Effect of Varying Nitrate Concentrations on Denitrifying Phosphorus Uptake by DPAOs With a Molecular Insight Into Pho Regulon Gene Expression

Chandan Mukherjee; Rajojit Chowdhury; Mst Momtaj Begam; Sayak Ganguli; Ritabrata Basak; Basab Chaudhuri; Krishna Ray

doi:10.3389/fmicb.2019.02586

Effect of Varying Nitrate Concentrations on Denitrifying Phosphorus Uptake by DPAOs With a Molecular Insight Into Pho Regulon Gene Expression

Front Microbiol. 2019 Nov 8:10:2586. doi: 10.3389/fmicb.2019.02586. eCollection 2019.

Authors

Chandan Mukherjee¹, Rajojit Chowdhury¹, Mst Momtaj Begam¹, Sayak Ganguli², Ritabrata Basak³, Basab Chaudhuri⁴, Krishna Ray¹

Affiliations

¹ Environmental Biotechnology Group, Department of Botany, West Bengal State University, Kolkata, India.
² Theoretical and Computational Biology Division, AIIST and The Biome, Kolkata, India.
³ Department of Biochemistry, Ballygunge Science College, University of Calcutta, Kolkata, India.
⁴ West Bengal State University, Kolkata, India.

Abstract

Bacterial Pho regulon is a key regulator component in biological phosphorus-uptake. Poly-phosphate accumulating bacteria used in enhanced biological phosphorus removal (EBPR) system encounter negative regulation of the Pho regulon, resulting in reduced phosphorus-uptake from phosphorus-replete waste effluents. This study demonstrates possible trends of overcoming the PhoU negative regulation, resulting in excessive PO₄ ^3--P uptake at varying concentrations of NO₃ ^--N through denitrifying phosphorus removal process. We investigated the Pho regulon gene expression pattern and kinetic studies of P-removal by denitrifying phosphate accumulating organisms (DPAOs) which are able to remove both PO₄ ^3--P and NO₃ ^--N in single anoxic stage with the utilization of external carbon sources, without the use of stored polyhydroxyalkanoate (PHA) and without any anaerobic-aerobic or anaerobic-anoxic switches. Our study establishes that a minimum addition of 100 ppm NO₃ ^--N leads to the withdrawal of the negative regulation of Pho regulon and results in ∼100% P-removal with concomitant escalated poly-phosphate accumulation by our established DPAO isolates and their artificially made consortium, isolated from sludge sample of PO₄ ^3- -rich parboiled rice mill effluent, in a settling tank within 12 h of treatment. The same results were obtained when a phosphate rich effluent (stillage from distillery) mixed with a nitrate rich effluent (from explosive industry) was treated together in a single phase anoxic batch reactor, eliminating the need for alternating anaerobic/aerobic or anaerobic/anoxic switches for removing both the pollutants simultaneously. The highest poly-phosphate accumulation was observed to be more than 17% of cell dry weight. Our studies unequivocally establish that nitrate induction of Pho regulon is parallely associated with the repression of PhoU gene transcription, which is the negative regulator of Pho regulon. Based on earlier observations where similar nitrate mediated transcriptional repression was cited, we hypothesize the possible involvement of NarL/NarP transcriptional regulator proteins in PhoU repression. At present, we propose this denitrifying phosphorus removal endeavor as an innovative methodology to overcome the negative regulation of Pho regulon for accelerated unhindered phosphorus remediation from phosphate rich wastewater in India and the developing world where the stringency of EBPR and other reactors prevent their use due to financial reasons.

Keywords: DPAO; Pho regulon; denitrifying phosphorus removal; negative regulation; poly-phosphate accumulation; single stage anoxic reactor; transcriptional repression.