Global Patterns of Metal and Other Element Enrichment in Bog and Fen Peatlands.

Peatlands are found on all continents, covering 3% of the global land area. However, the spatial extent and causes of metal enrichment in peatlands is understudied and no attempt has been made to evaluate global patterns of metal enrichment in bog and fen peatlands, despite that certain metals and rare earth elements (REE) arise from anthropogenic sources. We analyzed 368 peat cores sampled in 16 countries across five continents and measured metal and other element concentrations at three depths down to 70 cm as well as estimated cumulative atmospheric S deposition (1850-2009) for each site. Sites were assigned to one of three distinct broadly recognized peatland categories (bog, poor fen, and intermediate-to-moderately rich fen) that varied primarily along a pH gradient. Metal concentrations differed among peatland types, with intermediate-to-moderately rich fens demonstrating the highest concentrations of most metals. Median enrichment factors (EFs; a metric comparing natural and anthropogenic metal deposition) for individual metals were similar among bogs and fens (all groups), with metals likely to be influenced by anthropogenic sources (As, Cd, Co, Cu, Hg, Pb, and Sb) demonstrating median enrichment factors (EFs) > 1.5. Additionally, mean EFs were substantially higher than median values, and the positive correlation (< 0.40) with estimated cumulative atmospheric S deposition, confirmed some level of anthropogenic influence of all pollutant metals except for Hg that was unrelated to S deposition. Contrary to expectations, high EFs were not restricted to pollutant metals, with Mn, K and Rb all exhibiting elevated median EFs that were in the same range as pollutant metals likely due to peatland biogeochemical processes leading to enrichment of these nutrients in surface soil horizons. The global patterns of metal enrichment in bogs and fens identified in this study underscore the importance of these peatlands as environmental archives of metal deposition, but also illustrates that biogeochemical processes can enrich metals in surface peat and EFs alone do not necessarily indicate atmospheric contamination.

Soil carbon pools and fluxes following the regreening of a mining and smelting degraded landscape.

Increasing forest cover by regreening mining and smelting degraded landscapes provides an opportunity for global carbon (C) sequestration, however, the reported effects of regreening on soil C processes are mixed. One of the world's largest regreening programs is in the City of Greater Sudbury, Canada and has been ongoing since 1978. Prior to regreening, soils in the City of Greater Sudbury area were highly eroded, acidic, rich in metals, and poor in nutrients. This study used a chronosequence approach to investigate how forest soil C pools and fluxes have changed with stand age in highly "eroded" sites with minimal soil cover (n = 6) and "stable" sites covered by soil (n = 6). Encouragingly, the relationship between stand age and soil C processes (litterfall, litter decomposition, soil respiration, fine root growth) at both stable and eroded sites were comparable to observations reported for jack pine (Pinus banksiana Lamb.) and red pine (Pinus resinosa Ait.) plantations that have not been subject to over a century of industrial impacts. There was a strong "home-field advantage" for local decomposers, where litter decomposition rates were higher using a site-specific pine litter compared with a common pine litter. Higher soil respiration at eroded sites was linked to higher soil temperature, likely because of a more open tree canopy. Forest floor C pools increased with stand age while mineral soil C and aggregate C concentrations decreased with stand age. This loss of soil C is small relative to the substantial increases in aboveground tree and forest floor C pools, leading to a sizeable increase in total ecosystem C pools following regreening.

Recovery of Smelter-Impacted Peat and Sphagnum Moss: a Microbial Perspective.

