Strong selective effects of mitochondrial DNA on the nuclear genome.

Oxidative phosphorylation, the primary source of cellular energy in eukaryotes, requires gene products encoded in both the nuclear and mitochondrial genomes. As a result, functional integration between the genomes is essential for efficient adenosine triphosphate (ATP) generation. Although within populations this integration is presumably maintained by coevolution, the importance of mitonuclear coevolution in key biological processes such as speciation and mitochondrial disease has been questioned. In this study, we crossed populations of the intertidal copepod Tigriopus californicus to disrupt putatively coevolved mitonuclear genotypes in reciprocal F2 hybrids. We utilized interindividual variation in developmental rate among these hybrids as a proxy for fitness to assess the strength of selection imposed on the nuclear genome by alternate mitochondrial genotypes. Developmental rate varied among hybrid individuals, and in vitro ATP synthesis rates of mitochondria isolated from high-fitness hybrids were approximately two-fold greater than those of mitochondria isolated from low-fitness individuals. We then used Pool-seq to compare nuclear allele frequencies for high- or low-fitness hybrids. Significant biases for maternal alleles were detected on 5 (of 12) chromosomes in high-fitness individuals of both reciprocal crosses, whereas maternal biases were largely absent in low-fitness individuals. Therefore, the most fit hybrids were those with nuclear alleles that matched their mitochondrial genotype on these chromosomes, suggesting that mitonuclear effects underlie individual-level variation in developmental rate and that intergenomic compatibility is critical for high fitness. We conclude that mitonuclear interactions can have profound impacts on both physiological performance and the evolutionary trajectory of the nuclear genome.

Thermal tolerance of Acartia tonsa: In relation to acclimation temperature and life stage.

The Acartia tonsa, a calanoid copepod species, has high survival and thermal acclimation capacity in aquatic environments characterized by temperature variations. Dynamic and static thermal polygon areas of this species are 495 °C2 and 267 °C2 for nauplii stage, while adult stage has 747 °C2 and 411 °C2 dynamic and static thermal polygon area, respectively. In addition, Acartia tonsa is a copepod species which is more resistant to both high and low lethal temperatures, with its resistance zone of 105 °C2 and 131 °C2 for nauplii and adults, respectively. Acartia tonsa nauplii acclimated to 18 °C, 23 °C and 28 °C have lover and upper thermal limit (CTMin-CTMax) of 6.82-26.15 °C, 8.65-29.49 °C, and 11.70-34.10 °C, respectively. This species in the adult stage has a CTMin-CTMax of 4.47-30.30 °C, 6.35-33.94 °C, and 9.92-35.90 °C at acclimation temperatures mentioned above. Its broad dynamic and static thermal tolerance polygon areas and, accordingly, its significant thermal limits allow Acartia tonsa to survive at warm or cold extremes in their natural environment.

Nitric oxide mediates oxylipin production and grazing defense in diatoms.

Diatom blooms are important features of productive marine ecosystems and are known to support higher trophic levels. However, when stressed or wounded, diatoms can produce oxylipin molecules known to inhibit the reproduction and development of copepods and decrease microzooplankton growth rates. Using oxylipin chemical treatments, lipidomic analysis and functional genomic approaches, we provide evidence that nitric oxide (NO) and oxylipin signalling pathways in diatoms respond to protist grazers, resulting in increased defence fitness and survival. Exposure of the diatom Phaeodactylum tricornutum to the dinoflagellate Oxyrrhis marina resulted in NO production by P. tricornutum and pronounced change in its dissolved oxylipin profile. Experimentally elevating levels of NO also resulted in increased oxylipin production, and lower overall grazing rates. Furthermore, O. marina preferentially grazed on P. tricornutum prey with lower levels of NO, suggesting that this molecule and its effect on oxylipin pathways play a key role in prey selection. Exposure of O. marina grazing on P. tricornutum to exogenous oxylipins also decreased grazing rates, which is consistent with a grazing deterrence role for these molecules. These results suggest that NO and oxylipin production help to structure diatom communities, in part by modulating interactions with microzooplankton predators.

