Ontogeny influences sensitivity to climate change stressors in an endangered fish.

Coastal ecosystems are among the most human-impacted habitats globally, and their management is often critically linked to recovery of declining native species. In the San Francisco Estuary, the Delta Smelt (Hypomesus transpacificus) is an endemic, endangered fish strongly tied to Californian conservation planning. The complex life history of Delta Smelt combined with dynamic seasonal and spatial abiotic conditions result in dissimilar environments experienced among ontogenetic stages, which may yield stage-specific susceptibility to abiotic stressors. Climate change is forecasted to increase San Francisco Estuary water temperature and salinity; therefore, understanding the influences of ontogeny and phenotypic plasticity on tolerance to these critical environmental parameters is particularly important for Delta Smelt and other San Francisco Estuary fishes. We assessed thermal and salinity limits in several ontogenetic stages and acclimation states of Delta Smelt, and paired these data with environmental data to evaluate sensitivity to climate-change stressors. Thermal tolerance decreased among successive stages, with larval fish exhibiting the highest tolerance and post-spawning adults having the lowest. Delta Smelt had limited capacity to increase tolerance through thermal acclimation, and comparisons with field temperature data revealed that juvenile tolerance limits are the closest to current environmental conditions, which may make this stage especially susceptible to future climate warming. Maximal water temperatures observed in situ exceeded tolerance limits of juveniles and adults. Although these temperature events are currently rare, if they increase in frequency as predicted, it could result in habitat loss at these locations despite other favourable conditions for Delta Smelt. In contrast, Delta Smelt tolerated salinities spanning the range of expected environmental conditions for each ontogenetic stage, but salinity did impact survival in juvenile and adult stages in exposures over acute time scales. Our results underscore the importance of considering ontogeny and phenotypic plasticity in assessing the impacts of climate change, particularly for species adapted to spatially and temporally heterogeneous environments

Effects of Aberrant Pax6 Gene Dosage on Mouse Corneal Pathophysiology and Corneal Epithelial Homeostasis

Background: Altered dosage of the transcription factor PAX6 causes multiple human eye pathophysiologies. PAX6(+/-) heterozygotes suffer from aniridia and aniridia-related keratopathy (ARK), a corneal deterioration that probably involves a limbal epithelial stem cell (LESC) deficiency. Heterozygous Pax6(+/Sey-Neu) (Pax6(+/-)) mice recapitulate the human disease and are a good model of ARK. Corneal pathologies also occur in other mouse Pax6 mutants and in PAX77(Tg/-) transgenics, which over-express Pax6 and model human PAX6 duplication. Methodology/Principal Findings: We used electron microscopy to investigate ocular defects in Pax6(+/-) heterozygotes (low Pax6 levels) and PAX77(Tg/-) transgenics (high Pax6 levels). As well as the well-documented epithelial defects, aberrant Pax6 dosage had profound effects on the corneal stroma and endothelium in both genotypes, including cellular vacuolation, similar to that reported for human macular corneal dystrophy. We used mosaic expression of an X-linked LacZ transgene in X-inactivation mosaic female (XLacZ(Tg/-)) mice to investigate corneal epithelial maintenance by LESC clones in Pax6(+/-) and PAX77(Tg/-) mosaic mice. PAX77(Tg/-) mosaics, over-expressing Pax6, produced normal corneal epithelial radial striped patterns (despite other corneal defects), suggesting that centripetal cell movement was unaffected. Moderately disrupted patterns in Pax6(+/-) mosaics were corrected by introducing the PAX77 transgene (in Pax6(+/-), PAX77(Tg/-) mosaics). Pax6(Leca4/+), XLacZ(Tg/-) mosaic mice (heterozygous for the Pax6(Leca4) missense mutation) showed more severely disrupted mosaic patterns. Corrected corneal epithelial stripe numbers (an indirect estimate of active LESC clone numbers) declined with age (between 15 and 30 weeks) in wild-type XLacZ(Tg/-) mosaics. In contrast, corrected stripe numbers were already low at 15 weeks in Pax6(+/-) and PAX77(Tg/-) mosaic corneas, suggesting Pax6 under-and over-expression both affect LESC clones. Conclusions/Significance: Pax6(+/-) and PAX77(Tg/-) genotypes have only relatively minor effects on LESC clone numbers but cause more severe corneal endothelial and stromal defects. This should prompt further investigations of the pathophysiology underlying human aniridia and ARK

