Genetic relationships among the shads (Alosa) revealed by mitochondrial DNA analysis

We surveyed restriction site differences in mitochondrial DNA. (mtDNA) among five species of shad ( Alosa ) from North America and Europe. Allis shad, Alosa alosa and twaite shad, Alosa fallax shared two divergent genotype groups, suggesting that the two forms are either a single species, or are distinct species that have hybridized. Phenetic and cladistic analyses of the relationships among the mitochondrial genotypes defined two groups of shad, corresponding to the subgenera, Alosa and Pomolobus . The mean estimated sequence divergence between the mtDNAs of these two groups of shad was 6.5%. Taken in conjunction with fossil data, this divergence estimate suggests that the rate of mtDNA divergence between the two subgenera has been almost 10-fold lower than the 'conventional' clock calibration for mtDNA.     

Isolation and characterization of eight dinucleotide microsatellite loci from two closely related clupeid species (Alosa alosa and A. fallax)

Eight dinucleotide microsatellite loci were developed through an enrichment protocol for allis shad (Alosa alosa) and twaite shad (A. fallax). Cross‐species amplification was successful for all loci isolated. The number of alleles per locus ranged from three to nine for A. alosa and from two to seven for A. fallax, while the observed heterozygosity ranged from 0.267 to 0.926 and from 0.240 to 0.727, respectively. These markers will constitute useful tools for studies of population structure and gene flow between two closely related hybridizing species.     

Morphometric, meristic character and electrophoretic analyses of two Irish populations of twaite shad, Alosa fallax (Lacépède)

Since 1911 a rare lacustrine form of twaite shad, Alosa fallax killarnensis Regan (1916), has been known to occur in the Killarney Lakes. During the summers of 1985 and 1986 a gill netting programme in Lough Leane for brown trout, Salmo trutta L., yielded 32 shad (1985) and 64 shad (1986) as a by-catch. These fish were deep frozen and compared with a sample of 59 marine twaite shad, taken on rod and line from St Mullin's, Co. Carlow, which had entered the River Barrow to spawn.
Morphometric, meristic and electrophoretic character analyses were carried out on the two populations. The three analyses, particularly the isoelectric focusing, confirmed a high degree of genetic similarity. The two major differences found were the dwarfed size of the Killarney shad and the increased numbers of gill rakers carried on the first branchial arch. The merits and demerits of assigning the term subspecies to the Killarney shad are discussed in the light of the available evidence produced.     

Development of hypoosmoregulatory ability in allis shad Alosa alosa

When different ages of juvenile allis shad Alosa alosa (18–74 days post-fertilization) were exposed for 48 h to elevated salinity (25 or 30), all ages could tolerate moderates salinity (25) whereas only the older stages could tolerate the higher salinity. In 88 days post-fertilization allis shad, acclimatation to full-strength sea water for 2 weeks did not lead to significant changes in total water content and gill Na⁺/K⁺-ATPase activity whereas total Na⁺ content increased within 5 days to values which remained stable thereafter. Together, these data suggest that juvenile allis shad develop a limited euryhalinity, which allows them to fully adapt to hyperosmotic environment immediately after transfer.     

Photoresponse in allis shad larvae

Allis shad Alosa alosa photoresponse was investigated in 3 day-old to 1 month-old larvae. Regardless of age, larvae strongly preferred the lit compartment in aquaria experiments. Such response was consistent with other experimental studies which have suggested a pelagic behaviour of allis shad larvae from the time of hatch.     

Growth and movements of the twaite shad, Alosa fallax (Lacépède), in the Severn Estuary

Between July 1974 and April 1977, appreciable numbers of 0 group twaite shad, Alosa fallax , have been collected from the cooling water intake screens of the Nuclear Power Stations at Oldbury-upon-Severn and Berkeley on the Severn Estuary and at Hinkley Point on the Bristol Channel, England. Young of the year first appeared at Oldbury in July at a size of approximately 32 mm standard length, and grew to approximately 61 mm by October. The downstream migration in the late summer and autumn appeared to be closely correlated with water temperature since movement reached a peak soon after the temperature in the Estuary had declined below 19° C, and ceased altogether below 9° C. Thereafter, there was virtually no movement of shad until temperatures rose sharply above 7° C in the spring. Maturing adult twaite shad were captured in late April and May as they moved into freshwater to spawn, probably in mid-June. Males were recorded slightly earlier during the migration and were generally younger than females. Furthermore, mean lengths and weights for individuals belonging to the different age classes indicated that males were also smaller. Logarithmic relationships between length and weight of young and adult A. fallax and for the variation with size in the number of rakers on the first gill arch are presented.     

