Abstract
The goal of the present work was to investigate the physicochemical and radiochemical conditions and the composition of the microbial community in the groundwater of a suspended surface repository for radioactive waste (Russia) and to determine the possibility of in situ groundwater bioremediation by removal of nitrate ions. Groundwater in the repository area (10-m depth) had elevated concentrations of strontium, tritium, nitrate, sulfate, and bicarbonate ions. High-throughput sequencing of the V3–V4/V4 region of the 16S rRNA gene revealed the presence of members of the phyla Proteobacteria (genera Acidovorax, Simplicispira, Thermomonas, Thiobacillus, Pseudomonas, Brevundimonas, and uncultured Oxalobacteraceae), Firmicutes (genera Bacillus and Paenibacillus), and Actinobacteria (Candidatus Planktophila, Gaiella). Canonical correspondence analysis suggested that major contaminant – nitrate, uranium, and sulfate shaped the composition of groundwater microbial community. Groundwater samples contained culturable aerobic organotrophic, as well as anaerobic fermenting, iron-reducing, and denitrifying bacteria. Pure cultures of 33 bacterial strains belonging to 15 genera were isolated. Members of the genera Pseudomonas, Rhizobium, Cupriavidus, Shewanella, Ensifer, and Thermomonas reduced nitrate to nitrite and/or dinitrogen. Application of specific primers revealed the nirS and nirK genes encoding nitrite reductases in bacteria of the genera Pseudomonas, Rhizobium, and Ensifer. Nitrate reduction by pure bacterial cultures resulted in decreased ambient Eh. Among the organic substrates tested, sodium acetate and milk whey were the best for stimulation of denitrification by the microcosms with groundwater microorganisms. Injection of these substrates into the subterranean horizon (single-well push-pull test) resulted in temporary removal of nitrate ions in the area of the suspended radioactive waste repository and confirmed the possibility for in situ application of this method for bioremediation.
Highlights
The methods used in Russia, United States, and other countries for disposal of radioactive waste (RAW) during the 20th century resulted in numerous repositories which represent a certain environmental danger (Rybal’chenko et al, 1998; Wall and Krumholz, 2006)
Nitrate was the major contaminant and its concentration exceeded significantly the maximal permissible concentrations (MPCs) value (45 mg l−1 for groundwater in Russia), which necessitated the procedures for its removal from groundwater
Nitrate and concomitant contaminants significantly linked to the overall microbial communities of groundwater from B, D, and E wells and determined the presence of members of the genera Acidivorax, Simplicispira, Pseudomonas, Brevundimonas, Thermomonas, Acinetobacter, capable of denitrification with dinitrogen release or of nitrate reduction to nitrite (Figure 7) (Elomari et al, 1996; Mergaert et al, 2003; Grabovich et al, 2006; Choi et al, 2010). These results indicate that a unique microbial community formed in groundwater at the area of a suspended surface repository for liquid RAW, which differed from the community of natural groundwater
Summary
The methods used in Russia, United States, and other countries for disposal of radioactive waste (RAW) during the 20th century resulted in numerous repositories which represent a certain environmental danger (Rybal’chenko et al, 1998; Wall and Krumholz, 2006). Non-radioactive waste components migrate with groundwater, covering significant distances and creating the risk of contamination of surrounding areas. The microorganisms of nitrate- and radionuclide-contaminated subterranean water-bearing strata and sediments have been studied in detail (Anderson et al, 2003; Elias et al, 2003; Nevin et al, 2003; Yan et al, 2003; North et al, 2004; Fields et al, 2005; Vrionis et al, 2005; Hwang et al, 2009). Denitrifying bacteria of the genera Afipia, Hyphomicrobium, Rhodanobacter, Intrasporangium, and Bacillus were isolated from Oak Ridge sediments, Oak Ridge, TN, United States (The Field Research Center; OR-FRC1) (Green et al, 2010). Analysis of the nirK and nirS nitrite reductase functional genes revealed predominance of Rhodanobacter spp. in these sediments (Green et al, 2012)
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