First global approach: morphological and biological variability in a genetically homogeneous population of the European pilchard, Sardina pilchardus (Walbaum, 1792) in the North Atlantic coast

The European pilchard Sardina pilchardus represents the most commercially relevant fisheries resource in many countries bordering north Atlantic coasts and the Mediterranean Sea, being especially significant along the coast of Morocco. The continuous exploitation of this pelagic species for several decades places Morocco as the leader in sardine production. However, the conditions of exploitation of this resource underwent a great change during the last recent years. In order to identify the populations of the European pilchard sardine (Sardina pilchardus, Walbaum, 1792) in the Atlantic coast of Morocco and Spain, we have combined the truss network data to conduct multivariate analysis with biologic parameters and genetic analysis based on Microsatellite and mitochondrial control region data. Sardine morphometrics data truss variables from 10 samples spanning the Atlantic coast of Morocco were analysed by multivariate analysis. Thirteen morphometric measurements and some biological parameters such as the sex and the age of fishes were made for each individual. Discriminant analysis on size-corrected truss variables and cluster analysis of mean fishes shape using landmark data indicate, that the shape of north Moroccan sardines is distinct from the shape of sardines from south Morocco. However the analysis of the mitochondrial region and four microsatellites loci (Sp2, Sp7, Sp8 and SpI5) demonstrated an homogeneity population in the Moroccan Atlantic coast, with a low but significant genetic differentiation, which followed an isolation-by-distance pattern according to Mantel test.     

Behavioural responses of sardines Sardina pilchardus to simulated purse‐seine capture and slipping

The behavioural effects of confinement of sardine Sardina pilchardus in a purse seine were evaluated through three laboratory experiments simulating the final stages of purse seining; the process of slipping (deliberately allowing fishes to escape) and subsequent exposure to potential predators. Effects of holding time (the time S. pilchardus were held or entangled in the simulation apparatus) and S. pilchardus density were investigated. Experiment 1 compared the effect of a mild fishing stressor (20 min in the net and low S. pilchardus density) with a control (fishing not simulated) while the second and third experiments compared the mild stressor with a severe stressor (40 min in the net and high S. pilchardus density). In all cases, sea bass Dicentrarchus labrax were used as potential predators. Results indicated a significant effect of crowding time and density on the survival and behaviour of slipped S. pilchardus. After simulated fishing, S. pilchardus showed significant behavioural changes including lower swimming speed, closer approaches to predators and higher nearest‐neighbour distances (wider school area) than controls, regardless of stressor severity. These results suggest that, in addition to the delayed and unobserved mortality caused by factors related to fishing operations, slipped pelagic fishes can suffer behavioural impairments that may increase vulnerability to predation. Possible sub‐lethal effects of behavioural impairment on fitness are discussed, with suggestions on how stock assessment might be modified to account for both unobserved mortality and sub‐lethal effects, and possible approaches to provide better estimates of unobserved mortality in the field are provided.     

Comparative phylogeography of two co‐distributed but ecologically distinct rainbowfishes of far‐northern Australia

Aim

To test the influence of historical and contemporary environment in shaping the genetic diversity of freshwater fauna we contrast genetic structure in two co‐distributed, but ecologically distinct, rainbowfish; a habitat generalist (Melanotaenia splendida) and a habitat specialist (M. trifasciata).

Location

Fishes were sampled from far northern Australia (Queensland and Northern Territory).

Methods

We used sequence data from one mitochondrial gene and one nuclear gene to investigate patterns of genetic diversity in M. splendida and M. trifasciata to determine how differences in habitat preference and historical changes in drainage boundaries affected patterns of connectivity.

Results

Melanotaenia splendida showed high levels of genetic diversity and little population structure across its range. In contrast, M. trifasciata showed high levels of population structure. Whereas phylogeographic patterns differed, both species showed a strong relationship between geographical distance and genetic differentiation between populations. Melanotaenia splendida showed a shallower relationship with geographical distance, and genetic differentiation was best explained by stream length and a lower scaled ocean distance (11.98 times coast length). For M. trifasciata, genetic differentiation was best explained by overwater distance between catchments and ocean distance scaled at 1.16 × 10⁶ times coast length.

