No observed developmental effects in early life stages of capelin (Mallotus villosus) exposed to a water-soluble fraction of crude oil during embryonic development.

The rise in offshore oil and gas operations, maritime shipping, and tourism in northern latitudes enhances the risk of oil spills to sub-Arctic and Arctic coastal environments. Therefore, there is a need to understand the potential adverse effects of petroleum on key species in these areas. Here, we investigated the effects of oil exposure on the early life stages of capelin (Mallotus villosus), an ecologically and commercially important Barents Sea forage fish species that spawns along the coast of Northern Norway. Capelin embryos were exposed to five different concentrations (corresponding to 0.5-19 µg/L total PAHs) of water-soluble fraction (WSF) of crude oil from 6 days post fertilization (dpf) until hatch (25 dpf), and development of larvae in clean seawater was monitored until 52 dpf. None of the investigated endpoints (embryo development, larval length, heart rate, arrhythmia, and larval mortality) showed any effects. Our results suggest that the early life stages of capelin may be more robust to crude oil exposure than similar life stages of other fish species.

Genome-wide DNA methylation predicts environmentally driven life history variation in a marine fish.

Epigenetic modifications are thought to be one of the molecular mechanisms involved in plastic adaptive responses to environmental variation. However, studies reporting associations between genome-wide epigenetic changes and habitat-specific variations in life history traits (e.g., lifespan, reproduction) are still scarce, likely due to the recent application of methylome resequencing methods to non-model species. In this study, we examined associations between whole genome DNA methylation and environmentally driven life history variation in 2 lineages of a marine fish, the capelin (Mallotus villosus), from North America and Europe. In both lineages, capelin harbor 2 contrasting life history tactics (demersal vs. beach-spawning). Performing whole genome and methylome sequencing, we showed that life history tactics are associated with epigenetic changes in both lineages, though the effect was stronger in European capelin. Genetic differentiation between the capelin harboring different life history tactics was negligible, but we found genome-wide methylation changes in both lineages. We identified 9,125 European and 199 North American differentially methylated regions (DMRs) due to life history. Gene ontology (GO) enrichment analysis for both lineages revealed an excess of terms related to neural function. Our results suggest that environmental variation causes important epigenetic changes that are associated with contrasting life history tactics in lineages with divergent genetic backgrounds, with variable importance of genetic variation in driving epigenetic variation. Our study emphasizes the potential role of genome-wide epigenetic variation in adaptation to environmental variation.

Characterizing the stress response in juvenile Delta smelt exposed to multiple stressors.

The Delta Smelt (Hypomesus transpacificus), once an abundant fish endemic to the Sacramento-San Joaquin Estuary, is now on the brink of extinction. Due to the high sensitivity of this species, knowledge of their stress response will be vital to their future survival and sustainability. Understanding the magnitude and kinetics of cortisol induction in Delta Smelt will provide valuable information when interpreting the degree of environmentally relevant stressors, such as warming and predator exposure. As little is known about the primary stress response and cortisol dynamics in Delta Smelt, the first aim of this study was to measure basal and maximal whole-body cortisol prior to and following exposure to a sublethal and significant netting stress at 17 and 21 °C. Our findings reveal that juvenile Delta Smelt held at 21 °C display an exacerbated stress response and a reduction in available energy compared to fish held at 17 °C. There was no evidence of the secondary stress response to the netting stress as whole-body glucose and lactate levels in treatment groups remained similar to basal values. The second aim of this study was to investigate the effect of a largemouth bass (Micropterus salmoides) predator cue, which was found to induce a significant increase in cortisol relative to control levels in juvenile Delta Smelt. Indices such as cortisol can be used as bioindicators of stress in the field and results from this study suggest that moderate temperatures and reduced predation are optimal release conditions during hatchery-based supplementation to minimize stress to this highly sensitive species.

Unpacking the complexity of longitudinal movement and recruitment patterns of facultative amphidromous fish.

