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The killifish Fundulus heteroclitus is an estuarine species with broad physiological plasticity, enabling acclimation to diverse stressors. Previous work suggests that freshwater populations expanded their physiology to accommodate low salinity environments; however, it is unknown whether this compromises their tolerance to high salinity. We used a comparative approach to investigate the mechanisms of a derived freshwater phenotype and the fate of an ancestral euryhaline phenotype after invasion of a freshwater environment. We compared physiological and transcriptomic responses to high- and low-salinity stress in fresh and brackish water populations and found an enhanced plasticity to low salinity in the freshwater population coupled with a reduced ability to acclimate to high salinity. Transcriptomic data identified genes with a conserved common response, a conserved salinity-dependent response and responses associated with population divergence. Conserved common acclimation responses revealed stress responses and alterations in cell-cycle regulation as important mechanisms in the general osmotic response. Salinity-specific responses included the regulation of genes involved in ion transport, intracellular calcium, energetic processes and cellular remodeling. Genes diverged between populations were primarily those showing salinity-specific expression and included those regulating polyamine homeostasis and the cell cycle. Additionally, when populations were matched with their native salinity, expression patterns were consistent with the concept of 'transcriptomic resilience', suggesting local adaptation. These findings provide insight into the fate of a plastic phenotype after a shift in environmental salinity and help to reveal mechanisms allowing for euryhalinity., (© 2015. Published by The Company of Biologists Ltd.)

Environmental salinity presents a key barrier to dispersal for most aquatic organisms, and adaptation to alternate osmotic environments likely enables species diversification. Little is known of the functional basis for derived tolerance to environmental salinity. We integrate comparative physiology and functional genomics to explore the mechanistic underpinnings of evolved variation in osmotic plasticity within and among two species of killifish; Fundulus majalis harbours the ancestral mainly salt-tolerant phenotype, whereas Fundulus heteroclitus harbours a derived physiology that retains extreme salt tolerance but with expanded osmotic plasticity towards the freshwater end of the osmotic continuum. Common-garden comparative hypo-osmotic challenge experiments show that F. heteroclitus is capable of remodelling gill epithelia more quickly and at more extreme osmotic challenge than F. majalis. We detect an unusual pattern of baseline transcriptome divergence, where neutral evolutionary processes appear to govern expression divergence within species, but patterns of divergence for these genes between species do not follow neutral expectations. During acclimation, genome expression profiling identifies mechanisms of acclimation-associated response that are conserved within the genus including regulation of paracellular permeability. In contrast, several responses vary among species including those putatively associated with cell volume regulation, and these same mechanisms are targets for adaptive physiological divergence along osmotic gradients within F. heteroclitus. As such, the genomic and physiological mechanisms that are associated with adaptive fine-tuning within species also contribute to macro-evolutionary divergence as species diversify across osmotic niches., (© 2013 John Wiley & Sons Ltd.)

The Gulf killifish (Fundulus grandis) is a euryhaline fish found in coastal marsh along the entire of Gulf of Mexico and southern Atlantic of coast of the United States. The objective of this study was to investigate the effects of salinity on embryogenesis in the Gulf killifish. Four recirculation systems at salinities of 0.4, 7, 15, and 30 g/L were maintained at a static temperature with flow-through trays, containing embryos (n = 39) placed in triplicate into each system. Throughout embryogenesis, the rate of development, ammonia and urea excretion, and heart rate were monitored. Percent hatch was recorded, and morphological parameters were measured for larvae at hatch. As salinity was increased, the rate of embryogenesis decreased. Salinity significantly affected percent hatch with an 80.0% ± 2.6% for 7 g/L and 39.1 ± 4.3, 45.4 ± 4.5, and 36.3% ± 12.0% for 0.4, 15, and 30 g/L, respectively. Salinity and stage of development significantly affected production of ammonia and urea. As salinity increased, the dominate metabolite end product changed from urea to ammonia. However, the 15 g/L salinity treatment had the two highest levels of urea recorded. Heart rate was unaffected by salinity but increased throughout embryogenesis and remained constant once embryos reached stages where hatching has been recorded. While mean total length was not affected by salinity, embryos incubated in 30 g/L produced larvae with significantly thicker body depth at hatch. The 0.4, 7, and 15 g/L salinity treatments all had similar mean hours to hatch. The 30 g/L treatment resulted in a significantly longer mean time to hatch and smaller body cavity area at hatch.

