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A recurring trend in evidence scrutinized over the past few decades is that disease outbreaks will become more frequent, intense, and widespread on land and in water, due to climate change. Pathogens and the diseases they inflict represent a major constraint on seafood production and yield, and by extension, food security. The risk(s) for fish and shellfish from disease is a function of pathogen characteristics, biological species identity, and the ambient environmental conditions. A changing climate can adversely influence the host and environment, while augmenting pathogen characteristics simultaneously, thereby favoring disease outbreaks. Herein, we use a series of case studies covering some of the world's most cultured aquatic species (e.g., salmonids, penaeid shrimp, and oysters), and the pathogens (viral, fungal, bacterial, and parasitic) that afflict them, to illustrate the magnitude of disease-related problems linked to climate change., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

Non-genetic sources of phenotypic variation, such as the epigenome and the microbiome, could be important contributors to adaptive variation for species with low genetic diversity. However, little is known about the complex interaction between these factors and the genetic diversity of the host, particularly in wild populations. Here, we examine the skin microbiome composition of two closely-related mangrove killifish species with different mating systems (self-fertilising and outcrossing) under sympatric and allopatric conditions. This allows us to partition the influence of the genotype and the environment on their microbiome and (previously described) epigenetic profiles. We find the diversity and community composition of the skin microbiome are strongly shaped by the environment and, to a lesser extent, by species-specific influences. Heterozygosity and microbiome alpha diversity, but not epigenetic variation, are associated with the fluctuating asymmetry of traits related to performance (vision) and behaviour (aggression). Our study identifies that a proportion of the epigenetic diversity and microbiome differentiation is unrelated to genetic variation, and we find evidence for an associative relationship between microbiome and epigenetic diversity in these wild populations. This suggests that both mechanisms could potentially contribute to variation in species with low genetic diversity., (© 2024. The Author(s).)

Colorectal cancer (CRC) remains a leading cause of cancer-related deaths worldwide, despite advancements in diagnosis. The main reason for this is that many newly diagnosed CRC patients will suffer from metastasis to other organs. Thus, the development of new therapies is of critical importance. Claudin-1 protein is a component of tight junctions in epithelial cells, including those found in the lining of the colon. It plays a critical role in the formation and maintenance of tight junctions, which are essential for regulating the passage of molecules between cells. In CRC, claudin-1 is often overexpressed, leading to an increase in cell adhesion, which can contribute to the development and progression of the disease. Studies show that high levels of claudin-1 are associated with poor prognosis in CRC patients and targeting claudin-1 may have therapeutic potential for the treatment of CRC. Previously, we have identified a small molecule that inhibits claudin-1 dependent CRC progression. Reported herein are our lead optimization efforts around this scaffold to identify the key SAR components and the discovery of a key new compound that exhibits enhanced potency in SW620 cells., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Experimental exposures aimed at assessing the risks posed by estrogens in waste-water treatment work (WwTW) effluents to fish populations have rarely considered whether populations differ in their sensitivity to estrogenic compounds. This is despite evidence that selection at genes involved in the estrogen response has occurred in wild populations, and evidence that genotype can influence estrogen-response. In this study we compare the effects of a two-year exposure to a low measured concentration (1.3 ng/L) of ethinylestradiol (EE2) on the sexual development of roach (Rutilus rutilus) whose parental generation was sampled from two river stretches heavily contaminated with WwTW effluent and from two without any known WwTW effluent contamination. Exposure to EE2 significantly reduced the proportion of genetic males and induced a range of feminized phenotypes in males. Significantly, exposure also increased the proportion of genetic females with vitellogenic oocytes from 51 to 96%, raising the possibility that estrogen pollution could impact populations of annually spawning fish species through advancing female reproduction by at least a year. However, there was no evidence that river origin affected sensitivity to estrogens in either sex. Thus, we conclude that chronic exposure to low level EE2 has reproductive health outcomes for both male and female roach, but we find no evidence that the nature or magnitude of the response is affected by the population origin., (Copyright © 2022. Published by Elsevier B.V.)

Abstract: Webster, TM, Comfort, P, and Jones, PA. Relationship between physical fitness and the physical demands of 50-over cricket in fast bowlers. J Strength Cond Res 36(3): e66-e72, 2022-Professional cricket is constantly evolving and resulting in increased physiological demand placed on players. Fast bowlers experience the greatest physical demand during match-play; despite this, research has overlooked the importance of specific physical attributes to optimizing physical match performance. The aim of this study was to investigate the relationships between 50-Over physical match performance and tests of physical qualities in fast bowlers. Fifteen professional male fast bowlers (age = 23.8 ± 4.0 years; height = 183.4 ± 6.7 cm; body mass = 82.9 ± 9.2 kg) participated in the study. Subjects completed the following physical fitness tests; sum of skinfolds, countermovement jump (CMJ), 20-m sprint, and the Yo-Yo intermittent recovery test level 1. Global positioning system data were collected during 6 professional 50-Over fixtures. Moderate correlations were established between CMJ height to total distance covered (r = 0.585; p = 0.022), sprinting distance (ρ = 0.554; p = 0.032), and maximal velocity (r = 0.567; p = 0.027); 20-m sprint time to total distance covered (ρ = -0.519; p = 0.047) and PlayerLoad (ρ = -0.561; p = 0.03); and Yo-Yo distance to total match distance (r = 0.520; p = 0.047) and sprinting distance (ρ = 0.524; p = 0.045). These findings demonstrate the predictive importance of prolonged intermittent running ability, jump, and sprint performance to match running performance of elite fast bowlers in 1-day cricket. Strength and conditioning coaches should use such information to design training to ensure fast bowlers are adequately conditioned to meet the demands of match running performance., (Copyright © 2020 National Strength and Conditioning Association.)

Farmed fish are commonly exposed to stress in intensive aquaculture systems, often leading to immune impairment and increased susceptibility to disease. As microbial communities associated with the gut and skin are vital to host health and disease resilience, disruption of microbiome integrity could contribute to the adverse consequences of stress exposure. Little is known about how stress affects the fish microbiome, especially during sensitive early life stages when initial colonisation and proliferation of host-associated microbial communities take place. Therefore, we compared the effects of two aquaculture-relevant early-life stressors on the gut and skin microbiome of Atlantic salmon fry (four months post hatching) using 16S rRNA amplicon sequencing. Acute cold stress applied during late embryogenesis had a pronounced, lasting effect on the structure of the skin microbiome, as well as a less consistent effect on the gut microbiome. Follow-up targeted qPCR assays suggested that this is likely due to disruption of the egg shell microbial communities at the initial stages of microbiome colonisation, with persistent effects on community structure. In contrast, chronic post hatching stress altered the structure of the gut microbiome, but not that of the skin. Both types of stress promoted similar Gammaproteobacteria ASVs, particularly within the genera Acinetobacter and Aeromonas, which include several important opportunistic fish pathogens. Our results demonstrate the sensitivity of the salmon microbiome to environmental stressors during early life, with potential associated health impacts on the host. We also identified common signatures of stress in the salmon microbiome, which may represent useful microbial stress biomarkers., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Microbes that thrive in premise plumbing can have potentially important effects on human health. Yet, how and why plumbing-associated microbial communities vary across broad spatial scales remain undetermined. We characterized the bacterial communities in 496 showerheads collected from across the continental United States. The overall community structure, determined by 16S rRNA gene amplicon sequencing, revealed high levels of bacterial diversity. Although a large fraction of the observed variation in community composition could not be explained, differences in bacterial community composition were associated with water supply (private well water vs public municipal water), water source (groundwater vs surface water), and associated differences in water chemistry (pH and chlorine). Most notably, showerheads in homes supplied with public water had higher abundances of Blastomonas , Mycobacterium , and Porphyrobacter , while Pseudorhodoplanes , Novosphingobium , and Nitrospira were more abundant in those receiving private well water. We conducted shotgun metagenomic analyses on 92 of these samples to assess differences in genomic attributes. Public water-sourced showerheads had communities enriched in genes related to lipid and xenobiotic metabolisms, virulence factors, and antibiotic resistance. In contrast, genes associated with oxidative stress and membrane transporters were over-represented in communities from private well water-sourced showerheads compared to those supplied by public water systems. These results highlight the broad diversity of bacteria found in premise plumbing across the United States and the role of the water source and treatment in shaping the microbial community structure and functional potential.