Longitudinal movement plays fundamental role in habitat colonization and population establishment of many riverine fish species. Movement patterns of amphidromous fish species at fine-scales that would allow characterizing the direction of movement and factors associated with the establishment of specific life-history strategies (resident or amphidromous) in rivers are still poorly understood. We assess fine-scale longitudinal movement variability patterns of facultative amphidromous fish species Galaxias maculatus in order to unfold its life-history variation and associated recruitment habitats. Specifically, we analyzed multi-elemental composition along core to edge transects in ear-bones (otoliths) of each fish using recursive partitions that divides the transect along signal discontinuities. Fine-scale movement assessment in five free-flowing river systems allowed us to identify movement direction and potential recruitment habitats. As such, resident recruitment of G. maculatus in freshwater (71%) and estuarine (24%) habitats was more frequent than amphidromous recruitment (5%), and was linked to availability of slow-flowing lotic or lentic habitats that produce or retain small-bodied prey consumed by their larvae. We postulate that life-history variation and successful recruitment of facultative amphidromous fish such as G. maculatus in river systems is driven by availability of suitable recruitment habitats and natural hydrologic connectivity that allows fish movement to these habitats.

Gross morphology, histology, and ultrastructure of the olfactory rosette of a critically endangered indicator species, the Delta Smelt, Hypomesus transpacificus.

The Delta Smelt (Hypomesus transpacificus) is a small, semi-anadromous fish native to the San Francisco Bay-Delta Estuary and has been declared as critically endangered. Their olfactory biology, in particular, is poorly understood and a basic description of their sensory anatomy is needed to advance our understanding of the sensory ecology of species to inform conservation efforts to manage and protect them. We provide a description of the gross morphology, histological, immunohistochemical, and ultrastructural features of the olfactory rosette in this fish and discuss some of the functional implications in relation to olfactory ability. We show that Delta Smelt have a multilamellar olfactory rosette with allometric growth. Calretinin immunohistochemistry revealed a diffuse distribution of olfactory receptor neurons within the epithelium. Ciliated, microvillous and crypt neurons were clearly identified using morphological and immunohistochemical features. The olfactory neurons were supported by robust ciliated and secretory sustentacular cells. Although the sense of smell has been overlooked in Delta Smelt, we conclude that the olfactory epithelium has many characteristics of macrosmatic fish. With this study, we provide a foundation for future research into the sensory ecology of this imperiled fish.

Reproductive ecology of a critically endangered alpine galaxiid.

Stocky galaxias Galaxias tantangara is a newly described freshwater fish restricted to a single population, occupying a 3 km reach of a small headwater stream in the upper Murrumbidgee River catchment of south-eastern Australia. This species is listed as critically endangered under IUCN Red List criteria, and knowledge of the species' ecology is critical for future conservation efforts to establish additional populations by translocation and captive breeding. This study details the first account of spawning and reproductive ecology of G. tantangara, including reproductive development, timing of spawning and a description of one spawning site. Peak gonadosomatic index was observed in March/April in males and in October in females. Absolute fecundity ranged from 211 oocytes for a 76 mm length to caudal fork (LCF) fish to 810 oocytes for a 100 mm LCF fish. The observation of spent females in mid-November 2017 and discovery of an egg mass 8 days later suggest that spawning had occurred, and over a relatively short period. Larvae were subsequently detected in monthly electrofishing surveys in December 2017. Findings from this study provide new understanding of existing and future threats to G. tantangara and have important implications for conservation management of not only this species but also other closely related threatened Galaxias species.

Understanding seasonal migration of Shishamo smelt in coastal regions using environmental DNA.