Peatlands store approximately one-half of terrestrial soil carbon and one-tenth of non-glacial freshwater. Some of these important ecosystems are located near heavy metal emitting smelters. To improve the understanding of smelter impacts and potential recovery after initial pollution controls in the 1970s (roughly 50 years of potential recovery), we sampled peatlands along a distance gradient of 134 km from a smelter in Sudbury, Ontario, Canada, an area with over a century of nickel (Ni) and copper (Cu) mining activity. This work is aimed at evaluating potential shifts in bacterial and archaeal community structures in Sphagnum moss and its underlying peat within smelter-impacted poor fens. In peat, total Ni and Cu concentrations were higher (0.062-0.067 and 0.110-0.208 mg/g, respectively) at sites close to the smelter and exponentially dropped with distance from the smelter. This exponential decrease in Ni concentrations was also observed in Sphagnum. 16S rDNA amplicon sequencing showed that peat and Sphagnum moss host distinct microbiomes with peat accommodating a more diverse community structure. The microbiomes of Sphagnum were dominated by Proteobacteria (62.5%), followed by Acidobacteria (11.9%), with no observable trends with distance from the smelter. Dominance of Acidobacteria (32.4%) and Proteobacteria (29.6%) in peat was reported across all sites. No drift in taxonomy was seen across the distance gradient or from the reference sites, suggesting a potential microbiome recovery toward that of the reference peatlands microbiomes after decades of pollution controls. These results advance the understanding of peat and Sphagnum moss microbiomes, as well as depict the sensitivities and the resilience of peatland ecosystems.

How arsenic contamination influences downslope wetland plant and microbial community structure and function.

Mine tailings are prevalent worldwide and can adversely impact adjacent ecosystems, including wetlands. This study investigated the impact of gold (Au) mine tailings contamination on peatland soil and pore water geochemistry, vegetation and microbial communities, and microbial carbon (C) cycling. Maximum arsenic (As) concentrations in peat and pore water reached 20,137 mg kg-1 and 16,730 µg L-1, respectively, but decreased by two orders of magnitude along a 128 m gradient extending from the tailings into the wetland. Carbon and other macronutrient (N, P, K) concentrations in peat and pore water significantly increased with distance from contamination. Relative percent cover and species richness of vascular and non-vascular plants significantly increased with distance into the wetland, with higher non-vascular richness being found at intermediate distances before transitioning to a vascular plant dominated community. Bacterial and archaeal community composition exhibited a decreased proportion of members of the phylum Acidobacteria (notably of the order Acidobacteriales) and increased diversity and richness of methanogens across a larger range of orders farther from the tailings source, an indication of microbial C-cycling potential. Consistent with changes in microbial communities, in vitro microbial CH4 production potential significantly increased with distance from the contaminant source. This study demonstrates both the profound negative impact that metalliferous tailings contamination can have on above and belowground communities in peatlands, and the value of wetland preservation and restoration.

Variation in carbon and nitrogen concentrations among peatland categories at the global scale.

Peatlands account for 15 to 30% of the world's soil carbon (C) stock and are important controls over global nitrogen (N) cycles. However, C and N concentrations are known to vary among peatlands contributing to the uncertainty of global C inventories, but there are few global studies that relate peatland classification to peat chemistry. We analyzed 436 peat cores sampled in 24 countries across six continents and measured C, N, and organic matter (OM) content at three depths down to 70 cm. Sites were distinguished between northern (387) and tropical (49) peatlands and assigned to one of six distinct broadly recognized peatland categories that vary primarily along a pH gradient. Peat C and N concentrations, OM content, and C:N ratios differed significantly among peatland categories, but few differences in chemistry with depth were found within each category. Across all peatlands C and N concentrations in the 10-20 cm layer, were 440 ± 85.1 g kg-1 and 13.9 ± 7.4 g kg-1, with an average C:N ratio of 30.1 ± 20.8. Among peatland categories, median C concentrations were highest in bogs, poor fens and tropical swamps (446-532 g kg-1) and lowest in intermediate and extremely rich fens (375-414 g kg-1). The C:OM ratio in peat was similar across most peatland categories, except in deeper samples from ombrotrophic tropical peat swamps that were higher than other peatlands categories. Peat N concentrations and C:N ratios varied approximately two-fold among peatland categories and N concentrations tended to be higher (and C:N lower) in intermediate fens compared with other peatland types. This study reports on a unique data set and demonstrates that differences in peat C and OM concentrations among broadly classified peatland categories are predictable, which can aid future studies that use land cover assessments to refine global peatland C and N stocks.

