Not so dangerous? PET enviromental-like microplastics and nanoplastics effect on Escherichia coli and environmental bacteria

Microplastics and nanoplastics (MNPs) are increasingly recognized as biologically active environmental stressors, yet bacterial responses to environmentally relevant MNPs remain poorly characterized compared to algae and cyanobacteria commonly used in ecotoxicity testing. Here, we investigated the effects of environmentally aged polyethylene terephthalate envinronmental-like micro- and nanoplastics (PET-elMNPs) on a panel of Gram-positive ( Bacillus mycoides, Curtobacterium oceanosedimentum ) and Gram-negative ( Vibrio natriegens, Pseudoalteromonas rubra ) freshwater and saltwater bacteria, with Escherichia coli as a reference model. Growth inhibition by PET-elMNPs strongly depended on the cultivation format, with 24-well plates providing the most discriminative conditions (e.g., 33–45% inhibition for E. coli , 30–60% for V. natriegens and 34–38% for B. mycoides ). In contrast, obligate aerobes showed format-specific sensitivity ( P. rubra 36–60% in 96-well plates; C. oceanosedimentum 37–45% in 12-well plates), consistent with differences in oxygen availability and mixing. Confocal imaging revealed species-specific interactions within the plastisphere but only limited loss of viability. Common enzyme-activated fluorescent probes were confounded by strong PET-elMNP binding and PET-elMNPs-mediated dye activation, indicating unique enzyme-mimicking surface chemistry of the PET-MNPs. Quantitative proteomics showed highly species-specific stress responses centered on iron/metal homeostasis, cell-wall remodeling, and proteostasis/mRNA regulation, including the induction of secreted proteins (e.g. chondroitin AC-lyase in Vibrio and a βγ-crystallin-fold protein in Bacillus ) that may contribute to the modifying of MNPs surface. Together, our results highlight PET-elMNPs as biologically active interfaces that challenge standard bacterial toxicity assays and elicit complex, species-dependent adaptive responses.