Informing the Exposure Landscape: The Fate of Microplastics in a Large Pelagic In-Lake Mesocosm Experiment.

Understanding microplastic exposure and effects is critical to understanding risk. Here, we used large, in-lake closed-bottom mesocosms to investigate exposure and effects on pelagic freshwater ecosystems. This article provides details about the experimental design and results on the transport of microplastics and exposure to pelagic organisms. Our experiment included three polymers of microplastics (PE, PS, and PET) ranging in density and size. Nominal concentrations ranged from 0 to 29,240 microplastics per liter on a log scale. Mesocosms enclosed natural microbial, phytoplankton, and zooplankton communities and yellow perch (Perca flavescens). We quantified and characterized microplastics in the water column and in components of the food web (biofilm on the walls, zooplankton, and fish). The microplastics in the water stratified vertically according to size and density. After 10 weeks, about 1% of the microplastics added were in the water column, 0.4% attached to biofilm on the walls, 0.01% within zooplankton, and 0.0001% in fish. Visual observations suggest the remaining >98% were in a surface slick and on the bottom. Our study suggests organisms that feed at the surface and in the benthos are likely most at risk, and demonstrates the value of measuring exposure and transport to inform experimental designs and achieve target concentrations in different matrices within toxicity tests.

Our Data, Our Question: Public-Centric Approaches to Administrative Data Analysis.

There is growing recognition of the importance of patient, public and community engagement in health research, which has not been used widely in analyzing health administrative datasets. In Ontario, health data are stewarded by ICES, whose strategic decision making is guided by a diverse Public Advisory Council (PAC). In a first foray into publicly led projects, the ICES PAC undertook an analysis project on mental health and addiction health service use. Public members guided the project through all stages of research. This generated critical lessons for ICES on improving participation, collaboration and trust.

Understanding temporal variability across trophic levels and spatial scales in freshwater ecosystems.

A tenet of ecology is that temporal variability in ecological structure and processes tends to decrease with increasing spatial scales (from locales to regions) and levels of biological organization (from populations to communities). However, patterns in temporal variability across trophic levels and the mechanisms that produce them remain poorly understood. Here we analyzed the abundance time series of spatially structured communities (i.e., metacommunities) spanning basal resources to top predators from 355 freshwater sites across three continents. Specifically, we used a hierarchical partitioning method to disentangle the propagation of temporal variability in abundance across spatial scales and trophic levels. We then used structural equation modeling to determine if the strength and direction of relationships between temporal variability, synchrony, biodiversity, and environmental and spatial settings depended on trophic level and spatial scale. We found that temporal variability in abundance decreased from producers to tertiary consumers but did so mainly at the local scale. Species population synchrony within sites increased with trophic level, whereas synchrony among communities decreased. At the local scale, temporal variability in precipitation and species diversity were associated with population variability (linear partial coefficient, ß = 0.23) and population synchrony (ß = -0.39) similarly across trophic levels, respectively. At the regional scale, community synchrony was not related to climatic or spatial predictors, but the strength of relationships between metacommunity variability and community synchrony decreased systematically from top predators (ß = 0.73) to secondary consumers (ß = 0.54), to primary consumers (ß = 0.30) to producers (ß = 0). Our results suggest that mobile predators may often stabilize metacommunities by buffering variability that originates at the base of food webs. This finding illustrates that the trophic structure of metacommunities, which integrates variation in organismal body size and its correlates, should be considered when investigating ecological stability in natural systems. More broadly, our work advances the notion that temporal stability is an emergent property of ecosystems that may be threatened in complex ways by biodiversity loss and habitat fragmentation.

Freshwater connectivity transforms spatially integrated signals of biodiversity.

