Rapid on-site detection of harmful algal blooms: real-time cyanobacteria identification using Oxford Nanopore sequencing.

With the increasing occurrence and severity of cyanobacterial harmful algal blooms (cHAB) at the global scale, there is an urgent need for rapid, accurate, accessible, and cost-effective detection tools. Here, we detail the RosHAB workflow, an innovative, in-the-field applicable genomics approach for real-time, early detection of cHAB outbreaks. We present how the proposed workflow offers consistent taxonomic identification of water samples in comparison to traditional microscopic analyses in a few hours and discuss how the generated data can be used to deepen our understanding on cyanobacteria ecology and forecast HABs events. In parallel, processed water samples will be used to iteratively build the International cyanobacterial toxin database (ICYATOX; http://icyatox.ibis.ulaval.ca) containing the analysis of novel cyanobacterial genomes, including phenomics and genomics metadata. Ultimately, RosHAB will (1) improve the accuracy of on-site rapid diagnostics, (2) standardize genomic procedures in the field, (3) facilitate these genomics procedures for non-scientific personnel, and (4) identify prognostic markers for evidence-based decisions in HABs surveillance.

Microbial degradation of naphthenic acids using constructed wetland treatment systems: metabolic and genomic insights for improved bioremediation of process-affected water.

Naphthenic acids (NAs) are a complex mixture of organic compounds released during bitumen extraction from mined oil sands that are important contaminants of oil sands process-affected water (OSPW). NAs can be toxic to aquatic organisms and, therefore, are a main target compound for OSPW. The ability of microorganisms to degrade NAs can be exploited for bioremediation of OSPW using constructed wetland treatment systems (CWTS), which represent a possible low energy and low-cost option for scalable in situ NA removal. Recent advances in genomics and analytical chemistry have provided insights into a better understanding of the metabolic pathways and genes involved in NA degradation. Here, we discuss the ecology of microbial NA degradation with a focus on CWTS and summarize the current knowledge related to the metabolic pathways and genes used by microorganisms to degrade NAs. Evidence to date suggests that NAs are mostly degraded aerobically through ring cleavage via the beta-oxidation pathway, which can be combined with other steps such as aromatization, alpha-oxidation, omega-oxidation, or activation as coenzyme A (CoA) thioesters. Anaerobic NA degradation has also been reported via the production of benzoyl-CoA as an intermediate and/or through the involvement of methanogens or nitrate, sulfate, and iron reducers. Furthermore, we discuss how genomic, statistical, and modeling tools can assist in the development of improved bioremediation practices.

Sources of exposure to lead in Arctic and subarctic regions: a scoping review.

Understanding lead exposure pathways is a priority because of its ubiquitous presence in the environment as well as the potential health risks. We aimed to identify potential lead sources and pathways of lead exposure, including long-range transport, and the magnitude of exposure in Arctic and subarctic communities. A scoping review strategy and screening approach was used to search literature from January 2000 to December 2020. A total of 228 academic and grey literature references were synthesised. The majority of these studies (54%) were from Canada. Indigenous people in Arctic and subarctic communities in Canada had higher levels of lead than the rest of Canada. The majority of studies in all Arctic countries reported at least some individuals above the level of concern. Lead levels were influenced by a number of factors including using lead ammunition to harvest traditional food and living in close proximity to mines. Lead levels in water, soil, and sediment were generally low. Literature showed the possibility of long-range transport via migratory birds. Household lead sources included lead-based paint, dust, or tap water. This literature review will help to inform management strategies for communities, researchers, and governments, with the aim of decreasing lead exposure in northern regions.

Spatio-temporal connectivity of the aquatic microbiome associated with cyanobacterial blooms along a Great Lake riverine-lacustrine continuum.