Viruses direct carbon cycling in lake sediments under global change.

Global change is altering the vast amount of carbon cycled by microbes between land and freshwater, but how viruses mediate this process is poorly understood. Here, we show that viruses direct carbon cycling in lake sediments, and these impacts intensify with future changes in water clarity and terrestrial organic matter (tOM) inputs. Using experimental tOM gradients within sediments of a clear and a dark boreal lake, we identified 156 viral operational taxonomic units (vOTUs), of which 21% strongly increased with abundances of key bacteria and archaea, identified via metagenome-assembled genomes (MAGs). MAGs included the most abundant prokaryotes, which were themselves associated with dissolved organic matter (DOM) composition and greenhouse gas (GHG) concentrations. Increased abundances of virus-like particles were separately associated with reduced bacterial metabolism and with shifts in DOM toward amino sugars, likely released by cell lysis rather than higher molecular mass compounds accumulating from reduced tOM degradation. An additional 9.6% of vOTUs harbored auxiliary metabolic genes associated with DOM and GHGs. Taken together, these different effects on host dynamics and metabolism can explain why abundances of vOTUs rather than MAGs were better overall predictors of carbon cycling. Future increases in tOM quantity, but not quality, will change viral composition and function with consequences for DOM pools. Given their importance, viruses must now be explicitly considered in efforts to understand and predict the freshwater carbon cycle and its future under global environmental change.

Dark stress for improved lipid quantity and quality in bioprospected acid-tolerant green microalgae.

The cost of microalgae cultivation is one of the largest limitations to achieving sustainable, large-scale microalgae production of commercially desirable lipids. Utilizing CO2 as a 'free' carbon source from waste industrial flue gas emissions can offer wide-ranging cost savings. However, these gas streams typically create acidic environments, in which most microalgae cannot survive due to the concentration of CO2 and the presence of other acidic gasses such as NO2 and SO2. To address this situation, we investigated growth of a mixed acid-tolerant green microalgal culture (91% dominated by a single Coccomyxa sp. taxon) bioprospected at pH 2.8 from an acid mine drainage impacted water body. The culture was grown at pH 2.5 and fed with a simulated flue gas containing 6% CO2 and 94% N2. On reaching the end of the exponential growth phase, the culture was exposed to either continued light-dark cycle conditions or continual dark conditions. After three days in the dark, the biomass consisted of 28% of lipids, which was 42% higher than at the end of the exponential phase and 55% higher than the maximum lipid content achieved under light/dark conditions. The stress caused by being continually in the dark also favoured the production of omega-3 and omega-6 polyunsaturated fatty acids (PUFAs; 19.47% and 21.04%, respectively, after 7 days) compared to 7-days of light-dark treatment (1.94% and 9.53%, respectively) and showed an increase in nitrogen content (C:N ratio of 6.4) compared to light-dark treatment (C:N ratio of 11.9). The results of the research indicate that use of acid tolerant microalgae overcomes issues using flue gasses that will create an acidic environment and that applying dark stress is a low-cost stressor stimulates production of desirable dietary lipids.

Forest soil biotic communities show few responses to wood ash applications at multiple sites across Canada.

There is interest in utilizing wood ash as an amendment in forestry operations as a mechanism to return nutrients to soils that are removed during harvesting, with the added benefit of diverting this bioenergy waste material from landfill sites. Existing studies have not arrived at a consensus on what the effects of wood ash amendments are on soil biota. We collected forest soil samples from studies in managed forests across Canada that were amended with wood ash to evaluate the effects on arthropod, bacterial and fungal communities using metabarcoding of F230, 16S, 18S and ITS2 sequences as well as enzyme analyses to assess its effects on soil biotic function. Ash amendment did not result in consistent effects across sites, and those effects that were detected were small. Overall, this study suggests that ash amendment applied to managed forest systems in amounts (up to 20 Mg ha-1) applied across the 8 study sties had little to no detectable effects on soil biotic community structure or function. When effects were detected, they were small, and site-specific. These non-results support the application of wood ash to harvested forest sites to replace macronutrients (e.g., calcium) removed by logging operations, thereby diverting it from landfill sites, and potentially increasing stand productivity.