Aquatic ecosystems offer a continuum of water flow from headwater streams to inland lakes and coastal marine systems. This spatial connectivity influences the structure, function and dynamics of aquatic communities, which are among the most threatened and degraded on the Earth. Here, we determine the spatial resolution of environmental DNA (eDNA) in dendritic freshwater networks, which we use as a model for connected metacommunities. Our intensive sampling campaign comprised over 420 eDNA samples across 21 connected lakes, allowing us to analyse detections at a variety of scales, from different habitats within a lake to entire lake networks. We found strong signals of within-lake variation in eDNA distribution reflective of typical habitat use by both fish and zooplankton. Most importantly, we also found that connecting channels between lakes resulted in an accumulation of downstream eDNA detections in lakes with a higher number of inflows, and as networks increased in length. Environmental DNA achieves biodiversity surveys in these habitats in a high-throughput, spatially integrated way. These findings have profound implications for the interpretation of eDNA detections in aquatic ecosystems in global-scale biodiversity monitoring observations.

Valsartan, Losartan and Irbesartan use in the USA, UK, Canada and Denmark after the nitrosamine recalls: a descriptive cohort study.

OBJECTIVES: To examine valsartan, losartan and irbesartan usage and switching patterns in the USA, UK, Canada and Denmark before and after July 2018, when the first Angiotensin-Receptor-Blocker (ARB) (valsartan) was recalled. DESIGN: Retrospective cohort study. SETTING: USA, Canadian administrative healthcare data, Danish National Prescription Registry and UK primary care electronic health records. PARTICIPANTS: Patients aged 18 years and older between January 2014 and December 2020. INTERVENTION: Valsartan, losartan and irbesartan. MAIN OUTCOME: Monthly percentages of individual ARB episodes, new users and switches to another ARB, ACE inhibitors (ACEI) or calcium channel blockers containing products. RESULTS: We identified 10.8, 3.2, 1.8 and 1.2 million ARB users in the USA, UK, Canada and Denmark, respectively. Overall proportions of valsartan, losartan and irbesartan use were 18.4%, 67.9% and 5.2% in the USA; 3.1%, 48.3% and 10.2% in the UK, 16.3%, 11.4% and 18.3% in Canada, 1%, 93.5% and 0.6% in Denmark. In July 2018, we observed an immediate steep decline in the proportion of valsartan use in the USA and Canada. A similar trend was observed in Denmark; however, the decline was only minimal. We observed no change in trends of ARB use in the UK. Accompanying the valsartan decline was an increase in switching to other ARBs in the USA, Canada and Denmark. There was a small increase in switching to ACEI relative to the valsartan-to-other-ARBs switch. We also observed increased switching from other affected ARBs, losartan and irbesartan, to other ARBs throughout 2019, in the USA and Canada, although the usage trends in the USA remained unchanged. CONCLUSION: The first recall notice for valsartan resulted in substantial decline in usage due to increased switching to other ARBs. Subsequent notices for losartan and irbesartan were also associated with increased switching around the time of the recall, however, overall usage trends remained unchanged.

Large Fractionation in Iron Isotopes Implicates Metabolic Pathways for Iron Cycling in Boreal Shield Lakes.

Stable Fe isotopes have only recently been measured in freshwater systems, mainly in meromictic lakes. Here we report the δ56Fe of dissolved, particulate, and sediment Fe in two small dimictic boreal shield headwater lakes: manipulated eutrophic Lake 227, with annual cyanobacterial blooms, and unmanipulated oligotrophic Lake 442. Within the lakes, the range in δ56Fe is large (ca. -0.9 to +1.8‰), spanning more than half the entire range of natural Earth surface samples. Two layers in the water column with distinctive δ56Fe of dissolved (dis) and particulate (spm) Fe were observed, despite differences in trophic states. In the epilimnia of both lakes, a large Δ56Fedis-spm fractionation of 0.4-1‰ between dissolved and particulate Fe was only observed during cyanobacterial blooms in Lake 227, possibly regulated by selective biological uptake of isotopically light Fe by cyanobacteria. In the anoxic layers in both lakes, upward flux from sediments dominates the dissolved Fe pool with an apparent Δ56Fedis-spm fractionation of -2.2 to -0.6‰. Large Δ56Fedis-spm and previously published metagenome sequence data suggest active Fe cycling processes in anoxic layers, such as microaerophilic Fe(II) oxidation or photoferrotrophy, could regulate biogeochemical cycling. Large fractionation of stable Fe isotopes in these lakes provides a potential tool to probe Fe cycling and the acquisition of Fe by cyanobacteria, with relevance for understanding biogeochemical cycling of Earth's early ferruginous oceans.

