Pond Water Microbiome Taxa Profiles and Antibiotic Resistance Genes Associated with Acidity and Tannins

Microbial communities of small freshwater bodies are poorly understood. Four ponds in Knox County, Ohio, were sampled over two years to investigate the relationship between the microbial taxa profiles, antibiotic resistance genes (ARGs), and environmental factors such as pH and tannin concentrations. For each site, microbial communities were collected by filtration and metagenomes were analyzed by short-read sequencing. Taxa profiles were predicted by the Kraken2/Bracken pipelines. Bacterial taxa with high abundance in these ponds included Betaproteobacteria (Polynucleobacter and Methylopumilus) and Actinobacteria (Planktophila, Nanopelagicus, and Mycolicibacterium). One pond, a former quarry with elevated pH, showed high prevalence of Cyanobacteria with a seasonal shift from Synechococcus to Planktothrix in the fall. Planktothrix increase was associated with acidification. ARGs were quantified using the ShortBRED pipeline to detect and quantify hits to a marker set derived from the Comprehensive Antibiotic Resistance Database (CARD). The top two ARGs with the largest marker hits encode components of a Stenotrophomonas drug efflux pump powered by proton-motive force (smeABC) and a mycobacterial global regulator that activates a drug pump and other cell defenses (mtrA). Pump function and global activation of transcription incur large energy expenditures, whose fitness cost may increase at high external pH where the cell's proton-motive force is diminished. The smeABC and mtrA prevalence showed a modest correlation with acidifying conditions (low pH and high tannins) which contribute a large transmembrane pH difference to the proton-motive force, thus increasing the cell's energy available for pump function and global gene expression. Compared to rivers and lakes, pond microbial ecosystems are understudied despite close contact with agriculture and recreation. Environmental microbes offer health benefits as well as hazards for human contact. Small water bodies may act as reservoirs for drug-resistant organisms and transfer of antibiotic resistance genes. Yet, the public is rarely aware of the potential for exposure to ARG-carrying organisms in recreational water bodies. Little is known about the capacity for freshwater microbial communities to remediate drug pollution and which biochemical factors may select against antibiotic resistance genes. This study analyzes the bacterial taxa composition and ARG prevalence including possible influence of factors such as pH and tannic acid levels.