Methane production potential of pulp mill sludges: microbial community and substrate constraints.

Sludges from pulp and paper mills represent a major ecological and environmental cost, and anaerobic digestion represents a method of waste reduction and energy recovery for these mills. This study compared methane production potential and microbial communities across 11 primary- and biosludges from five pulp and paper mills using various mill processes. We measured methane production from sludges in anaerobic batch reactor experiments over 64 days. Sludges were incubated with and without added substrate to test for organic substrate limitation versus inhibition of methanogens. Initial microbial communities and changes to community composition were determined using Illumina MiSeq for metabarcoding of bacterial and archaeal 16S rRNA genes. Mean methane production potential varied greatly between sludges (0.002-79 mL CH4 g-1 TS). Among primary sludges, kraft mill sludge produced more methane than other mill types. For these other mills, biosludge produced more methane than primary sludge, which had evidence of methanogen inhibition. Microbial communities and diversity were influenced by the initial community composition, and high methane production was only seen in sludges with high diversity. A number of sludges innately produced substantial methane and may be targets for further modelling and larger scale testing of anaerobic digestion.

Native plants facilitate vegetation succession on amended and unamended mine tailings.

Facilitating the establishment of native pioneer plant species on mine tailings with inherent metal and/or acid tolerance is important to speed up natural succession at minimal cost, especially in remote areas where phytoremediation can be labor intensive. We investigated vegetation community dynamics after ∼48 years of succession along two legacy Ni-Cu mine tailings and waste rock deposits in the Sudbury Basin, Ontario, Canada with and without various site amendments (i.e. liming and fertilization) and planting. Metal/acid tolerant pioneer plants (Betula papyrifera, Populus tremuloides, Pohlia nutans) appeared to facilitate the establishment of less tolerant species. Conifers and nitrogen-fixers less tolerant to site conditions were planted at the fully amended (limed, fertilized, planted) mine tailings site in the 1970s, but conifers were not propagating at the site or facilitating understory succession. The planted nitrogen-fixing leguminous species Lotus corniculatus was, however, associated with increased diversity. These findings have implications for long-term reclamation strategies in acidic mine waste deposits utilizing native species, as primary colonizing tree species are only recently emerging as candidates for phytoremediation. Novelty statement The potential for native species to act as facilitators for vegetation colonization has rarely been investigated on tailings, despite wide use in remediation of less toxic sites. This study provides a retrospective of over 40 years of plant growth following initial treatment of toxic tailings. We observed that regardless of tailings geochemical conditions, acid/metal tolerant pioneer plants were facilitating ecological succession on acidic Ni-Cu mine tailings sites.

Beyond the usual suspects: methanogenic communities in eastern North American peatlands are also influenced by nickel and copper concentrations.

Peatlands both accumulate carbon and release methane, but their broad range in environmental conditions means that the diversity of microorganisms responsible for carbon cycling is still uncertain. Here, we describe a community analysis of methanogenic archaea responsible for methane production in 17 peatlands from 36 to 53 N latitude across the eastern half of North America, including three metal-contaminated sites. Methanogenic community structure was analysed through Illumina amplicon sequencing of the mcrA gene. Whether metal-contaminated sites were included or not, metal concentrations in peat were a primary driver of methanogenic community composition, particularly nickel, a trace element required in the F430 cofactor in methyl-coenzyme M reductase that is also toxic at high concentrations. Copper was also a strong predictor, likely due to inhibition at toxic levels and/or to cooccurrence with nickel, since copper enzymes are not known to be present in anaerobic archaea. The methanogenic groups Methanocellales and Methanosarcinales were prevalent in peatlands with low nickel concentrations, while Methanomicrobiales and Methanomassiliicoccales were abundant in peatlands with higher nickel concentrations. Results suggest that peat-associated trace metals are predictors of methanogenic communities in peatlands.

Temperature, moisture and freeze-thaw controls on CO2 production in soil incubations from northern peatlands.