Lake Erie is subject to recurring events of cyanobacterial harmful algal blooms (cHABs), but measures of nutrients and total phytoplankton biomass seem to be poor predictors of cHABs when taken individually. A more integrated approach at the watershed scale may improve our understanding of the conditions that lead to bloom formation, such as assessing the physico-chemical and biological factors that influence the lake microbial community, as well as identifying the linkages between Lake Erie and the surrounding watershed. Within the scope of the Government of Canada's Genomics Research and Development Initiative (GRDI) Ecobiomics project, we used high-throughput sequencing of the 16S rRNA gene to characterize the spatio-temporal variability of the aquatic microbiome in the Thames River-Lake St. Clair-Detroit River-Lake Erie aquatic corridor. We found that the aquatic microbiome was structured along the flow path and influenced mainly by higher nutrient concentrations in the Thames River, and higher temperature and pH downstream in Lake St. Clair and Lake Erie. The same dominant bacterial phyla were detected along the water continuum, changing only in relative abundance. At finer taxonomical level, however, there was a clear shift in the cyanobacterial community, with Planktothrix dominating in the Thames River and Microcystis and Synechococcus in Lake St. Clair and Lake Erie. Mantel correlations highlighted the importance of geographic distance in shaping the microbial community structure. The fact that a high proportion of microbial sequences found in the Western Basin of Lake Erie were also identified in the Thames River, indicated a high degree of connectivity and dispersal within the system, where mass effect induced by passive transport play an important role in microbial community assembly. Nevertheless, some cyanobacterial amplicon sequence variants (ASVs) related to Microcystis, representing less than 0.1% of relative abundance in the upstream Thames River, became dominant in Lake St. Clair and Erie, suggesting selection of those ASVs based on the lake conditions. Their extremely low relative abundances in the Thames suggest additional sources are likely to contribute to the rapid development of summer and fall blooms in the Western Basin of Lake Erie. Collectively, these results, which can be applied to other watersheds, improve our understanding of the factors influencing aquatic microbial community assembly and provide new perspectives on how to better understand the occurrence of cHABs in Lake Erie and elsewhere.

Structural shifts in sea ice prokaryotic communities across a salinity gradient in the subarctic.

Current knowledge of the processes that shape prokaryotic community assembly in sea ice across polar ecosystems is scarce. Here, we coupled culture-dependent (bacterial isolation on R2A medium) and culture-independent (high-throughput 16S rRNA gene sequencing) approaches to provide the first comprehensive assessment of prokaryotic communities in the late winter ice and its underlying water along a natural salinity gradient in coastal Hudson Bay, an iconic cryo-environment that marks the ecological transition between Canadian Subarctic and Arctic biomes. We found that prokaryotic community assembly processes in the ice were less selective at low salinity since typical freshwater taxa such as Frankiales, Burkholderiales, and Chitinophagales dominated both the ice and its underlying water. In contrast, there were sharp shifts in community structure between the ice and underlying water samples at sites with higher salinity, with the orders Alteromonadales and Flavobacteriales dominating the ice, while the abovementioned freshwater taxa dominated the underlying water communities. Moreover, primary producers including Cyanobium (Cyanobacteria, Synechococcales) may play a role in shaping the ice communities and were accompanied by known Planctomycetes and Verrucomicrobiae taxa. Culture-dependent analyses showed that the ice contained pigment-producing psychrotolerant or psychrophilic bacteria from the phyla Proteobacteria, Actinobacteriota, and Bacteroidota, likely favored by the combination of low temperatures and the seasonal increase in sunlight. Our findings suggest that salinity, photosynthesis and dissolved organic matter are the main drivers of prokaryotic community structure in the late winter ice of coastal Hudson Bay, the ecosystem with the fastest sea ice loss rate in the Canadian North.

Frogs Respond to Commercial Formulations of the Biopesticide Bacillus thuringiensis var. israelensis, Especially Their Intestine Microbiota.

