Sex and gender analysis improves science and engineering.

The goal of sex and gender analysis is to promote rigorous, reproducible and responsible science. Incorporating sex and gender analysis into experimental design has enabled advancements across many disciplines, such as improved treatment of heart disease and insights into the societal impact of algorithmic bias. Here we discuss the potential for sex and gender analysis to foster scientific discovery, improve experimental efficiency and enable social equality. We provide a roadmap for sex and gender analysis across scientific disciplines and call on researchers, funding agencies, peer-reviewed journals and universities to coordinate efforts to implement robust methods of sex and gender analysis.

How do abiotic environmental conditions influence shrimp susceptibility to disease? A critical analysis focussed on White Spot Disease.

White Spot Syndrome Virus (WSSV) causes White Spot Disease (WSD) and is historically the most devastating disease in the shrimp industry. Global losses from this disease have previously exceeded $3 bn annually, having a major impact on a global industry worth US$19 bn per annum. Shrimp are cultured predominantly in enclosed ponds that are subject to considerable fluctuations in abiotic conditions and WSD outbreaks are increasingly linked to periods of extreme weather, which may cause major fluctuations in pond culture conditions. Combined with the intensity of production in these systems, the resulting suboptimal physicochemical conditions have a major bearing on the susceptibility of shrimp to infection and disease. Current knowledge indicates that pond temperature and salinity are major factors determining outbreak severity. WSSV appears to be most virulent in water temperatures between 25 and 28 °C and salinities far removed from the isoosmotic point of shrimp. Elevated temperatures (>30 °C) may protect against WSD, depending on the stage of infection, however the mechanisms mediating this effect have not been well established. Other factors relating to water quality that may play key roles in determining outbreak severity include dissolved oxygen concentration, nitrogenous compound concentration, partial pressure of carbon dioxide and pH, but data on their impacts on WSSV susceptibility in cultured shrimps is scarce. This illustrates a major research gap in our understanding of the influence of environmental conditions on disease. For example, it is not clear whether temperature manipulations can be used effectively to prevent or mitigate WSD in cultured shrimp. Therefore, developing our understanding of the impact of environmental conditions on shrimp susceptibility to WSSV may provide insight for WSD mitigation when, even after decades of research, there is no effective practical prophylaxis or treatment.

Lessons from two high CO2 worlds - future oceans and intensive aquaculture.

Exponentially rising CO2 (currently ~400 µatm) is driving climate change and causing acidification of both marine and freshwater environments. Physiologists have long known that CO2 directly affects acid-base and ion regulation, respiratory function and aerobic performance in aquatic animals. More recently, many studies have demonstrated that elevated CO2 projected for end of this century (e.g. 800-1000 µatm) can also impact physiology, and have substantial effects on behaviours linked to sensory stimuli (smell, hearing and vision) both having negative implications for fitness and survival. In contrast, the aquaculture industry was farming aquatic animals at CO2 levels that far exceed end-of-century climate change projections (sometimes >10 000 µatm) long before the term 'ocean acidification' was coined, with limited detrimental effects reported. It is therefore vital to understand the reasons behind this apparent discrepancy. Potential explanations include 1) the use of 'control' CO2 levels in aquaculture studies that go beyond 2100 projections in an ocean acidification context; 2) the relatively benign environment in aquaculture (abundant food, disease protection, absence of predators) compared to the wild; 3) aquaculture species having been chosen due to their natural tolerance to the intensive conditions, including CO2 levels; or 4) the breeding of species within intensive aquaculture having further selected traits that confer tolerance to elevated CO2 . We highlight this issue and outline the insights that climate change and aquaculture science can offer for both marine and freshwater settings. Integrating these two fields will stimulate discussion on the direction of future cross-disciplinary research. In doing so, this article aimed to optimize future research efforts and elucidate effective mitigation strategies for managing the negative impacts of elevated CO2 on future aquatic ecosystems and the sustainability of fish and shellfish aquaculture.

Does sex really matter? Explaining intraspecies variation in ocean acidification responses.