Experimental evidence for recovery of mercury-contaminated fish populations.

Anthropogenic releases of mercury (Hg)1-3 are a human health issue4 because the potent toxicant methylmercury (MeHg), formed primarily by microbial methylation of inorganic Hg in aquatic ecosystems, bioaccumulates to high concentrations in fish consumed by humans5,6. Predicting the efficacy of Hg pollution controls on fish MeHg concentrations is complex because many factors influence the production and bioaccumulation of MeHg7-9. Here we conducted a 15-year whole-ecosystem, single-factor experiment to determine the magnitude and timing of reductions in fish MeHg concentrations following reductions in Hg additions to a boreal lake and its watershed. During the seven-year addition phase, we applied enriched Hg isotopes to increase local Hg wet deposition rates fivefold. The Hg isotopes became increasingly incorporated into the food web as MeHg, predominantly from additions to the lake because most of those in the watershed remained there. Thereafter, isotopic additions were stopped, resulting in an approximately 100% reduction in Hg loading to the lake. The concentration of labelled MeHg quickly decreased by up to 91% in lower trophic level organisms, initiating rapid decreases of 38-76% of MeHg concentration in large-bodied fish populations in eight years. Although Hg loading from watersheds may not decline in step with lowering deposition rates, this experiment clearly demonstrates that any reduction in Hg loadings to lakes, whether from direct deposition or runoff, will have immediate benefits to fish consumers.

Using zooplankton metabarcoding to assess the efficacy of different techniques to clean-up an oil-spill in a boreal lake.

Regulators require adequate information to select best practices with less ecosystem impacts for remediation of freshwater ecosystems after oil spills. Zooplankton are valuable indicators of aquatic ecosystem health as they play pivotal roles in biochemical cycles while stabilizing food webs. Compared with morphological identification, metabarcoding holds promise for cost-effective, high-throughput, and benchmarkable biomonitoring of zooplankton communities. The objective of this study was to apply DNA and RNA metabarcoding of zooplankton for ecotoxicological assessment and compare it with traditional morphological identification in experimental shoreline enclosures in a boreal lake. These identification methods were also applied in context of assessing response of the zooplankton community exposed to simulated spills of diluted bitumen (dilbit), with experimental remediation practices (enhanced monitored natural recovery and shoreline cleaner application). Metabarcoding detected boreal zooplankton taxa up to the genus level, with a total of 24 shared genera, and while metabarcoding-based relative abundance served as an acceptable proxy for biomass inferred by morphological identification (ρ ≥ 0.52). Morphological identification determined zooplankton community composition changes due to treatments at 11 days post-spill (PERMANOVA, p = 0.0143) while metabarcoding methods indicated changes in zooplankton richness and communities at 38 days post-spill (T-test, p < 0.05; PERMANOVA, p ≤ 0.0429). Shoreline cleaner application overall seemed to have the largest impact on zooplankton communities relative to enhanced monitored natural recovery, regardless of zooplankton identification method. Both metabarcoding and morphological identification were able to discern the differences between the two experimental remediation practices. Metabarcoding of zooplankton could provide informative results for ecotoxicological assessment of the remediation practices of dilbit, advancing our knowledge of best practices for remediating oil-impacted aquatic ecosystems while serving to accelerate the assessment of at-risk freshwater ecosystems.

Trends in the Prevalence and Incidence of Psoriasis and Psoriatic Arthritis in Ontario, Canada: A Population-Based Study.