It is generally believed that Bacillus thuringiensis var. israelensis (Bti) biopesticides are harmless to non-target organisms; however, new research shows controversial results. We exposed acutely and chronicallyLithobates sylvaticusandAnaxyrus americanus tadpoles until metamorphic climax to VectoBac 200G (granules) and VectoBac 1200L (aqueous suspension) at 300-20,000 ITU/L covering field-relevant concentrations and higher. The data show that the exposure parameters tested did not affect significantly the survival, total length, total weight, hepatosomatic index, gonadosomatic index, the expression of genes of interest (i.e., related to xenobiotic exposure, oxidative stress, and metamorphosis), and the intestine tissue layer detachment ofL. sylvaticusandA. americanus in a concentration-response pattern. In contrast, VectoBac 200G significantly increased the median time to metamorphosis ofL. sylvaticus tadpoles by up to 3.5 days and decreased the median by up to 1 day inA. americanus. VectoBac 1200L significantly increased the median time to metamorphosis ofL. sylvaticusandA. americanustadpoles by up to 4.5 days. Also, the exposure to VectoBac 200G and 1200L altered the intestine bacterial community composition inA. americanus at application rates recommended by the manufacturer, which led to an increase in the relative abundance of Verrucomicrobia, Firmicutes, Bacteroidetes, and Actinobacteria. Changes in the intestine microbiota might impact the fitness of individuals, including the susceptibility to parasitic infections. Our results indicate that the effect of Bti commercial products is limited; however, we recommend that Bti-spraying activities in amphibian-rich ecosystems should be kept minimal until there is more conclusive research to assess if the changes in the time to metamorphosis and microbiota can lead to negative outcomes in amphibian populations and, eventually, the functioning of ecosystems.

The Ecobiomics project: Advancing metagenomics assessment of soil health and freshwater quality in Canada.

Transformative advances in metagenomics are providing an unprecedented ability to characterize the enormous diversity of microorganisms and invertebrates sustaining soil health and water quality. These advances are enabling a better recognition of the ecological linkages between soil and water, and the biodiversity exchanges between these two reservoirs. They are also providing new perspectives for understanding microorganisms and invertebrates as part of interacting communities (i.e. microbiomes and zoobiomes), and considering plants, animals, and humans as holobionts comprised of their own cells as well as diverse microorganisms and invertebrates often acquired from soil and water. The Government of Canada's Genomics Research and Development Initiative (GRDI) launched the Ecobiomics Project to coordinate metagenomics capacity building across federal departments, and to apply metagenomics to better characterize microbial and invertebrate biodiversity for advancing environmental assessment, monitoring, and remediation activities. The Project has adopted standard methods for soil, water, and invertebrate sampling, collection and provenance of metadata, and nucleic acid extraction. High-throughput sequencing is located at a centralized sequencing facility. A centralized Bioinformatics Platform was established to enable a novel government-wide approach to harmonize metagenomics data collection, storage and bioinformatics analyses. Sixteen research projects were initiated under Soil Microbiome, Aquatic Microbiome, and Invertebrate Zoobiome Themes. Genomic observatories were established at long-term environmental monitoring sites for providing more comprehensive biodiversity reference points to assess environmental change.

Microbial connectivity and sorting in a High Arctic watershed.

Aquatic ecosystems in the High Arctic are facing unprecedented changes as a result of global warming effects on the cryosphere. Snow pack is a central feature of northern landscapes, but the snow microbiome and its microbial connectivity to adjacent and downstream habitats have been little explored. To evaluate these aspects, we sampled along a hydrologic continuum at Ward Hunt Lake (latitude 83°N) in the Canadian High Arctic, from snow banks, water tracks in the permafrost catchment, the upper and lower strata of the lake, and the lake outlet and its coastal marine mixing zone. The microbial communities were analyzed by high-throughput sequencing of 16 and 18S rRNA to determine the composition of potentially active Bacteria, Archaea and microbial Eukarya. Each habitat had distinct microbial assemblages, with highest species richness in the subsurface water tracks that connected the melting snow to the lake. However, up to 30% of phylotypes were shared along the hydrologic continuum, showing that many taxa originating from the snow can remain in the active fraction of downstream microbiomes. The results imply that changes in snowfall associated with climate warming will affect microbial community structure throughout all spatially connected habitats within snow-fed polar ecosystems.

Diversity and potential activity of methanotrophs in high methane-emitting permafrost thaw ponds.

