An electrophoretic investigation of populations of Atherina boyeri Risso, 1810 and A. presbyter Cuvier, 1829 (Teleostei: Atherinidae): genetic evidence in support of the two species

The data obtained using electrophoresis strongly support the specific status of Atherina boyeri Risso, 1810 and A. presbyfer Cuvier, 1829, and thuscontradict the recently proposedsynonymyof the two species. Four populations of A. boyeri and six populations of A. presbyter were assayed for 11 enzymes and general protein using muscle and liver extracts. Eight of the 11 enzymes were shown to be polymorphic at the 95% level. Sixteen loci, encoding 40 putative alleles were consistently resolved in all 10 populations.
The two species were fixed for different alleles at the EST-3 locus. At the G3PDH locus. with the exception of two heterozygotes, all individuals of each species were also homozygous for different alleles. At the PGM locus the common allele was unique to each species.
The mean Nei's genetic distance ( ), over all loci, calculated between populations of A. boyeri (= 0.10 ± 0.06); between populations of A. presbyter ( = 0.02 0.02) and between populations of A. boyeri and A. presbyter ( = 0.42 0.09) indicated the separateness of the two species.
UPGMA cluster analysis based on genetic distances produced a dendrogram whose principal dichotomy resulted in the formation of two clusters. The ordination of populations in the UPGMA cluster analysis strongly reflected the geographic distribution of populations in both species.     

A multivariate morphometric investigation of Atherina boyeri Risso, 1810 and A. presbyter Cuvier, 1829 (Teleostei: Atherinidae): morphometric evidence in support of the two species

Two recently developed morphometric methodologies, multiple group principal components analysis (MGPCA) and the box truss method of body form transformation are used to re-address whether Atherina boyeri Risso, 1810 and A. presbyter Cuvier, 1829 are morphometrically distinct. The results revealed that morphometric differences are present between A. boyeri and A. presbyter , consistent with expectations of the existence of two species.
Females of the two species were clearly discriminated into two separate groups because the within-group covariances of several characters within populations of each species were different between species i.e., the covariances were heteroscedastic. The occurrence of heteroscedasticity within the data, although statistically invalidating the results of canonical variates anslysis, indicated there to be a large amount of morphological variation between females of the two species. Males were differentiated as a result of differences in body 'shape', particularly the relationship between head 'shape' and body 'shape'.
Several probable reasons are given for the failure of a previous study to differentiate the two species. The data rejects the synonymy of the two species in support of the continued use of the taxa A. boyeri Risso, 1810 and A. presbyter Cuvier, 1829.     

Multilocus phylogenetic analysis of the genus Atherina (Pisces: Atherinidae)

Sand-smelts are small fishes inhabiting inshore, brackish and freshwater environments and with a distribution in the eastern Atlantic and Mediterranean Sea, extending south into the Indian Ocean. Here, we present a broad phylogenetic analysis of the genus Atherina using three mitochondrial (control region, 12S and 16S) and two nuclear markers (rhodopsin and 2nd intron of S7). Phylogenetic analyses fully support the monophyly of the genus. Two anti-tropical clades were identified, separating the South African Atherina breviceps from the north-eastern Atlantic and Mediterranean Atherina' species. In European waters, two groups were found. The first clade formed by a well supported species-pair: Atherina presbyter (eastern Atlantic) and Atherina hepsetus (Mediterranean), both living in marine waters; a second clade included Atherina boyeri (brackish and freshwater environments) and two independent lineages of marine punctated and non-punctated fishes, recently proposed as separate species. Sequence divergence values strongly suggest multiple species within the A. boyeri complex.     

