Impact of aeration rate on the transfer range of antibiotic-resistant plasmids during manure composting

Conjugative plasmids are important vectors of mobile antibiotic resvistance genes (ARGs), facilitating their horizontal transfer within the environment. While composting is recognized as an effective method to reduce antibiotics and ARGs in animal manure, its impact on the bacterial host communities containing antibiotic- resistant plasmids remains unclear. In this study, we investigated the permissiveness of bacterial community during composting when challenged with multidrug-resistant conjugative RP4 plasmids, employing Pseudomonas putida as the donor strain. Ultimately, this represents the first exploration of the effects of aeration rates on the range of RP4 plasmid transfer hosts. Transconjugants were analyzed through fluorescent reporter gene-based fluorescence-activated cell sorting and Illumina sequencing. Overall, aeration rates were found to influence various physicochemical parameters of compost, including temperature, pH, total organic matter, total nitrogen, and potassium. Regarding RP4 plasmid host bacteria, the dominant phylum was determined to shift from Bacteroidetes in the raw material to Proteobacteria in the compost. Notably, a moderate-intensity aeration rate (0.05 L/min/L) was found to be more effective in reducing the diversity and richness of the RP4 plasmid host bacterial community. Following composting, the total percentage of dominant transconjugant-related genera decreased by 66.15-76.62%. Ultimately, this study determined that the aeration rate negatively impacts RP4 plasmid host abundance primarily through alterations to the environmental factors during composting. In summary, these findings enhance our understanding of plasmid host bacterial communities under varying composting aeration rates and offer novel insights into preventing the dissemination of ARGs from animal manure to farmland.