Growth hormone and Prolactin-1 gene transcription in natural populations of the black-chinned tilapia Sarotherodon melanotheron acclimatised to different salinities

The effects of salinity on the expression of genes coding for growth hormone (GH) and prolactin-1 (PRL1) were studied in various natural populations of the black-chinned tilapia Sarotherodon melanotheron from West Africa. Individuals were sampled in June 2005 in six locations in Senegal and the Gambia, at various salinities between 0 and 101. The poorest condition factors were recorded in the most saline sampling site and the best growth in the fish from a marine environment. The pituitary GH mRNA levels were significantly higher in fish adapted to seawater, whereas the PRL1 mRNA levels were highest in fish adapted to fresh- and brackish water. These results show that the PRL1 mRNA levels seem to reflect relatively well the differences in environmental salinity, in contrast to those of GH, which would tend instead to reflect the individual growth in each environment. However, no relation could be found between growth in the hypersaline areas and the expression profile of GH. Although the fish analysed were morphologically identical, the expression of genes coding for GH and PRL1 showed large differences between individuals. This inter-individual variation in gene expression remains poorly understood.     

Comparative analysis of reproductive traits in black-chinned tilapia females from various coastal marine, estuarine and freshwater ecosystems

The black-chinned tilapia Sarotherodon melanotheron is a marine teleost characterised by an extreme euryhalinity. However, beyond a certain threshold at very high salinity, the species exhibits impaired growth and precocious reproduction. In this study, the relationships between reproductive parameters, environmental salinity and condition factor were investigated in wild populations of this species that were sampled in two consecutive years (2003 and 2004) from three locations in Senegal with different salinities: Guiers lake (freshwater, 0 psu), Hann bay (seawater, 37 psu) and Saloum estuary (hypersaline water, 66-127 psu). The highest absolute fecundity and spawning weight were recorded in seawater by comparison to either freshwater or hypersaline water whereas the poorest condition factors were observed in the most saline sampling site. These results reflect higher resource allocation to the reproduction due to the lowest costs of adaptation to salinity in seawater (the natural environment of this species) rather than differences in food resources at sites and/or efficiency at foraging and prey availability. Fecundities, oocyte size as well as spawning weight were consistent from year to year. However, the relative fecundity in the Saloum estuary varied significantly between the dry and rainy raisons with higher values in the wet season, which seems to reflect seasonal variations in environmental salinity. Such a reproductive tactic of producing large amounts of eggs in the rainy season when the salinity in the estuary was lower, would give the fry a better chance at survival and therefore assures a high larval recruitment. An inverse correlation was found between relative fecundity and oocyte size at the two extreme salinity locations, indicating that S. melanotheron has different reproductive strategies in these ecosystems. The adaptive significance of these two reproductive modes is discussed in regard to the heavy osmotic constraint imposed by extreme salinities and high inter-specific competition.     

Osmoregulatory strategies in natural populations of the black-chinned tilapia Sarotherodon melanotheron exposed to extreme salinities in West African estuaries

The effect of salinity was studied in natural populations of the black-chinned tilapia (Sarotherodon melanotheron) from West Africa. This euryhaline species colonizes nearly all coastal environments from bays to lagoons characterized by salinities ranging from fresh water to hypersaline water over 100 ‰. Individuals were sampled during the dry season at several locations characterized by different levels of salinity (3-102 ‰). Their osmotic status and their gills were analyzed. The branchial mitochondria-rich cells (MRC), localized at the basis of the filaments and along the lamellae in fish taken from the saline stations, showed a wide plasticity with significant differences in their number and size. The most striking results were a significant larger area (≈3x) and a higher number (≈55x) of MRC at high salinity (102 ‰) compared to low salinity (3 ‰). The major ion transporters and channels were localized by immunocytochemistry and different expression patterns have been recorded between stations. Despite an increased Na?/K?-ATPase (NKA) α-subunit expression and NKA activity, pointing to an increased monovalent ion excretion, a severe osmotic imbalance was recorded in animals living in hypersaline environments.     

