metabolic studies, since this organism is naturally transformable, genetically malleable, and nutritionally diverse [1,2,3]. Laboratory studies using A. baylyi ADP1 are facilitated by the capacity of the organism to be transformed with exogenous DNA, in either linear or plasmid form, by incubating the DNA samples with logarithmicphase A. baylyi ADP1 cells [2]. Integration of foreign DNA into the A. baylyi ADP1 chromosome occurs at high frequencies, without the need for extensive regions of homology to be present on transfected DNA molecules [1]. Despite occupying diverse environmental niches, most of the 31 described
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