Accurate assessment of larval community composition in spawning areas is essential for fisheries management and conservation but is often hampered by the cryptic nature of many larvae, which renders them difficult to identify morphologically. Metabarcoding is a rapid and cost-effective method to monitor early life stages for management and environmental impact assessment purposes but its quantitative capability is under discussion. We compared metabarcoding with traditional morphological identification to evaluate taxonomic precision and reliability of abundance estimates, using 332 fish larvae from multinet hauls (0-50 m depth) collected at 14 offshore sampling sites in the Irish and Celtic seas. To improve quantification accuracy (relative abundance estimates), the amount of tissue for each specimen was standardized and mitochondrial primers (12S gene) with conserved binding sites were used. Relative family abundance estimated from metabarcoding reads and morphological assessment were positively correlated, as well as taxon richness (R S  = 0.81, P = 0.007) and diversity (R S  = 0.90, P = 0.002). Spatial patterns of community composition did not differ significantly between metabarcoding and morphological assessments. Our results show that DNA metabarcoding of bulk tissue samples can be used to monitor changes in fish larvae abundance and community composition. This represents a feasible, efficient, and faster alternative to morphological methods that can be applied to terrestrial and aquatic habitats., (© 2021 The Authors. Ecological Applications published by Wiley Periodicals LLC on behalf of Ecological Society of America.)

Exposure of male fish to estrogenic substances from wastewater treatment works (WwTWs) results in feminization and reduced reproductive fitness. Nevertheless, self-sustaining populations of roach ( Rutilus rutilus ) inhabit river stretches polluted with estrogenic WwTW effluents. In this study, we examine whether such roach populations have evolved adaptations to tolerate estrogenic pollution by comparing frequency differences in single-nucleotide polymorphisms (SNPs) between populations sampled from rivers receiving either high- or low-level WwTW discharges. SNPs within 36 "candidate" genes, selected for their involvement in estrogenic responses, and 120 SNPs in reference genes were genotyped in 465 roaches. There was no evidence for selection in highly estrogen-dependent candidate genes, including those for the estrogen receptors, aromatases, and vitellogenins. The androgen receptor ( ar ) and cytochrome P450 1A genes were associated with large shifts in allele frequencies between catchments and in individual populations, but there is no clear link to estrogen pollution. Selection at ar in the effluent-dominated River Lee may have resulted from historical contamination with endocrine-disrupting pesticides. Critically, although our results suggest population-specific selection including at genes related to endocrine disruption, there was no strong evidence that the selection resulted from exposure to estrogen pollution.

Infectious diseases represent an important barrier to sustainable aquaculture development. Rearing density can substantially impact fish productivity, health and welfare in aquaculture, including growth rates, behaviour and, crucially, immune activity. Given the current emphasis on aquaculture diversification, stress-related indicators broadly applicable across species are needed. Utilising an interspecific comparative transcriptomic (RNAseq) approach, we compared gill gene expression responses of Atlantic salmon (Salmo salar) and Nile tilapia (Oreochromis niloticus) to rearing density and Saprolegnia parasitica infection. Salmon reared at high-density showed increased expression of stress-related markers (e.g. c-fos and hsp70), and downregulation of innate immune genes. Upon pathogen challenge, only salmon reared at low density exhibited increased expression of inflammatory interleukins and lymphocyte-related genes. Tilapia immunity, in contrast, was impaired at low-density. Using overlapping gene ontology enrichment and gene ortholog analyses, we found that density-related stress similarly impacted salmon and tilapia in key immune pathways, altering the expression of genes vital to inflammatory and Th 17 responses to pathogen challenge. Given the challenges posed by ectoparasites and gill diseases in fish farms, this study underscores the importance of optimal rearing densities for immunocompetence, particularly for mucosal immunity. Our comparative transcriptomics analyses identified density stress impacted immune markers common across different fish taxa, providing key molecular targets with potential for monitoring and enhancing aquaculture resilience in a wide range of farmed species., (Copyright © 2020. Published by Elsevier Ltd.)

Exposure to environmental stressors can compromise fish health and fitness. Little is known about how stress-induced microbiome disruption may contribute to these adverse health effects, including how cortisol influences fish microbial communities. We exposed juvenile Atlantic salmon to a mild confinement stressor for two weeks. We then measured cortisol in the plasma, skin-mucus, and feces, and characterized the skin and fecal microbiome. Fecal and skin cortisol concentrations increased in fish exposed to confinement stress, and were positively correlated with plasma cortisol. Elevated fecal cortisol was associated with pronounced changes in the diversity and structure of the fecal microbiome. In particular, we identified a marked decline in the lactic acid bacteria Carnobacterium sp. and an increase in the abundance of operational taxonomic units within the classes Clostridia and Gammaproteobacteria. In contrast, cortisol concentrations in skin-mucus were lower than in the feces, and were not related to any detectable changes in the skin microbiome. Our results demonstrate that stressor-induced cortisol production is associated with disruption of the gut microbiome, which may, in turn, contribute to the adverse effects of stress on fish health. They also highlight the value of using non-invasive fecal samples to monitor stress, including simultaneous determination of cortisol and stress-responsive bacteria., (Copyright © 2020 Uren Webster, Rodriguez-Barreto, Consuegra and Garcia de Leaniz.)

Microbial communities associated with the gut and the skin are strongly influenced by environmental factors, and can rapidly adapt to change. Historical processes may also affect the microbiome. In particular, variation in microbial colonisation in early life has the potential to induce lasting effects on microbial assemblages. However, little is known about the relative extent of microbiome plasticity or the importance of historical colonisation effects following environmental change, especially for nonmammalian species. To investigate this we performed a reciprocal translocation of Atlantic salmon between artificial and semi-natural conditions. Wild and hatchery-reared fry were transferred to three common garden experimental environments for 6 weeks: standard hatchery conditions, hatchery conditions with an enriched diet, and simulated wild conditions. We characterized the faecal and skin microbiome of individual fish before and after the environmental translocation, using a BACI (before-after-control-impact) design. We found evidence of extensive microbiome plasticity for both the gut and skin, with the greatest changes in alpha and beta diversity associated with the largest changes in environment and diet. Microbiome richness and diversity were entirely determined by environment, with no detectable effects of fish origin, and there was also a near-complete turnover in microbiome structure. However, we also identified, for the first time in fish, evidence of historical colonisation effects reflecting early-life experience, including ASVs characteristic of captive rearing. These results have important implications for host adaptation to local selective pressures, and highlight how conditions experienced during early life can have a long-term influence on the microbiome and, potentially, host health., (© 2020 The Authors. Molecular Ecology published by John Wiley & Sons Ltd.)

Arsenic field test kits are widely used to measure arsenic levels in drinking water sources, especially in countries like Bangladesh, where water supply is highly decentralized and water quality testing infrastructure is limited. From a public health perspective, the ability of a measurement technique to distinguish samples above and below relevant and actionable drinking water standards is paramount. In this study, the performance of eight commercially available field test kits was assessed by comparing kit estimates to hydride generation atomic absorption spectroscopy (HG-AAS) analyses. The results of tests that control for user-dependent color matching errors showed that two kits (LaMotte and Quick II kits) provided accurate and precise estimates of arsenic, four kits (Econo-Quick, Quick, Wagtech and Merck kits) were either accurate or precise, but not both, and two kits (Hach and Econo-Quick II kits) were neither accurate nor precise. Tests were performed for arsenic concentration ranges commonly found in natural waters and treated waters (such as community drinking water filter systems), and also on laboratory generated arsenic standards in DI water. For those kits that did not perform well, test strips often produced colors too light compared to manufacturer-provided arsenic color calibration charts. Based on these results, we recommend stakeholders carefully re-consider the use of poorly performing field test kits until better quality control of components of these kits is implemented. In addition, we recommend that field test kit manufacturers provide suitable internal standards in every kit box for users to verify the veracity of manufacturer provided color charts., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Plant-microbe interactions are mediated by signaling compounds that control vital plant functions, such as nodulation, defense, and allelopathy. While interruption of signaling is typically attributed to biological processes, potential abiotic controls remain less studied. Here, we show that higher organic carbon (OC) contents in soils repress flavonoid signals by up to 70%. Furthermore, the magnitude of repression is differentially dependent on the chemical structure of the signaling molecule, the availability of metal ions, and the source of the plant-derived OC. Up to 63% of the signaling repression occurs between dissolved OC and flavonoids rather than through flavonoid sorption to particulate OC. In plant experiments, OC interrupts the signaling between a legume and a nitrogen-fixing microbial symbiont, resulting in a 75% decrease in nodule formation. Our results suggest that soil OC decreases the lifetime of flavonoids underlying plant-microbe interactions., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Biosand filtration systems are widely used for drinking water treatment, from household-level, intermittently operated filters to large-scale continuous municipal systems. While it is well-established that microbial activity within the filter is essential for the removal of potential pathogens and other contaminants, the microbial ecology of these systems and how microbial succession relates to their performance remain poorly resolved. We determined how different source waters influence the composition, temporal dynamics, and performance of microbial communities in intermittently operated biosand filters. We operated lab-scale biosand filters, adding daily inputs from two contrasting water sources with differing nutrient concentrations and found that total coliform removal increased and became less variable after 4 weeks, regardless of water source. Total effluent biomass was also lower than total influent biomass for both water sources. Bacterial community composition, assessed via cultivation-independent DNA sequencing, varied by water source, sample type (influent, effluent, or sand), and time. Despite these differences, we identified specific taxa that were consistently removed, including common aquatic and wastewater bacteria. In contrast, taxa consistently more abundant in the sand and effluent included predatory, intracellular, and symbiotic bacteria. IMPORTANCE Although microbial activities are known to contribute to the effectiveness of biosand filtration for drinking water treatment, we have a limited understanding of what microbial groups are most effectively removed, colonize the sand, or make it through the filter. This study tracked the microbial communities in the influent, sand, and effluent of lab-scale, intermittently operated biosand filters over 8 weeks. These results represent the most detailed and time-resolved investigation of the microbial communities in biosand filters typical of those implemented at the household level in many developing countries. We show the importance of the microbial food web in biosand filtration, and we identified taxa that are preferentially removed from wastewater-impacted water sources. We found consistent patterns in filter effectiveness from source waters with differing nutrient loads and, likewise, identified specific bacterial taxa that were consistently more abundant in effluent waters, taxa that are important targets for further study and posttreatment., (Copyright © 2019 American Society for Microbiology.)

