Isolation and characterization of denitrifiers from woodchip bioreactors for bioaugmentation application

Authors: ANDERSON, EMILY - UNIVERSITY OF MINNESOTA; JANG, JEONGHWAN - UNIVERSITY OF MINNESOTA; Venterea, Rodney - Rod; Feyereisen, Gary; ISHII, SATOSHI - UNIVERSITY OF MINNESOTA

Submitted to: Journal of Applied Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/27/2020
Publication Date: 4/7/2020
Citation: Anderson, E., Jang, J., Venterea, R.T., Feyereisen, G.W., Ishii, S. 2020. Isolation and characterization of denitrifiers from woodchip bioreactors for bioaugmentation application. Journal of Applied Microbiology. https://doi.org/10.1111/jam.14655.
DOI: https://doi.org/10.1111/jam.14655

Interpretive Summary: Woodchip bioreactor technology is a promising approach to remove nitrate from agricultural subsurface drainage water by using denitrifying microorganisms. Under cold temperatures, however, bioreactor efficiency could decrease due to inhibited microbial activity. Biostimulation and bioaugmentation have been proposed to enhance nitrate removal by adding labile carbon and psychrophilic denitrifiers, respectively. This study was done to obtain psychrophilic denitrifiers that could be used in future bioaugmentation. We isolated psychrophilic denitrifiers from four different bioreactors in Minnesota, and characterized the strains by measuring their denitrification rates and analyzing their whole genomes. A total of 206 bacteria were isolated from woodchips and biofilms that formed in the bioreactors, 76 of which were able to reduce nitrate at 15°C. Among those, nine potential denitrifying strains were identified, all of which were isolated from the woodchip samples. Although many nitrate- reducing strains were isolated from the biofilm samples, none were categorized as denitrifiers but instead as carrying out dissimilatory nitrate reduction to ammonium (DNRA). Among the denitrifiers confirmed by 15N stable isotope analysis and genome analysis, Cellulomonas cellasea strain WB94 and Microvirgula aerodenitrificans strain BE2.4 appear to be promising for bioreactor bioaugmentation due to their potential for both aerobic and anaerobic denitrification, and the ability of strain WB94 to degrade cellulose. Aerobic denitrification ability may be beneficial in treating water in woodchip bioreactors where water and oxygen levels tend to fluctuate widely, and the ability to degrade cellulose could enhance denitrification by providing carbon and electron donors. Inoculating these strains could enhance nitrate removal in woodchip bioreactors at low temperature conditions. This research helps to develop a new technology that has the potential to improve water quality in the upper Midwest USA and will be of interest to producers, policy makers and the general public.

Technical Abstract: Woodchip bioreactor technology is a promising approach to remove nitrate from agricultural subsurface drainage water by using denitrifying microorganisms. Under cold temperatures, however, bioreactor efficiency could decrease due to inhibited microbial activity. Biostimulation and bioaugmentation have been proposed to enhance nitrate removal by adding labile carbon and psychrophilic denitrifiers, respectively. This study was done to obtain psychrophilic denitrifiers that could be used in future bioaugmentation. We isolated psychrophilic denitrifiers from four different bioreactors in Minnesota, and characterized the strains by measuring their denitrification rates and analyzing their whole genomes. A total of 206 bacteria were isolated from woodchips and biofilms that formed in the bioreactors, 76 of which were able to reduce nitrate at 15°C. Among those, nine potential denitrifying strains were identified, all of which were isolated from the woodchip samples. Although many nitrate- reducing strains were isolated from the biofilm samples, none were categorized as denitrifiers but instead as carrying out dissimilatory nitrate reduction to ammonium (DNRA). Among the denitrifiers confirmed by 15N stable isotope analysis and genome analysis, Cellulomonas cellasea strain WB94 and Microvirgula aerodenitrificans strain BE2.4 appear to be promising for bioreactor bioaugmentation due to their potential for both aerobic and anaerobic denitrification, and the ability of strain WB94 to degrade cellulose. Aerobic denitrification ability may be beneficial in treating water in woodchip bioreactors where water and oxygen levels tend to fluctuate widely, and the ability to degrade cellulose could enhance denitrification by providing carbon and electron donors. Inoculating these strains could enhance nitrate removal in woodchip bioreactors at low temperature conditions.