Molecular characterization of the ?-subunit of Na?/K? ATPase from the euryhaline barnacle Balanus improvisus reveals multiple genes and differential expression of alternative splice variants.
ABSTRACT: The euryhaline bay barnacle Balanus improvisus has one of the broadest salinity tolerances of any barnacle species. It is able to complete its life cycle in salinities close to freshwater (3 PSU) up to fully marine conditions (35 PSU) and is regarded as one of few truly brackish-water species. Na?/K? ATPase (NAK) has been shown to be important for osmoregulation when marine organisms are challenged by changing salinities, and we therefore cloned and examined the expression of different NAKs from B. improvisus. We found two main gene variants, NAK1 and NAK2, which were approximately 70% identical at the protein level. The NAK1 mRNA existed in a long and short variant with the encoded proteins differing only by 27 N-terminal amino acids. This N-terminal stretch was coded for by a separate exon, and the two variants of NAK1 mRNAs appeared to be created by alternative splicing. We furthermore showed that the two NAK1 isoforms were differentially expressed in different life stages and in various tissues of adult barnacle, i.e the long isoform was predominant in cyprids and in adult cirri. In barnacle cyprid larvae that were exposed to a combination of different salinities and pCO2 levels, the expression of the long NAK1 mRNA increased relative to the short in low salinities. We suggest that the alternatively spliced long variant of the Nak1 protein might be of importance for osmoregulation in B. improvisus in low salinity conditions.
Project description:Barnacles are sessile macro-invertebrates, found along rocky shores in coastal areas worldwide. The euryhaline bay barnacle Balanus improvisus (Darwin, 1854) (= Amphibalanus improvisus) can tolerate a wide range of salinities, but the molecular mechanisms underlying the osmoregulatory capacity of this truly brackish species are not well understood. Aquaporins are pore-forming integral membrane proteins that facilitate transport of water, small solutes and ions through cellular membranes, and that have been shown to be important for osmoregulation in many organisms. The knowledge of the function of aquaporins in crustaceans is, however, limited and nothing is known about them in barnacles. We here present the repertoire of aquaporins from a thecostracan crustacean, the barnacle B. improvisus, based on genome and transcriptome sequencing. Our analyses reveal that B. improvisus contains eight genes for aquaporins. Phylogenetic analysis showed that they represented members of the classical water aquaporins (Aqp1, Aqp2), the aquaglyceroporins (Glp1, Glp2), the unorthodox aquaporin (Aqp12) and the arthropod-specific big brain aquaporin (Bib). Interestingly, we also found two big brain-like proteins (BibL1 and BibL2) constituting a new group of aquaporins not yet described in arthropods. In addition, we found that the two water-specific aquaporins were expressed as C-terminal splice variants. Heterologous expression of some of the aquaporins followed by functional characterization showed that Aqp1 transported water and Glp2 water and glycerol, agreeing with the predictions of substrate specificity based on 3D modeling and phylogeny. To investigate a possible role for the B. improvisus aquaporins in osmoregulation, mRNA expression changes in adult barnacles were analysed after long-term acclimation to different salinities. The most pronounced expression difference was seen for AQP1 with a substantial (>100-fold) decrease in the mantle tissue in low salinity (3 PSU) compared to high salinity (33 PSU). Our study provides a base for future mechanistic studies on the role of aquaporins in osmoregulation.
Project description:Understanding the ecological and evolutionary forces that determine the genetic structure and spread of invasive species is a key component of invasion biology. The bay barnacle, Balanus improvisus (= Amphibalanus improvisus), is one of the most successful aquatic invaders worldwide, and is characterised by broad environmental tolerance. Although the species can spread through natural larval dispersal, human-mediated transport through (primarily) shipping has almost certainly contributed to the current global distribution of this species. Despite its worldwide distribution, little is known about the phylogeography of this species. Here, we characterize the population genetic structure and model dispersal dynamics of the barnacle B. improvisus, and describe how human-mediated spreading via shipping as well as natural larval dispersal may have contributed to observed genetic variation. We used both mitochondrial DNA (cytochrome c oxidase subunit I: COI) and nuclear microsatellites to characterize the genetic structure in 14 populations of B. improvisus on a global and regional scale (Baltic Sea). Genetic diversity was high in most populations, and many haplotypes were shared among populations on a global scale, indicating that long-distance dispersal (presumably through shipping and other anthropogenic activities) has played an important role in shaping the population genetic structure of this cosmopolitan species. We could not clearly confirm prior claims that B. improvisus originates from the western margins of the Atlantic coasts; although there were indications that Argentina could be part of a native region. In addition to dispersal via shipping, we show that natural larval dispersal may play an important role for further colonisation following initial introduction.