Lakes and ponds derived from thawing permafrost are strong emitters of carbon dioxide and methane to the atmosphere, but little is known about the methane oxidation processes in these waters. Here we investigated the distribution and potential activity of aerobic methanotrophic bacteria in thaw ponds in two types of eroding permafrost landscapes in subarctic Québec: peatlands and mineral soils. We hypothesized that methanotrophic community composition and potential activity differ regionally as a function of the landscape type and permafrost degradation stage, and locally as a function of depth-dependent oxygen conditions. Our analysis of pmoA transcripts by Illumina amplicon sequencing and quantitative PCR showed that the communities were composed of diverse and potentially active lineages. Type I methanotrophs, particularly Methylobacter, dominated all communities, however there was a clear taxonomic separation between the two landscape types, consistent with environmental control of community structure. In contrast, methanotrophic potential activity, measured by pmoA transcript concentrations, did not vary with landscape type, but correlated with conductivity, phosphorus and total suspended solids. Methanotrophic potential activity was also detected in low-oxygen bottom waters, where it was inversely correlated with methane concentrations, suggesting methane depletion by methanotrophs. Methanotrophs were present and potentially active throughout the water column regardless of oxygen concentration, and may therefore be resilient to future mixing and oxygenation regimes in the warming subarctic.

Comparison of Antioxidant Evaluation Assays for Investigating Antioxidative Activity of Gallic Acid and Its Alkyl Esters in Different Food Matrices.

The addition of antioxidants is one of the strategies to inhibit lipid oxidation, a major cause of lipid deterioration in foods leading to rancidity development and nutritional losses. However, several studies have been reported that conventional antioxidant assays, e.g., TPC, ABTS, FRAP, and ORAC could not predict antioxidant performance in several foods. This study aimed to investigate the performance of two recently developed assays, e.g., the conjugated autoxidizable triene (CAT) and the apolar radical-initiated conjugated autoxidizable triene (ApoCAT) assays to predict the antioxidant effectiveness of gallic acid and its esters in selected food models in comparison with the conventional antioxidant assays. The results indicated that the polarities of the antioxidants have a strong impact on antioxidant activities. In addition, different oxidant locations demonstrated by the CAT and ApoCAT assays influenced the overall antioxidant performances of the antioxidants with different polarities. To validate the predictability of the assays, the antioxidative performance of gallic acid and its alkyl esters was investigated in oil-in-water (O/W) emulsions, bulk soybean oils, and roasted peanuts as the lipid food models. The results showed that only the ApoCAT assay could be able to predict the antioxidative performances in O/W emulsions regardless of the antioxidant polarities. This study demonstrated that the relevance of antioxidant assays to food models was strongly dependent on physical similarities between the tested assays and the food structure matrices.

Contribution of different bacterial dispersal sources to lakes: Population and community effects in different seasons.

The diversity and composition of lake bacterial communities are driven by the interplay between local contemporary environmental conditions and dispersal of cells from the surroundings, i.e. the metacommunity. Still, a conceptual understanding of the relative importance of the two types of factors is lacking. For instance, it is unknown which sources of dispersal are most important and under which circumstances. Here, we investigated the seasonal variation in the importance of dispersal from different sources (mixing, precipitation, surface runoff and sediment resuspension) for lake bacterioplankton community and population dynamics. For that purpose, two small forest lakes and their dispersal sources were sampled over a period of 10 months. The influence of dispersal on communities and populations was determined by 454 sequencing of the 16S rRNA gene and SourceTracker analysis. On the community level direct effects of dispersal were questionable from all sources. Instead we found that the community of the preceding sampling occasion, representing growth of resident bacteria, was of great importance. On the population level, however, dispersal of individual taxa from the inlet could be occasionally important even under low water flow. The effect of sediment resuspension and precipitation appeared small.

Contribution of different dispersal sources to the metabolic response of lake bacterioplankton following a salinity change.

Dispersal can modify how bacterial community composition (BCC) changes in response to environmental perturbations, yet knowledge about the functional consequences of dispersal is limited. Here we hypothesized that changes in bacterial community production in response to a salinity disturbance depend on the possibility to recruit cells from different dispersal sources. To investigate this, we conducted an in situ mesocosm experiment where bacterial communities of an oligotrophic lake were exposed to different salinities (0, 18, 36 psu) for 2 weeks and subjected to dispersal of cells originating from sediments, air (mesocosms open to air deposition), both or none. BCC was determined using 454 pyrosequencing of the 16S rRNA gene and bacterial production was measured by 3 H leucine uptake. Bacterial production differed significantly among salinity treatments and dispersal treatments, being highest at high salinity. These changes were associated with changes in BCC and it was found that the identity of the main functional contributors differed at different salinities. Our results further showed that after a salinity perturbation, the response of bacterial communities depended on the recruitment of taxa, including marine representatives (e.g., Alphaproteobacteria Loktanella, Erythrobacter and the Gammaproteobacterium Rheiheimera) from dispersal sources, in which atmospheric deposition appeared to play a major role.