Phylogenetic relationships and demographic histories of the Atherinidae in the Eastern Atlantic and Mediterranean Sea re-examined by Bayesian inference

The aim of our study is to examine the phylogenetic relationship, divergence times and demographic history of the five close-related Mediterranean and North-eastern Atlantic species/forms of Atherina using the full Bayesian framework for species tree estimation recently implemented in ?BEAST. The inference is made possible by multilocus data using three mitochondrial genes (12S rRNA, 16S rRNA, control region) and one nuclear gene (rhodopsin) from multiple individuals per species available in GenBank. Bayesian phylogenetic analysis of the complete gene dataset produced a tree with strong support for the monophyly of each species, as well as high support for higher level nodes. An old origin of the Atherina group was suggested (19.2 MY), with deep split events within the Atherinidae predating the Messinian Salinity Crisis. Regional genetic substructuring was observed among populations of A. boyeri, with AMOVA and MultiDimensional Scaling suggesting the existence of five groupings (Atlantic/West Mediterranean, Adriatic, Greece, Black Sea and Tunis). The level of subdivision found might be consequence of the hydrographic isolation within the Mediterranean Sea. Bayesian inference of past demographic histories showed a clear signature of demographic expansion for the European coast populations of A. presbyter, possibly linked to post-glacial colonizations, but not for the Azores/Canary Islands, which is expected in isolated populations because of the impossibility of finding new habitats. Within the Mediterranean, signatures of recent demographic expansion were only found for the Adriatic population of A. boyeri, which could be associated with the relatively recent emergence of the Adriatic Sea.     

Isolation, characterization and cross-species testing of microsatellites obtained from a sand smelt (Atherina boyeri) genomic library

This study reports the isolation and characterization of 11 polymorphic microsatellites from a sand smelt (Atherina boyeri) genomic library. Enrichment was performed with di-, tri- and tetranucleotide motifs following the FIASCO procedure (fast isolation by AFLP of sequences containing repeats). All loci were found to be in linkage and in Hardy-Weinberg equilibrium. This represents the first microsatellite isolation for the family Atherinidae and the isolated loci were accordingly tested on four additional species of the family: two recognized (A. presbyter and A. hepsetus) and two proposed ('punctata' and 'non-punctata' forms). Moreover their cross-species suitability on Menidia menidia, belonging to the same order but to the family Atherinopsidae, was also tested.     

Phylogenetic relationships of Atherina hepsetus and Atherina boyeri (Pisces: Atherinidae) populations from Greece, based on mtDNA sequences

The genetic divergence and the phylogenetic relationships of six Atherina boyeri (freshwater and marine origin) and five Atherina hepsetus populations from Greece were investigated using partial sequence analysis of 12s rRNA, 16s rRNA and control region mtDNA segments. Three different well divergent groups were revealed; the first one includes A. boyeri populations living in the sea, the second includes A. boyeri populations living in the lakes and lagoons whereas the third one includes all A. hepsetus populations. Fifty-seven different haplotypes were detected among the populations studied. In all three mtDNA segments examined, sequence analysis revealed the existence of fixed haplotypic differences discriminating A. boyeri populations inhabiting the lagoon and the lakes from both the coastal A. boyeri and the A. hepsetus populations. The genetic divergence values estimated between coastal (marine) A. boyeri populations and those living in the lagoon and the lakes are of the same order of magnitude as those observed among coastal A. boyeri and A. hepsetus populations. The results obtained by different phylogenetic methods were identical. The deep sequence divergence with the fixed different haplotypes observed suggests the occurrence of a cryptic or sibling species within A. boyeri complex.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 92 , 151–161.     

A study of the population biology of Athevina boyeri Risso, 1810 in Aberthaw Lagoon, on the Bristol Channel, in South Wales

The biology of the sixth population of A. boyeri Risso, 1810 to be recorded outside of the Mediterranean basin is described. The 2 year life cycle was similar to that of other populations of the species that have been investigated. A maximum standard length of 93 mm was recorded for 2-year-old fish. Sexual maturity was attained by females at a length of 39 mm. A temporal lag in ovarian development was noted between the two spawning age groups, with the older age group spawning first. Details are provided of batch spawning in A. boyeriand the difficulties of assessing fecundity in batch spawners is discussed. Discrepancies between direct and indirect methods of assessing batch size indicated that the eggs around the largest mode may not represent the next batch to be spawned. Evidence is presented to suggest that in A. boyeri these eggs represent a ' store' from which batches of eggs are subsequently released.     