Changes in the Microbiome of Mariculture Feed Organisms after Treatment with a Potentially Probiotic Strain of Phaeobacter inhibens.

The Phaeobacter genus has been explored as probiotics in mariculture as a sustainable strategy for the prevention of bacterial infections. Its antagonistic effect against common fish pathogens is predominantly due to the production of the antibacterial compound tropodithietic acid (TDA), and TDA-producing strains have repeatedly been isolated from mariculture environments. Despite many in vitro trials targeting pathogens, little is known about its impact on host-associated microbiomes in mariculture. Hence, the purpose of this study was to investigate how the addition of a TDA-producing Phaeobacter inhibens strain affects the microbiomes of live feed organisms and fish larvae. We used 16S rRNA gene sequencing to characterize the bacterial diversity associated with live feed microalgae (Tetraselmis suecica), live feed copepod nauplii (Acartia tonsa), and turbot (Scophthalmus maximus) eggs/larvae. The microbial communities were unique to the three organisms investigated, and the addition of the probiotic bacterium had various effects on the diversity and richness of the microbiomes. The structure of the live feed microbiomes was significantly changed, while no effect was seen on the community structure associated with turbot larvae. The changes were seen primarily in particular taxa. The Rhodobacterales order was indigenous to all three microbiomes and decreased in relative abundance when P. inhibens was introduced in the copepod and turbot microbiomes, while it was unaffected in the microalgal microbiome. Altogether, the study demonstrates that the addition of P. inhibens in higher concentrations, as part of a probiotic regime, does not appear to cause major imbalances in the microbiome, but the effects were specific to closely related taxa.IMPORTANCE This work is an essential part of the risk assessment of the application of roseobacters as probiotics in mariculture. It provides insights into the impact of TDA-producing Phaeobacter inhibens on the commensal bacteria related to mariculture live feed and fish larvae. Also, the study provides a sequencing-based characterization of the microbiomes related to mariculture-relevant microalga, copepods, and turbot larvae.

Hatching and survival of the salmon 'gill maggot' Salmincola californiensis (Copepoda: Lernaeopodidae) reveals thermal dependence and undocumented naupliar stage.

Salmincola californiensis is a Lernaeopodid copepod parasitizing Pacific salmon and trout of the genus Oncorhynchus. Salmincola californiensis is of increasing concern in both native and introduced ranges because of its potential fish health impacts and high infection prevalence and intensity in some systems. Discrepancies in the documented life history phenology of S. californiensis with the sister species Salmincola edwardsii, as well as our laboratory observations, led us to question the existing literature. We documented a naupliar stage, thought lost for S. californiensis. In addition, we found a high degree of thermal sensitivity in egg development, with eggs developing faster under warmer conditions. Survival of copepodids was also highly dependent on temperature, with warmer conditions reducing lifespan. The longest lived copepodid survived 18 days at 4°C in stark contrast to the generally accepted <48 h survival for that life stage. We also note a consistent relationship between egg sac size and the number of eggs contained. However, egg sac sizes were highly variable. Our findings demonstrate that revisiting old assumptions for S. californiensis and related taxa will be a necessary step to improving our knowledge of the parasite life history and development that will be critical to disease management.

A Next-Generation Approach to Calculate Source-Sink Dynamics in Marine Metapopulations.

In marine systems, adult populations confined to isolated habitat patches can be connected by larval dispersal. Source-sink theory provides effective tools to quantify the effect of specific habitat patches on the dynamics of connected populations. In this paper, we construct the next-generation matrix for a marine metapopulation and demonstrate how it can be used to calculate the source-sink dynamics of habitat patches. We investigate the effect of environmental variables on the source-sink dynamics and demonstrate how the next-generation matrix can provide useful biological insight into transient as well as asymptotic dynamics of the model.

