Expanding hybrid zone between Solea aegyptiaca and Solea senegalensis: genetic evidence over two decades

Several marine hybrid zones have been described and studied during the last years. Assessing the movements of extending hybrid zones is central to improve our understanding of evolutionary processes. We have re-examined the hybrid zone between Solea aegyptiaca and Solea senegalensis that was first described 22 years ago in northern Tunisia when introgressed S. senegalensis individuals were found in the Gulf of Tunis, whereas locally caught S. aegyptiaca were genetically pure. Six population samples harvested both inside and outside the area where the two fish species coexist were genotyped for allozymes and exon-primed intron length polymorphism. Both types of markers were congruent and revealed that introgression takes place indeed in both directions. A high introgression level (36.4%) in the Bizerta lagoon and much less outside indicate that this is the main area where hybridization occurs while introgression clines towards the south in S. aegyptiaca and towards the north in S. senegalensis plead in favour of the existence of a unimodal hybrid zone. The higher introgression level calculated in the current study (when compared to 16% reported formerly) and the newly found introgressed S. aegyptiaca in Bizerta lagoon seem to indicate that the genetic exchanges occurring between the two taxa are evolving and not stabilized yet.     

Molecular systematics of the Atlanto-Mediterranean Solea species

Within the genus Solea , four sister lineages have been evolved that actually correspond to the species S. vulgaris, S. senegalensis, S. kleini and S. lascaris . The existence of S. aegyptiaca and S. impar is not supported and these species can be synonymized under S. vulgaris and S. lascaris , respectively. This pattern of taxonomy agrees fully with that proposed by Ben-Tuvia (1990; Journal of Fish Biology 36, 947–960) based on morphological features. In addition, mtDNA data contradict the widely accepted relatedness between S. kleini and S. lascaris based on the shared enlarged forenostril on the blind side. A sample of soles clearly related to S. senegalensis was caught in the Gulf of Taranto (Ionian Sea), representing the most western finding of this species in the Mediterranean Sea.     

Substantial loss of genetic variation in a single generation of Senegalese sole (Solea senegalensis) culture: implications in the domestication process

The flatfish Solea senegalensis represents an important resource in modern mariculture and is largely raised in South Spain and Portugal. Substantial progress has been achieved in its domestication, though suitable reproduction and zootechnical conditions still remain unknown. Difficulties to obtain breeders from wild lead many companies to set up broodstocks with first generation (G1) progeny without genetic guidelines. To study the genetic processes underlying the early stage of domestication of this species, the genetic structure of four representative broodstocks from Southern Spain has been assessed by means of eight microsatellite loci . Data revealed a substantial reduction in levels of genetic variability on just one generation in stocks totally or partially composed of G1 individuals when compared with those solely integrated by individuals from wild. The genetic relatedness within the four stocks has been established, having detected close relationships between individuals from cultured origin, thus suggesting that the loss of variability is apparently due to setting up broodstocks by blind selecting of family related G1 individuals. The high proportion of siblings in these stocks can have negative consequences in future generations due to inbreeding effects. The relationships between the four broodstocks were traced, having found a common origin between two of them, which come from the same donor hatchery, thus enhancing the homogenization of the existent resources for S. senegalensis among the various stocks. This finding alerts about the risks in exchanging fish between hatcheries without knowing their genetic origin. From this study, the use of microsatellites is strongly recommended to control the genetic composition of S. senegalensis broodstocks, aimed to maintain standards of genetic health and improve their reproduction capacity, which is a key issue in the domestication process.     

A taxonomic reappraisal of the Atlanto-Mediterranean soles Solea solea, S. senegalensis and S. lascaris

Examination of morphometric characters of Solea aegyptiaca shows that it cannot be separated from Solea solea . Diagnostic characters are given to separate this species from S. senegalensis . The available data show that Solea impar and S. nasuta are junior synonyms of S. lascaris . Meristic characters of S. solea and S. lascaris vary greatly according to the geographical origin of the examined fishes, indicating the influence of temperature on the number of vertebrae and fin rays. A key is provided for the identification of the four species of the genus Solea known in the Atlanto-Mediterranean region.     