Ocean acidification (OA) poses a major threat to marine ecosystems globally, having significant ecological and economic importance. The number and complexity of experiments examining the effects of OA has substantially increased over the past decade, in an attempt to address multi-stressor interactions and long-term responses in an increasing range of aquatic organisms. However, differences in the response of males and females to elevated pCO2 have been investigated in fewer than 4% of studies to date, often being precluded by the difficulty of determining sex non-destructively, particularly in early life stages. Here we highlight that sex can significantly impact organism responses to OA, differentially affecting physiology, reproduction, biochemistry and ultimately survival. What is more, these impacts do not always conform to ecological theory based on differential resource allocation towards reproduction, which would predict females to be more sensitive to OA owing to the higher production cost of eggs compared with sperm. Therefore, non-sex-specific studies may overlook subtle but ecologically significant differences in the responses of males and females to OA, with consequences for forecasting the fate of natural populations in a near-future ocean.

Ocean acidification and host-pathogen interactions: blue mussels, Mytilus edulis, encountering Vibrio tubiashii.

Ocean acidification (OA) can shift the ecological balance between interacting organisms. In this study, we have used a model system to illustrate the interaction between a calcifying host organism, the blue mussel Mytilus edulis and a common bivalve bacterial pathogen, Vibrio tubiashii, with organisms being exposed to a level of acidification projected to occur by the end of the 21st century. OA exposures of the mussels were carried out in relative long-term (4 months) and short-term (4 days) experiments. We found no effect of OA on the culturability of V. tubiashii, in broth or in seawater. OA inhibited mussel shell growth and impaired crystalline shell structures but did not appear to affect mussel immune parameters (i.e haemocyte counts and phagocytotic capacity). Despite no evident impact on host immunity or growth and virulence of the pathogen, V. tubiashii was clearly more successful in infecting mussels exposed to long-term OA compared to those maintained under ambient conditions. Moreover, OA exposed V. tubiashii increased their viability when exposed to haemocytes of OA-treated mussel. Our findings suggest that even though host organisms may have the capacity to cope with periods of OA, these conditions may alter the outcome of host-pathogen interactions, favouring the success of the latter.

Ocean acidification increases copper toxicity to the early life history stages of the polychaete Arenicola marina in artificial seawater.

The speciation and therefore bioavailability of the common pollutant copper is predicted to increase within the pH range anticipated under near-future ocean acidification (OA), hence the potential exists for copper toxicity to marine organisms to also increase. We investigated the impact of OA (seawater pH values of 7.77 (pCO2 1400 µatm) and 7.47 (pCO2 3000 µatm)) upon copper toxicity responses in early life history stages of the polychaete Arenicola marina and found both synergistic and additive toxicity effects of combined exposures depending on life history stage. The toxicity of copper on sperm DNA damage and early larval survivorship was synergistically increased under OA conditions. Larval survival was reduced by 24% when exposed to both OA and copper combined compared to single OA or copper exposures. Sperm motility was negatively affected by both OA and copper singularly with additive toxicity effects of the two stressors when combined. Fertilization success was also negatively affected by both OA and copper individually, but no additive effects when exposed as combined stressors were present for this stage. These findings add to the growing body of evidence that OA will act to increase the toxicity of copper to marine organisms, which has clear implications for coastal benthic ecosystems suffering chronic metal pollution as pCO2 levels rise and drive a reduction in seawater pH.

(1)H NMR metabolomics reveals contrasting response by male and female mussels exposed to reduced seawater pH, increased temperature, and a pathogen.

Human activities are fundamentally altering the chemistry of the world's oceans. Ocean acidification (OA) is occurring against a background of warming and an increasing occurrence of disease outbreaks, posing a significant threat to marine organisms, communities, and ecosystems. In the current study, (1)H NMR spectroscopy was used to investigate the response of the blue mussel, Mytilus edulis, to a 90-day exposure to reduced seawater pH and increased temperature, followed by a subsequent pathogenic challenge. Analysis of the metabolome revealed significant differences between male and female organisms. Furthermore, males and females are shown to respond differently to environmental stress. While males were significantly affected by reduced seawater pH, increased temperature, and a bacterial challenge, it was only a reduction in seawater pH that impacted females. Despite impacting males and females differently, stressors seem to act via a generalized stress response impacting both energy metabolism and osmotic balance in both sexes. This study therefore has important implications for the interpretation of metabolomic data in mussels, as well as the impact of environmental stress in marine invertebrates in general.