The killifish Fundulus heteroclitus is abundant in osmotically dynamic estuaries and it can quickly adjust to extremes in environmental salinity. We performed a comparative osmotic challenge experiment to track the transcriptomic and physiological responses to two salinities throughout a time course of acclimation, and to explore the genome regulatory mechanisms that enable extreme osmotic acclimation. One southern and one northern coastal population, known to differ in their tolerance to hypo-osmotic exposure, were used as our comparative model. Both populations could maintain osmotic homeostasis when transferred from 32 to 0.4 p.p.t., but diverged in their compensatory abilities when challenged down to 0.1 p.p.t., in parallel with divergent transformation of gill morphology. Genes involved in cell volume regulation, nucleosome maintenance, ion transport, energetics, mitochondrion function, transcriptional regulation and apoptosis showed population- and salinity-dependent patterns of expression during acclimation. Network analysis confirmed the role of cytokine and kinase signaling pathways in coordinating the genome regulatory response to osmotic challenge, and also posited the importance of signaling coordinated through the transcription factor HNF-4α. These genome responses support hypotheses of which regulatory mechanisms are particularly relevant for enabling extreme physiological flexibility.

Adaptive variation tends to emerge clinally along environmental gradients or discretely among habitats with limited connectivity. However, in Atlantic killifish (Fundulus heteroclitus), a population genetic discontinuity appears in the absence of obvious barriers to gene flow along parallel salinity clines and coincides with a physiologically stressful salinity. We show that populations resident on either side of this discontinuity differ in their abilities to compensate for osmotic shock and illustrate the physiological and functional genomic basis of population variation in hypoosmotic tolerance. A population native to a freshwater habitat, upstream of the genetic discontinuity, exhibits tolerance to extreme hypoosmotic challenge, whereas populations native to brackish or marine habitats downstream of the discontinuity lose osmotic homeostasis more severely and take longer to recover. Comparative transcriptomics reveals a core transcriptional response associated with acute and acclimatory responses to hypoosmotic shock and posits unique mechanisms that enable extreme osmotic tolerance. Of the genes that vary in expression among populations, those that are putatively involved in physiological acclimation are more likely to exhibit nonneutral patterns of divergence between freshwater and brackish populations. It is not the well-known effectors of osmotic acclimation, but rather the lesser-known immediate-early responses, that appear important in contributing to population differences.

Genomic and physiological responses in Gulf killifish (Fundulus grandis) in the northern Gulf of Mexico have confirmed oil exposure of resident marsh fish following the Macondo blowout in 2010. Using these same fish, we evaluated otolith microchemistry as a method for assessing oil exposure history. Laser-ablation inductively-coupled-plasma mass spectrometry was used to analyze the chemical composition of sagittal otoliths to assess whether a trace metal signature could be detected in the otoliths of F. grandis collected from a Macondo-oil impacted site in 2010, post-spill relative to pre-spill, as well as versus fish from areas not impacted by the spill. We found no evidence of increased concentrations of two elements associated with oil contamination (nickel and vanadium) in F. grandis otoliths regardless of Macondo oil exposure history. One potential explanation for this is that Macondo oil is relatively depleted of those metals compared to other crude oils globally. During and after the spill, however, elevated levels of barium, lead, and to a lesser degree, copper were detected in killifish otoliths at the oil-impacted collection site in coastal Louisiana. This may reflect oil contact or other environmental perturbations that occurred concomitant with oiling. For example, increases in barium in otoliths from oil-exposed fish followed (temporally) freshwater diversions in Louisiana in 2010. This implicates (but does not conclusively demonstrate) freshwater diversions from the Mississippi River (with previously recorded higher concentrations of lead and copper), designed to halt the ingress of oil, as a mechanism for elevated elemental uptake in otoliths of Louisiana marsh fishes. These results highlight the potentially complex and indirect effects of the Macondo oil spill and human responses to it on Gulf of Mexico ecosystems, and emphasize the need to consider the multiple stressors acting simultaneously on inshore fish communities. [ABSTRACT FROM AUTHOR]

