Adaptation of a wastewater bacterial community to a typical antibiotic analyzed by flow cytometry

The impacts of environmentally widespread antibiotics on natural microbial communities have attracted the public’s attention for years. Reliable and efficient technologies for detecting these effects are much required. To study the microbial ecotoxicology of an antibiotic widely used for bladder infections treatment--fosfomycin sodium (FOM), which fails to be removed by routine wastewater treatment from the wastewater of a pharmaceutical factory, we used a gfp-labeled marker Pseudomonas putida strain and microbial community flow-cytometry. The response of the wastewater bacterial community to FOM was measured on single-cell level and on daily basis and evaluated by standard cytometric bioinformatics tools. The wastewater community was cultivated in a continuous bioreactor amended with increased concentration of FOM (0, 10, and 268 mg/L). The fate of the fluorescent P. putida strain after being added (0.5% or 2.5%) to the bacterial community was tracked and found to lead to disappearance, indicating a superior growth of bacterial types from the wastewater community that adapted to FOM stress. A significant turnover in community composition was observed, which was strongly correlated with FOM concentration, chemical oxygen demand (COD), and cell number. The combination of cell sorting and 16S rRNA amplicon sequencing showed that the major subcommunities that persisted at 268 mg/L were dominated, for example, by the genus Elizabethkingia, which is described to harbor a large number of multidrug resistant emerging pathogens. The study showed that the community under FOM exposure changed its composition, with susceptible species being displaced by organisms able to withstand antibiotic pressure.