Effects of Dispersal and Initial Diversity on the Composition and Functional Performance of Bacterial Communities.

Natural communities are open systems and consequently dispersal can play an important role for the diversity, composition and functioning of communities at the local scale. It is, however, still unclear how effects of dispersal differ depending on the initial diversity of local communities. Here we implemented an experiment where we manipulated the initial diversity of natural freshwater bacterioplankton communities using a dilution-to-extinction approach as well as dispersal from a regional species pool. The aim was further to test whether dispersal effects on bacterial abundance and functional parameters (average community growth rates, respiration rates, substrate utilisation ability) differ in dependence of the initial diversity of the communities. First of all, we found that both initial diversity and dispersal rates had an effect on the recruitment of taxa from a regional source, which was higher in communities with low initial diversity and at higher rates of dispersal. Higher initial diversity and dispersal also promoted higher levels of richness and evenness in local communities and affected, both, separately or interactively, the functional performance of communities. Our study therefore suggests that dispersal can influence the diversity, composition and functioning of bacterial communities and that this effect may be enhanced if the initial diversity of communities is depleted.

Remnants of marine bacterial communities can be retrieved from deep sediments in lakes of marine origin.

Some bacteria can be preserved over time in deep sediments where they persist either in dormant or slow-growing vegetative stages. Here, we hypothesized that such cells can be revived when exposed to environmental conditions similar to those before they were buried in the sediments. To test this hypothesis, we collected bacteria from sediment samples of different ages (140-8500 calibrated years before present, cal BP) from three lakes that differed in the timing of their physical isolation from the Baltic Sea following postglacial uplift. After these bacterial communities were grown in sterile water from the Baltic Sea, we determined the proportion of 16S rRNA sequence reads associated with marine habitats by extracting the environment descriptive terms of homologous sequences retrieved from public databases. We found that the proportion of reads associated with marine descriptive term was significantly higher in cultures inoculated with sediment layers formed under Baltic conditions and where salinities were expected to be similar to current levels. Moreover, a similar pattern was found in the original sediment layers. Our study, therefore, suggests that remnants of marine bacterial communities can be preserved in sediments over thousands of years and can be revived from deep sediments in lakes of marine origin.

Bacterial community structure across environmental gradients in permafrost thaw ponds: methanotroph-rich ecosystems.

Permafrost thawing leads to the formation of thermokarst ponds that potentially emit CO2 and CH4 to the atmosphere. In the Nunavik subarctic region (northern Québec, Canada), these numerous, shallow ponds become well-stratified during summer. This creates a physico-chemical gradient of temperature and oxygen, with an upper oxic layer and a bottom low oxygen or anoxic layer. Our objective was to determine the influence of stratification and related limnological and landscape properties on the community structure of potentially active bacteria in these waters. Samples for RNA analysis were taken from ponds in three contrasting valleys across a gradient of permafrost degradation. A total of 1296 operational taxonomic units were identified by high throughput amplicon sequencing, targeting bacterial 16S rRNA that was reverse transcribed to cDNA. ß-proteobacteria were the dominant group in all ponds, with highest representation by the genera Variovorax and Polynucleobacter. Methanotrophs were also among the most abundant sequences at most sites. They accounted for up to 27% of the total sequences (median of 4.9% for all samples), indicating the importance of methane as a bacterial energy source in these waters. Both oxygenic (cyanobacteria) and anoxygenic (Chlorobi) phototrophs were also well-represented, the latter in the low oxygen bottom waters. Ordination analyses showed that the communities clustered according to valley and depth, with significant effects attributed to dissolved oxygen, pH, dissolved organic carbon, and total suspended solids. These results indicate that the bacterial assemblages of permafrost thaw ponds are filtered by environmental gradients, and are complex consortia of functionally diverse taxa that likely affect the composition as well as magnitude of greenhouse gas emissions from these abundant waters.

