AbstractAcinetobacter baumannii is a feared, drug-resistant pathogen, characterized by its ability to resist extreme environmental and nutrient-deprived conditions. Previously, we showed that human serum albumin (HSA) can increase foreign DNA acquisition specifically and alter the expression of genes associated with pathogenicity. Moreover, in a recent genome-wide transcriptomic study, we observed that pleural fluid (PF), an HSA-containing fluid, increases DNA acquisition, can modulate cytotoxicity, and control immune responses by eliciting changes in the A. baumannii metabolic profile. In the present work, using more stringent criteria and focusing on the analysis of genes related to pathogenicity and response to stress, we analyzed our previous RNA-seq data and performed phenotypic assays to further explore the impact of PF on A. baumannii’s microbial behavior and the strategies used to overcome environmental stress. We observed that PF triggered differential expression of genes associated with motility, efflux pumps, antimicrobial resistance, biofilm formation, two-component systems (TCSs), capsule synthesis, osmotic stress, and DNA-damage response, among other categories. Phenotypic assays of A. baumannii A118 and two other clinical A. baumannii strains, revealed differences in their responses to PF in motility, biofilm formation, antibiotic susceptibility, osmotic stress, and outer membrane vesicle (OMV) production, suggesting that these changes are strain specific. We conclude that A. baumannii’s pathoadaptive responses is induced by HSA-containing fluids and must be part of this bacterium armamentarium to persist in hostile environments.