To meet future global demand for fish protein, more fish will need to be farmed using fewer resources, and this will require the selection of nonaggressive individuals that perform well at high densities. Yet, the genetic changes underlying loss of aggression and adaptation to crowding during aquaculture intensification are largely unknown. We examined the transcriptomic response to aggression and crowding in Nile tilapia, one of the oldest and most widespread farmed fish, whose social structure shifts from social hierarchies to shoaling with increasing density. A mirror test was used to quantify aggression and skin darkening (a proxy for stress) of fish reared at low and high densities, and gene expression in the hypothalamus was analysed among the most and least aggressive fish at each density. Fish reared at high density were darker, had larger brains, were less active and less aggressive than those reared at low density and had differentially expressed genes consistent with a reactive stress-coping style and activation of the hypothalamus-pituitary-interrenal (HPI) axis. Differences in gene expression among aggressive fish were accounted for by density and the interaction between density and aggression levels, whereas for nonaggressive fish differences in gene expression were associated with individual variation in skin brightness and social stress. Thus, the response to crowding in Nile tilapia is context dependent and involves different neuroendocrine pathways, depending on social status. Knowledge of genes associated with the response to crowding may pave the way for more efficient fish domestication, based on the selection of nonaggressive individuals with increasing tolerance to chronic stress necessary for aquaculture intensification., Competing Interests: The authors declare no conflict of interest.

Bacteria within the genus Mycobacterium can be abundant in showerheads, and the inhalation of aerosolized mycobacteria while showering has been implicated as a mode of transmission in nontuberculous mycobacterial (NTM) lung infections. Despite their importance, the diversity, distributions, and environmental predictors of showerhead-associated mycobacteria remain largely unresolved. To address these knowledge gaps, we worked with citizen scientists to collect showerhead biofilm samples and associated water chemistry data from 656 households located across the United States and Europe. Our cultivation-independent analyses revealed that the genus Mycobacterium was consistently the most abundant genus of bacteria detected in residential showerheads, and yet mycobacterial diversity and abundances were highly variable. Mycobacteria were far more abundant, on average, in showerheads receiving municipal water than in those receiving well water and in U.S. households than in European households, patterns that are likely driven by differences in the use of chlorine disinfectants. Moreover, we found that water source, water chemistry, and household location also influenced the prevalence of specific mycobacterial lineages detected in showerheads. We identified geographic regions within the United States where showerheads have particularly high abundances of potentially pathogenic lineages of mycobacteria, and these "hot spots" generally overlapped those regions where NTM lung disease is most prevalent. Together, these results emphasize the public health relevance of mycobacteria in showerhead biofilms. They further demonstrate that mycobacterial distributions in showerhead biofilms are often predictable from household location and water chemistry, knowledge that advances our understanding of NTM transmission dynamics and the development of strategies to reduce exposures to these emerging pathogens. IMPORTANCE Bacteria thrive in showerheads and throughout household water distribution systems. While most of these bacteria are innocuous, some are potential pathogens, including members of the genus Mycobacterium that can cause nontuberculous mycobacterial (NTM) lung infection, an increasing threat to public health. We found that showerheads in households across the United States and Europe often harbor abundant mycobacterial communities that vary in composition depending on geographic location, water chemistry, and water source, with households receiving water treated with chlorine disinfectants having particularly high abundances of certain mycobacteria. The regions in the United States where NTM lung infections are most common were the same regions where pathogenic mycobacteria were most prevalent in showerheads, highlighting the important role of showerheads in the transmission of NTM infections., (Copyright © 2018 Gebert et al.)

Background: Captive animal populations, be it for food production or conservation programmes, are often maintained at densities far beyond those in natural environments, which can have profound effects on behaviour, immune and stress levels, and ultimately welfare. How such alterations impact transcriptional responses to pathogen infection is a 'different kettle of fish' and remains poorly understood. Here, we assessed survival and gene expression profiles of infected fish reared at two different densities to elucidate potential functional genomic mechanisms for density-related differences in disease susceptibility., Results: Utilising a whole-transcriptome sequencing (RNAseq) approach, we demonstrate that rearing density in tilapia (Oreochromis niloticus) significantly impacts susceptibility to the oomycete Saprolegnia parasitica, via altered transcriptional infection responses. Tilapia held at low densities have increased expression of genes related to stress, likely due to increased aggressive interactions. When challenged with Saprolegnia, low-density fish exhibit altered expression of inflammatory gene responses and enhanced levels of adaptive immune gene suppression compared to fish reared at higher density, resulting in significantly increased mortality rates. In addition, Saprolegnia infection substantially perturbs expression of circadian clock genes, with fish reared at low-density having higher levels of molecular clock dysregulation., Conclusions: Our results reveal the wide-scale impact of stocking density on transcriptional responses to infection and highlight the need to incorporate circadian biology into our understanding of disease dynamics in managed animals.

The microbiome has a crucial influence on host phenotype and is of broad interest to ecological and evolutionary research. Yet, the extent of variation that occurs in the microbiome within and between populations is unclear. We characterized the skin and gut microbiomes of seven populations of juvenile Atlantic salmon ( Salmo salar ) inhabiting a diverse range of environments, including hatchery-reared and wild populations. We found shared skin operational taxonomic units (OTUs) across all populations and core gut microbiota for all wild fish, but the diversity and structure of both skin and gut microbial communities were distinct between populations. There was a marked difference between the gut microbiomes of wild and captive fish. Hatchery-reared fish had lower intestinal microbial diversity, lacked core microbiota found in wild fish, and showed altered community structure and function. Skin and gut microbiomes were also less varied within captive populations, reflecting more uniform artificial rearing conditions. The surrounding water influenced the microbiome of the gut and, especially, the skin, but could not explain the degree of variation observed between populations. For both gut and skin, we found that there was greater difference in microbiome structures between more genetically distinct fish populations, and that population genetic diversity was positively correlated with microbiome diversity. However, diet is likely to be the major factor contributing to the large differences in gut microbiota between wild and captive fish. Our results highlight the scope of interpopulation variation in the Atlantic salmon microbiome and offer insights into the deterministic factors contributing to microbiome diversity and structure. IMPORTANCE Variation in the microbiome has a fundamental influence on host health, ecology, and evolution, but the scope and basis of this variation are not fully understood. We identified considerable variation in skin and gut microbial communities between seven wild and captive populations of Atlantic salmon, reflecting divergent environmental conditions and fish genetic diversity. In particular, we found very pronounced differences in the intestinal microbiomes of wild and hatchery-reared fish, likely reflecting differences in diet. Our results offer an insight into how the microbiome potentially contributes to the generation of local adaptations in this species and how domestication alters intestinal microbial communities, highlighting future research directions in these areas., (Copyright © 2018 Uren Webster et al.)