Project description:BACKGROUND: Salinity plays an important role in shaping coastal marine communities. Near-future climate predictions indicate that salinity will decrease in many shallow coastal areas due to increased precipitation; however, few studies have addressed this issue. The ability of ecosystems to cope with future changes will depend on species' capacities to acclimatise or adapt to new environmental conditions. Here, we investigated the effects of a strong salinity gradient (the Baltic Sea system--Baltic, Kattegat, Skagerrak) on plasticity and adaptations in the euryhaline barnacle Balanus improvisus. We used a common-garden approach, where multiple batches of newly settled barnacles from each of three different geographical areas along the Skagerrak-Baltic salinity gradient were exposed to corresponding native salinities (6, 15 and 30 PSU), and phenotypic traits including mortality, growth, shell strength, condition index and reproductive maturity were recorded. RESULTS: We found that B. improvisus was highly euryhaline, but had highest growth and reproductive maturity at intermediate salinities. We also found that low salinity had negative effects on other fitness-related traits including initial growth and shell strength, although mortality was also lowest in low salinity. Overall, differences between populations in most measured traits were weak, indicating little local adaptation to salinity. Nonetheless, we observed some population-specific responses--notably that populations from high salinity grew stronger shells in their native salinity compared to the other populations, possibly indicating adaptation to differences in local predation pressure. CONCLUSIONS: Our study shows that B. improvisus is an example of a true brackish-water species, and that plastic responses are more likely than evolutionary tracking in coping with future changes in coastal salinity.
Project description:Barnacles form a globally ubiquitous group of sessile crustaceans that are particularly common in the coastal intertidal. Several barnacle species are described as highly euryhaline and a few species even have the ability to colonize estuarine and brackish habitats below 5 PSU. However, the physiological and/or morphological adaptations that allow barnacles to live at low salinities are poorly understood and current knowledge is largely based on classical eco-physiological studies offering limited insight into the molecular mechanisms. This review provides an overview of available knowledge of salinity tolerance in barnacles and what is currently known about their osmoregulatory strategies. To stimulate future studies on barnacle euryhalinity, we briefly review and compare barnacles to other marine invertebrates with known mechanisms of osmoregulation with focus on crustaceans. Different mechanisms are described based on the current understanding of molecular biology and integrative physiology of osmoregulation. We focus on ion and water transport across epithelial cell layers, including transport mechanisms across cell membranes and paracellular transfer across tight junctions as well as on the use of intra- and extracellular osmolytes. Based on this current knowledge, we discuss the osmoregulatory mechanisms possibly present in barnacles. We further discuss evolutionary consequences of barnacle osmoregulation including invasion-success in new habitats and life-history evolution. Tolerance to low salinities may play a crucial role in determining future distributions of barnacles since forthcoming climate-change scenarios predict decreased salinity in shallow coastal areas. Finally, we outline future research directions to identify osmoregulatory tissues, characterize physiological and molecular mechanisms, and explore ecological and evolutionary implications of osmoregulation in barnacles.
Project description:Barnacle settlement involves sensing of a variety of exogenous cues. A pair of antennules is the main sensory organ that the cyprid larva uses to explore the surface. Antennules are equipped with a number of setae that have both chemo- and mechanosensing function. The current study explores the repertoire of sensory receptors in Balanus improvisus cyprid antennules with the goal to better understand sensory systems involved in the settling behavior of this species. We carried out transcriptome sequencing of dissected B. improvisus cyprid antennules. The generated transcriptome assembly was used to search for sensory receptors using HMM models. Among potential chemosensory genes, we identified the ionotropic receptors IR25a, IR8a and IR93a, and several divergent IR candidates to be expressed in the cyprid antennules. We found one gustatory-like receptor but no odorant receptors, chemosensory or odorant-binding proteins. Apart from chemosensory receptors, we also identified 13 potential mechanosensory genes represented by several transient receptor potential channels (TRP) subfamilies. Furthermore, we analyzed changes in expression profiles of IRs and TRPs during the B. improvisus settling process. Several of the sensory genes were differentially expressed during the course of larval settlement. This study gives expanded knowledge about the sensory systems present in barnacles, a taxonomic group for which only limited information about receptors is currently available. It furthermore serves as a starting point for more in depth studies of how sensory signaling affects settling behavior in barnacles with implications for preventing biofouling.
Project description:Temperature and salinity shape the distribution and genetic structure of marine communities. Future warming and freshening will exert an additional stress to coastal marine systems. The extent to which organisms respond to these shifts will, however, be mediated by the tolerances of all life-stages and populations of species and their potential to adapt. We investigated nauplius and cypris larvae of the barnacle Balanus (Amphibalanus) improvisus from the Swedish west coast with respect to temperature (12, 20, and 28 °C) and salinity (5, 15, and 30) tolerances. Warming accelerated larval development and increased overall survival and subsequent settlement success. Nauplii developed and metamorphosed best at intermediate salinity. This was also observed in cypris larvae when the preceding nauplii stages had been reared at a salinity of 30. Direct comparisons of the present findings with those on a population from the more brackish Baltic Sea demonstrate contrasting patterns. We conclude that i) B. improvisus larvae within the Baltic region will be favoured by near-future seawater warming and freshening, that ii) salinity tolerances of larvae from the two different populations reflect salinities in their native habitats, but are nonetheless suboptimal and that iii) this species is generally highly plastic with regard to salinity.