The filter-feeding bivalve Mytilus chilensis capture pelagic stages of Caligus rogercresseyi: A potential controller of the sea lice fish parasites.

The copepod Caligus rogercresseyi is an ectoparasite of several salmonid species. The pumping activity of filter-feeding molluscs could reduce the abundance of copepod dispersive larval stages in the water column. In this research, nauplius II and copepodid larvae of C. rogercresseyi were exposed to filtering mussels (Mytilus chilensis) of different sizes. These mussels were able to filter both larval stages, although they were more efficient in catching nauplius II. The fact that nauplius II were ingested more efficiently could be explained by their smaller size, lower swimming velocity (escape) and longer resting times between movements, when they were exposed to the influx of water around the inhalant area of the mussels. Larger mussels were more effective filtering C. rogercresseyi larvae due to their larger inhalant area and the related water influx. Additionally, the results suggest that larvae captured by the mussels can be incorporated into pseudofaeces or ingested and then released as part of the faeces. Thus, high concentrations of M. chilensis surrounding salmon farms may act as biological barriers, reducing the density of copepod dispersive larval stages and, thus, salmon infestations.

A biological characteristic extrapolation of compound toxicity for different developmental stage species with toxicokinetic-toxicodynamic model.

Intraspecific difference in toxicity brings uncertainty to ecological risk assessment (ERA) and water quality criteria (WQC) of chemicals. Here, we compared intraspecies sensitivity to toxicants for Mesocyclops leuckarti of which toxicity data was obtained from published literatures, and zebrafish Danio rerio of which toxicity data was done in this study). Due to the internal concentration of chemicals not measured, simplified toxicokinetic-toxicodynamic (TK-TD) models were used, and we investigated whether TK-TD parameters estimated by Bayesian method might represent the differences in sensitivity between life-stages of 2 species. The results demonstrated that the difference in TK-TD parameters (background mortality m0, no effect concentration NEC, the killing rate ks, and the dominant rate kd) could represent the toxicity difference between life-stages of individual species. The TK-TD model could predict toxicity in individual species (Cyprinus carpio L., Enchytraeus crypticus, Folsomia candida, Hyalella Azteca) exposed to different chemical concentrations and successfully extrapolate toxicity between different life stages of Mesocyclops leuckarti and Danio rerio by scaling several TK-TD parameters. The modified TK-TD model on the extrapolation toxicity of chemicals between life stages for species could be useful for the ERA and for deriving and revising WQC for chemicals.

Effects of a glyphosate-based herbicide on the development and biochemical biomarkers of the freshwater copepod Notodiaptomus carteri (Lowndes, 1934).

In this work we analyzed the effects of Sulfosato Touchdown®, a glyphosate-based herbicide, on the ontogenic development and biochemical markers of the freshwater copepod Notodiaptomus carteri. A 30-days life-cycle experiment was carried out with three different glyphosate concentrations (0, 0.38, and 0.81 mg L-1) to analyze the developmental time from nauplii to adult copepods and their individual growth. An additional 10-days experiment with the same glyphosate concentrations was designed to evaluate the energy reserves (glycogen, proteins and lipids) and the activity of three antioxidant enzymes, superoxide dismutase (SOD), catalase, and glutathione-S-transferase (GST) in adult copepods, separately for females and males. We found that the lowest glyphosate concentration increased the nauplii and total development time. The highest glyphosate concentration prevented copepods from reaching the adult stage, inhibited the growth of the first copepodite stage and increased the GST and SOD activity in adult females. According to our results, the presence of this herbicide in freshwater systems could impose a risk in the ecological role of copepods in nature. This study will contribute to propose the Notodiaptomus genus as model specie for monitoring purposes in the Neotropical aquatic systems.

The Effects of Microplastics on Dolioletta gegenbauri (Tunicata, Thaliacea).