Reproduction is an essential process for life and is regulated by complex hormone networks and environmental factors. To date, little is known about the contribution of epigenetic mechanisms to the regulation of reproduction, particularly in lower vertebrates. We used the zebrafish (Danio rerio) model to investigate the sex-specific transcription and DNA methylation profiles for genes involved in the regulation of reproduction and in epigenetic signalling in the livers and gonads. We found evidence for associations between DNA promotor methylation and transcription for esr1 (gonads and female livers), amh (gonads) and dnmt1 (livers). In the liver, esr1 was shown to be significantly over-expressed in females compared to males, and its promoter was significantly hypo-methylated in females compared to males. In the gonads, genes involved in epigenetic processes including dnmt1, dnmt3 and hdac1 were over-expressed in the ovary compared to the testis. In addition, dnmt1 and dnmt3 transcription in the testis was found to be strongly correlated with global DNA methylation. These data provide evidence of the sex-specific epigenetic regulation and transcription of genes involved in reproduction and epigenetic signalling in a commonly used vertebrate model., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The data presented here corresponds to the research paper "Simultaneous detection of invasive signal crayfish, endangered white-clawed crayfish and the crayfish plague using environmental DNA". A crayfish-specific assay was designed and optimised using three real-time PCR supermixes (SYBR™ Green, SsoFast™ EvaGreen® and HOT FIREPol® EvaGreen®). Diagnostic high resolution melt (HRM) data from direct application of assay on both ex-situ eDNA water samples and field samples from four catchments (two in Wales, two in England) is presented in this article, displaying positive HRM profiles for invasive signal crayfish ( Pacifastacus leniusculus), native white-clawed crayfish ( Austropotamobius pallipes ) and crayfish plague causal agent ( Aphanomyces astaci ).

Inhibition of anaerobic digestion (AD) due to perturbation caused by substrate composition and/or operating conditions can significantly reduce performance. Such perturbations could be limited by elucidating microbial community response to inhibitors and devising strategies to increase community resilience. To this end, advanced molecular methods are increasingly being applied to study the AD microbiome, a diverse community of microbial populations with complex interactions. This literature review of AD inhibition studies indicates that inhibitory concentrations are highly variable, likely stemming from differences in community structure or activity profile and previous exposure to inhibitors. More recent molecular methods such as 'omics' tools, substrate mapping, and real-time sequencing are helping to unravel the complexity of AD inhibition by elucidating physiological and ecological significance of key microbial populations. The AD community must strive towards developing predictive abilities to avoid system failure (e.g., real-time tracking of an indicator species) to improve resilience of AD systems., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Parasites are strong drivers of evolutionary change and the genetic variation of both host and parasite populations can co-evolve as a function of parasite virulence and host resistance. The role of transcriptome variation in specific interactions between host and parasite genotypes has been less studied and can be confounded by differences in genetic variation. We employed two naturally inbred lines of a self-fertilizing fish to estimate the role of host genotype in the transcriptome response to parasite infection using RNA-seq. In addition, we targeted several differentially expressed immune-related genes to further investigate the relative role of individual variation in the immune response using RT-qPCR, taking advantage of the genomic uniformity of the self-fertilizing lines. We found significant differences in gene expression between lines in response to infection both in the transcriptome and in individual gene RT-qPCR analyses. Individual RT-qPCR analyses of gene expression identified significant variance differences between lines for six genes but only for three genes between infected and control fish. Our results indicate that although the genetic background plays an important role in the transcriptome response to parasites, it cannot fully explain individual differences within genetically homogeneous lines, which can be important for determining the response to parasites., (© The Author(s) 2018. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Stress experienced during early life may have lasting effects on the immune system, with impacts on health and disease dependent on the nature and duration of the stressor. The epigenome is especially sensitive to environmental stimuli during early life and represents a potential mechanism through which stress may cause long-lasting health effects. However, the extent to which the epigenome responds differently to chronic vs acute stressors is unclear, especially for non-mammalian species. We examined the effects of acute stress (cold-shock during embryogenesis) and chronic stress (absence of tank enrichment during larval-stage) on global gene expression (using RNA-seq) and DNA methylation (using RRBS) in the gills of Atlantic salmon (Salmo salar) four months after hatching. Chronic stress induced pronounced transcriptional differences, while acute stress caused few lasting transcriptional effects. However, both acute and chronic stress caused lasting and contrasting changes in the methylome. Crucially, we found that acute stress enhanced transcriptional immune response to a pathogenic challenge (bacterial lipopolysaccharide, LPS), while chronic stress suppressed it. We identified stress-induced changes in promoter and gene-body methylation that were associated with altered expression for a small proportion of immune-related genes, and evidence of wider epigenetic regulation within signalling pathways involved in immune response. Our results suggest that stress can affect immuno-competence through epigenetic mechanisms, and highlight the markedly different effects of chronic larval and acute embryonic stress. This knowledge could be used to harness the stimulatory effects of acute stress on immunity, paving the way for improved stress and disease management through epigenetic conditioning.

Competing Interests: Competing interests: The authors report no competing personal or financial interests.

Some fish populations inhabiting contaminated environments show evidence of increased chemical tolerance, however the mechanisms contributing to this tolerance, and whether this is heritable, are poorly understood. We investigated the responses of two populations of wild three-spined stickleback (Gasterosteus aculeatus) with different histories of contaminant exposure to an oestrogen and copper, two widespread aquatic pollutants. Male stickleback originating from two sites, the River Aire, with a history of complex pollution discharges, and Siblyback Lake, with a history of metal contamination, were depurated and then exposed to copper (46μg/L) and the synthetic oestrogen ethinyloestradiol (22ng/L). The hepatic transcriptomic response was compared between the two populations and to a reference population with no known history of exposure (Houghton Springs, Dorset). Gene responses included those typical for both copper and oestrogen, with no discernable difference in response to oestrogen between populations. There was, however, some difference in the magnitude of response to copper between populations. Siblyback fish showed an elevated baseline transcription of genes encoding metallothioneins and a lower level of metallothionein induction following copper exposure, compared to those from the River Aire. Similarly, a further experiment with an F1 generation of Siblyback fish bred in the laboratory found evidence for elevated transcription of genes encoding metallothioneins in unexposed fish, together with an altered transcriptional response to 125μg/L copper, compared with F1 fish originating from the clean reference population exposed to the same copper concentration. These data suggest that the stickleback from Siblyback Lake have a differential response to copper, which is inherited by the F1 generation in laboratory conditions, and for which the underlying mechanism may include an elevation of baseline transcription of genes encoding metallothioneins. The genetic and/or epigenetic mechanisms contributing to this inherited alteration of metallothionein transcription have yet to be established., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Many fish populations are exposed to harmful levels of chemical pollution and selection pressures associated with these exposures have led to the evolution of tolerance. Our understanding of the physiological basis for these adaptations is limited, but they are likely to include processes involved with the absorption, distribution, metabolism and/or excretion of the target chemical. Other potential adaptive mechanisms include enhancements in antioxidant responses, an increased capacity for DNA and/or tissue repair and alterations to the life cycle of fish that enable earlier reproduction. Analysis of single-nucleotide polymorphism frequencies has shown that tolerance to hydrocarbon pollutants in both marine and estuarine fish species involves alteration in the expression of the xenobiotic metabolism enzyme CYP1A. In this review, we present novel data showing also that variants of the CYP1A gene have been under selection in guppies living in Trinidadian rivers heavily polluted with crude oil. Potential costs associated with these adaptations could reduce fitness in unpolluted water conditions. Integrating knowledge of local adaptation to pollution is an important future consideration in conservation practices such as for successful restocking, and improving connectivity within river systems.This article is part of the themed issue 'Human influences on evolution, and the ecological and societal consequences'., (© 2016 The Authors.)

The removal of arsenic from drinking water sources produces arsenic-bearing wastes, which are disposed of in a variety of ways. Several disposal options involve anaerobic environments, including mixing arsenic waste with cow dung, landfills, anaerobic digesters, and pond sediments. Though poorly understood, the production of gaseous arsenic species in these environments can be a primary goal (cow dung mixing) or an unintended consequence (anaerobic digesters). Once formed, these gaseous arsenic species are readily diluted in the atmosphere. Arsenic volatilization can be mediated by the enzyme arsenite S-adenosylmethionine methyltransferase (ArsM) or through the enzymes involved in methanogenesis. In this study, methanogenic mesocosms with arsenic-bearing ferric iron waste from an electrocoagulation drinking water treatment system were used to evaluate the role of methanogenesis in arsenic volatilization using methanogen inhibitors. Arsenic volatilization was highest in methanogenic mesocosms, but represented <0.02% of the total arsenic added. 16S rRNA cDNA sequencing, qPCR of mcrA transcripts, and functional gene array-based analysis of arsM expression, revealed that arsenic volatilization correlated with methanogenic activity. Aqueous arsenic concentrations increased in all mesocosms, indicating that unintended contamination may result from disposal in anaerobic environments. This highlights that more research is needed before recommending anaerobic disposal intended to promote arsenic volatilization.