Genetic polymorphism of adenosine deaminase (ADA; E.C. 3.5.4.4.) in allis shad, Alosa alosa and twaite shad, Alosa fallax

A genetic polymorphism of adenosine deaminase (ADA), is described for the first time in European shad species, Alosa alosa and Alosa fallax . Significant heterogeneity in gene frequencies was found, both between species and within species.     

Genetic data on endangered twaite shad (Clupeidae) assessed in landlocked and anadromous populations: one or more species?

Seven Italian populations of twaite shad Alosa fallax from Northern and Central Italy were investigated to assess genetic diversity by Cytochrome b (Cytb) gene sequencing. The two ecotypes historically referred to landlocked and anadromous populations were investigated for the first time from a genetic point of view, to clarify their phylogenetic relationships. Moreover, results obtained from populations coming from separated Adriatic and Tyrrhenian basins were compared with data assessed in samples of allis shad Alosa alosa from the Atlantic basin. All the Italian samples were recognized at species level as A. fallax, differing for five mutations from A. alosa. The analyses confirmed the occurrence of a single phylogenetic lineage and of a single species within Italian waters, in both landlocked and migratory populations. The minimum spanning network identified six haplotypes for A. fallax and two haplotypes for A. alosa. The neighbour-joining tree and the maximum likelihood on the Cytb gene sequences confirmed two distinct lineages for A. alosa and A. fallax, without evidence of a separation at specific level within the A. fallax group. A weak separation due to incipient population differentiation was detected between anadromous and landlocked Italian populations, supporting the idea of a recent separation. The molecular data herein collected do not support the existence of the already controversial incipient species Alosa agone. Despite this, the two ecotypes could be considered as different management units from a conservation viewpoint.     

A review of the past and present status of anadromous fish species in the Netherlands: is restocking the Rhine feasible?

The paper reviews the past, present and future of eight anadromous fish species inhabiting the Lower Rhine (The Netherlands), viz. -sturgeon (Acipenser sturio), whitefish and houting (Coregonus lavaretus, C. oxyrinchus), smelt (Osmerus eperlanus), allis and twaite shad (Alosa alosa, A. fallax), sea trout (Salmo trutta) and salmon (Salmo salar). All species are under threat or became extinct (e.g. sturgeon, allis shad). It is not possible to single out a specific factor for the decline or disappearance. A combination of factors is responsible, as the degradation of the spawning and nursery areas, river correction for shipping, building of sluices and hydropower dams, extraction of sand and gravel and river pollution. The likelihood that a species will return via natural recovery, or restocking is assessed. The return of the sturgeon is unlikely. Present observations of sturgeon can be attributed to releases of unwanted sturgeon hybrids. A natural stock of coregonids in Dutch waters seems not feasible any more due to irreversible habitat degradation. Present day catches originate from German releases. The anadromous smelt, heavily reduced in numbers, still inhabits some of our waters, the non-migratory smelt is still very common. The allis shad is extinct and unlikely to recover. The species never spawned in the Dutch part of the Rhine. Twaite shad, declining in numbers, are still observed in the lower reaches of Rhine and Meuse. Sea trout is presumably still present in the same varying numbers as before. Spawning in our waters has not been documented. The salmon, once fished in large numbers, is now the subject of restocking programmes in Germany. Observations of individuals can partly be attributed to these programmes but also to straying salmon. Restocking programmes should be considerably improved before noticeable success is to be met.     

A highly permeable species boundary between two anadromous fishes

Meristic identification, mitochondrial DNA and a suite of microsatellite markers were employed to estimate the incidence of hybridization in wild populations of anadromous Allis shad Alosa alosa and twaite shad Alosa fallax in southern Irish riverine and estuarine waters. It was shown that 16% of the fishes examined were misclassified using meristic count of gill rakers. Next, a significant proportion of fishes that were robustly assigned to a species using nuclear markers were shown to possess the mtDNA of the other. The genomes of A. alosa and A. fallax in Ireland are extensively introgressed, which suggests a complex history of hybridization between these species, which can only partially be explained by recent man-made habitat changes.     