Project description:Non-indigenous species (NIS) can impact marine biodiversity and ecosystem structure and function. Once introduced into a new region, secondary dispersal is limited by the physiology of the organism in relation to the ambient environment and by complex interactions between a suite of ecological factors such as presence of predators, competitors, and parasites. Early prediction of dispersal potential and future 'area of impact' is challenging, but also a great asset in taking appropriate management actions. Aerobic scope (AS) in fish has been linked to various fitness-related parameters, and may be valuable in determining dispersal potential of aquatic invasive species in novel environments. Round goby, Neogobius melanostomus, one of the most wide-ranging invasive fish species in Europe and North America, currently thrives in brackish and fresh water, but its ability to survive in high salinity waters is unknown to date. We show that AS in round goby is reduced by 30% and blood plasma osmolality increased (indicating reduced capacity for osmoregulation) at salinities approaching oceanic conditions, following slow ramping (5 PSU per week) and subsequent long-term acclimation to salinities ranging between 0 and 30 PSU (8 days at final treatment salinities before blood plasma osmolality measurements, 12-20 additional days before respirometry). Survival was also reduced at the highest salinities yet a significant proportion (61%) of the fish survived at 30 PSU. Reduced physiological performance at the highest salinities may affect growth and competitive ability under oceanic conditions, but to what extent reduced AS and osmoregulatory capacity will slow the current 30 km year-1 rate of advance of the species through the steep salinity gradient from the brackish Baltic Sea and into the oceanic North Sea remains speculative. An unintended natural experiment is in progress to test whether the rate of advance slows down. At the current rate of advance the species will reach the oceanic North Sea by 2018/2019, therefore time for taking preventative action is short.
Project description:In estuaries, salinity is believed to limit the colonization of brackish water habitats by freshwater species. Blue catfish Ictalurus furcatus, recognized as a freshwater species, is an invasive species in tidal rivers of the Chesapeake Bay. Salinity tolerance of this species, though likely to determine its potential range expansion and dispersal in estuarine habitats, is not well-known. To address this issue, we subjected blue catfish to a short-term salinity tolerance experiment and found that this species tolerates salinities higher than most freshwater fishes and that larger blue catfish tolerate elevated salinities for longer periods compared with smaller individuals. Our results are supported by spatially extensive, long-term fisheries surveys in the Chesapeake Bay region, which revealed a gradual (1975-2017) down-estuary range expansion of blue catfish from tidal freshwater areas to habitats exceeding 10 psu [practical salinity units] and that large blue catfish (> 200 mm fork length) occur in salinities greater than 10 psu in Chesapeake Bay tributaries. Habitat suitability predictions based on our laboratory results indicate that blue catfish can use brackish habitats to colonize new river systems, particularly during wet months when salinity decreases throughout the tidal rivers of the Chesapeake Bay.
Project description:The invasive Asian shore crab, Hemigrapsus sanguineus, is ubiquitous in the rocky intertidal zone of the western North Atlantic. A likely contributor to this colonization is that H. sanguineus is able to handle a wide range of salinities, and is thus more likely to spread through a greater geographic area of estuaries. This study investigated the salinity effects on this animal by observing survival across a range of salinities, the maintenance of hemolymph osmolality under different salinities, and behavioral preference for and avoidance of salinities. H. sanguineus showed high survival across a broad range of salinities, had little change in hemolymph osmolality over a short-term salinity shock, and behaviorally distinguished between salinities when presented with a choice, under both acclimation salinities of 5 PSU or 35 PSU. Such results suggest H. sanguineus has a hardiness for the rapid changes in salinity that happen in the intertidal zone, yet is capable of physically moving to a more optimal salinity. This enhances their competitiveness as an invader, particularly surviving lower salinities that present challenges during high-precipitation events in rocky intertidal areas, and partially explains this species' dominance in this habitat type.
Project description:Disturbances and environmental change are important factors determining the diversity, composition, and functioning of communities. However, knowledge about how natural bacterial communities are affected by such perturbations is still sparse. We performed a whole ecosystem manipulation experiment with freshwater rock pools where we applied salinity disturbances of different intensities. The aim was to test how the compositional and functional resistance and resilience of bacterial communities, alpha- and beta-diversity and the relative importance of stochastic and deterministic community assembly processes changed along a disturbance intensity gradient. We found that bacterial communities were functionally resistant to all salinity levels (3, 6, and 12 psu) and compositionally resistant to a salinity increase to 3 psu and resilient to increases of 6 and 12 psu. Increasing salinities had no effect on local richness and evenness, beta-diversity and the proportion of deterministically vs. stochastically assembled communities. Our results show a high functional and compositional stability of bacterial communities to salinity changes of different intensities both at local and regional scales, which possibly reflects long-term adaptation to environmental conditions in the study system.