Peat accumulation in high latitude wetlands represents a natural long-term carbon sink, resulting from the cumulative excess of growing season net ecosystem production over non-growing season (NGS) net mineralization in soils. With high latitudes experiencing warming at a faster pace than the global average, especially during the NGS, a major concern is that enhanced mineralization of soil organic carbon will steadily increase CO2 emissions from northern peatlands. In this study, we conducted laboratory incubations with soils from boreal and temperate peatlands across Canada. Peat soils were pretreated for different soil moisture levels, and CO2 production rates were measured at 12 sequential temperatures, covering a range from - 10 to + 35 °C including one freeze-thaw event. On average, the CO2 production rates in the boreal peat samples increased more sharply with temperature than in the temperate peat samples. For same temperature, optimum soil moisture levels for CO2 production were higher in the peat samples from more flooded sites. However, standard reaction kinetics (e.g., Q10 temperature coefficient and Arrhenius equation) failed to account for the apparent lack of temperature dependence of CO2 production rates measured below 0 °C, and a sudden increase after a freezing event. Thus, we caution against using the simple kinetic expressions to represent the CO2 emissions from northern peatlands, especially regarding the long NGS period with multiple soil freeze and thaw events.

Selection and re-acclimation of bioprospected acid-tolerant green microalgae suitable for growth at low pH.

For mass culture of photosynthetic green microalgae, industrial flue gases can represent a low-cost resource of CO2. However, flue gases are often avoided, because they often also contain high levels of SO2 and/or NO2, which cause significant acidification of media to below pH 3 due to production of sulfuric and nitric acid. This creates an unsuitable environment for the neutrophilic microalgae commonly used in large-scale commercial production. To address this issue, we have looked at selecting acid-tolerant microalgae via growth at pH 2.5 carried out with samples bioprospected from an active smelter site. Of the eight wild samples collected, one consisting mainly of Coccomyxa sp. grew at pH 2.5 and achieved a density of 640 mg L-1. Furthermore, three previously bioprospected green microalgae from acidic waters (pH 3-4.5) near abandoned mine sites were also re-acclimated down to their in-situ pH environment after approximately 4 years spent at neutral pH. Of those three, an axenic culture of Coccomyxa sp. was the most successful at re-acclimating and achieved the highest density of 293.1 mg L-1 and maximum daily productivity of 38.8 mg L-1 day-1 at pH 3. Re-acclimation of acid-tolerant species is, therefore, achievable when directly placed at their original pH, but gradual reduction in pH is recommended to give the cells time to acclimate.

Blended municipal compost and biosolids materials for mine reclamation: Long-term field studies to explore metal mobility, soil fertility and microbial communities.

Application of stable soil amendments is often the key to successful phytostabilization and rehabilitation of mine tailings, and microbial guilds are primary drivers of many geochemical processes promoted by these amendments. Field studies were set up at a tailings management area near Sudbury, Ontario to examine performance of blends of lime stabilized municipal biosolids and compost at nine different rates over thick (1 m) municipal compost covers planted with agricultural crops. Based on biogeochemical variability of the substrates four and ten years after application of the initial compost cover, the experimental plots could be classified into three categories: "Low" rate (0-100 t ha-1 biosolids), "Medium" rate (200-800 t ha-1), and "High" rate (1600-3200 t ha-1) treatments. The addition of biosolids materials to the thick compost cover at rates higher than 100 t ha-1 significantly reduced C:N ratio of the substrates, available phosphorus, and some of the nutrient cations, while notably increasing inorganic carbon and the potential solubility of Ni and Cu. This suggests that increasing biosolids application rates may not equivalently ameliorate soil quality and geochemical stability. Correspondingly, microbial communities were altered by biosolids additions, further intensifying the negative impacts of biosolids on long-term efficiency of the initial compost cover. Abundance of cellulose, hemicellulose, and lignocellulose decomposers (as key drivers of mineralization and humification) was significantly reduced by "Medium" and "High" rate treatments. Most DNA sequences with high affinity to denitrifiers were detected in "High" rate treatments where geochemical conditions were optimal for higher microbial denitrification activities. These findings have implications for improving the long-term efficiency of reclamation and environmental management programs in mine tailings of northern temperate climates.