SNP discovery and genetic structure in blue mussel species using low coverage sequencing and a medium density 60 K SNP-array.

Blue mussels from the genus Mytilus are an abundant component of the benthic community, found in the high latitude habitats. These foundation species are relevant to the aquaculture industry, with over 2 million tonnes produced globally each year. Mussels withstand a wide range of environmental conditions and species from the Mytilus edulis complex readily hybridize in regions where their distributions overlap. Significant effort has been made to investigate the consequences of environmental stress on mussel physiology, reproductive isolation, and local adaptation. Yet our understanding on the genomic mechanisms underlying such processes remains limited. In this study, we developed a multi species medium-density 60 K SNP-array including four species of the Mytilus genus. SNPs included in the platform were called from 138 mussels from 23 globally distributed mussel populations, sequenced using a whole-genome low coverage approach. The array contains polymorphic SNPs which capture the genetic diversity present in mussel populations thriving across a gradient of environmental conditions (~59 K SNPs) and a set of published and validated SNPs informative for species identification and for diagnosis of transmissible cancer (610 SNPs). The array will allow the consistent genotyping of individuals, facilitating the investigation of ecological and evolutionary processes in these taxa. The applications of this array extend to shellfish aquaculture, contributing to the optimization of this industry via genomic selection of blue mussels, parentage assignment, inbreeding assessment and traceability. Further applications such as genome wide association studies (GWAS) for key production traits and those related to environmental resilience are especially relevant to safeguard aquaculture production under climate change.

Methods for successful inactivation of Rift Valley fever virus in infected mosquitoes.

Ensuring the successful inactivation of select agent material is critical for maintaining compliance with federal regulations and safeguarding laboratory personnel from exposure to dangerous pathogens. Rift Valley fever virus (RVFV), naturally transmitted by mosquitoes, is classified as a select agent by the CDC and USDA due to its potential to cause significant economic losses to the livestock industry and its demonstrated potential to emerge into naïve geographic areas. Herein we describe several effective inactivation procedures for RVFV infected mosquito samples. We also demonstrate the vaccine strain MP-12 can be used as an appropriate analog for inactivation testing and describe a method of validating inactivation using Amicon filters. Briefly, we show the following inactivation methods are all effective at inactivating RVFV and MP-12 by following the manufacturers'/established protocols: 4 % paraformaldehyde, Trizol LS (ThermoFisher Scientific), MagMAX™-96 Viral RNA Isolation Kit (ThermoFisher Scientific), and Mag-Bind® Viral DNA/RNA 96 Kit (Omega Bio-Tek).

New insights into organ-specific oxidative stress mechanisms using a novel biosensor zebrafish.

BACKGROUND: Reactive oxygen species (ROS) arise as a result from, and are essential in, numerous cellular processes. ROS, however, are highly reactive and if left unneutralised by endogenous antioxidant systems, can result in extensive cellular damage and/or pathogenesis. In addition, exposure to a wide range of environmental stressors can also result in surplus ROS production leading to oxidative stress (OS) and downstream tissue toxicity. OBJECTIVES: Our aim was to produce a stable transgenic zebrafish line, unrestricted by tissue-specific gene regulation, which was capable of providing a whole organismal, real-time read-out of tissue-specific OS following exposure to a wide range of OS-inducing environmental contaminants and conditions. This model could, therefore, serve as a sensitive and specific mechanistic in vivo biomarker for all environmental conditions that result in OS. METHODS: To achieve this aim, we exploited the pivotal role of the electrophile response element (EpRE) as a globally-acting master regulator of the cellular response to OS. To test tissue specificity and quantitative capacity, we selected a range of chemical contaminants known to induce OS in specific organs or tissues, and assessed dose-responsiveness in each using microscopic measures of mCherry fluorescence intensity. RESULTS: We produced the first stable transgenic zebrafish line Tg (3EpRE:hsp70:mCherry) with high sensitivity for the detection of cellular RedOx imbalances, in vivo in near-real time. We applied this new model to quantify OS after exposure to a range of environmental conditions with high resolution and provided quantification both of compound- and tissue-specific ROS-induced toxicity. DISCUSSION: Our model has an extremely diverse range of potential applications not only for biomonitoring of toxicants in aqueous environments, but also in biomedicine for identifying ROS-mediated mechanisms involved in the progression of a number of important human diseases, including cancer.

