Abundant and diverse arsenic-metabolizing microorganisms in peatlands treating arsenic-contaminated mining wastewaters.

Mining operations produce large quantities of wastewater. At a mine site in Northern Finland, two natural peatlands are used for the treatment of mining-influenced waters with high concentrations of sulphate and potentially toxic arsenic (As). In the present study, As removal and the involved microbial processes in those treatment peatlands (TPs) were assessed. Arsenic-metabolizing microorganisms were abundant in peat soil from both TPs (up to 108 cells gdw -1 ), with arsenate respirers being about 100 times more abundant than arsenite oxidizers. In uninhibited microcosm incubations, supplemented arsenite was oxidized under oxic conditions and supplemented arsenate was reduced under anoxic conditions, while little to no oxidation/reduction was observed in NaN3 -inhibited microcosms, indicating high As-turnover potential of peat microbes. Formation of thioarsenates was observed in anoxic microcosms. Sequencing of the functional genemarkers aioA (arsenite oxidizers), arrA (arsenate respirers) and arsC (detoxifying arsenate reducers) demonstrated high diversity of the As-metabolizing microbial community. The microbial community composition differed between the two TPs, which may have affected As removal efficiencies. In the present situation, arsenate reduction is likely the dominant net process and contributes substantially to As removal. Changes in TP usage (e.g. mine closure) with lowered water tables and heightened oxygen availability in peat might lead to re-oxidation and re-mobilization of bound arsenite.

Resistant ammonia-oxidizing archaea endure, but adapting ammonia-oxidizing bacteria thrive in boreal lake sediments receiving nutrient-rich effluents.

Climate change along with anthropogenic activities changes biogeochemical conditions in lake ecosystems, modifying the sediment microbial communities. Wastewater effluents introduce nutrients and organic material but also novel microbes to lake ecosystems, simulating forthcoming increases in catchment loadings. In this work, we first used 16s rRNA gene sequencing to study how the overall sediment microbial community responds to wastewater in six boreal lakes. To examine forthcoming changes in the lake biogeochemistry, we focused on the ammonia-oxidizing archaea (AOA) and bacteria (AOB), and examined their functional and compositional community response to wastewater. Although we found the least diverse and least resistant prokaryotic communities from the most wastewater-influenced sediments, the community changed fast toward the natural composition with the diminishing influence of wastewater. Each lake hosted a unique resistant AOA community, while AOB communities were adapting, responding to environmental conditions as well as receiving new members from WWTPs. In general, AOB dominated in numbers in wastewater-influenced sediments, while the ratio between AOA and AOB increased when moving toward pristine conditions. Our results suggest that although future climate-change-driven increases in nutrient loading and microbial migration might significantly disrupt lake sediment microbiomes, they can promote nitrification through adapting and abundant AOB communities.

Simplified MPN method for enumeration of soil naphthalene degraders using gaseous substrate.

We describe a simplified microplate most-probable-number (MPN) procedure to quantify the bacterial naphthalene degrader population in soil samples. In this method, the sole substrate naphthalene is dosed passively via gaseous phase to liquid medium and the detection of growth is based on the automated measurement of turbidity using an absorbance reader. The performance of the new method was evaluated by comparison with a recently introduced method in which the substrate is dissolved in inert silicone oil and added individually to each well, and the results are scored visually using a respiration indicator dye. Oil-contaminated industrial soil showed slightly but significantly higher MPN estimate with our method than with the reference method. This suggests that gaseous naphthalene was dissolved in an adequate concentration to support the growth of naphthalene degraders without being too toxic. The dosing of substrate via gaseous phase notably reduced the work load and risk of contamination. The result scoring by absorbance measurement was objective and more reliable than measurement with indicator dye, and it also enabled further analysis of cultures. Several bacterial genera were identified by cloning and sequencing of 16S rRNA genes from the MPN wells incubated in the presence of gaseous naphthalene. In addition, the applicability of the simplified MPN method was demonstrated by a significant positive correlation between the level of oil contamination and the number of naphthalene degraders detected in soil.

Response of Soil Bacterial Community Diversity and Composition to Time, Fertilization, and Plant Species in a Sub-Boreal Climate.

