Plasmid Backbone Impacts Conjugation Rate, Transconjugant Fitness, and Community Assembly of Genetically Bioaugmented Soil Microbes for PAH Bioremediation.

Abstract

Many polycyclic aromatic hydrocarbons (PAHs) in the environment resulting from crude oil spills and the incomplete combustion of organic matter are highly toxic, mutagenic, or carcinogenic to microorganisms and humans. Bioremediation of PAHs using microorganisms that encode biodegradative genes is a promising approach for environmental PAH cleanup. However, the viability of exogenous microorganisms is often limited due to competition with the native microbial community. Instead of relying on the survival of one or a few species of bacteria, genetic bioaugmentation harnesses conjugative plasmids that spread functional genes to native microbes. In this study, two plasmid backbones that differ in copy number regulation, replication, and mobilization genes were engineered to contain a PAH dioxygenase gene (bphC) and conjugated to soil bacteria including Bacillus subtilis, Pseudomonas putida, and Acinetobacter sp., as well as a synthetic community assembled from these bacteria. Fitness effects of the plasmids in transconjugants significantly impacted the rates of conjugative transfer and biotransformation rates of a model PAH (2,3-dihydroxybiphenyl). A synergistic effect was observed in which synthetic communities bioaugmented with bphC had significantly higher PAH degradation rates than bacteria grown in monocultures. Finally, conjugation rates were significantly associated with the relative abundances of bacteria in synthetic communities, underscoring how fitness impacts of plasmids can shape the microbial community structure and function.