Microbial diversity in a constructed wetland system for treatment of acid sulfate water
Lee, Yong Jin
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Acid sulfate waste waters are produced from industrial and mining activities, which have serious impacts on public health and our environment. A constructed treatment wetland system has been developed to treat low pH, ferric iron-dominated acid sulfate water drained off from two coal storage facilities at the DOE Savannah River Site, near Aiken, SC. Since microbial sulfate reduction plays a pivotal role in raising pH and removing metal contaminants, it is necessary to obtain microbial profiles especially sulfate-reducing bacteria (SRB) to understand their ecological niches and roles in the processes. A large number of sequences obtained from PCR-based culture-independent molecular method were identified as uncultured or unidentified bacteria. The sequences retrieved with a universal primer set were distributed to several main clades, such as: the Cytophaga-Flexibacter-Bacteroides group, OP11, Chloroflexi, and the delta-subclass Proteobacteria. The sequences identified using six sets of SRB-specific primers fell into each target species, but the specificity and diversity of each primer group varied. To identify the predominant sulfate-reducing bacterial community, 16S rDNA sequence analysis was performed with the highest positive MPN dilution tubes. The obtained data suggested Desulfovibrio-like species are the predominant SRB and play a major role in the sulfate reduction process. Heterotrophic bacteria were also abundant and may play a fundamental role in the anaerobic community of the system. The sequencing data showed the addition of acetate to enhance microbial sulfate reduction altered the composition of the predominant community from Gram-type positive spore-forming bacteria to Gram-type negative sulfate-reducing bacteria. Spatial and temporal changes of the predominant microbial communities in the system were also detected using denaturing gradient gel electrophoresis. A parallel approach was directed to isolate, identify, and characterize known and novel bacteria obtained from the highest MPN cultures to determine their roles in various biogeochemical processes in such acidic and metal-contaminated environment. The majority of isolates belonged to low G+C Gram-type positive bacteria including sulfate-reducing Desulfotomaculum and Desulfosporosinus species. Among the isolates, a novel heterotrophic bacterium proposed as Gracilibacter thermotolerans gen. nov. sp. nov. has been described.