The presence of anadromous twaite shad (Alosa fallax) in the Ebro River (western Mediterranean,Spain): an indicator of the population's recovery?

Although the Ebro River is one of the largest Mediterranean rivers, the presence of anadromous fish species was near to complete extirpation since the second half of the 20th century. However, many mature twaite shads (Alosa fallax) were captured at the lower reaches of this river (Xerta's weir site, 63 river km) in spring 2005, suggesting a natural recovery of this population. In this study, the first biological data on twaite shad from the Ebro River are presented, and their significance for the species conservation and recovery are discussed. The historical reasons for the decline of this species are reviewed, and data supports the proposal for urgent construction of a proper fish passage for anadromous species at the Xerta's weir. This action would allow fish access to a further 60 km of river upstream of the weir, representing a 200% increase in the amount of freshwater habitat and potential spawning areas. Improved access together with restrictive fishery regulations will assist with renewed interest in the fishery and conservation of the species.     

Primary and secondary oocyte growth dynamics in anadromous semelparous Allis shad Alosa alosa

We analysed the ovarian dynamics of the anadromous semelparous allis shad Alosa alosa for which our working hypothesis was that mature pre-spawning females would have very low or even exhausted primary growth (PG) oocyte reserves; semelparity has been linked with the depletion of the pool of PG oocytes. To test this hypothesis, the PG oocytes were enumerated, their recruitment pattern to the secondary growth (SG) phase was analysed and their potential replenishment from the pool of oogonia was examined in females caught very close to the Mondego River mouth, in central Portugal and along the river. The development of the SG oocytes was also analysed, the fecundity (batch, total and annual) values were estimated and the intensity of atresia was quantified. Ovarian samples and histological sections were investigated in parallel. A dynamic recruitment pattern of PG oocytes to the SG phase was revealed, where all PG oocytes were recruited and were not replenished by oogonia. Annual fecundity was subject to down-regulation due to atresia prior to spawning and its size was multiple times higher than the size of batch fecundity. Lack of population synchronicity in ovarian development and spawning migration was also observed. This multifaceted analysis of the ovarian dynamics of this species will contribute to management efforts for this critically endangered and economically important fish throughout its geographical distribution. The results reported in this study will also assist in unravelling the complexity of the early processes of oogenesis in fish.     

The biology of the twaite shad, Alosa fallax fallax (Lacépède), in the Severn Estuary

The estuarine biology of the twaite shad was studied in the Severn Estuary. Adults enter the estuary at the start of the freshwater phase of their spawning migration between April and June. Peak immigration generally occurs in May and is associated with temperatures in the range 10.6–12.3°C. The mean (± s.d. ) instantaneous mortality rate for the mature population was 0.53±0.18. The effect of additional mortality on the spawning population was modelled assuming constant recruitment and no density-dependent effects.
Juvenile twaite shad are present in the estuary from July until they emigrate seaward during the autumn. A portion of these fish re-enter the estuary the following April–May and remain until late summer/early autumn before once more migrating seaward.
The 0 + age group feed mainly on harpacticoid and calanoid copepods and mysids, the relative preponderance of these in the diet being apparently related to tidal conditions. The possible implications of the proposed tidal power barrage in the Severn Estuary on the twaite shad population are discussed in relation to movement, diet and additional mortality of the mature population.     

Genetic structure of the spawning population of Danube shad, Alosa pontica, Eichwaldt 1838 (Clupeiformes, Alosiinae)

The genetic structure of the population of Danube shad, Alosa pontica, has been studied by means of analysis of 3 polymorphic biochemical gene loci. The results of the study provide evidence of its genetic heterogeneity which is expressed by: firstly, an unbalanced ratio of genotype loci manifested by an excess of rare homozygotes and deficite of the correspoding heterozygotes, and secondly, differences in the allele frequencies between shad populations arriving for spawning in March-April and May-June. It is suggested that there could be two causes of heterogeneity: one due to the introgressive hybridization between various forms of A. pontica and another to the differences between early and late spring races of shad which to a certain extent are reproductively autonomous.     