Impact of environmental DDT concentrations on gill adaptation to increased salinity in the tilapia Sarotherodon melanotheron

Estuaries of tropical developing countries suffering from severe droughts induced by climate change are habitats to fish, which face drastic salinity variations and the contact with pollutants. The Western Africa tilapia Sarotherodon melanotheron is highly resistant to hypersalinity, but the effect of human-released xenobiotics on its adaptation is barely known. Controlled experiments were conducted to observe S. melanotheron gill adaptation to abrupt salinity variations in the presence of waterborne DDT, at concentrations detected in their natural habitat. The gills appeared as an important site of DDT conversion to DDD and/or depuration. A 12-days DDT exposure resulted in decreased gill epithelium thickness at all salinities (from fresh- to hypersaline-water), and the structure of gills from freshwater fish was particularly altered, relative to controls. No unbalance in tilapia blood osmolality was observed following DDT exposure, which however caused a decrease in branchial Na(+)-K(+)-ATPase (NKA) activity. Gill cellular NKA expression was reduced in salt-water, together with the expression of the CFTR chloride channel in hypersaline water. Although S. melanotheron seems very resistant (especially in seawater) to short-term waterborne DDT contamination, the resulting alterations of the gill tissue, cells and enzymes might affect longer term respiration, toxicant depuration and/or osmoregulation in highly fluctuating salinities.     

Functional microsatellite and possible selective sweep in natural populations of the black-chinned tilapia Sarotherodon melanotheron (Teleostei, Cichlidae)

The Prolactin I (PrlI) gene is a key locus involved in fish osmotic regulation. Two microsatellites, PrlI GT and PrlI AC, are present in the promoter region of this gene in tilapias. One of these microsatellite is associated to PrlI gene expression and growth rate. Aquaria reared individuals homozygous for long microsatellite alleles at the PrlI AC locus expressed less PrlI in fresh water than fishes with other genotypes. To assess the occurrence of selection acting on these microsatellites we carried out a survey of their variability compared with four others microsatellites in natural populations. Samples of two tilapia species were studied, Oreochromis niloticus (6 samples) a typically freshwater species inhabiting osmotically stable environments and Sarotherodon melanotheron (5 samples) a brackish water species obliged and able to adapt very rapidly to any salinity changes. For both species the different populations studied exhibited a high amount of differentiation (all F(st) value were statistically significant) at every loci. Only one locus, PrlI AC, was monomorphic for the same allele in all S. melanotheron populations. We have hypothesized that this outlier locus with its unique allele could have experienced a selective sweep that took place in the early stages of the species origin. S. melanotheron represents an example of adaptation (to fluctuant salinity environment) acting indirectly on gene product through its promoter.     

The Baltic Sea as a model system for studying postglacial colonization and ecological differentiation,exemplified by the red alga Ceramium tenuicorne

The Baltic Sea provides a unique model system for studying genetic effects of postglacial colonization and ecological differentiation, because all marine organisms must have immigrated after the opening of the Danish Straits 8000 years ago and responded to the development of the steep Skagerrak-Baltic salinity gradient. The red alga Ceramium tenuicorne shows conspicuous variation in growth and reproduction along this gradient. Herein we obtained reproductive data coupled with two types of molecular markers, one organellar (cox2-3 spacer sequences of mitochondrial DNA; mtDNA) and one mainly nuclear (random amplified polymorphic DNAs; RAPDs). Nine main populations were sampled in a nested spatial hierarchy including three salinity regions (Oslofjorden, Kattegat, and the Baltic Sea), and nine additional populations were sampled for the mtDNA analysis. Asexuality was frequent at low (Baltic) and medium (Kattegat) salinities but virtually absent at the highest salinity (Oslofjorden). Five mtDNA haplotypes were observed, of which two highly divergent ones were common. One was restricted to and fixed in Oslofjorden, and the other, which was closely related to the three rare haplotypes, was found from southernmost Norway via Kattegat into the Baltic. The RAPD data revealed, on the other hand, a continuous cline corresponding to the salinity gradient, with 27.4% divergence among salinity regions and most of the variation stored at the smallest spatial scale analysed (64.2%; within 1 m2 subpopulations). The combined data suggest colonization from a diverse Atlantic glacial gene pool followed by (1) lineage sorting of ancestral mtDNA polymorphisms and (2) strong differential selection among nuclear genotypes along the salinity gradient, including selection for nonrecombinant multiplication of those best fit to the marginal low-salinity habitats.     