Growth variability of juvenile soles Solea solea and Solea senegalensis, and comparison with RNA : DNA ratios in the Tagus estuary, Portugal

Growth variability and condition of juvenile soles Solea solea and Solea senegalensis , were assessed through RNA : DNA estimates and compared to absolute growth rates. Higher mean cohort RNA : DNA ratios were observed for cohort I at the beginning of estuarine occurrence for both species (4·42 and 4·87, for S. solea and S. senegalensis respectively). Despite different estuarine colonization habits, no significant differences were observed between RNA : DNA monthly variation for both sole species within the same year ( P  > 0·05 for 2003 and 2004). Juvenile S. senegalensis showed significant differences between RNA : DNA ratios obtained for the two nursery areas ( P  < 0·001). The decrease of seasonal growth rates with fish age was similar to seasonal variation of mean RNA : DNA values. Thus the RNA : DNA pattern of juvenile S. solea and S. senegalensis reflected growth and estuarine colonization patterns.     

Comparative analysis of AFLPs and SSRs efficiency in resolving population genetic structure of Mediterranean Solea vulgaris

The performance of different molecular markers in the assessment of population structure was tested using samples of Solea vulgaris collected in the Mediterranean within and outside the hypothetical dispersal ability of the species. A total of 172 individuals belonging to four population samples were analysed using 15 microsatellites [simple sequence repeats (SSRs)] and 153 amplified fragment length polymorphisms (AFLPs). Considering the global qualitative patterns, we found a correlation between SSRs and AFLPs in detecting genetic differentiation among samples. However, on a small geographical scale, AFLPs were able to discriminate individuals from neighbouring populations whereas SSRs were not, and the percentage of individuals correctly assigned to their population of origin was higher with AFLPs than with SSRs. The high number of loci analysed with the AFLP technique could increase the probability to include outlier loci in the analysis; however, the neutrality test performed on our data set did not show evidence of selection acting on the S. vulgaris samples. Even if the choice of the molecular marker depends mainly on the biological question to be addressed, the higher power of discrimination and the comparative technical ease of obtaining data from AFLPs with respect to SSRs suggest the use of AFLPs for many population genetics studies.     

Comparative feeding ecology of sympatric Solea solea and S. senegalensis, within the nursery areas of the Tagus estuary, Portugal

Among the stomach contents of 609 individuals of Solea solea and 1104 of S. senegalensis the main food items of S. solea were Corophium spp. and Hediste diversicolor, and of S. senegalensis were Corophium spp., H. diversicolor and Scrobicularia plana. For both species, the importance of larger prey items in the diet, namely H. diversicolor and Crangon crangon, increased with fish size. Feeding activity of S. solea and S. senegalensis increased in spring and summer. Short-term variations were particularly related to the tidal cycle and the two species fed in intertidal areas. Dietary dierences between the two nursery areas reflected prey availability mainly. Although intra- and interspecific length classes overlapped in diet, potential interspecific competition was probably minimized by a dierential habitat use pattern.     

Validation and comparison of microsatellite markers derived from Senegalese sole (Solea senegalensis, Kaup) genomic and expressed sequence tags libraries

In this work, we tested 100 potential new microsatellites (SSRs) equally derived from expressed sequence tag (EST) and enriched genomic-DNA libraries from Senegalese sole (Solea senegalensis, Kaup), a valuable cultured flatfish species. A final set of 69 new polymorphic microsatellites were validated after a population analysis, 37 of which corresponded to the first EST library constructed for Senegalese sole (EST-SSR). Although differences were not significant, EST sequences provided a higher proportion of quality markers (74%) than anonymous ones (64%). Most of the rejected anonymous SSRs (17 loci) were discarded because they did not generate PCR products; only one was monomorphic. On the contrary, all EST-SSRs gave PCR products, although monomorphism was more frequent (26%). Altogether, the number of alleles per locus was fairly similar in both SSR types, ranging from 2 to 19. The observed and expected heterozygosities varied from 0.105 to 1 and from 0.108 to 0.937, respectively. The main difference between the two sets was the percentage of annotated loci, being higher in EST-SSRs, as expected. Within the EST-SSRs, 46% of them showed flanking regions that significantly matched with EST sequences from other three flatfish species; however, the microsatellite itself was present only on half of these cases. These two new SSR sets constitute a suitable tool for fingerprinting, gene flow, genetic diversity, genome mapping studies and molecular-assisted breeding in this species.     