Methanogenic archaea in peatlands.

Methane emission feedbacks in wetlands are predicted to influence global climate under climate change and other anthropogenic stressors. Herein, we review the taxonomy and physiological ecology of the microorganisms responsible for methane production in peatlands. Common in peat soils are five of the eight described orders of methanogens spanning three phyla (Euryarchaeota, Halobacterota and Thermoplasmatota). The phylogenetic affiliation of sequences found in peat suggest that members of the thus-far-uncultivated group Candidatus Bathyarchaeota (representing a fourth phylum) may be involved in methane cycling, either anaerobic oxidation of methane and/or methanogenesis, as at least a few organisms within this group contain the essential gene, mcrA, according to metagenomic data. Methanogens in peatlands are notoriously challenging to enrich and isolate; thus, much remains unknown about their physiology and how methanogen communities will respond to environmental changes. Consistent patterns of changes in methanogen communities have been reported across studies in permafrost peatland thaw where the resulting degraded feature is thermokarst. However much remains to be understood regarding methanogen community feedbacks to altered hydrology and warming in other contexts, enhanced atmospheric pollution (N, S and metals) loading and direct anthropogenic disturbances to peatlands like drainage, horticultural peat extraction, forestry and agriculture, as well as post-disturbance reclamation.

Lake characteristics influence how methanogens in littoral sediments respond to terrestrial litter inputs.

Shallow lake sediments harbor methanogen communities that are responsible for large amounts of CH4 flux to the atmosphere. These communities play a major role in degrading in-fluxed terrestrial organic matter (t-OM)-much of which settles in shallow near-shore sediments. Little work has examined how sediment methanogens are affected by the quantity and quality of t-OM, and the physicochemical factors that shape their community. Here, we filled mesocosms with artificial lake sediments amended with different ratios and concentrations of coniferous and deciduous tree litter. We installed them in three boreal lakes near Sudbury, Canada that varied in trophic status and water clarity. We found that higher endogenous nutrient concentrations led to greater CH4 production when sediment solar irradiance was similar, but high irradiance of sediments also led to higher CH4 concentrations regardless of nutrient concentrations, possibly due to photooxidation of t-OM. Sediments with t-OM had overall higher CH4 concentrations than controls that had no t-OM, but there were no significant differences in CH4 concentrations with different t-OM compositions or increasing concentrations over 25%. Differences among lakes also explained variation in methanogen community structure, whereas t-OM treatments did not. Therefore, lake characteristics are important modulators of methanogen communities fueled by t-OM.

Edaphic factors influencing vegetation colonization and encroachment on arsenical gold mine tailings near Sudbury, Ontario.

Mine tailings are found worldwide and can have significant impacts on ecosystem and human health. In this study, natural vegetation patterns on arsenical (As) gold (Au) mine tailings located in Sudbury, Ontario were assessed using transects located at the edge of the tailings and on the tailings. Vegetation communities were significantly different between the edge and open tailings areas of the site. Arsenic concentrations in both areas were extremely variable (from 285-17,567 mg/kg) but were not significantly correlated with vegetation diversity at the site. Nutrients (carbon (C), phosphorus (P)) and organic matter concentrations were associated with higher diversity and with the presence of climax vegetation on the tailings, but there were no significant relationships between tailings chemistry and vegetation indices on the edge. Encroachment onto the tailings from the edge occurred in conventional succession patterns, with a clear gradient from grasses (Agrostis gigantea) to trees such as Picea glauca. On the tailings, a nucleation pattern was visible, distinct from conventional succession. Trees and shrubs such as Betula papyrifera and Diervilla lonicera were associated with higher diversity and higher nutrient concentrations in the underlying tailings, whereas grasses such as A. gigantea were not. We concluded that at all areas of the site, vegetation - particularly trees - was facilitating amelioration of the underlying tailings. Despite high concentrations of As, nutrients appeared to have a greater influence than metals on vegetation diversity.