Life history strategies of four small-size fishes in the Suez Canal, Egypt

Of 35 species collected from the shores of the Suez Canal and its lakes, five were abundant year round. Sihouettea aegyptia and Aphanius dispar are of Red Sea (warm water) origin, whereas Pomatoschistus marmoratus, Aphanius fasciatus and Atherina boyeri are Mediterranean species. S. aegyptia and A. dispar are distributed throughout the Suez system whereas A. fasciatus is restricted to the northern canal. P. marmoratus and A. boyeri have not spread southward beyond the Bitter Lakes. Salinity is the main limiting factor for the distribution of Mediterranean species. S. aegyptia and A. dispar are dominant in the Bitter Lakes (salinity 44‰), whereas P. marmoratus and A. boyeri are abundant in Timsah Lake (salinity 7·8–41·6‰). There was considerable interannual and monthly variation in the relative abundance of each species. The populations were dominated by a single age group, and life spans were no more than 2 years. Rapid growth was evident during the first spring in P. marmoratus and A. boyeri and during the first summer, early autumn in S. aegyptia and A. dispar . The relative abundance of each food item in the diet varied with fish size and season. S. aegyptia, P. marmoratus and A. dispar fed mostly on harpacticoid copepods, polychaetes, demersal eggs, diatoms and blue-green algae whereas A. boyeri fed mostly on planktonic copepods. The warm water species are summer spawners, whereas the temperate species are autumn-winter spawners.     

Atherina punctata and Atherina lagunae (Pisces,Atherinidae), new species found in the Mediterranean sea. 2. Molecular investigations of three Atherinid species

On the basis of morphoanatomical parameters, the sand smelt species (Atherina boyeri Risso, 1810) is viewed as a highly polymorphic complex. In this study, intraspecific sequence variation in a portion of the cytochrome b gene was examined in 88 individuals from Tunisia and France. The correlation between the results of statistical analysis of the sequence data using a variety of tree-building algorithms and morphoanatomical analyses demonstrated the subdivision into three putative species: A. boyeri, which only includes non-punctuated fishes, A. punctata, which corresponds to punctuated fishes and A. lagunae, which corresponds to atherines living in lagoons.     

Stable carbon and nitrogen ratios in the sand smelt from a Mediterranean coastal area: feeding habits and effect of season and size

Enriched δ¹³C values (annual average – 11.2‰) suggest that in the study site (Stagnone di Marsala, Italy) the sand smelt Atherina boyeri exploited benthic prey throughout the year. In particular, Mysidacea and Isopoda seemed to dominate the diet. Stomach content data from the literature was in partial agreement with the present isotopic analysis suggesting that, while Mysidacea and Isopoda are exploited by A. boyeri , Copepoda Harpacticoida and Amphipoda are the dominant food items of the sand smelt diet. Several hypotheses are proposed to explain such partially different results from the two techniques. Both season and size influenced the δ¹³C and δ¹⁵N values of A. boyeri . Although throughout the sampling year, A. boyeri showed very enriched δ¹³C and δ¹⁵N values consistent with a diet based on benthic resources, it showed significant seasonal differences in both nitrogen and carbon isotope ratios. Such a finding suggests that the food preferences of A. boyeri undergo small seasonal differences, probably as a consequence of prey availability. Size also affected the isotopic composition of the sand-smelt indicating that this species shows ontogenetic diet shifts in the study site. Compared with other Mediterranean trophic data of the sand smelt these results substantiate the great adaptability of A. boyeri to the local environmental resources and its trophic plasticity.     