Impacts of ocean acidification on sperm develop with exposure time for a polychaete with long lived sperm.

The majority of marine invertebrate species release eggs and sperm into seawater for external fertilisation. Seawater conditions are currently changing at an unprecedented rate as a consequence of ocean acidification (OA). Sperm are thought to be particularly vulnerable to these changes and may be exposed to external environmental conditions for variable periods of time between spawning and fertilisation. Here, we undertook a mechanistic investigation of sperm swimming performance in the coastal polychaete Arenicola marina during an extended exposure to OA conditions (pHNBS 7.77, 1000 µatm pCO2). We found that key fitness-related aspects of sperm functioning declined faster under OA conditions i.e. impacts became apparent with exposure time. Sperm swimming speed (VCL), the number of motile sperm and sperm path linearity all dropped significantly after 4 h under OA conditions whilst remaining constant under ambient conditions at this time point. Our results highlight the importance of sperm exposure duration in ocean acidification experiments and may help towards explaining species specific differences in response.

Ocean acidification increases copper toxicity differentially in two key marine invertebrates with distinct acid-base responses.

Ocean acidification (OA) is expected to indirectly impact biota living in contaminated coastal environments by altering the bioavailability and potentially toxicity of many pH-sensitive metals. Here, we show that OA (pH 7.71; pCO2 1480 µatm) significantly increases the toxicity responses to a global coastal contaminant (copper ~0.1 µM) in two keystone benthic species; mussels (Mytilus edulis) and purple sea urchins (Paracentrotus lividus). Mussels showed an extracellular acidosis in response to OA and copper individually which was enhanced during combined exposure. In contrast, urchins maintained extracellular fluid pH under OA by accumulating bicarbonate but exhibited a slight alkalosis in response to copper either alone or with OA. Importantly, copper-induced damage to DNA and lipids was significantly greater under OA compared to control conditions (pH 8.14; pCO2 470 µatm) for both species. However, this increase in DNA-damage was four times lower in urchins than mussels, suggesting that internal acid-base regulation in urchins may substantially moderate the magnitude of this OA-induced copper toxicity effect. Thus, changes in metal toxicity under OA may not purely be driven by metal speciation in seawater and may be far more diverse than either single-stressor or single-species studies indicate. This has important implications for future environmental management strategies.

The history of the Conference of Research Workers in Animal Diseases (CRWAD) 1920-2014.

The following history has been compiled and written by the authors. The historical facts are available from the Conference of Research Workers in Animal Diseases (CRWAD) archives, dating back to letters and summaries written by the founders, and by a few of the Secretary-Treasurers from the early decades through 2014. THE ORGANIZATION AND PURPOSE: The CRWAD is a non-profit organization and has been since its origin. The sole purpose of CRWAD is to discuss and disseminate the most current research advances in animal diseases. Graduate students and industry and academic professionals present and discuss the most recent advances on subjects of interest to the CRWAD and of importance to the global livestock and companion animal industries. The oral and poster abstracts of new and unpublished data presented at the meeting sessions are published each year in the CRWAD Proceedings (formerly the CRWAD Abstracts). CRWAD publishes, copyrights, and distributes the Proceedings. The presentations are arranged into the following 10 sections, according to the primary topic of the presentation: Bacterial Pathogenesis, Biosafety and Biosecurity, Companion Animal Epidemiology, Ecology and Management of Foodborne Agents, Epidemiology and Animal Health Economics, Immunology, Pathobiology of Enteric and Foodborne Pathogens, Respiratory Diseases, Vector-Borne and Parasitic Diseases, and Viral Pathogenesis. Prospective members should be actively engaged in animal disease research or research administration. Meeting information and membership applications may be obtained by contacting the Executive Director or by visiting the CRWAD website. Annual abstracts are currently available on-line at the On-line Meeting Planner and Itinerary Builder, with access through the CRWAD website.

