Environmental DNA metabarcoding for benthic monitoring: A review of sediment sampling and DNA extraction methods.

Environmental DNA (eDNA) metabarcoding (parallel sequencing of DNA/RNA for identification of whole communities within a targeted group) is revolutionizing the field of aquatic biomonitoring. To date, most metabarcoding studies aiming to assess the ecological status of aquatic ecosystems have focused on water eDNA and macroinvertebrate bulk samples. However, the eDNA metabarcoding has also been applied to soft sediment samples, mainly for assessing microbial or meiofaunal biota. Compared to classical methodologies based on manual sorting and morphological identification of benthic taxa, eDNA metabarcoding offers potentially important advantages for assessing the environmental quality of sediments. The methods and protocols utilized for sediment eDNA metabarcoding can vary considerably among studies, and standardization efforts are needed to improve their robustness, comparability and use within regulatory frameworks. Here, we review the available information on eDNA metabarcoding applied to sediment samples, with a focus on sampling, preservation, and DNA extraction steps. We discuss challenges specific to sediment eDNA analysis, including the variety of different sources and states of eDNA and its persistence in the sediment. This paper aims to identify good-practice strategies and facilitate method harmonization for routine use of sediment eDNA in future benthic monitoring.

Temperature dependence of denitrification in phototrophic river biofilms.

Denitrification is an ecosystem service of nitrogen load regulation along the terrestrial-freshwater-marine continuum. The present study documents the short-term temperature sensitivity of denitrification enzyme activity in phototrophic river biofilms as a typical microbial assemblage of this continuum. Denitrification measurements were performed using the acetylene inhibition method at four incubation temperatures: 1.1, 12.1, 21.2 and 30.9°C. For this range of temperature, N(2)O production could be fitted to an exponential function of incubation temperature, yielding mean (±standard error) activation energy of 1.42 (±0.24) eV and Q(10) of 7.0 (±1.4). This first quantification of denitrification enzyme activity temperature dependence in phototrophic river biofilms compares with previous studies performed in soils and sediments. This demonstrates the high temperature dependence of denitrification as compared to other community-level metabolisms such as respiration or photosynthesis. This result suggests that global warming can unbalance natural community metabolisms in phototrophic river biofilms and affect their biogeochemical budget.

Associations among pathogenic bacteria, parasites, and environmental and land use factors in multiple mixed-use watersheds.

Over a five year period (2004-08), 1171 surface water samples were collected from up to 24 sampling locations representing a wide range of stream orders, in a river basin in eastern Ontario, Canada. Water was analyzed for Cryptosporidium oocysts and Giardia cyst densities, the presence of Salmonella enterica subspecies enterica, Campylobacter spp., Listeria monocytogenes, and Escherichia coli O157:H7. The study objective was to explore associations among pathogen densities/occurrence and objectively defined land use, weather, hydrologic, and water quality variables using CART (Classification and Regression Tree) and binary logistical regression techniques. E. coli O157:H7 detections were infrequent, but detections were related to upstream livestock pasture density; 20% of the detections were located where cattle have access to the watercourses. The ratio of detections:non-detections for Campylobacter spp. was relatively higher (>1) when mean air temperatures were 6% below mean study period temperature values (relatively cooler periods). Cooler water temperatures, which can promote bacteria survival and represent times when land applications of manure typically occur (spring and fall), may have promoted increased frequency of Campylobacter spp. Fifty-nine percent of all Salmonella spp. detections occurred when river discharge on a branch of the river system of Shreve stream order = 9550 was >83 percentile. Hydrological events that promote off farm/off field/in stream transport must manifest themselves in order for detection of Salmonella spp. to occur in surface water in this region. Fifty seven percent of L. monocytogenes detections occurred in spring, relative to other seasons. It was speculated that a combination of winter livestock housing, silage feeding during winter, and spring application of manure that accrued during winter, contributed to elevated occurrences of this pathogen in spring. Cryptosporidium and Giardia oocyst and cyst densities were, overall, positively associated with surface water discharge, and negatively associated with air/water temperature during spring-summer-fall. Yet, some of the highest Cryptosporidium oocyst densities were associated with low discharge conditions on smaller order streams, suggesting wildlife as a contributing fecal source. Fifty six percent of all detections of ≥ 2 bacteria pathogens (including Campylobacter spp., Salmonella spp., and E. coli O157:H7) in water was associated with lower water temperatures (<∼ 14 °C; primarily spring and fall) and when total rainfall the week prior to sampling was >∼ 27 mm (62 percentile). During higher water temperatures (>∼ 14 °C), a higher amount of weekly rainfall was necessary to promote detection of ≥ 2 pathogens (primarily summer; weekly rainfall ∼>42 mm (>77 percentile); 15% of all ≥ 2 detections). Less rainfall may have been necessary to mobilize pathogens from adjacent land, and/or in stream sediments, during cooler water conditions; as these are times when manures are applied to fields in the area, and soil water contents and water table depths are relatively higher. Season, stream order, turbidity, mean daily temperature, surface water discharge, cropland coverage, and nearest upstream distance to a barn and pasture were variables that were relatively strong and recurrent with regard to discriminating pathogen presence and absence, and parasite densities in surface water in the region.

Livestock waste treatment systems for reducing environmental exposure to hazardous enteric pathogens: some considerations.

Intensive livestock production systems produce significant quantities of excreted material that must be managed to protect water, air, and crop quality. Many jurisdictions mandate how livestock wastes are managed to protect adjacent water quality from microbial and chemical contaminants that pose an environmental and human health challenge. Here, we consider innovative livestock waste treatment systems in the context of multi-barrier strategies for protecting water quality from agricultural contamination. Specifically, we consider some aspects of how enteric bacterial populations can evolve during manure storage, how their fate following land application of manure can vary according to manure composition, and finally the challenge of distinguishing enteric pathogens of agricultural provenance from those of other sources of fecal pollution at a policy-relevant watershed scale. The beneficial impacts of livestock waste treatment on risk to humans via exposure to manured land are illustrated using quantitative microbial risk assessment (QMRA) scenarios. Overall, innovative livestock treatment systems offer a crucially important strategy for making livestock wastes more benign before they are released into the broader environment.