Skin darkness is related to cortisol, but not MSH, content in post‐larval Solea senegalensis

In this study it was shown that dark coloured post‐larvae of Senegalese sole Solea senegalensis , at two different ages, had elevated cortisol concentrations compared with lighter coloured individuals. As melanophore‐stimulating hormone levels were not elevated in dark coloured fish, it is possible that this hormone may not be the main melanotropic hormone involved in stress‐related skin darkening.     

Differential Use of Nursery Areas Within the Tagus Estuary by Sympatric Soles, Solea solea and Solea senegalensis

The spatio-temporal pattern of estuarine use by both Solea solea and Solea senegalensis was investigated between April 1995 and November 1996 by beam trawl surveys. Spatial and seasonal distribution and abundance were different for each species. The highest densities of S. solea were recorded in deeper, warmer, low salinity areas and where the sediment consisted of a high proportion of fine sand and presented a high abundance of amphipods. The 0-group individuals of this species first occurred on the nursery ground in April. Numbers peaked in June and July, and from October to April abundance was low. S. senegalensis had a wider distribution and its abundance was related to food availability. The 0-group individuals of this species occurred in the nursery areas later than S. solea, from June to August.     

The complete mitochondrial genome of the Senegal sole, Solea senegalensis Kaup. Comparative analysis of tandem repeats in the control region among soles.

The complete nucleotide sequence of the mitochondrial genome for the Senegal sole Solea senegalensis Kaup was determined. The mitochondrial DNA was 16,659 base pairs (bp) in length. Sequence features of the 13 protein-coding genes, two ribosomal RNAs and 22 transfer RNAs are described. The non-coding control region (1017 bp) was compared with those of the closely related soles Solea solea and Solea lascaris. The typical conservative blocks were identified. A cluster of 42 and 22 tandemly arrayed repeats was detected near the 3' end of control region in S. solea and S. lascaris, respectively. On the contrary, only two (93.8% of haplotypes) or three copies (6.2%) of an 8-bp repeated sequence motif was found in S. senegalensis. Phylogenetic analysis showed that 7 out of 9 of haplotypes bearing three copies grouped in a separate cluster. Possible mechanisms influencing the evolution of control region among soles are discussed.     

Development and characterization of microsatellites from Senegal sole (Solea senegalensis)

To assist in the domestication of the sole fish Solea senegalensis, 15 microsatellites were developed from its genome (13 di‐ and two trinucleotide). These loci, along with three from S. solea that cross‐amplified, were characterized on a sample of 20 individuals of S. senegalensis from a hatchery stock. All loci were polymorphic displaying from two to 15 alleles/locus (mean 6.5). The results obtained from the hatchery population screening indicate that they might be useful for studies of genetic structuring and/or kinship analysis in S. senegalensis.     

Introgressive hybridization and morphological transgression in the contact zone between two Mediterranean Solea species

Hybrid zones provide natural experiments where new combinations of genotypes and phenotypes are produced. Studying the reshuffling of genotypes and remodeling of phenotypes in these zones is of particular interest to document the building of reproductive isolation and the possible emergence of transgressive phenotypes that can be a source of evolutionary novelties. Here, we specifically investigate the morphological variation patterns associated with introgressive hybridization between two species of sole, Solea senegalensis and Solea aegyptiaca. The relationship between genetic composition at nuclear loci and individual body shape variation was studied in four populations sampled across the hybrid zone located in northern Tunisia. A strong correlation between genetic and phenotypic variation was observed among all individuals but not within populations, including the two most admixed ones. Morphological convergence between parental species was observed close to the contact zone. Nevertheless, the samples taken closest to the hybrid zone also displayed deviant segregation of genotypes and phenotypes, as well as transgressive phenotypes. In these samples, deviant body shape variation could be partly attributed to a reduced condition index, and the distorted genetic composition was most likely due to missing allelic combinations. These results were interpreted as an indication of hybrid breakdown, which likely contributes to postmating reproductive isolation between the two species.     

A review of the culture potential of Solea solea and S. senegalensis

Reviews in Fish Biology and Fisheries - A number of scientific studies have investigated aspects of soles(Solea soleaandS. senegalensis) ecology, population genetics and biology in their natural...     