Hemorrhagic bowel syndrome in dairy cattle: 22 cases (1997-2000).

OBJECTIVE: To determine signalment, history, and clinical, necropsy, and microbiologic findings in dairy cows with hemorrhagic bowel syndrome. DESIGN: Retrospective study. ANIMALS: 22 adult dairy cows from a single farm in Colorado. PROCEDURE: Medical records were reviewed for information on signalment, medical and reproductive history, the owner's chief complaints, results of physical examinations and ancillary diagnostic tests, treatment and response to treatment, results of microbiologic testing, and, if applicable, postmortem findings. RESULTS: Common clinical signs were acute signs of profound depression, decreased milk production, tachycardia, ruminal stasis, abdominal distention, and dark clotted blood in the feces. Rectal examination revealed distended loops of small intestine in 7 of 14 cows. Transabdominal ultrasonography revealed small intestinal ileus and distention in 12 of 12 cows and homogeneous echogenic intraluminal material compatible with intraluminal hemorrhage and clot formation in 4. Seven of 8 cows treated medically died; 9 of 13 cows that underwent surgery died or were euthanatized. Clostridium perfringens was isolated from fecal samples from 17 of 20 cows. The most common morphologic diagnosis at necropsy was severe necrohemorrhagic enteritis or jejunitis with intraluminal hemorrhage or blood clots. The most prominent histologic finding was severe, segmental submucosal hemorrhage and edema of the small intestine. CONCLUSIONS AND CLINICAL RELEVANCE: Results confirm that in adult cattle, hemorrhagic bowel syndrome is a sporadic acute intestinal disorder characterized by intraluminal hemorrhage and obstruction of the small intestine. Clostridium perfringens was consistently isolated from the feces of affected cows. The prognosis for affected cows was grave.

Population-based study of fecal shedding of Clostridium perfringens in broodmares and foals.

OBJECTIVE: To determine the percentage of broodmares and foals that shed Clostridium perfringens in their feces and classify the genotypes of those isolates. DESIGN: Prospective cross-sectional study. ANIMALS: 128 broodmares and their foals on 6 equine premises. PROCEDURES: Anaerobic and aerobic bacteriologic cultures were performed on feces collected 3 times from broodmares and foals. All isolates of C. perfringens were genotyped. RESULTS: Clostridium perfringens was isolated from the feces of 90% of 3-day-old foals and 64% of foals at 8 to 12 hours of age. A lower percentage of broodmares and 1- to 2-month-old foals shed C. perfringens in their feces, compared with neonatal foals. Among samples with positive results, C. perfringens type A was the most common genotype identified (85%); C. perfringens type A with the beta2 toxin gene was identified in 12% of samples, C. perfringens type A with the enterotoxin gene was identified in 2.1% of samples, and C. perfringens type C was identified in < 1% of samples. CONCLUSIONS AND CLINICAL RELEVANCE: Clostridium perfringens was identified from the feces of all but 6 foals by 3 days of age and is likely part of the normal microflora of neonatal foals. Most isolates from broodmares and foals are C. perfringens type A; thus, the clinical relevance of culture results alone is questionable. Clostridium perfringens type C, which has been associated with neonatal enterocolitis, is rarely found in the feces of horses.

Guidelines for Biosafety Training Programs for Workers Assigned to BSL-3 Research Laboratories.

The Guidelines for Biosafety Training Programs for Workers Assigned to BSL-3 Research Laboratories were developed by biosafety professionals who oversee training programs for the 2 national biocontainment laboratories (NBLs) and the 13 regional biocontainment laboratories (RBLs) that participate in the National Institute of Allergy and Infectious Diseases (NIAID) NBL/RBL Network. These guidelines provide a general training framework for biosafety level 3 (BSL-3) high-containment laboratories, identify key training concepts, and outline training methodologies designed to standardize base knowledge, understanding, and technical competence of laboratory personnel working in high-containment laboratories. Emphasis is placed on building a culture of risk assessment-based safety through competency training designed to enhance understanding and recognition of potential biological hazards as well as methods for controlling these hazards. These guidelines may be of value to other institutions and academic research laboratories that are developing biosafety training programs for BSL-3 research.