Main conclusions

Connectivity of freshwater populations inhabiting regions periodically interconnected during glacial periods appears to have been affected by ecological differences between species. Species‐specific differences are epitomized here by the contrast between co‐distributed congeners with different habitat requirements: for the habitat generalist, M. splendida, there was evidence for greater historical genetic connectivity with oceans as a weaker barrier to gene exchange in contrast with the habitat specialist, M. trifasciata.     

Chaotic genetic patchiness in the pelagic teleost fish Sardina pilchardus across the Siculo-Tunisian Strait

ABSTRACT

This investigation aims at assessing patterns of spatial genetic structure of the teleost fish Sardina pilchardus across the Siculo-Tunisian Strait (a well-known discontinuous biogeographic area) and delineating putative genetic stocks within the species. For this purpose, a total of 180 specimens, collected from 11 locations stretching across the western and eastern Mediterranean coasts of Tunisia, were analysed genetically by means of 18 nuclear allozyme loci. The outcome of this study revealed strong genetic differentiation among populations, with the marked genetic distinctiveness of the central Tunisian population at Mahdia. Despite the delineation of seven well-defined genetic groups, no significant correlation was found between genetic and geographic distances. Besides, the recorded population subdivision did not align with biogeographic boundaries, suggesting the presence of chaotic genetic patchiness. Recent genetic bottlenecks were evidenced in S. pilchardus populations. Patchy migration patterns were recorded among the examined pairs of sardine populations. Among the recorded 16 polymorphic loci, GPI-2 and SOD appeared to be subject to natural selection. Patterns of population genetic differentiation and structuring were not found to be driven by outlier loci that appeared to be under selection. Furthermore, the detected neutral GPI-1 locus was found to be responsible for most of the genetic variation among identified genetic clusters. Hence, natural selection cannot cause the detected genetic heterogeneity among sardine samples. Different explanations to the origin of chaotic genetic patterns, observed within S. pilchardus, were discussed.     

Isolation and characterization of polymorphic microsatellites for the sardine Sardina pilchardus (Clupeiformes: Clupeidae)

The sardine Sardina pilchardus Walbaum, 1792 is a small pelagic fish species that sustains an economically important fishery in the eastern Atlantic Ocean. Eight perfect microsatellite loci of sardine were characterized from a partial genomic library enriched for CA repeat sequences. All loci revealed high levels of polymorphism, with the number of alleles per locus ranging between 22 and 45, and an average observed heterozygosity of 0.772. The newly described loci can be employed to determine population genetic structure of the sardine, and may aid in fishery management of this species.     

Mitochondrial DNA Sequence Variation Suggests the Lack of Genetic Heterogeneity in the Adriatic and Ionian Stocks of Sardina pilchardus

A genetic stock structure analysis of 11 sardine samples from the Adriatic Sea and Ionian neighboring area was carried out through sequence variation analysis of a 307-bp cytochrome b gene fragment in order to identify self-recruiting units in the Adriatic Sardina pilchardus stock. The overall lack of genetic subdivision among samples detected by analysis of molecular variance, pairwise Φ_st values, and the exact test of population differentiation indicates this sardine stock is part of a larger self-recruiting population whose boundaries are larger than the investigated area. This conclusion is in agreement with preliminary allozymic and mitochondrial DNA restriction fragment length polymorphism data, but contradicts the previous identification of 2 subpopulations of sardines in the Adriatic Sea argued on morphologic differences, which could be rather attributed to different hydrographic or ecologic conditions occurring in different areas of the Adriatic Sea. The reduced gene flow observed between Adriatic-Ionian and Spanish sardine geographic samples (P < 0.001) suggests that reproductively isolated populations of sardines may occur in the Mediterranean Sea.     

Genetic structuring and migration patterns of Atlantic bigeye tuna, <Emphasis Type="Italic">Thunnus obesus</Emphasis>(Lowe, 1839)

Background  

Large pelagic fishes are generally thought to have little population genetic structuring based on their cosmopolitan distribution, large population sizes and high dispersal capacities. However, gene flow can be influenced by ecological (e.g. homing behaviour) and physical (e.g. present-day ocean currents, past changes in sea temperature and levels) factors. In this regard, Atlantic bigeye tuna shows an interesting genetic structuring pattern with two highly divergent mitochondrial clades (Clades I and II), which are assumed to have been originated during the last Pleistocene glacial maxima. We assess genetic structure patterns of Atlantic bigeye tuna at the nuclear level, and compare them with mitochondrial evidence.     

