Wastewater treatment plants (WWTPs) are recognized as significant contributors of paracetamol (APAP) into the environment due to their limited ability to degrade it. This study used a bioaugmentation strategy with Pseudomonas extremaustralis CSW01 and Stutzerimonas stutzeri CSW02 to achieve APAP biodegradation in solution in wide ranges of temperature (10-40oC) and pH (5-9), reaching DT50 values < 1.5hours to degrade 500mgL-1 APAP. Bacterial strains also mineralized APAP in solution (<30%), but when forming consortia with Mycolicibacterium aubagnense HPB1.1, mineralization significantly increased (up to 74% and 58% for CSW01+HPB1.1 and CSW02+HPB1.1, respectively), decreasing DT50 values to only 1 and 9 days. Despite the complete degradation of APAP and its high mineralization, residual toxicity throughout the process was observed. Three APAP metabolites were identified (4-aminophenol, hydroquinone and trans-2-hexenoic acid) that quickly disappeared, but residual toxicity remained, indicating the presence of other non-detected intermediates. CSW01 and CSW02 degraded also 100% APAP (50mgkg-1) adsorbed on sewage sludge, with DT50 values of only 0.7 and 0.3 days, respectively, but < 15% APAP was mineralized. A genome-based analysis of CSW01 and CSW02 revealed that amidases, deaminases, hydroxylases, and dioxygenases enzymes were involved APAP biodegradation, and a possible metabolic pathway was proposed. Environmental ImplicationThe wide use of paracetamol (APAP) involves ecological risks, being considered an emerging pollutant. APAP excreted by humans ends up in WWTPs, where its elimination is not complete, remaining in part in the effluent treated waters (discharged into surface waters used for soil irrigation) and in part adsorbed on sewage sludge (used as fertilizer in agricultural soils). APAP bioremediation in WWTPs is the unique opportunity to remove it from both matrices before its dispersion in the environment.The novelty of this study is the isolation of two bacterial strains from sewage sludge, P. extremaustralis CSW01 and S. stutzeri CSW02, that could degrade very high concentrations of APAP and their main formed metabolites, both in water and, for the first time, in sewage sludge, in very short periods of time, and even reach high percentages of APAP mineralization in water. Moreover, CSW01 and CSW02 were active under wide ranges of temperature and pH, and are therefore excellent candidates for the chance to achieve a reduction of APAP toxicity in the environment. In addition, by a genome-based analysis, enzymes possibly involved in APAP biodegradation have been identified and a possible degradation pathway has been proposed.
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