Ceratomyxa aegyptiaca n. sp. (Myxozoa: Myxosporea) from the gall-bladder of Solea aegyptiaca Chabanaud (Pleuronectiformes: Soleidae) in a Tunisian coastal lagoon

A new marine myxosporean species, Ceratomyxa aegyptiaca n. sp. is described from the gall-bladder of Solea aegyptiaca Chabanaud collected from the Ghar El Melh Lagoon in northeastern Tunisia. Mature spores are elongate and crescent-shaped, measuring 8-11?μm in length and 48-58?μm in width. The polar capsules are spherical, 3.2-4?μm in diameter and equal in size. Trophozoites are polysporous and float free in the bile or are attached on the epithelium of the gall-bladder. Morphological data and molecular analysis based on 18S rDNA sequences are provided. The 18S rDNA of C. aegyptiaca is readily distinguishable from that of other myxozoan species, as the genetically most similar myxozoan parasite, C. seriolae Yokoyama & Fukuda, 2001 (AB530265) collected from Seriola quinqueradiata Temminck & Schlegel in Japanese waters, shares with it only 67.5% identical nucleotides over a 1,680-bp long fragment of 18S rDNA.     

Lunar and daily spawning rhythms of Senegal sole Solea senegalensis

A periodicity of 29 days was observed in spawning rhythms in Senegal sole Solea senegalensis , with an acrophase around the last quarter and the new moon. In both spring and autumn, a very marked nocturnal spawning rhythm was registered, with spawning beginning after dusk and the acrophase occurring around 2300 hours. When the photoperiod was artificially extended (from 10L:14D to 14L:10D), S. senegalensis synchronized to the new photoperiod: spawning took place after the new 'dusk', the beginning gradually shifting from 2100 to 2300 hours and the acrophase from 2325 to 0032 hours. Under continuous light conditions, fish sustained rhythmicity for 2 days, with an acrophase at 2249 hours, which suggested the existence of an endogenous pacemaker controlling the daily spawning rhythm. These findings provided new insights for better understanding the reproductive physiology of this species and for optimizing the timing protocols of egg collection and larvae production in S. senegalensis aquaculture.     

Control region haplotype variation in the central Mediterranean common sole indicates geographical isolation and population structuring in Italian stocks

Nine common sole Solea vulgaris samples from five fishery Management Units (MUs) of the central Mediterranean exhibited differences in a control region sequence marker. Parsimony network showed two low-divergent haplotype phylogroups. The former predominated in samples from the Ligurian, Tyrrhenian, Adriatic and north-western Ionian Seas (MUs 9, 10, 17, 18 and northern part of 19, respectively), whereas the latter was abundant in the south-western Ionian (southern part of MU 19). The geographical clustering of maternal lineages accounts for population structuring and indicates geographical isolation of common sole stocks in the Mediterranean. Several life-history traits of common sole and hydro-geographical features of the basin might support this pattern of differentiation. Haplotype frequency differences were detected among samples either from different MUs or within the same MU. This situation indicates the presence of partially subdivided or nearly panmictic population units, whose exploitation should be based on appropriate bio-ecological features. The usefulness of control region sequence marker enhances the routinely use of the genetic stock structure analysis in low-dispersal demersal marine resources.     

Osmoregulatory response of Senegalese sole (Solea senegalensis) to changes in environmental salinity

The osmoregulatory response of Senegalese sole (Solea senegalensis, Kaup 1858) to 14-day exposure and throughout 17-day exposure to different environmental salinities was investigated. A linear relationship was observed between environmental salinity and gill Na(+),K(+)-ATPase activity whereas kidney Na(+),K(+)-ATPase activity was unaffected. Two osmoregulatory periods could be distinguished according to variations in plasma osmolality: an adjustment period and a chronic regulatory period. No major changes in plasma osmolality and ions levels were registered at the end of the 14- to 17-day exposure period, indicating an efficient adaptation of the osmoregulatory system. Plasma levels of glucose and lactate were elevated in hypersaline water, indicating the importance of these energy substrates in these environments. Glucose was increased during hyper-osmoregulation but only in the adjustment period. Cortisol proved to be a good indicator of chronic stress and stress induced by transfer to the different osmotic conditions. This work shows that S. senegalensis is able to acclimate to different osmotic conditions during short-term exposure.     

Biometric variations in Solea vulgaris acclimatized in Lake Quarun, Upper Egypt

A population of Solea vulgaris was transplanted in 1938 from the Mediterranean to a land locked brackish lake. In 1977, the morphology of these fishes was investigated and a number of measurements made and compared with those of fish caught in the Mediterranean. Among these variations was less vertebrae and fewer dorsal fin rays in the lake soles.