<p>Bioaerosols are highly relevant in medical centers (MCs), where factors such as seasonality, human occupancy (HO), and ventilation systems (VS) are proposed to influence the airborne bacterial communities and potential airborne human pathogenic bacteria (PAHPB). We investigated the compositions and spatiotemporal variations in the airborne bacterial communities, including those of PAHPB and antibiotic resistance genes (ARG), in two MCs in Chile during 2021 and 2022. 16S rRNA gene amplicon sequencing and qPCR revealed significant (<i>p</i> &lt; 0.05) variations in diversity, composition, and gene abundance across the studied years. Proteobacteria, Actinobacteria, and Firmicutes were dominant in bioaerosols, with gene abundances ranging from 10³ to 10⁷ copies m³ and were negatively correlated with diversity. The PAHPB and ARG genes were variable, with <i>Kleb</i>2 (<i>Klebsiella pneumoniae</i>) and <i>bla</i>TEM (β-lactam) being the most prevalent. Although HO and VS influenced the bacterial communities, no significant correlations (<i>p</i> &lt; 0.05) with bacterial diversity were found. Interestingly, 6 genera in the core microbiome were associated with humans, whereas 19 genera were associated with the environment, suggesting that outdoor air is a relevant factor influencing bacterial communities in MCs. We provide baseline data for research on bioaerosols in MCs, highlighting the complexity of airborne bacterial communities and suggesting some factors influencing their spatiotemporal variations.</p>