Spatial Analysis of Twaite Shad,Alosa fallax (Lacepède, 1803), in the Southern North Sea: Application of Non‐Linear Geostatistics as a Tool to Search for Special Areas of Conservation

This study aims to evaluate the suitability of non‐linear geostatistics and indicator kriging (IK) as a tool in environmental impact assessment and nature conservation, in particular to search for potential Special Areas of Conservation (SAC) for the endangered fish species twaite shad, Alosa fallax (Lacepède , 1803) within the German Exclusive Economical Zone (EEZ) of the North Sea. To analyse the spatial distribution of this fish species, data on standardised biomass index (catch per unit effort, c.p.u.e., kg × 30 min^–1) from 1996 to 2001 were used, regarding the third and fourth quarters of each year, respectively. Thereby we assume that the spatial distribution can be described as a time‐invariant process. This assumption is supported by information on annual sampling effort, allocation of hauls and spatial distribution of the positive catches. All indicator variograms obtained for different c.p.u.e. cut‐off values displayed distinct spatial structures, clearly indicating that the indicator variables were spatially autocorrelated. Gaussian models were fitted by least‐squares methods and were evaluated with a goodness‐of‐fit statistic. Subsequently, IK was employed to estimate the probability of exceeding the c.p.u.e. cut‐off values for the twaite shad in the investigation area. These were highest in the Weser‐ and Elbe‐estuary, probably because of migrations of twaite shad to and from estuaries at the time of investigation due to spawning, while within the German EEZ of the North Sea no such areas with increased probabilities could be discerned. Thus, although available data did not allow to identify and implement any SAC in the German EEZ, the methods employed here can be regarded as a promising management tool in biological conservation issues. (© 2004 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

The occurrence of Eubothrium fragile (Cestoda: Pseudophyllidae) in twaite shad, Alosa fallax (Lacépède) in the River Severn

The occurrence, size and maturity changes of Eubothrium fragile have been studied in postlarvae, juveniles and adult twaìte shad, Alosa fallax , from several locations in the River Severn. Parasites were only found in adult shad and not in post-larvae or juveniles. No juvenile or recently acquired cestodes were identified as such, but adults were present in shad throughout the whole period of their spawning migration. A large proportion of the parasites were gravid upon arrival in the river and, although eggs were subsequently released into fresh water, there was no loss of cestodes from the fish. It was concluded that E. fragile is a marine species, that the parasites found in adult shad in fresh water were the residue of a marine life cycle and that the eggs released in fresh water were part of the parasite's natural reproductive wastage. The distribution and biology of E. fragile were discussed and it is considered that it is typical of the marine species of the genus.     

Length–weight and length–length relationships for four shad species along the western Black Sea coast of Turkey

This study investigates length–weight and length–length relationships of four shad species (Alosa caspia caspia, Alosa immaculata, Alosa maeotica, Alosa tanaica) captured in the western Black Sea coast of Turkey. A total of 686 specimens, 355 females and 331 males, were caught by gill net, trammel net and longline from 2006 through 2007 along the western Black Sea coast of Turkey. The relationships of total length (TL), fork length (FL) and standard length (SL) are presented for inhabiting in the Black Sea four shad species and the relationships between total length (TL) and body weight (BW) from the two localities, Sile and Karasu in the western Black Sea coast of Turkey. The Length–weight relationships generally indicated positive allometric growth in Alosa species for both males and females and an isometric growth for A. immaculata females. The coefficient b ranged from a minimum 2.97 for females of A. immaculata to a maximum 3.75 for both sexes A. c. caspia. Results indicated that the length–length relationship between the three length measurements were highly correlated (r² > 0.99, P < 0.001).     