Oceanographic studies revealed the abundance of minute plastic particles in coastal regions. Such particles, called microplastics, are abundant in sizes smaller than 100 µm ESD (Equivalent Spherical Diameter) and can be collected and ingested by planktonic copepods. Those animals are the most abundant metazoans on our planet. Abundantly co-occurring with planktonic copepods in subtropical and temperate neritic waters are doliolids (Tunicata, Thaliacea), which can dominate subtropical shelves because of their high asexual reproductive performance. Our studies were designed to examine the effects of polystyrene beads at low abundance, compared with phytoplankton, on abundantly occurring gonozooids of Dolioletta gegenbauri. Our findings reveal that such abundance of microplastic particles, in the presence of environmental concentrations of phytoplankton, reduces rates of feeding, growth, and oxygen consumption of this tunicate. Feeding rates on phytoplankton in the presence of beads were reduced by up to 58%, growth rates by up to 85%, and oxygen consumption rates by up to 33%. We conclude that such microplastic particles could limit the often in situ encountered pronounced proliferation of this tunicate species (Deibel in: Bone (ed) The biology of pelagic tunicates, Oxford University Press, Oxford, 1998).

Molecular evidence for the retention of the Thaumatopsyllidae in the order Cyclopoida (Copepoda) and establishment of four suborders and two families within the Cyclopoida.

The classification of the Thaumatopsyllidae within the Copepoda has been an issue of ongoing discussion since the discovery of Thaumatopsyllus paradoxus G.O. Sars, 1913 from the Norwegian coast. The family has been formally placed in the Monstrilloida, the Cyclopoida and even in its own order, the Thaumatopsylloida, based on different morphological criteria. We examined for the first time, the phylogenetic position of the Thaumatopsyllidae using gene sequences of 28S and 18S rRNA, as well as COI mtDNA, obtained from two thaumatopsyllid species occurring off the coast of southern California. We also fortuitously explored the phylogenetic relationships of the Cyclopoida in more detail than Khodami et al. (2017) by including a wider sample of key families such as the Erebonasteridae and Gisellinidae. Both Maximum Likelihood and Bayesian analyses revealed the Thaumatopsyllidae is nested in the Cyclopoida and is related to the marine Speleoithonidae. In addition, 16 families of the Cyclopoida are supported to be monophyletic, but surprisingly, the Cyclopidae is paraphyletic. The Cyclopicinidae is the first monophyletic offshoot of the cyclopoid tree, followed by two derived clades. The first clade contains a monophylum consisting of the Schminkepinellidae + Giselinidae which is sister to a clade including the monophyletic Erebonasteridae and all other poecilostome families. The second clade is divided into two main, well-supported family clusters. One includes the Cyclopidae encompassing two subfamilies (Eucylopinae and Cyclopinae), but unexpectedly the parasitic Lernaeidae cluster as a sister-group to the brackish water Halicyclops (subfamily Halicyclopinae) and the Euryteinae is the sister to all the rest of Cyclopidae s. l., making the Cyclopidae paraphyletic. To resolve this conundrum, we erected two families, Euryteidae and Halicyclopidae. The Cyclopidae s. str. retains the subfamilies Eucyclopinae and Cyclopinae, although our phylogeny does not support the reciprocal monophyly of these subfamilies. Our results support the gradual invasion of fresh water by the four families in this cluster. The highly supported monophyletic marine Euryteidae is the first offshoot followed by the brackish-water, free-living Halicyclopidae and the freshwater, parasitic Lernaeidae. The Cyclopidae fulfilled the colonization of freshwater bodies. The other clade of families comprises 12 monophyletic families recovered by our analysis, including the Pterinopsyllidae (at first offshoot), the Smirnovipinidae sister to the Hemicyclopinidae + Psammocyclopinidae, the Thaumatopsyllidae + Speleoithonidae, an undescribed family sister to the Archinotodelphyidae + Notodelphyidae and the Cyclopinidae sister to the Oithonidae + Cyclopettidae. We propose suborder ranks for each of the four main phylogenetic subdivisions of the Cyclopoida. These are named Cyclopicinida, Ergasilida, Cyclopida and Oithonida after the type genus of the oldest described family in the respective group.