OBJECTIVE: To estimate the prevalence and incidence of psoriasis and psoriatic arthritis (PsA) over time in Ontario, Canada. METHODS: We performed a population-based study of Ontario health administrative data, using validated case definitions for psoriasis and PsA. We computed the crude and age- and sex-standardized cumulative prevalence and incidence of psoriasis from 2000 to 2015. RESULTS: Among the 10,774,802 individuals ages ≥20 years residing in Ontario in 2015, we identified 273,238 patients with psoriasis and 18,655 patients with PsA, equating to cumulative prevalence estimates of 2.54% and 0.17%, respectively. Correcting the prevalence estimates for imperfect sensitivity and specificity resulted in similar estimates. The male:female ratio was approximately 1.0 for both conditions. For psoriasis, the age- and sex-standardized cumulative prevalence increased from 1.74% in 2000 to 2.32% in 2015. For PsA, the age- and sex-standardized cumulative prevalence increased from 0.09% in 2008 to 0.15% in 2015. Between 2008 and 2015, annual incidence rates for psoriasis decreased, whereas those for PsA remained relatively stable. CONCLUSION: The prevalence and incidence of psoriasis and PsA in Ontario are similar to those observed in Europe and the US. The steady increase in the prevalence of psoriasis and PsA over the past decade may be due to a combination of population aging, population growth, and increasing life expectancy.

Accumulation of Silver in Yellow Perch ( Perca flavescens) and Northern Pike ( Esox lucius) From a Lake Dosed with Nanosilver.

A total of 15 kg of silver nanoparticles (AgNPs) was added continuously over two ice-free field seasons to a boreal lake (i.e., Lake 222) at the IISD Experimental Lakes Area in Canada. We monitored the accumulation of silver (Ag) in the tissues of yellow perch ( Perca flavescens) and northern pike ( Esox lucius) exposed to the AgNPs under environmentally relevant conditions. The greatest accumulation was observed in the liver tissues of pike, and a single pike sampled in the second year of additions had the highest concentration observed in liver of 5.1 micrograms per gram of wet weight. However, the Ag concentrations in gill and muscle tissue of both pike and perch did not exceed 0.35 micrograms per gram of wet weight. Following additions of AgNP, the Ag residues in fish tissues declined, with a half-life of Ag in pike liver of 119 days. Monitoring using passive sampling devices and single-particle inductively coupled plasma mass spectrometry during the AgNP addition phase confirmed that Ag nanoparticles were present in the water column and that estimated mean concentrations of Ag increased over time to a maximum of 11.5 µg/L. These data indicate that both a forage fish and a piscivorous fish accumulated Ag in a natural lake ecosystem dosed with AgNPs, leading to Ag concentrations in some tissues of the piscivorous species that were 3 orders of magnitude greater than the concentrations in the water.

Chronic and pulse exposure effects of silver nanoparticles on natural lake phytoplankton and zooplankton.

The increasing use of silver nanoparticles (AgNPs) in consumer products raises concerns regarding the environmental exposure and impact of AgNPs on natural aquatic environments. Here, we investigated the effects of environmentally relevant AgNP concentrations on the natural plankton communities using in situ enclosures. Using twelve lake enclosures, we tested the hypotheses that AgNP concentration, dosing regimen, and capping agent (poly-vinyl pyrrolidone (PVP) vs. citrate) exhibit differential effects on plankton communities. Each of the following six treatments was replicated twice: control (no AgNPs added), low, medium, and high chronic PVP treatments (PVP-capped AgNPs added continuously, with target nominal concentrations of 4, 16, and 64 µg/L, respectively), citrate treatment (citrate-capped AgNPs added continuously, target nominal concentrations of 64 µg/L), and pulse treatment (64 µg/L PVP-AgNPs added as a single dose). Although Ag accumulated in the phytoplankton, no statistically significant treatment effect was found on phytoplankton community structure or biomass. In contrast, as AgNP exposure rate increased, zooplankton abundance generally increased while biomass and species richness declined. We also observed a shift in the size structure of zooplankton communities in the chronic AgNP treatments. In the pulse treatments, zooplankton abundance and biomass were reduced suggesting short periods of high AgNP concentrations affect zooplankton communities differently than chronic exposures. We found no evidence that capping agent affected AgNP toxicity on either community. Overall, our study demonstrates variable AgNP toxicity between trophic levels with stronger AgNP effects on zooplankton. Such effects on zooplankton are troubling and indicate that AgNP contamination could affect aquatic food webs.