Enzymatic survey of four populations of Atherina boyeri based on electrophoresis and the occurrence of a microsporidiosis

Biochemical genetics of a Atherinid species, Atherina boyeri were undertaken to determine variability. Among the 17 loci studied, 7 were polymorphic, of which three were retained for a more detailed comparison of populations inhabiting the French Mediterranean coastal lagoons. A microsporidiosis is present in these fishes and is limited to certain lagoons. Comparison of allelic frequencies at the polymorphic loci between fish populations of infected and healthy lagoons yielded no significant differences. Several hypotheses are discussed and attempt to agree parasite distribution with genetical results.     

On the reproductive biology of the sand smelt Atherina boyeri Risso (Pisces: Atherinidae) and its evolutionary potential

The small, typically estuarine fish Atherina boyeri Risso forms local populations which can differ greatly in their population biology and morphology; this species is viewed as being on the brink of speciation. This analysis of the reproductive strategy shows that A. boyeri can rapidly adapt its life history and morphology to environments ranging from freshwater to polyhaline coastal lagoons and oceanic waters. The optimum strategy is shown to be tied to temperate seasonality: the cold winters act to lengthen the life cycle, and result in cyclic switching of energy between reproduction, somatic growth and fat storage. The adaptability of the fish to a range of environmental characters is viewed as being due to natural selection acting to produce a genotype capable of a flexible phenotypic response. This flexibility is essential for survival in the highly variable inshore/brackish habitat. It is argued that such preadaptive plasticity, coupled with the potential for populations to become isolated, produces ideal conditions for speciation. The importance of the coastal, estuarine and lagoonal environment as a springboard for teleost evolution is discussed.     

Feeding ecology of indigenous and non-indigenous fish species within the family Sphyraenidae

The feeding ecology of two common indigenous (Sphyraena viridensis and Sphyraena sphyraena) and one abundant non-indigenous sphyraenid species, Sphyraena chrysotaenia, of Indo-Pacific Ocean origin, was investigated in an area of the eastern Mediterranean Sea. The stomach contents of 738 individuals of varying size, collected during the period December 2008 to August 2009, were examined. The dietary analyses revealed that all three species were specialized piscivores with a diet consisting of >90% fish, both by number and mass. Concurrent sampling of the fish assemblage made it possible to calculate selectivity as well as diet breadth and overlap of these strict piscivores. Even though several prey species were found in the stomachs of the three predators examined, selectivity towards Atherina boyeri was highly significant. For all species examined, >70% of the diet by mass was made up by three indigenous species of commercial value: Spicara smaris, Boops boops and A. boyeri. Diet breadth and size of prey increased with increasing body size for all predators. With increased body size, the diet overlap between indigenous and non-indigenous species decreased. This could be attributed to increased diet breadth and the specific life-history characteristics of indigenous species developing into larger individuals. During winter, the condition factor of the non-indigenous species was significantly lower than that of the indigenous, indicating that winter conditions in the Mediterranean Sea may limit its further expansion north and westward. With this study, the gap in knowledge of the feeding preferences of the most abundant piscivorous species found in coastal areas of the study region is filled. Additionally, the results indicate that non-indigenous species familial affiliation to indigenous ones does not facilitate invasion success.     

Genetic Divergence Among Marine and Lagoon Atherina boyeri Populations in Greece Using mtDNA Analysis

Genetic differentiation and phylogenetic relationships among 15 Atherina boyeri populations from several marine and lagoon or lake sites in Greece were investigated using mtDNA analysis. PCR-RFLP analysis of 12s, 16s rRNA genes and D-loop revealed 23 haplotypes. All the lake or lagoon populations, as well as the Kymi and Kalymnos populations that originated from sites with lagoonlike environmental conditions, showed haplotypes 1-6, clearly distinguishable from the marine populations, which exhibited types 7-23. The genetic divergence values estimated between the lagoon and the marine populations ranged from 5.55 to 10.45%. The high genetic differentiation observed between these two types of populations is also highlighted by the dendrograms obtained using UPGMA and maximum parsimony methods.     