Differences in Salinity Tolerance and Gene Expression Between Two Populations of Atlantic Cod (Gadus morhua) in Response to Salinity Stress

Populations of marine fish, even from contrasting habitats, generally show low genetic differentiation at neutral genetic markers. Nevertheless, there is increasing evidence for differences in gene expression among populations that may be ascribed to adaptive divergence. Studying variation in salinity tolerance and gene expression among Atlantic cod (Gadus morhua) from two populations distributed across a steep salinity gradient, we observed high mortality (45% North Sea cod and 80% Baltic Sea cod) in a reciprocal common garden setup. Quantitative RT-PCR assays for expression of hsp70 and Na/K-ATPase α genes demonstrated significant differences in gene regulation within and between populations and treatment groups despite low sample sizes. Most interesting are the significant differences observed in expression of the Na/K-ATPase α gene in gill tissue between North Sea and Baltic cod. The findings strongly suggest that Atlantic cod are adapted to local saline conditions, despite relatively low levels of neutral genetic divergence between populations.     

Tissue specific haemoglobin gene expression suggests adaptation to local marine conditions in North Sea flounder (Platichthys flesus L.)

Recent genetic analyses of candidate genes and gene expression in marine fishes have provided evidence of local adaptation in response to environmental differences, despite the lack of strong signals of population structure from conventional neutral genetic markers. In this study expression of the haemoglobin alpha and beta subunit genes was studied in reciprocally transplanted European flounder Platichthys flesus from the highly saline North Sea and the brackish Baltic Sea. Clear differences in expression patterns of haemoglobin alpha and beta subunit genes were found among different types of tissue in flounder. In gill tissue a plastic response to salinity treatments was observed with general up-regulation of these genes concomitant with higher salinity. For liver tissue a population specific expression differences was observed with lower expression at simulated non-native compared to native salinities. Finally, for kidney tissue a stress response was observed in one population, with gene up-regulation when North Sea flounders were transplanted to low salinity. This study underlines the importance of tissue specific gene expression and the significance of gene expression for evolution of local adaptation in high gene flow marine fishes.     

Divergence with gene flow in a population of thermophilic bacteria: a potential role for spatially varying selection

A fundamental goal of evolutionary biology is to understand how ecological diversity arises and is maintained in natural populations. We have investigated the contributions of gene flow and divergent selection to the distribution of genetic variation in an ecologically differentiated population of a thermophilic cyanobacterium (Mastigocladus laminosus) found along the temperature gradient of a nitrogen‐limited stream in Yellowstone National Park. For most loci sampled, gene flow appears to be sufficient to prevent substantial genetic divergence. However, one locus (rfbC) exhibited a comparatively low migration rate as well as other signatures expected for a gene experiencing spatially varying selection, including an excess of common variants, an elevated level of polymorphism and extreme genetic differentiation along the gradient. rfbC is part of an expression island involved in the production of the polysaccharide component of the protective envelope of the heterocyst, the specialized nitrogen‐fixing cell of these bacteria. SNP genotyping in the vicinity of rfbC revealed a ~5‐kbp region including a gene content polymorphism that is tightly associated with environmental temperature and therefore likely contains the target of selection. Two genes have been deleted both in the predominant haplotype found in the downstream region of White Creek and in strains from other Yellowstone populations of M. laminosus, which may result in the production of heterocysts with different envelope properties. This study implicates spatially varying selection in the maintenance of variation related to thermal performance at White Creek despite on‐going or recent gene flow.     

Intraspecific variation in expression of candidate genes for osmoregulation, heme biosynthesis and stress resistance suggests local adaptation in European flounder (Platichthys flesus)