Stressor interactions in freshwater habitats: Effects of cold water exposure and food limitation on early-life growth and upper thermal tolerance in white sturgeon, Acipenser transmontanus

Limited food availability and altered thermal regimes (e.g. cold water releases from dams) are two common stressors threatening the persistence of fishes inhabiting anthropogenically disturbed freshwater systems. Yet, the combined effects of these stressors remain poorly characterised. To remedy this, we examined the isolated and combined effects of low temperature exposure and food restriction on specific growth rate (SGR, % body mass/day) and upper thermal tolerance (critical thermal maxima, CTMax) in larval white sturgeon (Acipenser transmontanus [Acipenseridae], 32 days post-hatch, body mass: 0.25 ± 0.03 g, mean ± standard deviation). A 2 × 2 factorial design was implemented with fish exposed to one of two ecologically-relevant acclimation temperatures (cold exposure: 11°C or a control temperature: 18°C) and one of two food restriction treatments designed to emulate observed declines in food availability (100% or 40% optimal feed rate) for 6 weeks (N: 3 replicate tanks/treatment, 50 fish/tank). Specific growth rate was affected by both low temperature exposure and food restriction in isolation; low temperature exposure reduced SGR by 56.5% and food restriction reduced SGR by 30.6%. Simultaneous exposure to low temperature and food restriction resulted in a greater but less than additive reduction in SGR (80.6%), indicating that the stressors interacted antagonistically. Critical thermal maxima were c. 2°C higher in 18°C-acclimated fish (CTMax = 30.7 ± 0.4°C, mean ± standard error) compared to 11°C-acclimated fish (CTMax = 28.6 ± 0.2°C, mean ± standard error); however, CTMax was independent of food restriction in both 11°C- and 18°C-acclimated fish. These data highlight the unpredictability of stressor interactions and may guide holistic conservation strategies, which target co-occurring stressors in freshwater habitats

Stressor interactions in freshwater habitats: Effects of cold water exposure and food limitation on early-life growth and upper thermal tolerance in white sturgeon, Acipenser transmontanus

Limited food availability and altered thermal regimes (e.g. cold water releases from dams) are two common stressors threatening the persistence of fishes inhabiting anthropogenically disturbed freshwater systems. Yet, the combined effects of these stressors remain poorly characterised. To remedy this, we examined the isolated and combined effects of low temperature exposure and food restriction on specific growth rate (SGR, % body mass/day) and upper thermal tolerance (critical thermal maxima, CTMax) in larval white sturgeon (Acipenser transmontanus [Acipenseridae], 32 days post-hatch, body mass: 0.25 ± 0.03 g, mean ± standard deviation). A 2 × 2 factorial design was implemented with fish exposed to one of two ecologically-relevant acclimation temperatures (cold exposure: 11°C or a control temperature: 18°C) and one of two food restriction treatments designed to emulate observed declines in food availability (100% or 40% optimal feed rate) for 6 weeks (N: 3 replicate tanks/treatment, 50 fish/tank). Specific growth rate was affected by both low temperature exposure and food restriction in isolation; low temperature exposure reduced SGR by 56.5% and food restriction reduced SGR by 30.6%. Simultaneous exposure to low temperature and food restriction resulted in a greater but less than additive reduction in SGR (80.6%), indicating that the stressors interacted antagonistically. Critical thermal maxima were c. 2°C higher in 18°C-acclimated fish (CTMax = 30.7 ± 0.4°C, mean ± standard error) compared to 11°C-acclimated fish (CTMax = 28.6 ± 0.2°C, mean ± standard error); however, CTMax was independent of food restriction in both 11°C- and 18°C-acclimated fish. These data highlight the unpredictability of stressor interactions and may guide holistic conservation strategies, which target co-occurring stressors in freshwater habitats

