Elucidating the Relative Roles of Ammonia Oxidizing and Heterotrophic Bacteria during the Biotransformation of 17α-Ethinylestradiol and Trimethoprim

The biological fate of 17α-ethinylestradiol (EE2; 500 ng/L to 1 mg/L) and trimethoprim (TMP; 1 μg/L to 1 mg/L) was evaluated with flow through reactors containing an ammonia oxidizing bacterial (AOB) culture, two enriched heterotrophic cultures devoid of nitrifier activity, and nitrifying activated sludge (NAS) cultures. AOBs biotransformed EE2 but not TMP, whereas heterotrophs mineralized EE2, biotransformed TMP, and mineralized EE2-derived metabolites generated by AOBs. Kinetic bioassays showed that AOBs biotransformed EE2 five times faster than heterotrophs. The basal expression of heterotrophic dioxygenase enzymes was sufficient to achieve the high degree of transformation observed at EE2 and TMP concentrations ≤ 1 mg/L, and enhanced enzyme expression was not necessary. The importance of AOBs in removing EE2 and TMP was evaluated further by performing NAS experiments at lower feed concentrations (500−1000 ng/L). EE2 removal slowed markedly after AOBs were inhibited, while TMP removal was not affected by AOB inhibition. Two key EE2 metabolites formed by AOB and heterotrophic laboratory-scale chemostats were also found in independent laboratory-scale mixed culture bioreactors; one of these, sulfo-EE2, was largely resistant to further biodegradation. AOBs and heterotrophs may cooperatively enhance the reliability of treatment systems where efficient removal of EE2 is desired.