Peatland Microbial Community Composition Is Driven by a Natural Climate Gradient.

Peatlands are important players in climate change-biosphere feedbacks via long-term net carbon (C) accumulation in soil organic matter and as potential net C sources including the potent greenhouse gas methane (CH4). Interactions of climate, site-hydrology, plant community, and groundwater chemical factors influence peatland development and functioning, including C dioxide (CO2) and CH4 fluxes, but the role of microbial community composition is not well understood. To assess microbial functional and taxonomic dissimilarities, we used high throughput sequencing of the small subunit ribosomal DNA (SSU rDNA) to determine bacterial and archaeal community composition in soils from twenty North American peatlands. Targeted DNA metabarcoding showed that although Proteobacteria, Acidobacteria, and Actinobacteria were the dominant phyla on average, intermediate and rich fens hosted greater diversity and taxonomic richness, as well as an array of candidate phyla when compared with acidic and nutrient-poor poor fens and bogs. Moreover, pH was revealed to be the strongest predictor of microbial community structure across sites. Predictive metagenome content (PICRUSt) showed increases in specific genes, such as purine/pyrimidine and amino-acid metabolism in mid-latitude peatlands from 38 to 45° N, suggesting a shift toward utilization of microbial biomass over utilization of initial plant biomass in these microbial communities. Overall, there appears to be noticeable differences in community structure between peatland classes, as well as differences in microbial metabolic activity between latitudes. These findings are in line with a predicted increase in the decomposition and accelerated C turnover, and suggest that peatlands north of 37° latitude may be particularly vulnerable to climate change.

Variations in terrestrial arthropod DNA metabarcoding methods recovers robust beta diversity but variable richness and site indicators.

Terrestrial arthropod fauna have been suggested as a key indicator of ecological integrity in forest systems. Because phenotypic identification is expert-limited, a shift towards DNA metabarcoding could improve scalability and democratize the use of forest floor arthropods for biomonitoring applications. The objective of this study was to establish the level of field sampling and DNA extraction replication needed for arthropod biodiversity assessments from soil. Processing 15 individually collected soil samples recovered significantly higher median richness (488-614 sequence variants) than pooling the same number of samples (165-191 sequence variants) prior to DNA extraction, and we found no significant richness differences when using 1 or 3 pooled DNA extractions. Beta diversity was robust to changes in methodological regimes. Though our ability to identify taxa to species rank was limited, we were able to use arthropod COI metabarcodes from forest soil to assess richness, distinguish among sites, and recover site indicators based on unnamed exact sequence variants. Our results highlight the need to continue DNA barcoding local taxa during COI metabarcoding studies to help build reference databases. All together, these sampling considerations support the use of soil arthropod COI metabarcoding as a scalable method for biomonitoring.

Enrichment of peat yields novel methanogens: approaches for obtaining uncultured organisms in the age of rapid sequencing.

Methanogens are among the oldest forms of life on Earth and are detectable in a wide range of environments, but our knowledge of their overall diversity and functioning is limited. Peatlands in particular host a broad range of methanogens that contribute large amounts of methane to the atmosphere, but are largely under-represented in pure cultures. Here, we anaerobically enriched peat with common growth substrates, supplements and antibiotics to identifying novel methanogen sequences and potential growth conditions. Over 3 years, we obtained 28 new mcrA sequences from taxa that have remained previously uncultured and undescribed beyond distantly related clones or sequences detected in environmental samples. Evidence suggests that the novel methanogens, representing five of the seven known orders, were capable of growing on H2 as well as acetate and at temperatures ranging from 6 to ca. 22°C. Methods involving the use of ampicillin proved useful, although obtaining high methane production in the absence of H2 was difficult. Our results also indicate that many methanogens may rely on bacterial symbionts (commonly Clostridium spp.). Such enrichment approaches represent a useful intermediary between maker-gene detection and isolation, allowing us to broaden our understanding of methanogen physiological ecology while potentially providing valuable sequence data.