Length and restriction site heteroplasmy in the mitochondrial DNA of american shad (alosa sapidissima)

Restriction endonuclease analysis was used to assess mitochondrial DNA (mtDNA) variation in American shad (Alosa sapidissima) collected from 14 rivers ranging from Florida to Quebec. Two types of heteroplasmy were observed, one involving a major length polymorphism and the other a single restriction site. Shad mtDNA occurred in two principal size classes, 18.3 and 19.8 kb. Of 244 shad examined, 30 were heteroplasmic and carried both size classes of mtDNA in varying proportions; the remainder were homoplasmic for the smaller size class of mtDNA. The large mtDNA variant occurred most frequently at the southern end of the range, and except for two individuals from Nova Scotia, was not detected among shad from rivers north of the Delaware. In contrast, ten shad heteroplasmic for a SalI restriction site originated from rivers ranging from South Carolina to Nova Scotia. DNA mapping and hybridization experiments indicated that the length polymorphism is in the D-loop-containing region and consists of a tandemly repeated 1.5-kb DNA sequence occurring in two and three copies, respectively, in the two major size classes of shad mtDNA. Continuous length variation up to approximately 40 bp occurs among copies of the repeat both within and among individuals. Restriction site data support the conclusion that both forms of heteroplasmy in shad mtDNA have originated more than once.     

Genetic monitoring of two decades of hybridization between allis shad (Alosa alosa) and twaite shad (Alosa fallax)

Habitat alteration has been implicated in driving hybridization between the sympatric migratory shads Alosa alosa and Alosa fallax. Morphological and molecular evidence is consistent with hybridization across the overlapping range of these species, but the temporal extent of hybrid occurrence and genetic consequences for populations have not been explored. Using eight nuclear microsatellite loci and samples collected between 1989 and 2008 in the Solway Firth (UK), we genetically identified hybrids, studied temporal changes in their frequency, and explored changes in allele frequencies of parental populations. These molecular data confirmed the hybrid status of individuals identified using morphology (number of rakers on the outer gill arch), and enabled separation of hybrids from purebred individuals. Mitochondrial cytochrome-b sequencing revealed the presence of two haplogroups, each predominantly occurring in one species. Heterospecific haplotypes were found in 22.3 and 12.8% of A. alosa and A. fallax individuals, respectively, consistent with backcrossing and suggesting that hybrids are fertile. On average, microsatellite-identified hybrids comprised 12.7% of all samples, but when individuals with cytonuclear discordance were also considered introgressed on average 25.4% of individuals were of hybrid ancestry. Overall, allelic richness remained largely unchanged within species, but there were declines in the inbreeding coefficient (F _IS) of both species and episodes of significant temporal allelic frequency change. Hybrids sampled between 2004 and 2008 showed no evidence of lower fecundity relative to purebred individuals. Together, results suggest that hybridization between shad species in northern Europe is prevalent, and has been ongoing over at least two decades. The challenge is now to understand the extent to which observed patterns are linked to immigration from other populations, and the mechanisms that have prevented species collapse despite apparent hybrid fertility and longstanding introgression of neutral markers.     

The Combined Use of Correlative and Mechanistic Species Distribution Models Benefits Low Conservation Status Species

Species can respond to climate change by tracking appropriate environmental conditions in space, resulting in a range shift. Species Distribution Models (SDMs) can help forecast such range shift responses. For few species, both correlative and mechanistic SDMs were built, but allis shad (Alosa alosa), an endangered anadromous fish species, is one of them. The main purpose of this study was to provide a framework for joint analyses of correlative and mechanistic SDMs projections in order to strengthen conservation measures for species of conservation concern. Guidelines for joint representation and subsequent interpretation of models outputs were defined and applied. The present joint analysis was based on the novel mechanistic model GR3D (Global Repositioning Dynamics of Diadromous fish Distribution) which was parameterized on allis shad and then used to predict its future distribution along the European Atlantic coast under different climate change scenarios (RCP 4.5 and RCP 8.5). We then used a correlative SDM for this species to forecast its distribution across the same geographic area and under the same climate change scenarios. First, projections from correlative and mechanistic models provided congruent trends in probability of habitat suitability and population dynamics. This agreement was preferentially interpreted as referring to the species vulnerability to climate change. Climate change could not be accordingly listed as a major threat for allis shad. The congruence in predicted range limits between SDMs projections was the next point of interest. The difference, when noticed, required to deepen our understanding of the niche modelled by each approach. In this respect, the relative position of the northern range limit between the two methods strongly suggested here that a key biological process related to intraspecific variability was potentially lacking in the mechanistic SDM. Based on our knowledge, we hypothesized that local adaptations to cold temperatures deserved more attention in terms of modelling, but further in conservation planning as well.