Impacts of biostimulation and bioaugmentation on woodchip bioreactor microbiomes

Authors: WANG, HAO - University Of Minnesota; Feyereisen, Gary; WANG, PING - University Of Minnesota; ROSEN, CARL - University Of Minnesota; SADOWSKY, MICHAEL - University Of Minnesota; ISHII, SATOSHI - University Of Minnesota

Submitted to: Microbiology Spectrum
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/26/2023
Publication Date: 9/25/2023
Citation: Wang, H., Feyereisen, G.W., Wang, P., Rosen, C.J., Sadowsky, M.J., Ishii, S. 2023. Impacts of biostimulation and bioaugmentation on woodchip bioreactor microbiomes. Microbiology Spectrum. 11(5). Article 04053-22. https://doi.org/10.1128/spectrum.04053-22.
DOI: https://doi.org/10.1128/spectrum.04053-22

Interpretive Summary: Woodchip bioreactor is a conservation practice that removes nitrate from agricultural tile water by directing the water into a bed of woodchips where microbes convert the nitrate to a gaseous form that goes back to atmosphere. The conversion process, called denitrification, is hindered by cool temperatures during springtime when tile flow in greatest. The makeup of the microbial community and the amount of carbon available to the microbes influence the effectiveness of denitrification. In this experiment, microbes were selected in the laboratory for cold-tolerance and inoculated into a pair of pilot-scale field bioreactors (“bioaugmentation”). Also, an easy-to-consume form of carbon, acetate, was added to a second pair of bioreactor units to stimulate denitrification (“biostimulation”). Microbes were collected before and after the experiments. Tests for gene abundances showed that 2.5% of the microbial community variations were due to the bioaugmentation and biostimulation treatments and 17% was due to seasonal shift in composition. A positive relationship was identified between one of the inoculant microbes and abundance of genes needed for denitrification, suggesting bioaugmentation and biostimulation can be effective at influencing denitrifying microbial populations and thus nitrate removal in woodchip bioreactors. This finding provides insight into how bioreactor performance may be improved and is pertinent to researchers, practitioners, academics, and agency personnel with interest in using denitrifying bioreactors to remove nitrate-N from water.

Technical Abstract: Woodchip bioreactors (WBRs) are used to remove nutrients, especially nitrate-nitrogen, from subsurface drainage. However, the nitrogen removal efficiency of WBRs is limited by low temperature and limited availability of labile carbon. Bioaugmentation (i.e., adding cold-adapted denitrifying microbes) and biostimulation (i.e., adding substrates to enhance the activities of indigenous microbial populations) are potential approaches to enhance nitrate removal of WBRs at cold conditions, but their effectiveness is still unclear. We clarified the effects of bioaugmentation and biostimulation on WBR microbiomes and examined the relationship between microbiome structures and the overall nitrate removal. As a bioaugmentation treatment, we inoculated WBR-borne cold-adapted denitrifying bacteria Cellulomonas sp. strain WB94 and Microvirgula sp. strain BE2.4 back into the WBRs located at Willmar, MN (i.e., bioaugmentation). As a biostimulation treatment, acetate was added into the WBRs to promote denitrification. Woodchip samples were collected before and after the bioaugmentation and biostimulation treatments to examine the impact of the two treatments on the WBR microbiomes. The 16S rRNA gene amplicon sequencing showed that the microbiomes changed only slightly by the treatments. Treatments accounted for 2.5% of the microbiome variations, suggesting that only small fraction of microbiomes were influenced by the treatments. In contrast, there was a seasonal shift in the composition of WBR microbiomes, which accounted for 17.1% of the microbiome variations. Based on the high-throughput nitrogen cycle evaluation (NiCE) chip analysis, the abundance of NorB and nosZ were shown to be useful in predicting the nitrate removal rate of WBRs, and the relative abundance of denitrification genes was higher at locations closer to the inlet of the WBRs, suggesting that the microbiomes inside WBRs were rather heterogenous. In addition, a positive relationship was identified between the abundance of inoculant Microvirgula sp. strain BE2.4 and the abundance of norB and nosZ in the WBRs. These results suggested that the bioaugmentation and biostimulation treatments could influence denitrifier populations, thereby influencing the nitrate removal of WBRs.