Migratory organisms have their own life histories that efficiently link multiple ecosystems. Therefore, comprehensive understanding of migration ecologies of these organisms is essential for both species conservation and ecosystem management. However, monitoring migration at fine spatiotemporal scales, especially in open marine systems, often requires huge costs and effort. Recently, environmental DNA (eDNA) techniques that utilize DNA released from living organisms into their environment became available for monitoring wild animals without direct handling. In this study, we conducted an eDNA survey for understanding marine migration of an endemic fish species, Shishamo smelt (Spirinchus lanceolatus). We examined 1) seasonal habitat changes in coastal regions and 2) environmental factors potentially driving the migration of this species. The eDNA concentrations along a 100 km-long coastline exhibited spatiotemporal variation, suggesting that this species shifts their habitat away from nearshore areas between spring and summer. We also found a significantly negative association between the eDNA concentration and sea surface temperature. That finding suggests that the offshore migration of this species is associated with increased sea surface temperature. This study reveals new aspects of S. lanceolatus life history in coastal regions. Together with our previous eDNA study on the freshwater migration of S. lanceolatus, this study illustrates the potential of eDNA techniques for understanding the whole life history of this migratory species.

Bifenthrin exposure causes hyperactivity in early larval stages of an endangered fish species at concentrations that occur during their hatching season.

Bifenthrin is a pyrethroid insecticide commonly used in agricultural and urban sectors, and is found in watersheds worldwide. As a sodium channel blocker, at sublethal concentrations it causes off-target effects, including disruption of calcium signaling and neuronal growth. At the whole organism level, sublethal concentrations of bifenthrin cause behavioral effects in fish species, raising concerns about the neurotoxic properties of the compound on fish populations. Here we describe the application of a high-throughput behavioral system to evaluate contaminant impacts on the sensitive early-life stages of Delta smelt (Hypomesus transpacificus), a critically endangered teleost species endemic to the San Francisco Bay Delta (SFBD), California, USA. Leveraging the natural behavior of early-larval Delta smelt, whereby they increase movement in bright light and decrease movement in the dark, we developed a test using a cycle of light and dark periods in a closed chamber to test hyper- or hypoactivity for this species. We show that early-larval Delta smelt have a significant preference to move toward light, and utilized the behavioral test to evaluate the impact of exposure to bifenthrin at concentrations found in habitats where Delta smelt reportedly spawn, ranging up to concentrations detected in tributaries to these habitats. All tested concentrations of bifenthrin (nominal 2, 10, or 100 ng/L) caused hyperactivity, over a 96 h exposure, with noted significance determined during the light period of the test. To further understand the impact of bifenthrin exposure, expression of a suite of genes relevant to neurodevelopment, the mechanistic target of rapamycin (mTOR) signaling pathway, and biotransformation in exposed larvae were also measured. Following exposure to picomolar concentrations of bifenthrin, expression of genes in the mTOR signaling and neurogenesis pathways were altered alongside behavior. This study demonstrates how light and dark cycle behavioral tests can be used to assess sensitive alterations in swimming activity in Delta smelt at early developmental stages and how gene expression can complement these assays. This approach can be used to assess the impact of multiple compounds that occur within the restricted habitat of Delta smelt, thus having the potential to greatly inform conservation management strategies for this critically sensitive life stage.

Morphological predictors of swimming speed performance in river and reservoir populations of Australian smelt Retropinna semoni.

Dam construction is a major driver of ecological change in freshwater ecosystems. Fish populations have been shown to diverge in response to different flow velocity habitats, yet adaptations of fish populations to river and reservoir habitats created by dams remains poorly understood. We aimed to evaluate divergence of morphological traits and prolonged swimming speed performance between lotic and lentic populations of Australian smelt Retropinna semoni and quantify the relationship between prolonged swimming speed performance and morphology. Prolonged swimming speed performance was assessed for 15 individuals from each of three river and two reservoir populations of R. semoni using the critical swimming speed test (Ucrit ). Body shape was characterized using geometric morphometrics, which was combined with fin aspect ratios and standard length to assess morphological divergence among the five populations. Best subsets model-selection was used to identify the morphological traits that best explain Ucrit variation among individuals. Our results indicate R. semoni from river populations had significantly higher prolonged swimming speed performance (Ucrit = 46.61 ± 0.98 cm s-1 ) than reservoir conspecifics (Ucrit = 35.57 ± 0.83 cm s-1 ; F1,74 = 58.624, Z = 35.938, P < 0.001). Similarly, R. semoni sampled from river populations had significantly higher fin aspect ratios (ARcaudal = 1.71 ± 0.04 and 1.29 ± 0.02 respectively; F(1,74) = 56.247, Z = 40.107, P < 0.001; ARpectoral = 1.85 ± 0.03 and 1.33 ± 0.02 respectively; F(1,74) = 7.156, Z = 4.055, P < 0.01). Best-subset analyses revealed Ucrit was most strongly correlated with pectoral and caudal fin aspect ratios (R2 adj = 0.973, AICc = 41.54). Body shape, however, was subject to a three-way interaction among population, habitat and sex effects (F3,74 = 1.038. Z = 1.982; P < 0.05). Thus sexual dimorphism formed a significant component of unique and complex variation in body shape among populations from different habitat types. This study revealed profound effects of human-altered flow environments on locomotor morphology and its functional link to changes in swimming performance of a common freshwater fish. While past studies have indicated body shape may be an important axis for divergence between lotic and lentic populations of several freshwater fishes, fin aspect ratios were the most important predictor of swimming speed in our study. Differences in body morphology here were inconsistent between river and reservoir populations, suggesting this aspect of phenotype may be more strongly influenced by other factors such as predation and sexual dimorphism.

