Conjugative antibiotic-resistant plasmids promote bacterial colonization of microplastics in water environments

Both microplastic and bacterial antibiotic resistance have attracted attention worldwide. When microplastics coexist with antibiotic-resistant bacteria (ARB), which carry antibiotic resistance genes (ARGs), ARB colonize the surface of microplastics, and a unique biofilm is formed. The ARB and ARGs in biofilms are denser and more difficult to remove. However, studies on the factors influencing the formation of microplastic biofilms are limited. In this study, plasmid RP4, which appeared in wastewater treatment plants, was found to be able to promote irreversible bacterial colonization of microplastics, and the hypothetical reason was conjugative pili expression. Then, the potential conjugative pili synthesis promoter "nanoalumina" and inhibitor "free nitrous acid" (FNA) were selected to test this hypothesis. Simultaneously, nanoalumina promoted and FNA inhibited bacterial colonization when RP4 existed. Combined with the gene expression and ATP analysis results, this hypothesis was confirmed, and the mechanism of RP4 on bacterial colonization was related mainly to conjugative pili protein synthesis and intracellular ATP. In this study, the effects of plasmid RP4, nanoalumina, and FNA on the formation of microplastic biofilms were reported, which has a certain reference value for other researchers exploring microplastic biofilms.