Methanogenic inhibitors are often used to study methanogenesis in complex microbial communities or inhibit methanogens in the gastrointestinal tract of livestock. However, the resulting structural and functional changes in archaeal and bacterial communities are poorly understood. We characterized microbial community structure and activity in mesocosms seeded with cow dung and municipal wastewater treatment plant anaerobic digester sludge after exposure to two methanogenic inhibitors, 2-bromoethanesulfonate (BES) and propynoic acid (PA). Methane production was reduced by 89% (0.5 mmol/L BES), 100% (10 mmol/LBES), 24% (0.1 mmol/LPA), and 95% (10 mmol/LPA). Using modified primers targeting the methyl-coenzyme M reductase (mcrA) gene, changes in mcrA gene expression were found to correspond with changes in methane production and the relative activity of methanogens. Methanogenic activity was determined by the relative abundance of methanogen 16S rRNA cDNA as a percentage of the total community 16S rRNA cDNA. Overall, methanogenic activity was lower when mesocosms were exposed to higher concentrations of both inhibitors, and aceticlastic methanogens were inhibited to a greater extent than hydrogenotrophic methanogens. Syntrophic bacterial activity, measured by 16S rRNA cDNA, was also reduced following exposure to both inhibitors, but the overall structure of the active bacterial community was not significantly affected., (© 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Bisphenol A (BPA) is a commercially important high production chemical widely used in epoxy resins and polycarbonate plastics, and is ubiquitous in the environment. Previous studies demonstrated that BPA activates estrogenic signaling pathways associated with adverse effects on reproduction in vertebrates and that exposure can induce epigenetic changes. We aimed to investigate the reproductive effects of BPA in a fish model and to document its mechanisms of toxicity. We exposed breeding groups of zebrafish (Danio rerio) to 0.01, 0.1, and 1 mg/L BPA for 15 d. We observed a significant increase in egg production, together with a reduced rate of fertilization in fish exposed to 1 mg/L BPA, associated with significant alterations in the transcription of genes involved in reproductive function and epigenetic processes in both liver and gonad tissue at concentrations representing hotspots of environmental contamination (0.1 mg/L) and above. Of note, we observed reduced expression of DNA methyltransferase 1 (dnmt1) at environmentally relevant concentrations of BPA, along with a significant reduction in global DNA methylation, in testes and ovaries following exposure to 1 mg/L BPA. Our findings demonstrate that BPA disrupts reproductive processes in zebrafish, likely via estrogenic mechanisms, and that environmentally relevant concentrations of BPA are associated with altered transcription of key enzymes involved in DNA methylation maintenance. These findings provide evidence of the mechanisms of action of BPA in a model vertebrate and advocate for its reduction in the environment.

Hypoxia is a global and increasingly important stressor in aquatic ecosystems, with major impacts on biodiversity worldwide. Hypoxic waters are often contaminated with a wide range of chemicals but little is known about the interactions between these stressors. We investigated the effects of hypoxia on the responses of zebrafish (Danio rerio) embryos to copper, a widespread aquatic contaminant. We showed that during continuous exposures copper toxicity was reduced by over 2-fold under hypoxia compared to normoxia. When exposures were conducted during 24 h windows, hypoxia reduced copper toxicity during early development and increased its toxicity in hatched larvae. To investigate the role of the hypoxia signaling pathway on the suppression of copper toxicity during early development, we stabilized the hypoxia inducible factor (HIF) pathway under normoxia using a prolyl-4-hydroxylase inhibitor, dimethyloxalylglycine (DMOG) and demonstrated that HIF activation results in a strong reduction in copper toxicity. We also established that the reduction in copper toxicity during early development was independent of copper uptake, while after hatching, copper uptake was increased under hypoxia, corresponding to an increase in copper toxicity. These findings change our understanding of the current and future impacts of worldwide oxygen depletion on fish communities challenged by anthropogenic toxicants.

The performance of a vinegar-amended anaerobic biosand filter was evaluated for future application as point-of-use water treatment in rural areas for the removal of arsenic and nitrate from groundwater containing common ions. Due to the importance of sulfate and iron in arsenic removal and their variable concentrations in groundwater, influent sulfate and iron concentrations were varied. Complete removal of influent nitrate (50 mg/L) and over 50% removal of influent arsenic (200 μg/L) occurred. Of all conditions tested, the lowest median effluent arsenic concentration was 88 μg/L. Iron removal occurred completely when 4 mg/L was added, and sulfate concentrations were lowered to a median concentration <2 mg/L from influent concentrations of 22 and 50 mg/L. Despite iron and sulfate removal and the establishment of reducing conditions, arsenic concentrations remained above the World Health Organization's arsenic drinking water standard. Further research is necessary to determine if anaerobic biosand filters can be improved to meet the arsenic drinking water standard and to evaluate practical implementation challenges., (Copyright © 2016. Published by Elsevier Ltd.)

Estrogenic chemicals are major contaminants of surface waters and can threaten the sustainability of natural fish populations. Characterization of the global molecular mechanisms of toxicity of environmental contaminants has been conducted primarily in model species rather than species with limited existing transcriptomic or genomic sequence information. We aimed to investigate the global mechanisms of toxicity of an endocrine disrupting chemical of environmental concern [17β-estradiol (E2)] using high-throughput RNA sequencing (RNA-Seq) in an environmentally relevant species, brown trout (Salmo trutta). We exposed mature males to measured concentrations of 1.94, 18.06, and 34.38 ng E2/l for 4 days and sequenced three individual liver samples per treatment using an Illumina HiSeq 2500 platform. Exposure to 34.4 ng E2/L resulted in 2,113 differentially regulated transcripts (FDR < 0.05). Functional analysis revealed upregulation of processes associated with vitellogenesis, including lipid metabolism, cellular proliferation, and ribosome biogenesis, together with a downregulation of carbohydrate metabolism. Using real-time quantitative PCR, we validated the expression of eight target genes and identified significant differences in the regulation of several known estrogen-responsive transcripts in fish exposed to the lower treatment concentrations (including esr1 and zp2.5). We successfully used RNA-Seq to identify highly conserved responses to estrogen and also identified some estrogen-responsive transcripts that have been less well characterized, including nots and tgm2l. These results demonstrate the potential application of RNA-Seq as a valuable tool for assessing mechanistic effects of pollutants in ecologically relevant species for which little genomic information is available., (Copyright © 2015 the American Physiological Society.)

The herbicide linuron is used worldwide, and has been detected in surface waters as well as in food and drinking water. Toxicological studies have reported that linuron acts as an antiandrogen in vitro and in vivo and disrupts mammalian male reproductive function. However, global mechanisms of linuron toxicity are poorly documented. We used RNA-seq to characterize the hepatic transcriptional response of mature male brown trout exposed for 4 days to 1.7, 15.3, and 225.9 μg/L linuron. We identified a striking decrease in the expression of transcripts encoding the majority of enzymes forming the cholesterol biosynthesis pathway. We also measured a very significant decrease in total hepatic cholesterol in fish exposed to 225.9 μg/L linuron and a negative correlation between total cholesterol and linuron treatment concentration. We hypothesize that inhibition of cholesterol biosynthesis may result from the disruption of androgen signaling by linuron. Additionally, there was increased expression of a number of transcripts involved in cellular stress responses, including cyp1a (up to 560-fold), molecular chaperones, and antioxidant enzymes. We found some evidence of similar patterns of transcriptional change in fish exposed to an environmentally relevant concentration of linuron, and further research should investigate the potential for adverse effects to occur following chronic environmental exposure.

Background: Glyphosate, the active ingredient in Roundup formulations, is the most widely used herbicide worldwide, and as a result contaminates surface waters and has been detected in food residues, drinking water and human urine, raising concerns for potential environmental and human health impacts. Research has shown that glyphosate and Roundup can induce a broad range of biological effects in exposed organisms, particularly via generation of oxidative stress. However, there has been no comprehensive investigation of the global molecular mechanisms of toxicity of glyphosate and Roundup for any species. We aimed to characterise and compare the global mechanisms of toxicity of glyphosate and Roundup in the liver of brown trout (Salmo trutta), an ecologically and economically important vertebrate species, using RNA-seq on an Illumina HiSeq 2500 platform. To do this, we exposed juvenile female brown trout to 0, 0.01, 0.5 and 10 mg/L of glyphosate and Roundup (glyphosate acid equivalent) for 14 days, and sequenced 6 replicate liver samples from each treatment., Results: We assembled the brown trout transcriptome using an optimised de novo approach, and subsequent differential expression analysis identified a total of 1020 differentially-regulated transcripts across all treatments. These included transcripts encoding components of the antioxidant system, a number of stress-response proteins and pro-apoptotic signalling molecules. Functional analysis also revealed over-representation of pathways involved in regulating of cell-proliferation and turnover, and up-regulation of energy metabolism and other metabolic processes., Conclusions: These transcriptional changes are consistent with generation of oxidative stress and the widespread induction of compensatory cellular stress response pathways. The mechanisms of toxicity identified were similar across both glyphosate and Roundup treatments, including for environmentally relevant concentrations. The significant alterations in transcript expression observed at the lowest concentrations tested raises concerns for the potential toxicity of this herbicide to fish populations inhabiting contaminated rivers.