Prescription Stimulant Use and Hospitalization for Psychosis or Mania: A Population-Based Study.

Small studies suggest that prescription stimulants can precipitate psychosis and mania. We conducted a population-based case-crossover study to examine whether hospitalization for psychosis or mania was associated with initiation of stimulant therapy. Between October 1, 1999 and March 31, 2013, we studied 12,856 young people who received a stimulant prescription and were subsequently hospitalized for psychosis or mania. Of these, 183 commenced treatment during 1 of 2 prespecified 60-day intervals (defined as the "risk interval" and "control interval," respectively) prior to admission. We found that stimulant initiation was associated with an increased risk of hospitalization for psychosis or mania in the subsequent 60 days (odds ratio, 1.86; 95% confidence interval, 1.39-2.56). The risk was marginally higher in patients treated with antipsychotic drugs (odds ratio, 2.06; 95% confidence interval, 1.38-3.28), but remained in patients with no such history (odds ratio, 1.66; 95% confidence interval, 1.09-2.66). One third of subjects received another stimulant prescription after hospital discharge. Of these, 45% were readmitted with psychosis or mania shortly thereafter. We conclude that initiation of prescription stimulants is associated with an increased risk of hospitalization for psychosis or mania. Resumption of therapy is common, which may reflect a lack of awareness of the potential causative role of these drugs.

Crystal structure of a novel two domain GH78 family α-rhamnosidase from Klebsiella oxytoca with rhamnose bound.

The crystal structure of the GH78 family α-rhamnosidase from Klebsiella oxytoca (KoRha) has been determined at 2.7 Å resolution with rhamnose bound in the active site of the catalytic domain. Curiously, the putative catalytic acid, Asp 222, is preceded by an unusual non-proline cis-peptide bond which helps to project the carboxyl group into the active centre. This KoRha homodimeric structure is significantly smaller than those of the other previously determined GH78 structures. Nevertheless, the enzyme displays α-rhamnosidase activity when assayed in vitro, suggesting that the additional structural domains found in the related enzymes are dispensible for function.

Direct and indirect responses of a freshwater food web to a potent synthetic oestrogen.

Endocrine-disrupting chemicals (EDCs) in municipal effluents directly affect the sexual development and reproductive success of fishes, but indirect effects on invertebrate prey or fish predators through reduced predation or prey availability, respectively, are unknown. At the Experimental Lakes Area in northwestern Ontario, Canada, a long-term, whole-lake experiment was conducted using a before-after-control-impact design to determine both direct and indirect effects of the synthetic oestrogen used in the birth control pill, 17α-ethynyloestradiol (EE2). Algal, microbial, zooplankton and benthic invertebrate communities showed no declines in abundance during three summers of EE2 additions (5-6 ng l(-1)), indicating no direct toxic effects. Recruitment of fathead minnow (Pimephales promelas) failed, leading to a near-extirpation of this species both 2 years during (young-of-year, YOY) and 2 years following (adults and YOY) EE2 additions. Body condition of male lake trout (Salvelinus namaycush) and male and female white sucker (Catostomus commersonii) declined before changes in prey abundance, suggesting direct effects of EE2 on this endpoint. Evidence of indirect effects of EE2 was also observed. Increases in zooplankton, Chaoborus, and emerging insects were observed after 2 or 3 years of EE2 additions, strongly suggesting indirect effects mediated through the reduced abundance of several small-bodied fishes. Biomass of top predator lake trout declined by 23-42% during and after EE2 additions, most probably an indirect effect from the loss of its prey species, the fathead minnow and slimy sculpin (Cottus cognatus). Our results demonstrate that small-scale studies focusing solely on direct effects are likely to underestimate the true environmental impacts of oestrogens in municipal wastewaters and provide further evidence of the value of whole-ecosystem experiments for understanding indirect effects of EDCs and other aquatic stressors.