Pastures are an important part of crop and food systems in cold climates. Understanding how fertilization and plant species affect soil bacterial community diversity and composition is the key for understanding the role of soil bacteria in sustainable agriculture. To study the response of soil bacteria to different fertilization and cropping managements, a 3-year (2013-2015) field study was established. In the split-plot design, fertilizer treatment (unfertilized control, organic fertilizer, and synthetic fertilizer) was the main plot factor, and plant treatment [clear fallow, red clover (Trifolium pratense), timothy (Phleum pratense), and a mixture of red clover and timothy] was the sub-plot factor. Soil bacterial community diversity and composition, soil properties, and crop growth were investigated through two growing seasons in 2014 and 2015, with different nitrogen input levels. The community diversity measures (richness, Shannon diversity, and Shannon evenness) and composition changed over time (P < 0.05) and at different time scales. The community diversity was lower in 2014 than in 2015. The temporal differences were greater than the differences between treatments. The overall correlations of Shannon diversity to soil pH, NO 3 - , NH 4 + , and surplus nitrogen were positive and that of bacterial richness to crop dry matter yield was negative (P < 0.05). The major differences in diversity and community composition were found between fallow and planted treatments and between organic and synthetic fertilizer treatments. The differences between the planted plots were restricted to individual operational taxonomic units (OTUs). Soil moisture, total carbon content, and total nitrogen content correlated consistently with the community composition (P < 0.05). Compared to the unfertilized control, the nitrogen fertilizer loading enhanced the temporal change of community composition in pure timothy and in the mixture more than that in red clover, which further emphasizes the complexity of interactions between fertilization and cropping treatments on soil bacteria.

Bacterial community changes in response to oil contamination and perennial crop cultivation.

We investigated bacterial community dynamics in response to used motor oil contamination and perennial crop cultivation by 16S rRNA gene amplicon sequencing in a 4-year field study. Actinobacteria, Proteobacteria, Chloroflexi, Acidobacteria, and Gemmatimonadetes were the major bacterial phyla, and Rhodococcus was the most abundant genus. Initially, oil contamination decreased the overall bacterial diversity. Actinobacteria, Betaproteobacteria, and Gammaproteobacteria were sensitive to oil contamination, exhibiting clear succession with time. However, bacterial communities changed over time, regardless of oil contamination and crop cultivation. The abundance difference of most OTUs between oil-contaminated and non-contaminated plots remained the same in later sampling years after the initial abundance difference induced by oil spike. The abundances of three oil-favored actinobacteria (Lysinimonas, Microbacteriaceae, and Marmoricola) and one betaproteobacterium (Aquabacterium) changed in different manner over time in oil-contaminated and non-contaminated soil. We propose that these taxa are potential bio-indicators for monitoring recovery from motor oil contamination in boreal soil. The effect of crop cultivation on bacterial communities became significant only after the crops achieved stable growth, likely associated with plant material decomposition by Bacteroidetes, Armatimonadetes and Fibrobacteres.

Microbial diversity along a gradient in peatlands treating mining-affected waters.

Peatlands are used for the purification of mining-affected waters in Northern Finland. In Northern climate, microorganisms in treatment peatlands (TPs) are affected by long and cold winters, but studies about those microorganisms are scarce. Thus, the bacterial, archaeal and fungal communities along gradients of mine water influence in two TPs were investigated. The TPs receive waters rich in contaminants, including arsenic (As), sulfate (SO42-) and nitrate (NO3-). Microbial diversity was high in both TPs, and microbial community composition differed between the studied TPs. Bacterial communities were dominated by Proteobacteria, Actinobacteria, Chloroflexi and Acidobacteria, archaeal communities were dominated by Methanomicrobia and the Candidate phylum Bathyarchaeota, and fungal communities were dominated by Ascomycota (Leotiomycetes, Dothideomycetes, Sordariomycetes). The functional potential of the bacterial and archaeal communities in TPs was predicted using PICRUSt. Sampling points affected by high concentrations of As showed higher relative abundance of predicted functions related to As resistance. Functions potentially involved in nitrogen and SO42- turnover in TPs were predicted for both TPs. The results obtained in this study indicate that (i) diverse microbial communities exist in Northern TPs, (ii) the functional potential of the peatland microorganisms is beneficial for contaminant removal in TPs and (iii) microorganisms in TPs are likely well-adapted to high contaminant concentrations as well as to the Northern climate.

Bacterial community structure along the subtidal sandy sediment belt of a high Arctic fjord (Kongsfjorden, Svalbard Islands).

Open fjords are subject to contrasting environmental conditions, owing to meltwater glacial inputs, terrestrial runoff, and marine water mass exchanges, which are exacerbated by anthropogenic and climate perturbations. Following a slope-dependent water circulation, the subtidal sandy sediment belt regulates the convergent transport of nutrients downward the fjord depths, and the effective entrapment of suspended particles and microorganisms. In this study, we aimed at testing how glacial and seawater inputs may influence the bacterial community structure of subtidal sand deposits in the Kongsfjorden. Through total and viable cell counting and an amplicon sequencing approach, we found relevant differences in bacterial community structure along the glacio-marine sampling transect. Viable and high nucleic acid content (HNA) cells represented an important fraction of the total community, generally decreasing toward the glacier front. Besides the predominance of Alpha- and Gammaproteobacteria, Bacteroidetes, Firmicutes and Parcubacteria, the bacterial community structure was likely affected by the glacial activity in the inner fjord, with the occurrence of distinctive phylotypes belonging to Gemmatimonadates, Nitrospirae, Acidobacteria, and Chloroflexi. Overall, our outcomes highlighted that exploring the bacterial community distribution and structure can provide new insights into the active role of sand deposits in coastal cold environments.