Oceanographic structure drives the assembly processes of microbial eukaryotic communities.

Arctic Ocean microbial eukaryote phytoplankton form subsurface chlorophyll maximum (SCM), where much of the annual summer production occurs. This SCM is particularly persistent in the Western Arctic Ocean, which is strongly salinity stratified. The recent loss of multiyear sea ice and increased particulate-rich river discharge in the Arctic Ocean results in a greater volume of fresher water that may displace nutrient-rich saltier waters to deeper depths and decrease light penetration in areas affected by river discharge. Here, we surveyed microbial eukaryotic assemblages in the surface waters, and within and below the SCM. In most samples, we detected the pronounced SCM that usually occurs at the interface of the upper mixed layer and Pacific Summer Water (PSW). Poorly developed SCM was seen under two conditions, one above PSW and associated with a downwelling eddy, and the second in a region influenced by the Mackenzie River plume. Four phylogenetically distinct communities were identified: surface, pronounced SCM, weak SCM and a deeper community just below the SCM. Distance-decay relationships and phylogenetic structure suggested distinct ecological processes operating within these communities. In the pronounced SCM, picophytoplanktons were prevalent and community assembly was attributed to water mass history. In contrast, environmental filtering impacted the composition of the weak SCM communities, where heterotrophic Picozoa were more numerous. These results imply that displacement of Pacific waters to greater depth and increased terrigenous input may act as a control on SCM development and result in lower net summer primary production with a more heterotroph dominated eukaryotic microbial community.

Can marine bacteria be recruited from freshwater sources and the air?

There is now clear evidence that microorganisms present biogeographic patterns, yet the processes that create and maintain them are still not well understood. In particular, the contribution of dispersal and its exact impact on local community composition is still unclear. For example, dispersing cells may not thrive in recipient environments, but may still remain part of the local species pool. Here, we experimentally tested if marine bacteria can be retrieved from freshwater communities (pelagic and sediment) and the atmosphere by exposing bacteria from three lakes, that differ in their proximity to the Norwegian Sea, to marine conditions. We found that the percentage of freshwater taxa decreased with increasing salinities, whereas marine taxa increased along the same gradient. Our results further showed that this increase was stronger for lake and sediment compared with air communities. Further, significant increases in the average niche breadth of taxa were found for all sources, and in particular lake water and sediment communities, at higher salinities. Our results therefore suggests that marine taxa can readily grow from freshwater sources, but that the response was likely driven by the growth of habitat generalists that are typically found in marine systems. Finally, there was a greater proportion of marine taxa found in communities originating from the lake closest to the Norwegian Sea. In summary, this study shows that the interplay between bacterial dispersal limitation and dispersal from internal and external sources may have an important role for community recovery in response to environmental change.

Total and methylated mercury in Arctic multiyear sea ice.

Mercury is one of the primary contaminants of concern in the Arctic marine ecosystem. While considerable efforts have been directed toward understanding mercury cycling in the Arctic, little is known about mercury dynamics within Arctic multiyear sea ice, which is being rapidly replaced with first-year ice. Here we report the first study on the distribution and potential methylation of mercury in Arctic multiyear sea ice. Based on three multiyear ice cores taken from the eastern Beaufort Sea and McClure Strait, total mercury concentrations ranged from 0.65 to 60.8 pM in bulk ice, with the highest values occurring in the topmost layer (∼40 cm) which is attributed to the dynamics of particulate matter. Methylated mercury concentrations ranged from below the method detection limit (<0.1 pM) to as high as 2.64 pM. The ratio of methylated to total mercury peaked, up to ∼40%, in the mid to bottom sections of the ice, suggesting the potential occurrence of in situ mercury methylation. The annual fluxes of total and methylated mercury into the Arctic Ocean via melt of multiyear ice are estimated to be 420 and 42 kg yr(-1), respectively, representing an important and changing source of mercury and methylmercury into the Arctic Ocean marine ecosystem.