Despite the recent discovery of significant genetic structuring in a large number of marine organisms, the evolutionary significance of these often minute genetic differences are still poorly understood. To elucidate the adaptive relevance of low genetic differentiation among marine fish populations, we studied expression differences of osmoregulatory and stress genes in genetically weakly differentiated populations of the European flounder (Platichthys flesus), distributed across a natural salinity gradient. Flounders were maintained in a long-term reciprocal transplantation experiment mimicking natural salinities in the North Sea and the Baltic Sea. Applying real-time quantitative PCR and microarray analysis we studied expression of four candidate genes (hsp70, angiotensinogen, Na/K-ATPase-alpha and 5-aminolevulinic acid synthase (ALAS)) in gill, kidney and liver tissues. Genes involved in osmoregulative processes (Na/K-ATPases-alpha and angiotensinogen) showed highly plastic but similar expression in the two populations dependent on environmental salinity. However, we observed a unique sixfold up-regulation of hsp70 in kidney tissue of flounder from the North Sea following long-term acclimation to Baltic salinities. Similarly, significant differences between North Sea and Baltic flounders in expression of ALAS in response to different salinities were found in gill and liver tissue. These findings strongly suggest that gene expression in flounders is shaped by adaptation to local environmental conditions. This identification of adaptive differences in high gene flow marine organisms adds a new dimension to our current understanding of evolutionary processes in the sea and is of paramount importance for identification, protection and sustainable management of marine biodiversity.     

Estimating salinity stress via hsp70 expression in the invasive round goby (Neogobius melanostomus): implications for further range expansion

Species invasions often occur on coasts and estuaries where abiotic conditions vary, e.g. salinity, temperature, runoff etc. Successful establishment and dispersal of non-indigenous species in many such systems are poorly understood, partially since the species tend to show genetic and ecological plasticity at population level towards many abiotic conditions, including salinity tolerance. Plasticity may be driven by shifting expression of heat shock proteins such as Hsp70, which is widely recognized as indicator of physical stress. In this study, we developed a qPCR assay for expression of the hsp70 gene in the invasive round goby (Neogobius melanostomus) and tested the expression response of fish collected from a brackish environment in the western Baltic Sea to three different salinities, 0, 10 and 30. hsp70 expression was highest in fresh water, indicating higher stress, and lower at brackish (ambient condition for the sampled population) and oceanic salinities, suggestive of low stress response to salinities above the population’s current distribution. The highest stress in fresh water was surprising since populations in fresh water exist, e.g. large European rivers and Laurentian Great Lakes. The results have implications to predictions for the species’ plasticity potential and possible range expansion of the species into other salinity regimes.

     

Local adaptation and oceanographic connectivity patterns explain genetic differentiation of a marine diatom across the North Sea–Baltic Sea salinity gradient

Drivers of population genetic structure are still poorly understood in marine micro‐organisms. We exploited the North Sea–Baltic Sea transition for investigating the seascape genetics of a marine diatom, Skeletonema marinoi. Eight polymorphic microsatellite loci were analysed in 354 individuals from ten locations to analyse population structure of the species along a 1500‐km‐long salinity gradient ranging from 3 to 30 psu. To test for salinity adaptation, salinity reaction norms were determined for sets of strains originating from three different salinity regimes of the gradient. Modelled oceanographic connectivity was compared to directional relative migration by correlation analyses to examine oceanographic drivers. Population genetic analyses showed distinct genetic divergence of a low‐salinity Baltic Sea population and a high‐salinity North Sea population, coinciding with the most evident physical dispersal barrier in the area, the Danish Straits. Baltic Sea populations displayed reduced genetic diversity compared to North Sea populations. Growth optima of low salinity isolates were significantly lower than those of strains from higher native salinities, indicating local salinity adaptation. Although the North Sea–Baltic Sea transition was identified as a barrier to gene flow, migration between Baltic Sea and North Sea populations occurred. However, the presence of differentiated neutral markers on each side of the transition zone suggests that migrants are maladapted. It is concluded that local salinity adaptation, supported by oceanographic connectivity patterns creating an asymmetric migration pattern between the Baltic Sea and the North Sea, determines genetic differentiation patterns in the transition zone.     