Low-level experimental selenite additions decrease mercury in aquatic food chains and fish muscle but increase selenium in fish gonads.

We investigated whether low-level addition of selenium (Se) could decrease mercury (Hg) in freshwater fish without imposing Se toxicity. Using a regression design, selenite was added to large mesocosms in a lake to achieve target concentrations ≤1.6 µg/L. (198)Hg (spike Hg) was added to mesocosms to determine changes in Hg bioaccumulation. Adding Se decreased spike total Hg (THg) in fish muscle, ambient THg in fish liver, and bioaccumulation of spike THg in muscle and spike methylmercury (MeHg) in zooplankton and Chironomid larvae relative to controls. Se decreased Hg in the food web but not in water, indicating that the dominant effect of Se on Hg cycling occurs in the food web. Concentrations of Se in gonads of fish were positively correlated with Se concentrations in water but did not exceed reproductive toxicity thresholds after 8 weeks. We conclude that low-level addition of Se decreases MeHg bioaccumulation and increases Se in gonads of fish; however, additions of Se to freshwater systems to decrease Hg in fish should be treated with caution because Se in fish gonads were likely to exceed toxic concentrations if exposed to increased Se for a longer period of time.

Comparative effectiveness of tiotropium and ipratropium in prevention of hospital readmission for COPD: a population-based cohort study.

BACKGROUND: In the treatment of chronic obstructive pulmonary disease (COPD), tiotropium bromide has a longer duration of action than ipratropium bromide; however, tiotropium bromide is a more expensive alternative treatment. At issue is whether tiotropium reduces the risk for hospital readmissions for COPD compared with ipratropium. OBJECTIVES: A population-based cohort study was conducted to assess whether tiotropium reduces the risk for hospital readmissions for COPD compared with ipratropium. METHODS: British Columbia (BC) linked provincial administrative health databases were used to identify new patients with COPD (aged ≥45 years) with a first hospital admission for COPD from 2003 to 2011. The study period was defined as the 30-day tiotropium or ipratropium treatment-initiation period after hospital discharge. Patients were followed up for ≤6 months from drug initiation to hospital readmission for COPD. In a subanalysis, the 2 treatment groups were matched on age, sex, and high-dimensional propensity scores derived from 200 empirically identified and predefined covariates. The risk for hospital readmission was estimated using multivariate Cox proportional hazards and logistic regression analyses. RESULTS: In total, 3723 patients with COPD were dispensed tiotropium (n = 992) or ipratropium (n = 2731) within 30 days from the index hospital admission for COPD. The mean age of these patients was 72.8 years, and 50.8% were women. Tiotropium-treated patients were more likely to be in a higher income category and were more likely to use a greater number of medications compared with ipratropium-treated patients. Among the subset of 1500 matched patients, 215 (14.3%) were readmitted to hospital within 6 months. There was no statistically significant group difference in hospital readmissions using either analytical approach (hazard ratio = 0.98 [95% CI, 0.72-1.34]; odds ratio = 0.97 [95% CI, 0.70-1.36]). CONCLUSIONS: In this select group of patients, neither tiotropium nor ipratropium was effective in significantly decreasing the risk for rehospitalization for COPD within 6 months.

Dietary and waterborne mercury accumulation by yellow perch: a field experiment.