Variation in developmental temperature alters adulthood plasticity of thermal tolerance in Tigriopus californicus.

In response to environmental change, organisms rely on both genetic adaptation and phenotypic plasticity to adjust key traits that are necessary for survival and reproduction. Given the accelerating rate of climate change, plasticity may be particularly important. For organisms in warming aquatic habitats, upper thermal tolerance is likely to be a key trait, and many organisms express plasticity in this trait in response to developmental or adulthood temperatures. Although plasticity at one life stage may influence plasticity at another life stage, relatively little is known about this possibility for thermal tolerance. Here, we used locally adapted populations of the copepod Tigriopus californicus to investigate these potential effects in an intertidal ectotherm. We found that low latitude populations had greater critical thermal maxima (CTmax) than high latitude populations, and variation in developmental temperature altered CTmax plasticity in adults. After development at 25°C, CTmax was plastic in adults, whereas no adulthood plasticity in this trait was observed after development at 20°C. This pattern was identical across four populations, suggesting that local thermal adaptation has not shaped this effect among these populations. Differences in the capacities to maintain ATP synthesis rates and to induce heat shock proteins at high temperatures, two likely mechanisms of local adaptation in this species, were consistent with changes in CTmax owing to phenotypic plasticity, which suggests that there is likely mechanistic overlap between the effects of plasticity and adaptation. Together, these results indicate that developmental effects may have substantial impacts on upper thermal tolerance plasticity in adult ectotherms.

Eggs of the copepod Acartia tonsa Dana require hypoxic conditions to tolerate prolonged embryonic development arrest.

BACKGROUND: Copepods make up the largest zooplankton biomass in coastal areas and estuaries and are pivotal for the normal development of fish larva of countless species. During spring in neritic boreal waters, the copepod pelagic biomass increases rapidly from near absence during winter. In the calanoid species Acartia tonsa, a small fraction of eggs are dormant regardless of external conditions and this has been hypothesized to be crucial for sediment egg banks and for the rapid biomass increase during spring. Other eggs can enter a state of induced arrest called quiescence when external conditions are unfavourable. While temperature is known to be a pivotal factor in the transition from developing to resting eggs and back, the role of pH and free Oxygen in embryo development has not been systematically investigated. RESULTS: Here, we show in a laboratory setting that hypoxic conditions are necessary for resting eggs to maintain a near-intact rate of survival after several months of induced resting. We further investigate the influence of pH that is realistic for natural sediments on the viability of resting eggs and document the effect that eggs have on the pH of the surrounding environment. We find that resting eggs acidify their immediate surroundings and are able to survive in a wide range of pH. CONCLUSIONS: This is the first study to demonstrate the importance of hypoxia on the survival capabilities of A. tonsa resting eggs in a controlled laboratory setting, and the first to show that the large majority of quiescent eggs are able to tolerate prolonged resting. These findings have large implications for the understanding of the recruitment of copepods from sediment egg banks, which are considered the primary contributor of nauplii seeded to pelagic populations in nearshore habitats in late spring.

Dynamics of Caligus rogercresseyi (Boxshall & Bravo, 2000) in farmed Atlantic salmon (Salmo salar) in southern Chile: Are we controlling sea lice?

Caligus rogercresseyi generates the greatest losses in the salmon industry in Chile. The relationship between salmon farming and sea lice is made up of various components: the parasite, host, environment and farming practices, which make it difficult to identify patterns in parasite population dynamics to define prevention and control strategies. The objectives of this study were to analyse and compare the effect of farming, sanitary practices and environmental variables on the abundance of gravid females (GF) and juveniles (JUV) of C. rogercresseyi on Salmo salar in three Salmon Neighborhood Areas (SNAs) in Region 10, south of Chile. Linear mixed-effects models of the negative binomial distribution were used to evaluate the effect of the different explanatory variables on GF and JUV. Productive variables were the key drivers explaining the abundance of GF and JUV. Results suggest that C. rogercresseyi is not controlled and JUV are persistent in the three SNAs, and sanitary practices do not control the dissemination of the parasite among sites. Environmental variables had a low impact on sea lice abundance. There is a need to perform analysis for modelling of parasite population dynamics to improve Integrated Pest Management, including changes in the governance to achieve an effective prevention and control.