Sugar beet (Beta vulgaris L.) is not currently a commercial crop in Georgia, but experimental plantings as a winter rotational crop are promising in terms of yield and industrial sugar production (T. Brenneman, personal communication). A disease outbreak of suspected bacterial origin occurred in sugar beet plots (experimental lines Beta Seed energy beet 'BTS ENC115,' 'BTS EGC184,' 'BTS EGC195,' and 'BTS 1EN6702') in Tift Co., GA, in December 2012, at ~35% incidence. Foliar symptoms included circular to irregular spots, each with a tan center and dark margin. Ten leaves/experimental line with leaf spot symptoms were collected, and bacterial isolations made on King's B agar medium. After 48 h of incubation, cream-colored, fluorescent yellow, round colonies with smooth margins were isolated. The isolates were each gram negative, oxidase negative, non-pectolytic on potato, arginine dihydrolase negative, produced levan, and gave a hypersensitivity response (HR) on tobacco. These characteristics indicated that the isolates belonged to Pseudomonas syringae van Hall LOPAT group Ia (3). The 16S-23S rRNA (internal transcribed regions) (1) from four foliar isolates (SB-1, SB-2, SB-3, and SB-4), one/experimental line, was amplified, and the resultant PCR products were sequenced and BLAST searched in GenBank. The 16S-23S rRNA sequences matched those of P. syringae pv. syingae (Pss) (KF023189) and P. syringae pv. aptata (Psa) (AY342167.1) with 96 to 98% and 97 to 99% sequence identity, respectively. Also, the percent similarity of the 16S-23S rRNA sequences among the four isolates was >99% (KJ922021 to 24 for SB-1 to SB-4, respectively). The four test isolates also had ≤89 and ≤99% similarity with Pss and Psa, respectively, when tested with BIOLOG (Hayward, CA). In addition, four sugarbeet isolates along with a type strain of Psa (NCPPB 3539) were amplified using a PCR primer pair that detected the presence of the avrPphE gene, an avirulence gene present in Psa but absent in Pss (2). The type strain of Pss (NCPPB 1770) was not amplified using this primer pair. BOX-PCR analysis gave identical banding patterns for the four isolates as that of a type strain of Psa. In two independent experiments, 3-week-old seedlings of the sugar beet cv. Beta EGR099 (n = 10 seedlings/isolate/experiment) were spray-inoculated with a sterilized water suspension of 1 × 10 8 CFU/ml of each of the isolates. All of the inoculated seedlings developed symptoms (water-soaked lesions that developed into necrotic spots) 10 days after inoculation (DAI) in greenhouse conditions (~30°C and ~80% RH). All of the seedlings inoculated with the type strain of Psa also produced typical bacterial blight symptoms at 10 DAI. In contrast, five control seedlings inoculated with sterilized water remained asymptomatic, and target bacterial colonies were not re-isolated from the leaves of these plants. Bacterial colonies were re-isolated from symptomatic seedlings, and showed similar characteristics based on physiological tests, BIOLOG profile, BOX-PCR analysis, and positive amplification with the avrPphE PCR assay, which indicated that these strains were Psa. To our knowledge, this is the first report of Psa in sugarbeet in Georgia. The fact that a Psa strain was also isolated from a sugar beet seed lot (data not shown) suggested that the pathogen may have been introduced on contaminated seeds. Knowledge of the presence of Psa in the agro-ecosystem of Georgia may encourage scientists to implement integrated management practices for this pathogen. References: (1) C. Guasp et al. Int. J. Syst. Evol. Microbiol. 50:1629, 2000. (2) Y. Inoue and Y. Takikawa. Page 687 in: Presentations 6th Int. Conf. Pseudomonas syringae Pathovars and Related Pathogens, 2003. (3) R. A. Lelliot et al. J. Appl. Bacteriol. 29:470, 1966.

Yellow bud, caused by Pseudomonas sp., is an emerging bacterial disease of onion. A polymerase chain reaction assay based on the coronafacate ligase (cfl) and HrpZ genes was used to detect initial suspected bacteria on weeds. Growth on an agar medium, ability to cause a hypersensitive response in tobacco, pathogenicity on onion, and sequence analysis of 16S ribosomal RNA and cfl genes were used to confirm the identity of Pseudomonas sp. recovered from 10 asymptomatic weed species in the Vidalia onion-growing zone (VOZ) of Georgia. Among the weeds identified as epiphytic hosts for Pseudomonas sp., Italian ryegrass (Lolium multiflorum) and curly dock (Rumex crispus) were prominent because ≥73% of the samples from five sample sites were positive for the bacterium. These weeds are commonly found throughout Georgia and, thus, were selected to assess their role in yellow bud epidemiology. Samples of the two weed species were collected from sites along the perimeter of and within the VOZ (n = 5 sites) during late June, August, and September 2012 and 2013, which represented the time interval between onion growing seasons. Samples (n = 10/weed species/site) were collected and processed for bacterial detection as described above. In June (2012 and 2013), Pseudomonas sp. was detected from Italian ryegrass and curly dock in 100 and 40% of the sample sites, respectively. During the months of August and September (2012), the bacterium was recovered from Italian ryegrass in 60 and 10% of the sample sites, respectively; whereas, in August (2013), Pseudomonas sp. was recovered from 40% of the sample sites. However, the bacterium was not recovered from any of the sites in September (2013). In contrast, during August and September (2012), Pseudomonas sp. was recovered from curly dock in 20 and 80% of the sample sites, respectively. Similarly, in August and September (2013), the bacterium was detected from 40 and 100% of the sample sites, respectively. These data demonstrated that the Pseudomonas sp. responsible for yellow bud can survive as an epiphyte on Italian ryegrass and curly dock between onion crops. Furthermore, using artificially infested onion seed, we demonstrated that Pseudomonas sp. can be transmitted through contaminated seed.

Roundup and its active ingredient glyphosate are among the most widely used herbicides worldwide and may contaminate surface waters. Research suggests both Roundup and glyphosate induce oxidative stress in fish and may also cause reproductive toxicity in mammalian systems. We aimed to investigate the reproductive effects of Roundup and glyphosate in fish and the potential associated mechanisms of toxicity. To do this, we conducted a 21-day exposure of breeding zebrafish (Danio rerio) to 0.01, 0.5, and 10 mg/L (glyphosate acid equivalent) Roundup and 10 mg/L glyphosate. 10 mg/L glyphosate reduced egg production but not fertilization rate in breeding colonies. Both 10 mg/L Roundup and glyphosate increased early stage embryo mortalities and premature hatching. However, exposure during embryogenesis alone did not increase embryo mortality, suggesting that this effect was caused primarily by exposure during gametogenesis. Transcript profiling of the gonads revealed 10 mg/L Roundup and glyphosate induced changes in the expression of cyp19a1 and esr1 in the ovary and hsd3b2, cat, and sod1 in the testis. Our results demonstrate that these chemicals cause reproductive toxicity in zebrafish, although only at high concentrations unlikely to occur in the environment, and likely mechanisms of toxicity include disruption of the steroidogenic biosynthesis pathway and oxidative stress.

Worldwide, a number of viable populations of fish are found in environments heavily contaminated with metals, including brown trout (Salmo trutta) inhabiting the River Hayle in South-West of England. This population is chronically exposed to a water-borne mixture of metals, including copper and zinc, at concentrations lethal to naïve fish. We aimed to investigate the molecular mechanisms employed by the River Hayle brown trout to tolerate high metal concentrations. To achieve this, we combined tissue metal analysis with whole-transcriptome profiling using RNA-seq on an Illumina platform. Metal concentrations in the Hayle trout, compared to fish from a relatively unimpacted river, were significantly increased in the gills, liver and kidney (63-, 34- and 19-fold respectively), but not the gut. This confirms that these fish can tolerate considerable metal accumulation, highlighting the importance of these tissues in metal uptake (gill), storage and detoxification (liver, kidney). We sequenced, assembled and annotated the brown trout transcriptome using a de novo approach. Subsequent gene expression analysis identified 998 differentially expressed transcripts and functional analysis revealed that metal- and ion-homeostasis pathways are likely to be the most important mechanisms contributing to the metal tolerance exhibited by this population.