It is well accepted that the majority of monomethylmercury (MMHg) in fish originates in their food; however, the additional contribution of water as a source to fish MMHg levels remains unclear. We used isotope enriched mercury (Hg) in a controlled field experiment to quantify the uptake of Hg from ingested and aqueous sources by young-of-year yellow perch (Perca flavescens). Water and zooplankton from a lake that had received (202)Hg-enriched additions (called spike Hg) for 7 y during a whole-ecosystem loading study (METAALICUS) provided natural, low-level Hg exposure. We achieved separation of exposure pathways by housing perch in one of four treatments: clean water + clean food; clean water + Hg spiked food; Hg spiked water + clean food; Hg spiked water + Hg spiked food. Fish accumulated MMHg directly from water, and this source accounted for at least 10% of MMHg in fish during the 27-d trial. Accumulation of spike Hg from water and food was additive, with food providing the majority of spike MMHg taken in by fish. Predictions from a bioenergetics model that excludes water as a source underestimated Hg in perch by 11%. This study illustrates the importance of acknowledging both food and water as sources of Hg to fish and suggests that aqueous Hg should be included as a source of contamination in bioaccumulation models and experiments.

Assessment of mercury bioaccumulation within the pelagic food web of lakes in the western Great Lakes region.

While mercury is a health hazard to humans and wildlife, the biogeochemical processes responsible for its bioaccumulation in pelagic food webs are still being examined. Previous studies have indicated both "bottom-up" control of piscivorous fish Hg content through methylmercury.(MeHg) supply, as well as site-specific trophic factors. We evaluated ten studies from the western Great Lakes region to examine the similarity of MeHg trophic transfer efficiency within the pelagic food web, and assessed regional-scale spatial variability. Analyses of bioaccumulation and biomagnification factors between water, seston, zooplankton, and preyfish indicated that the largest increases in MeHg occurred at the base of the food web, and that the relative extent of trophic transfer was similar between sites. Positive correlations were observed between aqueous unfiltered MeHg, total Hg, and dissolved organic carbon, and measures of the efficiency of MeHg trophic transfer were consistent across widely disparate systems (both natural and experimentally manipulated) throughout North America. Such similarity suggests that the aqueous supply of MeHg is largely controlling bioaccumulation in pelagic food webs, while local, lake-specific variability can result from an array of trophic (biological) factors.

A fluorescence lifetime-based assay for serine and threonine kinases that is suitable for high-throughput screening.

We describe the development of a novel method for the assay of serine/threonine protein kinases based on fluorescence lifetime. The assay consists of three generic peptides (which have been used by others in the assay of >140 protein kinases in various assay formats) labeled with a long lifetime fluorescent dye (14 or 17 ns) that act as substrates for protein kinases and an iron(III) chelate that modulates the fluorescence lifetime of the peptide only when it is phosphorylated. The decrease in average fluorescence lifetime as measured in a recently developed fluorescence lifetime plate reader (Edinburgh Instruments) is a measure of the degree of phosphorylation of the peptide. We present data showing that the assay performs as well as, and in some cases better than, the "gold standard" radiometric kinase assays with respect to Z' values, demonstrating its utility in high-throughput screening applications. We also show that the assay gives nearly identical results in trial screening to those obtained by radiometric assays and that it is less prone to interference than simple fluorescence intensity measurements.

Environmental stability and lake zooplankton diversity - contrasting effects of chemical and thermal variability.

Environmental variability in space and time is a primary mechanism allowing species that share resources to coexist. Fluctuating conditions are a double edged sword for diversity, either promoting coexistence through temporal niche partitioning or excluding species by stochastic extinctions. The net effect of environmental variation on diversity is largely unknown. We examined the association between zooplankton species richness in lakes and environmental variability on interannual, seasonal and shorter time scales, as well as long-term average conditions. We analyzed data on physical, chemical and biological limnology in 53 temperate zone lakes in North America and Europe sampled over a combined 1042 years. Large fluctuations in pH, phosphorus and dissolved organic carbon concentration on different time scales were associated with reduced zooplankton species richness. More species were found in lakes that showed greater temperature variation on all time scales. Environmental variability on different time scales showed similar or, in some cases, stronger associations with zooplankton species richness compared with long-term average conditions. Our results suggest that temporal fluctuations in the chemical environment tend to exclude zooplankton species while temperature variability promotes greater richness. The results indicate that anthropogenic increases in temporal variability of future climates may have profound effects on biodiversity.