Does migration promote or inhibit diversification? A case study involving the dominant radiation of temperate Southern Hemisphere freshwater fishes.

Although theory predicts that dispersal has a pivotal influence on speciation and extinction rates, it can have contradictory effects on each, such that empirical quantification of its role is required. In many studies, dispersal reduction appears to promote diversification, although some comparisons of migratory and nonmigratory species suggest otherwise. We tested for a relationship between migratory status and diversification rate within the dominant radiation of temperate Southern Hemisphere freshwater fishes, the Galaxiidae. We reconstructed a molecular phylogeny comprising >95% of extant taxa, and applied State-dependent Speciation Extinction models to estimate speciation, extinction, and diversification rates. In contrast to some previous studies, we revealed higher diversification rates in nonmigratory lineages. The reduced gene flow experienced by nonmigratory galaxiids appears to have increased diversification under conditions of allopatry or local adaptation. Migratory galaxiid lineages, by contrast, are genetically homogeneous within landmasses, but may also be rarely able to diversify by colonizing other landmasses in the temperate Southern Hemisphere. Apparent contradictions among studies of dispersal-diversification relationships may be explained by the spatial context of study systems relative to species dispersal abilities, by means of the "intermediate dispersal" model; the accurate quantification of dispersal abilities will aid in the understanding of these proposed interactions.

Shared ancestral polymorphisms and chromosomal rearrangements as potential drivers of local adaptation in a marine fish.

Gene flow has tremendous importance for local adaptation, by influencing the fate of de novo mutations, maintaining standing genetic variation and driving adaptive introgression. Furthermore, structural variation as chromosomal rearrangements may facilitate adaptation despite high gene flow. However, our understanding of the evolutionary mechanisms impending or favouring local adaptation in the presence of gene flow is still limited to a restricted number of study systems. In this study, we examined how demographic history, shared ancestral polymorphism, and gene flow among glacial lineages contribute to local adaptation to sea conditions in a marine fish, the capelin (Mallotus villosus). We first assembled a 490-Mbp draft genome of M. villosus to map our RAD sequence reads. Then, we used a large data set of genome-wide single nucleotide polymorphisms (25,904 filtered SNPs) genotyped in 1,310 individuals collected from 31 spawning sites in the northwest Atlantic. We reconstructed the history of divergence among three glacial lineages and showed that they probably diverged from 3.8 to 1.8 million years ago and experienced secondary contacts. Within each lineage, our analyses provided evidence for large Ne and high gene flow among spawning sites. Within the Northwest Atlantic lineage, we detected a polymorphic chromosomal rearrangement leading to the occurrence of three haplogroups. Genotype-environment associations revealed molecular signatures of local adaptation to environmental conditions prevailing at spawning sites. Our study also suggests that both shared polymorphisms among lineages, resulting from standing genetic variation or introgression, and chromosomal rearrangements may contribute to local adaptation in the presence of high gene flow.

Cuttlefish retrieve whether they smelt or saw a previously encountered item.

