Metagenomic Insights Into Functional and Taxonomic Compositions of an Activated Sludge Microbial Community Treating Leachate of a Completed Landfill: A Pathway-Based Analysis

Abstract

Upcycling wastes into valuable products by mixed microbial communities has recently received considerable attention. Sustainable production of high-value substances from one-carbon (C1) compounds, e.g., methanol supplemented as an external electron donor in bioreactors for wastewater treatment, is a promising application of upcycling. This study undertook a gene-centric approach to screen valuable production potentials from mixed culture biomass, removing organic carbon and nitrogen from landfill leachate. To this end, the microbial community of the activated sludge from a landfill leachate treatment plant and its metabolic potential for the production of seven valuable products were investigated. The DNA extracted from the activated sludge was subjected to shotgun metagenome sequencing to analyze the microbial taxonomy and functions associated with producing the seven products. The functional analysis confirmed that the activated sludge could produce six of the valuable products, ectoine, polyhydroxybutyrate (PHB), zeaxanthin, astaxanthin, acetoin, and 2,3-butanediol. Quantification of the detected functional gene hit numbers for these valuable products as a primary trial identified a potential rate-limiting metabolic pathway, e.g., conversion of L-2,4-diaminobutyrate into N-γ-acetyl-L2,4,-diaminobutyrate during the ectoine biosynthesis. Overall, this study demonstrated that primary screening by the proposed gene-centric approach can be used to evaluate the potential for the production of valuable products using mixed culture or single microbe in engineered systems. The proposed approach can be expanded to sites where water purification is highly required, but resource recovery, or upcycling has not been implemented.

Potential survival strategies of novel comammox and nitrite-oxidizingNitrospirapresent in a reactor treating high-ammonia brackish landfill leachate

“…The sedimented biomass was returned to the SBR to retain biomass concentration. The detailed system configuration is shown elsewhere [22]. The leachate (average flow rate: 74.2 ± 40.0 m 3 /day) containing NH 4 + at 151.31 ± 49.53 mg-N/L and chemical oxygen demand (COD) at 43.9 ± 9.2 mg-COD/L was mixed with groundwater (13.2 ± 11.1 m 3 /day) in a flow equalization tank, located before the SBR.…”

“…Comammox detection in brackish water areas [20] and sediments of mangrove forests close to the salt concentration of seawater (0.5%–3%) has been reported by detection of the amoA gene [21]. Meanwhile, our previous work employed gene-centric metagenomic analysis of the microbiome inhabiting a landfill leachate treatment plant, unveiling the presence of comammox Nitrospira in a brackish (salinity 1.23%) and high-ammonia (117.80 ± 30.13 mg-NH 4 + -N/L) environment [22]. However, these analyses do not comprehensively describe the functions of comammox Nitrospira .…”

“…In the later stage of the landfill (after several decades), leachate includes not only low levels of organic matter and a high concentration of ammonia, but also high salinity derived from incineration residues [24, 25]. Leachate treatment plants install a system to perform nitrification/denitrification, and methanol is often added as an electron donor to supplement the low-concentration organic matter to promote denitrification [22]. Biomass in the treatment plant was recycled from a sedimentation tank to a reaction tank.…”

“…The long-term enrichment culture selected microorganisms that could gradually adapt to this unique environment. However, the 16S rRNA gene amplicon sequencing and gene-centric metagenome analysis, applied in previous studies [22, 26] to investigate microbial community compositions and functions, have not fully succeeded in identifying nitrifiers, determining their abundance, and characterizing their individual genomic features.…”

“…This study used genome-centric metagenomics to investigate nitrifiers in the microbiome treating landfill leachate in a brackish environment with high ammonia concentration and added methanol as a carbon source. To this end, we asked the following research questions: What are the dominant nitrifiers in this unique environment? What are their unique genomic features? Is it possible to obtain a draft comammox Nitrospira genome, and, if so, what type is it and what genetic characteristics does it have? Our previous gene-centric analysis of the same microbial community revealed the presence of Nitrospira and Thaumarchaeota , indicating the potential for acquisition of draft genomes for Nitrospira , including comammox strains, and AOA [22]. Hybrid sequencing with long and short reads, and integration of three binning algorithms to calculate an optimized set of bins from a single assembly, were performed to acquire high-quality metagenomic assembled genomes (MAGs) of microorganisms involved in nitrification.…”