Mitochondrial DNA diversity in Atherina boyeri populations as determined by RFLP analysis of three mtDNA segments

The genetic differentiation and the phylogenetic relationships of eight Atherina boyeri Greek populations have been investigated at the mtDNA level. The populations studied are from two different lakes, a lagoon, the interface zone between the lagoon and the sea, and four marine sites. RFLP analysis of three mtDNA segments (12s rRNA, 16s rRNA and D-loop) amplified by PCR was used. Six, seven and eight restriction enzymes were found to have at least one recognition site at 12s rRNA, 16s rRNA and D-loop respectively. Twenty-one different haplotypes were detected among the populations studied. Several restriction patterns were revealed. These patterns can be used for the discrimination of the populations living in the sea ('marine' type populations) from the others inhabiting the lagoon and the lakes ('lagoon' type populations). The estimated net nucleotide sequence divergence between the populations examined ranged from 0 to 10.385%, while the Nst value of 0.92 indicates the existence of high interpopulation genetic differentiation. This high degree of differentiation detected between the 'lagoon' and 'marine' type populations makes the classification of these two types of populations as a single taxon questionable.     

Diplostomiasis in sand smelt, Atherina presbyter Cuvier, from the Fleet, Dorset and its use as a population indicator

The population of sand smelt, Atherina presbyter , breeding in the Fleet, Dorset, shows a high infection of diplostomiasis. The population was studied in 1983 to clarify aspects for this parasitic condition previously analysed for the sand smelt population at Fawley, Southampton Water. All age classes showed a higher percentage infection and mean number of metacercaria per fish than at Fawley. Analysis of postlarvae and juveniles showed that infection can occur at 1 week old, and verified the hypothesis that the scales of older fish inhibit cercarial settlement. Circumstantial evidence suggests that Potamopyrgus jenkinsi may be the first vector host for this parasite, and the densities of this species and of nesting little tern colonies would account for the differential infection between these two sand smelt populations. The increase in infection of 2 fish at Fawley cannot be attributed to mixing with the Fleet population, and the different infection levels demonstrate population isolation. High levels of infection are limited to this part of the English Channel: sand smelt samples from around the coasts of the British Isles show minimal infection rates elsewhere and suggest a southerly distribution of the parasite, away from Atlantic oceanic waters.     

Environmental factors controlling the spring immigration of two estuarine fishes Atherina boyeri and Pomatoschistus spp. into a Mediterranean lagoon

Atherina boyeri , a pelagic species, mainly entered a Mediterranean lagoon in the Rhone River Delta, during the day, whatever the flow direction. Flow velocity did not appear to be a significant factor and its swimming abilities seemed to be sufficient to cope with the observed velocity. The combined responses of Pomatoschistus spp., benthic species, to the different environmental factors clearly demonstrated a change in behavioural response during their ontogenic development, i.e. a shift from passive to active or active-passive migration. Young Pomatoschistus spp. (<20 mm L _T) exhibited a behaviour that correspond to the classical selective tidal stream transport of young fishes towards estuarine areas (passive drifting). Large Pomatoschistus spp. were also able to enter the lagoon with the flow, but they remained negatively affected by the flow velocity. The diel cycle appeared to be the main factor controlling the immigration of A. boyeri into the lagoon whereas Pomatoschistus spp. seemed more sensitive to hydrographic conditions prevailing at the connection.     