Physiological stress biomarkers reveal stocking density effects in late larval Delta Smelt (Hypomesus transpacificus)

Suboptimal fish stocking densities in experimental systems may elicit stress responses that can affect experimental results. Fish species, age and size, water chemistry and flow, and physical characteristics of the experimental system (e.g., tank, cage) are among the parameters to be considered when determining stocking densities. However, systematic studies to define fish densities minimizing stress in experimental systems are rarely performed. This is particularly true when working with species of low aquaculture value or a non-model test species such as the Delta Smelt (. Hypomesus transpacificus). The aim of this study was to use physiological stress biomarkers to determine suitable fish densities for specific experimental vessels routinely used for this species. We maintained late larval Delta Smelt (60. days post-hatch; dph) over a period of 24. h, at five different densities: 7, 14, 28, 42, and 56 fish per 8. L circular fish tank. We assessed whole body cortisol and transcriptomic biomarkers that lead to cortisol production to quantify stress levels. Both marker types delivered similar results. Cortisol levels were lowest at densities of 28 and 42 fish per tank, whereas lowest fish densities (7 and 14 fish per tank) evoked the highest stress levels. Genes such as Mineralocorticoid Receptor 1 and Glucocorticoid Receptor 2, as well as 11-Beta-Hydroxysteroid-Dehydrogenase-2 depicted the lowest expression levels at stocking densities 28 and 42, and elevated expression levels for stocking densities 7 and 14. Our data support the observations that late larval Delta Smelt should be exposed, acclimated, and cultured in groups rather than as individuals or in low numbers. This study indicates the importance of adequately defining experimental conditions that minimize stress, specifically when stress is measured as an endpoint. In addition to classical cortisol measurements, responses of the transcriptome also appear suitable in assessing stress responses in fish, and in determining optimal holding conditions, particularly if short-term responses are the study focus.Statement of relevance. The study highlights the importance of evaluating stress in order to determine species-specific stocking densities. The results are thus relevant to a wide audience in the field of aquaculture and experimental biology

Understanding risks and consequences of pathogen infections on the physiological performance of outmigrating Chinook salmon

The greatest concentration of at-risk anadromous salmonids is found in California (USA)-the populations that have been negatively impacted by the degradation of freshwater ecosystems. While climate-driven environmental changes threaten salmonids directly, they also change the life cycle dynamics and geographic distribution of pathogens, their resulting host-pathogen interactions and potential for disease progression. Recent studies have established the correlation between pathogen detection and salmonid smolt mortality during their migration to the ocean. The objective of the present study was to screen for up to 47 pathogens in juvenile Chinook salmon (Oncorhynchus tshawytscha) that were held in cages at two key sites of the Sacramento River (CA, USA) and measure potential consequences on fish health. To do so, we used a combination of transcriptomic analysis, enzymatic assays for energy metabolism and hypoxia and thermal tolerance measures. Results revealed that fish were infected by two myxozoan parasites: Ceratonova shasta and Parvicapsula minibicornis within a 2-week deployment. Compared to the control fish maintained in our rearing facility, infected fish displayed reduced body mass, depleted hepatic glycogen stores and differential regulation of genes involved in the immune and general stress responses. This suggests that infected fish would have lower chances of migration success. In contrast, hypoxia and upper thermal tolerances were not affected by infection, suggesting that infection did not impair their capacity to cope with acute abiotic stressors tested in this study. An evaluation of long-term consequences of the observed reduced body mass and hepatic glycogen depletion is needed to establish a causal relationship between salmon parasitic infection and their migration success. This study highlights that to assess the potential sublethal effects of a stressor, or to determine a suitable management action for fish, studies need to consider a combination of endpoints from the molecular to the organismal level