Na+ and Ca2+ regulation were compared in two euryhaline species, killifish (normally estuarine-resident) and rainbow trout (normally freshwater-resident) during an incremental salinity increase. Whole-body unidirectional fluxes of Na+ and Ca2+, whole body Na+ and Ca2+, and plasma concentrations (trout only), were measured over 1-h periods throughout a total 6-h protocol of increasing salinity meant to simulate a natural tidal flow. Killifish exhibited significant increases in both Na+ influx and efflux rates, with efflux slightly lagging behind efflux up to 60% SW, but net Na+ balance was restored by the time killifish reached 100% SW. Whole body Na+ did not change, in agreement with the capacity of this species to tolerate daily salinity fluctuations in its natural habitat. In contrast, rainbow trout experienced a dramatic increase in Na+ influx (50-fold relative to FW values), but not Na+ efflux between 40 and 60% SW, resulting in a large net loading of Na+ at higher salinities (60–100% SW), and increases in plasma Na+ and whole body Na+ at 100% SW. Killifish were in negative Ca2+ balance at all salinities, whereas trout were in positive Ca2+ balance throughout. Ca2+ influx rate increased two- to threefold in killifish at 80 and 100% SW, but there were no concomitant changes in Ca2+ efflux. Ca2+ flux rates were affected to a larger degree in trout, with twofold increases in Ca2+ influx at 40% SW and sevenfold increases at 100% SW. Again, there was no change in Ca2+ efflux with salinity, so plasma Ca2+ concentration increased in 100% SW. As the killifish is regularly submitted to increased salinity in its natural environment, it is able to rapidly activate changes in unidirectional fluxes in order to ensure ionic homeostasis, in contrast to the trout. [ABSTRACT FROM AUTHOR]

Following the Deepwater Horizon oil spill, approximately 7 million liters of the dispersant Corexit 9500A were released to promote oil biodegradation by breaking up surface oil slick formation. This process is accomplished via amphipathic anionic surfactants within dispersants that facilitate the mixing of aqueous and lipid phases. However, the amphipathicity of Corexit may also cause it to interact with biological membranes like the gill, impairing gill function and ultimately disrupting physiological processes mediated by it, such as osmoregulation. The goal of this study was to investigate the osmoregulatory effects and toxicity of Corexit in Gulf killifish. Killifish at the embryonic, larval, juvenile, and adult life stages were exposed to Corexit in water of different salinities to assess the interactive effects of ontogeny and salinity on Corexit toxicity. Corexit was not toxic to embryos except when exposed in hyperosmotic water where it had negligible effects; however, its toxicity to killifish increased dramatically following hatch, showing its greatest deleterious effects in adults. Corexit tended to increase sodium and chloride burdens in killifish when exposed in hyperosmotic waters and reduced whole-body and plasma ion concentrations in fish exposed to hypoosmotic waters. However, Corexit exposure at hyperosmotic salinities resulted in an increased differential accumulation of sodium over chloride as killifish matured. These findings suggest that Corexit may impair gill structure or alter specific components of osmoregulatory function, thus impacting osmoregulation in hypersosmotic and hypoosmotic waters, potentially impairing survival during osmotic challenges. Furthermore, the magnitude of these impacts continues to increase concomitant with gill ontogeny. • Sensitivity to Corexit increases as Gulf killifish mature from embryos to adults. • Corexit was more toxic to Gulf killifish in hyperosmotic waters relative to hypoosmotic waters at all life stages. • The effect of Corexit on osmoregulatory physiology of killifish appears to change with maturation. [ABSTRACT FROM AUTHOR]