According to the Source Monitoring Framework, the origin of a memory is remembered through the retrieval of specific features (e.g. perceptive, sensitive, affective signals). In two source discrimination tasks, we studied the ability of cuttlefish to remember the modality in which an item had been presented several hours ago. In Experiment 1, cuttlefish were able to retrieve the modality of presentation of a crab (visual vs olfactory) sensed before 1 h and 3 hrs delays. In Experiment 2, cuttlefish were trained to retrieve the modality of the presentation of fish, shrimp, and crabs. After training, cuttlefish performed the task with another item never encountered before (e.g. mussel). The cuttlefish successfully passed transfer tests with and without a delay of 3 hrs. This study is the first to show the ability to discriminate between two sensory modalities (i.e. see vs smell) in an animal. Taken together, these results suggest that cuttlefish can retrieve perceptual features of a previous event, namely whether they had seen or smelled an item.

Finding continuity and discontinuity in fish schools via integrated information theory.

Collective behaviours are known to be the result of diverse dynamics and are sometimes likened to living systems. Although many studies have revealed the dynamics of various collective behaviours, their main focus has been on the information processing performed by the collective, not on interactions within the collective. For example, the qualitative difference between three and four elements in a system has rarely been investigated. Tononi et al. proposed integrated information theory (IIT) to measure the degree of consciousness Φ. IIT postulates that the amount of information loss caused by the minimum information partition is equivalent to the degree of information integration in the system. This measure is not only useful for estimating the degree of consciousness but can also be applied to more general network systems. Here, we obtained two main results from the application of IIT (in particular, IIT 3.0) to the analysis of real fish schools (Plecoglossus altivelis). First, we observed that the discontinuity on 〈Φ(N)〉 distributions emerges for a school of four or more fish. This transition was not observed by measuring the mutual information or the sum of the transfer entropy. We also analysed the IIT on Boids simulations with respect to different coupling strengths; however, the results of the Boids model were found to be quite different from those of real fish. Second, we found a correlation between this discontinuity and the emergence of leadership. We discriminate leadership in this paper from its traditional meaning (e.g. defined by transfer entropy) because IIT-induced leadership refers not to group behaviour, as in other methods, but the degree of autonomy (i.e. group integrity). These results suggest that integrated information Φ can reveal the emergence of a new type of leadership which cannot be observed using other measures.

Behavioural crypsis in a South African galaxiid fish induced by predatory and non-predatory heterospecifics.

This study investigated recognition of, and behavioural responses to, predatory and non-predatory heterospecifics by a small cryptically coloured fish species, Galaxias 'nebula'. Nebula recognised and differentiated between predatory and non-predatory heterospecifics and altered its behaviour facultatively. With both predatory and non-predatory fishes, the proportion of time spent motionless increased, whereas refugia use was affected only by predators and neither heterospecific affected the time spent active. Although nebula appeared to face no conflict, in that their responses to predatory and non-predatory heterospecifics varied in the same direction and differed only in intensity, the presence of both heterospecifics together induced responses midway between those for each heterospecific separately. Non-predatory heterospecifics thus modified nebula's responses towards predators, potentially making time available for other essential activities such as foraging. This modified predator response may aid its survival in an increasingly threatened habitat.

Assessing the potential for post-ejaculatory female choice in a polyandrous beach-spawning fish.

In species with limited opportunities for pre-ejaculatory sexual selection (behavioural components), post-ejaculatory mechanisms may provide opportunities for mate choice after gametes have been released. Recent evidence from a range of taxa has revealed that cryptic female choice (i.e., female-mediated differential fertilization bias), through chemical cues released with or from eggs, can differentially regulate the swimming characteristics of sperm from various males and ultimately determine male fertilization success under sperm competition. We assessed the potential role that such female-modulated chemical cues play in influencing sperm swimming characteristics in beach-spawning capelin (Mallotus villosus), an externally fertilizing fish that mates as couples (one male and one female) or threesomes (two males and one female) with presumably limited opportunities for pre-ejaculatory sexual selection. We assayed sperm swimming characteristics under varying doses and donor origins of egg cues and also examined the possibility of assortative mating based on body size. We found mating groups were not associated by size, larger males did not produce better quality ejaculates, and egg cues (regardless of dosage or donor identity) did not influence sperm swimming characteristics. Our findings suggest that intersexual pre-ejaculatory sexual selection and cryptic female choice mediated by female chemical cues are poorly developed in capelin, possibly due to unique natural selection constraints on reproduction.