Objective: : Cancer Care Ontario has published an evidence-based guideline on their website "Guideline for Optimization of Surgical and Pathological Quality Performance for Radical Prostatectomy in Prostate Cancer Management: Surgical and Pathological Guidelines." The evidentiary base for this guideline was recently published in CUAJ. The CCO guideline proposes the following: a positive surgical margin (PSM) rate of <25% for organ-confined disease (pT2), a perioperative mortality of <1%, a rate of rectal injury <1%, and a blood transfusion rate <10% in non-anemic patients. The objective of this study was to review the radical prostatectomy practice at the Grey Bruce Health Services, an Ontario community hospital, and to compare our performance in relation to the Cancer Care Ontario guideline and the literature., Methods: : We conducted a retrospective review of all radical prostatectomies performed at the Grey Bruce Health Services from January 1, 2006 to December 31, 2007. The following data were obtained from clinical records and pathology reports: patient age, pre-biopsy prostate-specific antigen, biopsy Gleason score, resected prostate gland weight, radical prostatectomy Gleason score, surgical margin status, pathological tumour stage (pT), lymph node dissection status, perioperative incidence of transfusion of blood products and if the patient was anemic (hemoglobin <140 g/L) preoperatively, incidence of rectal injury, and perioperative mortality within 30 days following surgery., Results: : Using the method proposed by D'Amico, most patients undergoing radical prostatectomy were intermediate risk (62%), with a minority of low-risk (24%) and high-risk (14%) patients. The overall PSM rate was 37%. The rate of PSMs in organ-confined disease (pT2) was 26%. There was a statistically significant trend between increasing D'Amico risk category and increasing rate of PSM (Cochran-Armitage trend test, p = 0.023). There was a strong correlation between the pathological tumour stage and the rate of PSM (Cochran-Armitage trend test, p = 0.0003). The rate of blood transfusion in non-anemic patients was 6%. There was 1 patient (0.8%) who experienced a rectal injury. There were no perioperative deaths in our study group., Conclusion: : Our results show that a community hospital group can appropriately select patients to undergo radical prostatectomy, as well as achieve an acceptable rate of PSMs. We believe that ongoing critical appraisal and reflective practice are essential to improving surgical outcomes and providing quality care.

Intensive glyphosate use has contributed to the evolution and occurrence of glyphosate-resistant weeds that threaten production of many crops. Sustained use of this highly valued herbicide requires rotation and/or substitution of herbicides with different modes of action. Cotton growers have shown considerable interest in the protoporphyrinogen oxidase inhibitor, fomesafen. Following registration for cotton in 2008, use has increased rapidly. Environmental fate data in major use areas are needed to appropriately evaluate risks. Field-based rainfall simulation was used to evaluate fomesafen runoff potential with and without irrigation incorporation in a conventional tillage system (CT) and when conservation tillage (CsT) was practiced with and without cover crop residue rolling. Without irrigation incorporation, relatively high runoff, about 5% of applied, was measured from the CT system, indicating that this compound may present a runoff risk. Runoff was reduced by >50% when the herbicide was irrigation incorporated after application or when used with a CsT system. Data indicate that these practices should be implemented whenever possible to reduce fomesafen runoff risk. Results also raised concerns about leaching and potential groundwater contamination and crop injury due to rapid washoff from cover crop residues in CsT systems. Further work is needed to address these concerns.

Use of genetically modified cultivars resistant to the herbicide glyphosate (N-phosphonomethylglycine) is strongly associated with conservation-tillage (CsT) management for maize ( Zea mays L.), soybean ( Glycine max L.), and cotton ( Gossypium hirsutum L.) cultivation. Due to the emergence of glyphosate-resistant weed biotypes, alternate weed management practices are needed to sustain CsT use. This work focused on metolachlor use (2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide) in a CsT system. The fate and efficacy of granular and emulsifiable concentrate (EC) formulations or an EC surrogate were compared for CsT cotton production in the Atlantic Coastal Plain region of southern Georgia (USA). The granular formulation, a clay-alginate polymer, was produced in the authors' laboratory; EC was a commercial product. In field and laboratory dissipations the granular metolachlor exhibited 8-fold greater soil persistence. Rainfall simulation runoff assessments indicated that use of the granular formulation in a common CsT system, strip-tillage (ST), may reduce metolachlor runoff loss when compared to conventional tillage (CT) management or when EC formulations are used in the ST system. Metolachlor leaching assessments using field-deployed lysimeters showed some tillage (ST > CT) and formulation (EC > granular) differences. Overall leaching was generally small when compared to runoff loss. Finally, greenhouse bioassays showed control of two weed species with the granular was greater than or equal to that of the EC formulation; however, the granular formulation suppressed cotton growth to a greater extent. In sum, this metolachlor granular formulation has advantages for CsT cotton production; however, additional research is needed to assess impacts on crop injury.

Phthalates are ubiquitous in the aquatic environment and are known to adversely affect male reproductive health in mammals through interactions with multiple receptor systems. However, little is known about the risks they pose to fish. This project investigated the effects of di(2-ethylhexyl) phthalate (DEHP), the most commonly used phthalate, on the reproductive health of male zebrafish (Danio rerio). Males were treated with 0.5, 50 and 5000 mg DEHP kg(-1) (body weight) for a period of 10 days via intraperitoneal injection. The effects of the exposure were assessed by analysing fertilisation success, testis histology, sperm DNA integrity and transcript profiles of the liver and testis. A significant increase in the hepatosomatic index and levels of hepatic vitellogenin transcript were observed following exposure to 5000 mg DEHP kg(-1). Exposure to 5000 mg DEHP kg(-1) also resulted in a reduction in fertilisation success of oocytes spawned by untreated females. However, survival and development of the resulting embryos were unaffected by all treatments, and no evidence of DEHP-induced sperm DNA damage was observed. Exposure to 50 and 5000 mg DEHP kg(-1) caused alterations in the proportion of germ cells at specific stages of spermatogenesis in the testis, including a reduction in the proportion of spermatozoa and an increase in the proportion of spermatocytes, suggesting that DEHP may inhibit the progression of meiosis. In parallel, exposure to 5000 mg DEHP kg(-1) increased the levels of two peroxisome proliferator-activated receptor (PPAR) responsive genes (acyl-coenzyme A oxidase 1 (acox1) and enoyl-coenzyme A, hydratase/3-hydroxyacyl coenzyme A dehydrogenase (ehhadh). These data demonstrated that exposure to high concentrations of DEHP disrupts spermatogenesis in adult zebrafish with a consequent decrease in their ability to fertilise oocytes spawned by untreated females. Furthermore, our data suggest that the adverse effects caused by exposure to DEHP are likely to occur preferentially via PPAR signalling pathways in the testis and oestrogen signalling pathways in the liver. We found no evidence of adverse effects on zebrafish reproductive health following exposure to the concentrations occurring in most aquatic systems, indicating that DEHP alone may not be a causative agent of the reproductive abnormalities seen in wildlife, at least as a result of short-term exposures., (Copyright 2010 Elsevier B.V. All rights reserved.)

The herbicide glyphosate became widely used in the United States and other parts of the world after the commercialization of glyphosate-resistant crops. These crops have constitutive overexpression of a glyphosate-insensitive form of the herbicide target site gene, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). Increased use of glyphosate over multiple years imposes selective genetic pressure on weed populations. We investigated recently discovered glyphosate-resistant Amaranthus palmeri populations from Georgia, in comparison with normally sensitive populations. EPSPS enzyme activity from resistant and susceptible plants was equally inhibited by glyphosate, which led us to use quantitative PCR to measure relative copy numbers of the EPSPS gene. Genomes of resistant plants contained from 5-fold to more than 160-fold more copies of the EPSPS gene than did genomes of susceptible plants. Quantitative RT-PCR on cDNA revealed that EPSPS expression was positively correlated with genomic EPSPS relative copy number. Immunoblot analyses showed that increased EPSPS protein level also correlated with EPSPS genomic copy number. EPSPS gene amplification was heritable, correlated with resistance in pseudo-F(2) populations, and is proposed to be the molecular basis of glyphosate resistance. FISH revealed that EPSPS genes were present on every chromosome and, therefore, gene amplification was likely not caused by unequal chromosome crossing over. This occurrence of gene amplification as an herbicide resistance mechanism in a naturally occurring weed population is particularly significant because it could threaten the sustainable use of glyphosate-resistant crop technology.

