Acute lethality and stress-responsive phenotypes differ among three Acinetobacter baumannii strains: phenotypic, genomic and transcriptomic analyses

Abstract Background Acinetobacter baumannii exhibits substantial strain-level heterogeneity in antimicrobial resistance, virulence-associated phenotypes and responses to environmental stress. Whether acute lethality covaries with surface-associated and stress-responsive phenotypes across different strain backgrounds remains unclear. We compared three clinically distinct A. baumannii strains, LS01, AB5075 and XH386, using phenotypic assays, comparative genome annotation and transcriptomic profiling. Results The three strains displayed distinct phenotypic profiles. Following inoculation with 5 × 10^4 CFU in Galleria mellonella , 96-h survival was 20% for LS01, 45% for AB5075 and 60% for XH386. In immunocompetent C57BL/6J mice challenged by oropharyngeal aspiration with a common inoculum of 5 × 10^6 CFU, 120-h survival was 25% after LS01 infection, whereas all AB5075- and XH386-infected mice survived. XH386 yielded the highest HBE-cell-associated bacterial recovery in both independent experiments. OD600 trajectories during exposure to non-heat-inactivated human serum varied with serum concentration and donor; at 10% serum, XH386 showed a higher late-phase OD600 than LS01 in both donor experiments. Transmission electron microscopy showed the most prominent capsule-associated extracellular layer in AB5075, whereas LS01 showed a thinner layer and XH386 markedly attenuated capsule-associated material. Attached biomass varied with strain and environmental condition, with XH386 showing the highest biomass under 10% heat-inactivated serum in repeated experiments. Comparative genome annotation showed extensive overlap in major pathogenicity-, biofilm-, iron-acquisition-, secretion- and stress-associated systems despite these phenotypic differences. In XH386, DIP-mediated iron restriction produced the largest transcriptional response, with 1,059 genes meeting adjusted P < 0.05 and |log2 fold change| ≥ 1. Iron-acquisition-associated loci were strongly induced by iron restriction and more modestly by heat-inactivated serum, whereas sub-inhibitory colistin was associated with coordinated repression of ribosomal, respiratory and metabolic genes. Conclusions LS01, AB5075 and XH386 showed no common ordering across acute lethality, surface-associated phenotypes and environmental stress responses. At the common murine challenge dose, LS01 showed the greatest acute lethality, whereas XH386 yielded the highest HBE-cell-associated bacterial recovery and repeatedly showed the highest attached biomass under 10% heat-inactivated serum. Major pathogenicity- and stress-associated functional systems were broadly shared among the three genomes, while XH386 exhibited marked condition-dependent transcriptional remodeling. Together, these findings indicate that the experimentally measured phenotypic differences among the three strains were not attributable to broad pathway presence or absence alone.