Sexually selected traits are larger and more variable in male than female beach-spawning capelin, Mallotus villosus.

We evaluated whether morphological traits in capelin, Mallotus villosus, that appear to be sexually selected (pectoral fin, pelvic fin, anal fin, lateral ridge) were larger and more variable in males than females compared with naturally selected morphological traits (eyes, dorsal fin). Photographs were obtained of 136 capelin captured at two spawning sites and standardised measurements were taken of six morphological traits. Males had larger traits than females for a given body size and this was most pronounced in the traits thought to be sexually selected. Body size explained much of the variation in female traits but less variation in male traits, suggesting alternative selection pressures are involved. We suggest that larger male body size aids in endurance rivalry and sexually dimorphic traits help males to remain in physical contact with females while spawning on the beach.

Effects of water accommodated fraction of physically and chemically dispersed heavy fuel oil on beach spawning capelin (Mallotus villosus).

Due to a northward shift in off-shore activities, including increased shipping traffic and oil and gas exploration there is a growing focus on the potential effects of oil pollution on Arctic marine ecosystems. Capelin (Mallotus villosus) is a small fish and a member of the smelt family, and is a key species in the marine food chain. Capelin are seasonally abundant in the Northern Atlantic and in coastal Arctic waters, e.g. in western Greenland and in the Barents Sea, where it undertakes aggregated spawning in the intertidal and subtidal zone. To study the possible effects of oil pollution on the physiology and development of early life stages in capelin, freshly fertilised capelin eggs were exposed to a water accommodated fraction of physically (WAF) and chemically (CEWAF) dispersed heavy fuel oil (IFO30) for 72 h. Subsequent mortality, hatching success, larvae malformations, growth and CYP1A/EROD activity was measured over a 4-week period. The nominal exposure concentrations of WAF and CEWAF were between 0.02 and 14.5 mg total hydrocarbon content (THC) L-1 and 0.5-304 mg THC L-1, respectively. Egg mortality correlated significantly with WAF exposure concentration. The proportions of hatched eggs decreased with increasing CEWAF exposure concentration. Further, the percentage of malformed larvae with craniofacial abnormalities, body axis defects, generally under developed larvae, reduced total body length (dwarfs), correlated significantly with exposure concentrations in both CEWAF and WAF treatments. The four types of the predominant malformations were distributed differently in two parallel experiments. At the biochemical level, we observed a significant relationship between CEWAF exposure concentration and CYP1A/EROD activity in newly hatched larvae and this effect persisted for 3 weeks after the 72 h exposure. We conclude that even short-term exposure to both heavy fuel oil WAF and CEWAF, at environmentally relevant THC concentrations following an oil spill, may induce adverse developmental effects on the vulnerable early life stages of capelin. The mechanisms responsible for the observed effects on mortality, growth and embryo development in capelin eggs and embryos following WAF and CEWAF exposure require further studies.

Oxidative stress in the galaxiid fish, Galaxias maculatus, exposed to binary waterborne mixtures of the pro-oxidant cadmium and the anti-oxidant diclofenac.