Development of acute and chronic toxicity bioassays using the pelagic copepod Gladioferens pectinatus.

Well validated and reliable biological assays using local and native species are required to characterise the impacts of pollution on ecosystem health. We identified a native estuarine pelagic copepod species suitable for assessing the ecotoxicological impact of anthropogenic contaminants. Gladioferens pectinatus fulfilled the necessary-selection criteria of: wide distribution and abundance across New Zealand estuaries, ease of maintenance in the laboratory, short life cycle, sensitivity to toxicants with different modes of action, and providing reproducibility of biological response to toxicants. Measured endpoints were survival and larval development rate for the nauplii, and survival, realized offspring and total potential offspring for adults. LC50 values for the survival of G. pectinatus exposed to copper, phenanthrene and chlorpyrifos were 170 (143-193), 181.3 (131.3-231.3) and 4.3 (3.8-4.9) µg/L, respectively. The most sensitive chronic endpoint identified for G. pectinatus was the larval development rate, with EC50 values of 49.8 (45-55.3), 31.3 (24.8-44.7) and 1.97 (1.6-2.31) µg/L for copper, phenanthrene and chlorpyrifos, respectively. The acute and chronic responses obtained for G. pectinatus against the three reference toxicants are comparable with those reported for other copepod species and confirm its sensitivity and suitability to assess the toxicity of New Zealand estuarine samples.

Larval neurogenesis in the copepod Tigriopus californicus (Tetraconata, Multicrustacea).

Arthropod early neurogenesis shows distinct patterns that have been interpreted in an evolutionary framework. For instance, crustaceans and Hexapoda form the taxon Tetraconata and share the differentiation of specific neural precursors, the neuroblasts, a character which sets them apart from Chelicerata and Myriapoda. Neuroblasts are relatively large stem cells that generate ganglion mother cells by asymmetric divisions. Ganglion mother cells typically divide once to give rise to neurons and glia cells. In hexapods, neuroblasts segregate from the neuroectoderm before they begin their characteristic proliferative activity. In the crustaceans studied so far, neuroblasts remain in the neuroectoderm. Yet, detailed studies on early neurogenesis of crustaceans at the cellular level are largely restricted to some malacostracan and branchiopod species. Crustaceans are very diverse and likely paraphyletic with respect to hexapods. Hence, knowledge about neural differentiation in other crustacean taxa might contribute to the understanding of evolution of neurogenesis in Tetraconata. Here, we describe the early neurogenesis during naupliar development of the copepod Tigriopus californicus. We show that neuroblasts are present that generate ganglion mother cells, which in turn divide to give rise to neurons of the ventral nerve cord. These two neural precursor cell types and their specific arrangement correspond to what has been found in other crustaceans. One obvious difference concerns the relative size of the neuroblasts, which are not much larger than their progeny. Our results complement the picture of neural differentiation in crustaceans and suggest that superficially located neuroblasts are likely the ancestral condition in Tetraconata.

Warming of Subarctic waters accelerates development of a key marine zooplankton Calanus finmarchicus.