Objective: Transrectal ultrasound-guided core biopsies of the prostate gland and prostatectomies have become common procedures at many community hospitals in Canada, especially in the era of serum prostate-specific antigen (PSA) screening for prostate cancer. The Gleason grading of prostate cancer in biopsies and prostatectomies is a major determinant used for treatment planning. There is evidence in the literature that suggests important discordance between community hospital pathologists and urological pathologists with respect to the Gleason grading of prostate cancer. Our objective was to determine the diagnostic rates and Gleason scoring patterns for prostate gland biopsies and prostatectomies at our institution compared with the literature., Methods: We conducted a retrospective review of all prostate gland biopsies and prostatectomies performed at the Grey Bruce Health Services from January 2005 to September 2005. We collected data from 194 biopsies and 44 prostatectomies. We obtained prebiopsy serum PSA levels and digital rectal exam results for all patients from urologists' office records., Results: The average age for men having biopsies was 65.8 (standard deviation [SD] 8.6) years, and the average prebiopsy serum PSA level was 8.7 (median 7.1, SD 6.2) mug/L. The rates of diagnosis from prostate gland biopsies of benign (17.6%), high-grade prostatic intraepithelial neoplasia (11.0%), atypical small acinar proliferation suspicious for invasive malignancy (13.2%) and invasive prostatic adenocarcinoma (58.2%) at our institution were significantly different than those reported in the literature (p < 0.001). We observed a significant variation in the rates of these diagnoses among the community hospital pathologists in our study (p = 0.004). There was a strong correlation between the increasing number of positive core biopsy sites and increasing Gleason scores in biopsies (p < 0.001). There was also a strong correlation between increasing pre-biopsy serum PSA levels and increasing Gleason scores in biopsies (p < 0.001). A substantial proportion (21.9%) of the biopsies given the Gleason score of 6 had a Gleason score of 7 in the prostatectomy specimen., Conclusion: Our results showed a significant difference in prostate gland biopsy categorical diagnoses compared with the literature. There were also significant differences in categorical diagnoses of prostate gland biopsies among the community hospital pathologists in our study. The data identify a strong positive correlation between the increasing number of positive core biopsy sites and increasing Gleason scores in biopsies, as well as a strong positive correlation between increasing prebiopsy serum PSA levels and increasing Gleason scores in biopsies that revealed cancer. We would encourage other community hospital pathologists, in collaboration with their urologists, to review periodically their prostate gland pathology practices in an attempt to improve the uniformity of diagnoses.

Background and Purpose: Laparoscopic pyeloplasty (LP) is a highly successful minimally invasive management option for ureteropelvic junction obstruction (UPJO) with success rates equaling or surpassing open repair. Definitions of success have varied greatly among reports of series in the literature. We propose that true success or failure be strictly defined by resolution of obstruction on objective physiologic testing (diuretic renography or Whitaker testing) and accordingly present our results., Patients and Methods: We prospectively gathered data on all LP from December 2001 to November 2006. Ninety-two patients have matured to follow-up testing (> 3 months). Primary uncomplicated procedures were completed in 62/92 (67%) while secondary and/or complex repairs were performed in 30/92 (33%). Rigorous follow-up with physiologic testing was performed postoperatively. Patients with equivocal diuretic renal scans were evaluated with Whitaker pressure-flow studies. The subjective symptom score was assessed for our first 47 patients using a visual analog scale (VAS). Treatment in patients who lacked objective evidence of resolution of UPJO was considered a failure., Results: The mean age of the cohort was 39 years (range 18-69 yrs). Average operating room time was 250 minutes (range 138-488 min), and estimated blood loss was 63 mL (range 10-500 mL). Average hospital length of stay was 1.2 days (range 1-6 d). Crossing vessels were identified in 47/92 (51%) patients. Overall mean follow-up was 13 months (range 2-54 months). There were no intraoperative complications and three perioperative complications. Overall, 85/92 (92%) patients were found to have resolution of UPJO on postoperative physiologic testing. The primary uncomplicated cohort success rate was 98%, while that for the secondary or complex repair cohort was 80%. The mean overall VAS pain score was 84%., Conclusion: Objective physiologic testing should be used as the measurement of true success after LP. A uniform definition of success will allow improved comparison of results with literature series and comparison with alternative management modalities.

Runoff from farm fields is a common source of herbicide residues in surface waters. Incorporation by irrigation has the potential to reduce herbicide runoff risks. To assess impacts, rainfall was simulated on plots located in a peanut (Arachis hypogaea L.) field in Georgia's Atlantic Coastal Plain region after pre-emergence application of metolachlor (2-chloro-N-(2-ethyl-6-methylphenyl)-N-[(1S)-2-methoxy-1-methylethyl]-acetamide) and pendimethalin (N-(1-ethylpropyl)-3,4-dimethyl-2,6-dinitro-benzenamine). Runoff, sediment, and herbicide loss as function of strip tillage (ST) versus conventional tillage (CT) were compared with and without irrigation (12.5 mm) after application of an herbicide tank mixture. For the CT system, metolachlor runoff was reduced 2x and pendimethalin 1.2x when compared with the non-irrigated treatment. The difference in irrigated and non-irrigated metolachlor means was significant (P = 0.05). Irrigation reduced metolachlor runoff by 1.3x in the ST system, but there was a 1.4x increase for pendimethalin. Overall results indicated that irrigation incorporation reduces herbicide runoff with the greatest impact when CT is practiced and products like metolachlor, which have relatively low K(oc) and high water solubility, are used. The lower ST system response was likely due to a combination of spray interception and retention by the ST system cover crop mulch and higher ST soil organic carbon content and less total runoff. During the study, the measured K(oc) of both herbicides on runoff sediment was found to vary with tillage and irrigation after herbicide application. Generally, K(oc) was higher for ST sediment and when irrigation incorporation was used with the CT system. These results have significant implications for simulation model parametization.

Background: Patients with pineal lesions typically present with the classical signs of increased intracranial pressure (headache, nausea, vomiting) and/or Parinaud's syndrome. Rare symptomatology of secondary parkinsonism attributed to pineal lesions has been previously reported in the literature. We describe an unusual case of a pineal cyst in a patient with the presenting sign of a resting tremor., Case Description: We report an 18-year-old Caucasian female who presented with a 1-month history of a new-onset progressive, unilateral low-frequency right-hand resting tremor with associated headache, nausea, vomiting, and excessive diarrhea. Magnetic resonance imaging demonstrated an atypical appearance with enhancement of a mildly prominent pineal gland, possibly representing a pineal cyst. The patient did not exhibit radiographic signs of hydrocephalus. Based upon the radiographic appearance, one could not exclude with absolute certainty the presence of a malignancy. The patient ultimately underwent a bilateral suboccipital craniotomy with gross total resection of the lesion. Postoperatively, the patient exhibited immediate resolution of her preoperative resting tremor and continues to be symptom free at 1 year., Conclusions: A new-onset, resting tremor and/or other secondary parkinsonism symptoms should raise clinical suspicions of pineal lesions. Treatment can be guided based on tissue type and the presence or absence of hydrocephalus. We observed that complete surgical resection of the lesion provided the best treatment option for the total resolution of symptoms attributed to the disturbance of the microvasculature surrounding the nigro-striatal-pallidal system.

Pesticide runoff research relies heavily on rainfall simulation experiments. Most are conducted at a constant intensity, i.e., at a fixed rainfall rate; however, large differences in natural rainfall intensity is common. To assess implications we quantified runoff of two herbicides, fluometuron and pendimethalin, and applied preemergence after planting cotton on Tifton loamy sand. Rainfall at constant and variable intensity patterns representative of late spring thunderstorms in the Atlantic Coastal Plain region of Georgia (USA) were simulated on 6-m2 plots under strip- (ST) and conventional-tillage (CT) management. The variable pattern produced significantly higher runoff rates of both compounds from CT but not ST plots. However, on an event-basis, runoff totals (% applied) were not significantly different, with one exception: fluometuron runoff from CT plots. There was about 25% more fluometuron runoff with the variable versus the constant intensity pattern (P = 0.10). Study results suggest that conduct of simulations using variable intensity storm patterns may provide more representative rainfall simulation-based estimates of pesticide runoff and that the greatest impacts will be observed with CT. The study also found significantly more fluometuron in runoff from ST than CT plots. Further work is needed to determine whether this behavior may be generalized to other active ingredients with similar properties [low K(oc) (organic carbon partition coefficient) approximately 100 mL g(-1); high water solubility approximately 100 mg L(-1)]. If so, it should be considered when making tillage-specific herbicide recommendations to reduce runoff potential.