光肩星天牛种群间及其近缘种遗传关系的RAPD研究

利用RAPD技术对采自中国和美国的星天牛属Anoplophora 5个种及8个光肩星天牛Anoplophora glabripennis (Motschulsky)地理种群共13个样品进行了遗传相似性分析。选用了Operon公司生产的引物H系列20个,L系列20个,Q系列11个共51个引物,最后从40个引物中筛选出26个具有多态性的引物作为第一组用于星天牛属种间和光肩星天牛种群间分析,从31个引物中筛选出19个具有多态性的引物作为第二组单独用于光肩星天牛种群分析。根据第一组引物实验获得的RAPD聚类图及遗传距离表明,各个地理种群的光肩星天牛和黄斑星天牛A. nobilis都聚在一起,形成一个大的分枝,而四川星天牛A. freyi、楝星天牛A. horsfieldi和星天牛A. chinensis均在此分枝之外。来自美国纽约和芝加哥的光肩星天牛种群聚于中国光肩星天牛种群之外的另一个独立的分枝上。分布在我国宁夏、内蒙古和河北的光肩星天牛以及宁夏黄斑星天牛和山东、陕西的光肩星天牛分别聚在一起,而甘肃的光肩星天牛与甘肃的黄斑星天牛则聚于另一枝,且它们之间的遗传距离很近,仅为0.1324,说明这两者之间有着极其相近的亲缘关系,由此推断光肩星天牛和黄斑星天牛的差异很小,遗传关系难以区分,进一步证实了它们很可能是同一个种下的两个不同的型。第二组引物实验得到了相似的结果,来自中国的6个光肩星天牛种群全部聚于同一枝中并分成两小枝： 分布于我国宁夏、河北、山东、甘肃的光肩星天牛聚在一起,内蒙古和陕西的光肩星天牛则成另一枝,而分布于美国纽约和芝加哥的光肩星天牛仍聚于中国光肩星天牛种群之外的一个单独的分枝上。但是美国光肩星天牛与中国光肩星天牛之间的遗传距离最近的为0.4578, 最远的为0.5960。由此认为,本研究中采自美国的两个光肩星天牛种群的样本和采自中国的光肩星天牛种群的样本之间存在显著差异,遗传关系较远。有必要从中国和世界其他天牛分布地采集更多样本做进一步DNA 分析。     

Biology of the sand-smelt (Atherina presbyter Valenciennes) around Fawley power station

Past studies of fish impingement at Fawley power station have shown that sand-smelts, Atherina presbyter Valenciennes, are susceptible to impingement and may be a useful species for examining effects of impingement mortalities on population size and structure. A 21 month study of populations in the vicinity of Fawley power station was carried out to obtain data on the biology of the species for future population dynamics modelling studies. Aspects included in the study were growth, sex ratio, reproduction, survival and mortality rates and diet. Differences in apparent and true rates of growth indicated some size-selective mortality towards younger age groups, possibly caused by the power station. A seasonallyoscillating von Bertalanffy growth equation is given to predict mean length-at-age. Most female sand-smelts (73%) matured at the end of their first year of life, fecundities ranging on average from 1394 eggs per female for 1+ group fish to 6872 for III+ fish. The sex ratio was even (1 : 1). An annual survival rate of 10–8% was estimated for mature fish, and the consequent age structure of the population indicates that the bulk of egg production lies with the smaller (1+ group) fish. Since smaller fish are generally more vulnerable to impingement this factor may render populations sensitive to impingement mortalities. Gut content analysis indicated that sand-smelts probably actively select zooplankters from the general plankton when young and, once larger than 104 mm in length, increasingly feed upon natant macrofaunal species.     

Diagnostic features and taxonomy of the Mediterranean species of the big-scale sand smelts belonging to <Emphasis Type="Italic">Atherina boyeri</Emphasis> group (Atherinidae)

Morphological studies of the big-scale sand smelt from the group Atherina boyeri sensu lato were carried out using the materials of the museum collections from the Mediterranean Sea, the Black Sea, and the Sea of Azov. It has been shown that low-raker sand smelt belonging to the species A. boyeri sensu stricto, and multi-raker sand smelt, referred to the A. mochon species, are differentiated by a complex of morphological features: the number of gill rakers, the shape of scales, the length of the ascending process of praemaxillare, and the shape of the lower margin of maxillare. The key for their identification is presented. It is assumed that A. mochon is not a native Black Sea species but belongs to the invasive species originating from the Mediterranean Sea. The isolation of two cryptic species is discussed. The neotype of A. pontica is designated.