Recent observations confirm the rising temperatures of Atlantic waters transported into the Arctic Ocean via the West Spitsbergen Current (WSC). We studied the overall abundance and population structure of the North Atlantic keystone zooplankton copepod Calanus finmarchicus, which is the main prey for pelagic fish and some seabirds, in relation to selected environmental variables in this area between 2001 and 2011, when warming in the Arctic and Subarctic was particularly pronounced. Sampling within a 3-week time window each summer demonstrated that trends in the overall abundance of C. finmarchicus varied between years, with the highest values in "extreme" years, due to high numbers of nauplii and early copepodite stages in colder years (2001, 2004, 2010), and contrary to that, the fifth copepodite stage (C5) peaking in warm years (2006, 2007, 2009). The most influential environmental variable driving C. finmarchicus life cycle was temperature, which promoted an increased C5 abundance when the temperature was above 6°C, indicating earlier spawning and/or accelerated development, and possibly leading to their development to adults later in the summer and spawning for the second time, given adequate food supply. Based on the presented high interannual and spatial variability, we hypothesize that under a warmer climate, C. finmarchicus may annually produce two generations in the southern part of the WSC, what in turn could lead to food web reorganization of important top predators, such as little auks, and induce northward migrations of fish, especially the Norwegian herring.

Pushing the limit: Resilience of an Arctic copepod to environmental fluctuations.

Life history strategies such as multiyear life cycles, resting stages, and capital breeding allow species to inhabit regions with extreme and fluctuating environmental conditions. One example is the zooplankton species Calanus hyperboreus, whose life history is considered an adaptation to the short and unpredictable growth season in the central Arctic Ocean. This copepod is commonly described as a true Arctic endemic; however, by statistically analyzing compiled observational data, we show that abundances are relatively low and later stages and adults dominate in the central Arctic Ocean basins, indicating expatriation. Combining data analyses with individual-based modeling and energy requirement estimation, we further demonstrate that while C. hyperboreus can reach higher abundances in areas with greater food availability outside the central Arctic basins, the species' resilience to environmental fluctuations enables the life cycle to be completed in the central Arctic basins. Specifically, the energy level required to reach the first overwintering stage-a prerequisite for successful local production-is likely met in some-but not all-years. This fine balance between success and failure indicates that C. hyperboreus functions as a peripheral population in the central Arctic basins and its abundance will likely increase in areas with improved growth conditions in response to climate change. By illustrating a key Arctic species' resilience to extreme and fluctuating environmental conditions, the results of this study have implications for projections of future biogeography and food web dynamics in the Arctic Ocean, a region experiencing rapid warming and sea ice loss.

Gene silencing reveals multiple functions of Na+/K+-ATPase in the salmon louse (Lepeophtheirus salmonis).

Na+/K+-ATPase has a key function in a variety of physiological processes including membrane excitability, osmoregulation, regulation of cell volume, and transport of nutrients. While knowledge about Na+/K+-ATPase function in osmoregulation in crustaceans is extensive, the role of this enzyme in other physiological and developmental processes is scarce. Here, we report characterization, transcriptional distribution and likely functions of the newly identified L. salmonis Na+/K+-ATPase (LsalNa+/K+-ATPase) α subunit in various developmental stages. The complete mRNA sequence was identified, with 3003 bp open reading frame encoding a putative protein of 1001 amino acids. Putative protein sequence of LsalNa+/K+-ATPase revealed all typical features of Na+/K+-ATPase and demonstrated high sequence identity to other invertebrate and vertebrate species. Quantitative RT-PCR analysis revealed higher LsalNa+/K+-ATPase transcript level in free-living stages in comparison to parasitic stages. In situ hybridization analysis of copepodids and adult lice revealed LsalNa+/K+-ATPase transcript localization in a wide variety of tissues such as nervous system, intestine, reproductive system, and subcuticular and glandular tissue. RNAi mediated knock-down of LsalNa+/K+-ATPase caused locomotion impairment, and affected reproduction and feeding. Morphological analysis of dsRNA treated animals revealed muscle degeneration in larval stages, severe changes in the oocyte formation and maturation in females and abnormalities in tegmental glands. Thus, the study represents an important foundation for further functional investigation and identification of physiological pathways in which Na+/K+-ATPase is directly or indirectly involved.