Chemical mixtures represent environmentally-realistic exposures of contaminants to aquatic biota. However, there remains a limited understanding of how toxicant mixtures may impact biological function, relative to their individual components. In the current study, oxidative stress responses of the freshwater galaxiid fish inanga (Galaxias maculatus) were examined following exposure to the pro-oxidant trace metal cadmium (2 or 9 µg L-1), and the anti-oxidant pharmaceutical drug diclofenac (770 µg L-1), individually or in simple binary mixtures. Cadmium exposure in the absence of diclofenac significantly decreased renal catalase activity, increased hepatic catalase activity, decreased renal superoxide dismutase (SOD) and decreased glutathione-S-transferase activity, effects that are suggestive of anti-oxidant defense inhibition and/or generation of increased reactive oxygen species. Diclofenac exposure in the absence of cadmium resulted in a decreased renal lipid peroxidation, consistent with its known anti-oxidant properties. The presence of waterborne diclofenac altered the effects of cadmium on catalase activity in the liver, SOD activity in the gill, and lipid peroxidation in the liver. Co-exposure with cadmium modulated diclofenac effects on lipid peroxidation in the kidney. These data indicate the capacity of each of these toxicants to offset biological effects of the other when both co-occur in urban waters at specific concentrations. This study also demonstrates the complexity of outcomes in contaminant mixtures, even when these stressors are presented as simple binary combinations.

Role of freshwater floodplain-tidal slough complex in the persistence of the endangered delta smelt.

Seasonal floodplain wetland is one of the most variable and diverse habitats found in coastal ecosystems, yet it is also one of the most highly altered by humans. The Yolo Bypass, the primary floodplain of the Sacramento River in California's Central Valley, USA, has been shown to provide various benefits to native fishes when inundated. However, the Yolo Bypass exists as a tidal dead-end slough during dry periods and its value to native fishes has been less studied in this state. During the recent drought (2012-2016), we found higher abundance of the endangered Delta Smelt (Hypomesus transpacificus), than the previous 14 years of fish monitoring within the Yolo Bypass. Meanwhile, Delta Smelt abundance elsewhere in the estuary was at record lows during this time. To determine the value of the Yolo Bypass as a nursery habitat for Delta Smelt, we compared growth, hatch dates, and diets of juvenile Delta Smelt collected within the Yolo Bypass with fish collected among other putative nursery habitats in the San Francisco Estuary between 2010 and 2016. Our results indicated that when compared to other areas of the estuary, fish in the Yolo Bypass spawned earlier, and offspring experienced both higher quality feeding conditions and growth rates. The occurrence of healthy juvenile Delta Smelt in the Yolo Bypass suggested that the region may have acted as a refuge for the species during the drought years of 2012-2016. However, our results also demonstrated that no single region provided the best rearing habitat for juvenile Delta Smelt. It will likely require a mosaic of habitats that incorporates floodplain-tidal sloughs in order to promote the resilience of this declining estuarine fish species.

Involvement of ayu NOD2 in NF-κB and MAPK signaling pathways: Insights into functional conservation of NOD2 in antibacterial innate immunity.

Nucleotide oligomerization domain 2 (NOD2) is a major cytoplasmic sensor for pathogens and is critical for the clearance of cytosolic bacteria in mammals. However, studies regarding NOD2, especially the initiated signaling pathways, are scarce in teleost species. In this study, we identified a NOD2 molecule (PaNOD2) from ayu (Plecoglossus altivelis). Bioinformatics analysis showed the structure of NOD2 to be highly conserved during vertebrate evolution. Dual-luciferase reporter assays examined the activation of NF-κB signaling and Western blotting analysis detected the phosphorylation of three MAP kinases (p-38, Erk1/2, and JNK1/2). Functional study revealed that, like its mammalian counterparts, PaNOD2 was the receptor of the bacterial cell wall component muramyl dipeptide (MDP), and the leucine-rich repeat motif was responsible for the recognition and binding of PaNOD2 with the ligand. Overexpression of PaNOD2 activated the NF-κB signaling pathway, leading to the upregulation of inflammatory cytokines, including TNF-α and IL-1ß in HEK293T cells and ayu head kidney-derived monocytes/macrophages (MO/MΦ). Particularly, we found that PaNOD2 activated the MAPK signaling pathways, as indicated by the increased phosphorylation of p-38, Erk1/2, and JNK1/2, which have not been characterized in any teleost species previously. Our findings proved that the NOD2 molecule and initiated pathways are conserved between mammals and ayu. Therefore, ayu could be used as an animal model to investigate NOD2-based diseases and therapeutic applications.