Successful aerobic bioremediation of groundwater contaminated with higher chlorinated phenols by indigenous degrader bacteria.

The xenobiotic priority pollutant pentachlorophenol has been used as a timber preservative in a polychlorophenol bulk synthesis product containing also tetrachlorophenol and trichlorophenol. Highly soluble chlorophenol salts have leaked into groundwater, causing severe contamination of large aquifers. Natural attenuation of higher-chlorinated phenols (HCPs: pentachlorophenol + tetrachlorophenol) at historically polluted sites has been inefficient, but a 4-year full scale in situ biostimulation of a chlorophenol-contaminated aquifer by circulation and re-infiltration of aerated groundwater was remarkably successful: pentachlorophenol decreased from 400 µg L-1 to <1 µg L-1 and tetrachlorophenols from 4000 µg L-1 to <10 µg L-1. The pcpB gene, the gene encoding pentachlorophenol hydroxylase - the first and rate-limiting enzyme in the only fully characterised aerobic HCP degradation pathway - was present in up to 10% of the indigenous bacteria already 4 months after the start of aeration. The novel quantitative PCR assay detected the pcpB gene in situ also in the chlorophenol plume of another historically polluted aquifer with no remediation history. Hotspot groundwater HCPs from this site were degraded efficiently during a 3-week microcosm incubation with one-time aeration but no other additives: from 5400 µg L-1 to 1200 µg L-1 and to 200 µg L-1 in lightly and fully aerated microcosms, respectively, coupled with up to 2400% enrichment of the pcpB gene. Accumulation of lower-chlorinated metabolites was observed in neither in situ remediation nor microcosms, supporting the assumption that HCP removal was due to the aerobic degradation pathway where the first step limits the mineralisation rate. Our results demonstrate that bacteria capable of aerobic mineralisation of xenobiotic pentachlorophenol and tetrachlorophenol can be present at long-term polluted groundwater sites, making bioremediation by simple aeration a viable and economically attractive alternative.

Sample Preservation, DNA or RNA Extraction and Data Analysis for High-Throughput Phytoplankton Community Sequencing.

Phytoplankton is the basis for aquatic food webs and mirrors the water quality. Conventionally, phytoplankton analysis has been done using time consuming and partly subjective microscopic observations, but next generation sequencing (NGS) technologies provide promising potential for rapid automated examination of environmental samples. Because many phytoplankton species have tough cell walls, methods for cell lysis and DNA or RNA isolation need to be efficient to allow unbiased nucleic acid retrieval. Here, we analyzed how two phytoplankton preservation methods, three commercial DNA extraction kits and their improvements, three RNA extraction methods, and two data analysis procedures affected the results of the NGS analysis. A mock community was pooled from phytoplankton species with variation in nucleus size and cell wall hardness. Although the study showed potential for studying Lugol-preserved sample collections, it demonstrated critical challenges in the DNA-based phytoplankton analysis in overall. The 18S rRNA gene sequencing output was highly affected by the variation in the rRNA gene copy numbers per cell, while sample preservation and nucleic acid extraction methods formed another source of variation. At the top, sequence-specific variation in the data quality introduced unexpected bioinformatics bias when the sliding-window method was used for the quality trimming of the Ion Torrent data. While DNA-based analyses did not correlate with biomasses or cell numbers of the mock community, rRNA-based analyses were less affected by different RNA extraction procedures and had better match with the biomasses, dry weight and carbon contents, and are therefore recommended for quantitative phytoplankton analyses.

Enrichment, isolation and biodegradation potential of psychrotolerant polychlorinated-biphenyl degrading bacteria from the Kongsfjorden (Svalbard Islands, High Arctic Norway).

Persistent organic pollutants (POPs), such as polychlorinated biphenyls (PCBs), have been detected in abiotic Arctic matrices: surface sediments and seawater from coastal areas in the Kongsfjorden were collected and analyzed. Levels of PCBs varied depending on the sampling site. Total PCB concentrations were between 11.63 (site C2W) and 27.69pgl-1 (site AW). These levels were comparable to those reported previously in lake sediments from the northern Svalbard. The occurrence and biodegradation potential of cold-adapted PCB-oxidizing bacteria in seawater and sediment along the fjord was also evaluated. After enrichment with biphenyl, 246 isolates were obtained with 45 of them that were able to grow in the presence of the PCB mixture Aroclor 1242, as the sole carbon source. The catabolic gene bphA was harbored by 17 isolates with affiliates to the genera Algoriphagus, Devosia and Salinibacterium that have been never reported as able to utilize PCBs, thus deserving further investigation. The total removal of Aroclor 1242 and selected PCB congeners was evaluated at 4 and 15°C for eight bphA-harboring isolates and Gelidibacter sp. DS-10. With few exceptions, tested strains showed greater efficiency at 15 than at 4°C. Isolates were able to reduce most chromatographic peaks by >50%, with some di- and trichlorobiphenyls that were quite totally removed (>90%).