Matching molecular diversity and ecophysiology of benthic cyanobacteria and diatoms in communities along a salinity gradient

The phylogenetic diversity of oxygenic phototrophic microorganisms in hypersaline microbial mats and their distribution along a salinity gradient were investigated and compared with the halotolerances of closely related cultivated strains. Segments of 16S rRNA genes from cyanobacteria and diatom plastids were retrieved from mat samples by DNA extraction and polymerase chain reaction (PCR), and subsequently analysed by denaturing gradient gel electrophoresis (DGGE). Sequence analyses of DNA from individual DGGE bands suggested that the majority of these organisms was related to cultivated strains at levels that had previously been demonstrated to correlate with characteristic salinity responses. Proportional abundances of amplified 16S rRNA gene segments from phylogenetic groupings of cyanobacteria and diatoms were estimated by image analysis of DGGE gels and were generally found to correspond to abundances of the respective morphotypes determined by microscopic analyses. The results indicated that diatoms accounted for low proportions of cells throughout, that the cyanobacterium Microcoleus chthonoplastes and close relatives dominated the communities up to a salinity of 11% and that, at a salinity of 14%, the most abundant cyanobacteria were related to highly halotolerant cultivated cyanobacteria, such as the recently established phylogenetic clusters of Euhalothece and Halospirulina . Although these organisms in cultures had previously demonstrated their ability to grow with close to optimal rates over a wide range of salinities, their occurrence in the field was restricted to the highest salinities investigated.     

Stable isotope analyses provide new insights into ecological plasticity in a mixohaline population of European eel

Recent studies have shown that anguillid eel populations in habitats spanning the marine–freshwater ecotone can display extreme plasticity in the range of catadromy expressed by individual fishes. Carbon and nitrogen stable isotope analysis was used to differentiate between European eels (Anguilla anguilla) collected along a short (2 km) salinity gradient ranging from <1‰ to ~30‰ in Lough Ahalia, a tidal Atlantic lake system. Significant differences were recorded in mean δ¹³C, δ¹⁵N and C:N values from eels collected from fresh, brackish and marine-dominated basins. A discriminant analysis using these predictor variables correctly classified ca. 85% of eels to salinity zone, allowing eels to be classified as freshwater (FW), brackish (BW) or marine (MW) residents. The results of the discriminant analysis also suggested that a significant proportion of eels moved between habitats (especially between FW and BW). Comparisons of several key population parameters showed significant variation between eels resident in different salinity zones. Mean condition and estimated age was significantly lower in MW eels, whilst observed length at age (a correlate of growth) was significantly higher in MW eels, intermediate in BW and lowest in FW eels. This study has demonstrated that the ecology of eels found along a short salinity gradient can be extremely plastic and that stable isotope analysis has considerable utility in demonstrating intra-population variation in diadromous fishes.     

Distribution of picophytoplankton communities from brackish to hypersaline waters in a South Australian coastal lagoon

Background

Picophytoplankton (i.e. cyanobacteria and pico-eukaryotes) are abundant and ecologically critical components of the autotrophic communities in the pelagic realm. These micro-organisms colonized a variety of extreme environments including high salinity waters. However, the distribution of these organisms along strong salinity gradient has barely been investigated. The abundance and community structure of cyanobacteria and pico-eukaryotes were investigated along a natural continuous salinity gradient (1.8% to 15.5%) using flow cytometry.

Results

Highest picophytoplankton abundances were recorded under salinity conditions ranging between 8.0% and 11.0% (1.3 × 10⁶ to 1.4 × 10⁶ cells ml^-1). Two populations of picocyanobacteria (likely Synechococcus and Prochlorococcus) and 5 distinct populations of pico-eukaryotes were identified along the salinity gradient. The picophytoplankton cytometric-richness decreased with salinity and the most cytometrically diversified community (4 to 7 populations) was observed in the brackish-marine part of the lagoon (i.e. salinity below 3.5%). One population of pico-eukaryote dominated the community throughout the salinity gradient and was responsible for the bloom observed between 8.0% and 11.0%. Finally only this halotolerant population and Prochlorococcus-like picocyanobacteria were identified in hypersaline waters (i.e. above 14.0%). Salinity was identified as the main factor structuring the distribution of picophytoplankton along the lagoon. However, nutritive conditions, viral lysis and microzooplankton grazing are also suggested as potentially important players in controlling the abundance and diversity of picophytoplankton along the lagoon.

Conclusions

The complex patterns described here represent the first observation of picophytoplankton dynamics along a continuous gradient where salinity increases from 1.8% to 15.5%. This result provides new insight into the distribution of pico-autotrophic organisms along strong salinity gradients and allows for a better understanding of the overall pelagic functioning in saline systems which is critical for the management of these precious and climatically-stress ecosystems.