A comparative, multi-study analysis of plastisphere resistomes, plasmid dynamics, and antibiotic resistance genes

ABSTRACT Microplastics are widespread in aquatic environments and support surface-associated microbial communities. Although antimicrobial resistance in the plastisphere has been reported, the organization of resistance genes across plasmid types and mobility categories on microplastic surfaces remains incompletely characterized. In this study, we performed a comparative analysis of published microplastic biofilm metagenomes to examine plasmid replicon diversity, predicted mobility, and antimicrobial resistance gene (ARG) distributions across various microplastic polymers. Phylum-level taxonomic profiles varied across polymer types, but most plastisphere communities were dominated by Pseudomonadota . Plasmid reconstruction revealed differences in predicted mobility profiles, with conjugative plasmids more frequently associated with polyethylene, polypropylene, and polyvinyl chloride. Network analyses linking plasmid mobility categories, replicon types, and ARG classes showed that conjugative plasmids connected a broader range of replicons to multiple ARG classes than mobilizable or non-mobilizable plasmids. Replicons from the Inc family, especially IncFIB and IncFII, were prominent and carried high ARG loads, consistent with their capacity to harbor multiple accessory genes. Additionally, several less-characterized rep_cluster replicons were detected across microplastic types, indicating diverse and understudied plasmid backbones within plastisphere communities. The co-occurrence of ARGs with mobile plasmid architectures underscores the importance of considering plasmid context when evaluating plastisphere resistomes. IMPACT STATEMENT Microplastics are increasingly recognized as microbial habitats that can concentrate antibiotic resistance genes (ARGs) in aquatic environments. While many studies have documented the presence of ARGs within microplastic-associated biofilms, far less is known about the genomic context of these genes. This study advances the field by shifting the focus from simple ARG inventories to the plasmid architectures associated with these ARGs. By integrating metagenomic data from previously published studies spanning freshwater, estuarine, and marine systems, we provide the first comparative, cross-system assessment of plasmid replicon diversity, mobility potential, and ARG co-occurrence across different microplastic polymers. This plasmid-centric perspective reveals that ARGs in microplastic biofilms are often associated with conjugative plasmids, which can facilitate their horizontal transfer within these biofilms. Importantly, this work identifies microplastics as environments where clinically relevant ARGs are linked to plasmid architectures commonly observed in pathogenic bacteria. This conceptual advance supports more mechanistic risk assessments of plastic pollution and informs One Health-oriented strategies to address antibiotic resistance across environmental, animal, and human systems.