A comparison of variation between a MHC pseudogene and microsatellite loci of the little greenbul (<Emphasis Type="Italic">Andropadus virens</Emphasis>)

Background  

We investigated genetic variation of a major histcompatibility complex (MHC) pseudogene (Anvi -DAB1) in the little greenbul (Andropadus virens) from four localities in Cameroon and one in Ivory Coast, West Africa. Previous microsatellite and mitochondrial DNA analyses had revealed little or no genetic differentiation among Cameroon localities but significant differentiation between localities in Cameroon and Ivory Coast.     

Comparative population structure of two dominant species,Shinkaia crosnieri (Munidopsidae: Shinkaia) and Bathymodiolus platifrons (Mytilidae: Bathymodiolus), inhabiting both deep‐sea vent and cold seep inferred from mitochondrial multi‐genes

Deep‐sea hydrothermal vents and cold seeps, limited environments without sunlight, are two types of extreme habitat for marine organisms. The differences between vents and cold seeps may facilitate genetic isolation and produce population heterogeneity. However, information on such chemosynthetic fauna taxa is rare, especially regarding the population diversity of species inhabiting both vents and cold seeps. In this study, three mitochondrial DNA fragments (the cytochrome c oxidase submit I (COI), cytochrome b gene (Cytb), and 16S) were concatenated as a mitochondrial concatenated dataset (MCD) to examine the genetic diversity, population structure, and demographic history of Shinkaia crosnieri and Bathymodiolus platifrons. The genetic diversity differences between vent and seep populations were statistically significant for S. crosnieri but not for B. platifrons. S. crosnieri showed less gene flow and higher levels of genetic differentiation between the vent and seep populations than B. platifrons. In addition, the results suggest that all the B. platifrons populations, but only the S. crosnieri vent populations, passed through a recent expansion or bottleneck. Therefore, different population distribution patterns for the two dominant species were detected; a pattern of population differentiation for S. crosnieri and a homogeneity pattern for B. platifrons. These different population distribution patterns were related to both extrinsic restrictive factors and intrinsic factors. Based on the fact that the two species were collected in almost identical or adjacent sampling sites, we speculated that the primary factors underlying the differences in the population distribution patterns were intrinsic. The historical demographics, dispersal ability, and the tolerance level of environmental heterogeneity are most likely responsible for the different distribution patterns.     

Lineage-specific late pleistocene expansion of an endemic subtropical gossamer-wing damselfly, Euphaea formosa, in Taiwan

Background  

Pleistocene glacial oscillations have significantly affected the historical population dynamics of temperate taxa. However, the general effects of recent climatic changes on the evolutionary history and genetic structure of extant subtropical species remain poorly understood. In the present study, phylogeographic and historical demographic analyses based on mitochondrial and nuclear DNA sequences were used. The aim was to investigate whether Pleistocene climatic cycles, paleo-drainages or mountain vicariance of Taiwan shaped the evolutionary diversification of a subtropical gossamer-wing damselfly, Euphaea formosa.     

Pleistocene glacial cycle effects on the phylogeography of the Chinese endemic bat species, <Emphasis Type="Italic">Myotis davidii</Emphasis>

Background  

Global climatic oscillations, glaciation cycles and the unique geographic topology of China have profoundly influenced species population distributions. In most species, contemporary distributions of populations cannot be fully understood, except in a historical context. Complex patterns of Pleistocene glaciations, as well as other physiographic changes have influenced the distribution of bat species in China. Until this study, there had been no phylogeographical research on Myotis davidii, an endemic Chinese bat. We used a combination of nuclear and mitochondrial DNA markers to investigate genetic diversity, population structure, and the demographic history of M. davidii. In particular, we compared patterns of genetic variation to glacial oscillations, topography, and environmental variation during the Pleistocene in an effort to explain current distributions in light of these historical processes.     

Mitochondrial haplotype diversity in the tortoise species <Emphasis Type="Italic">Testudo graeca</Emphasis> from North Africa and the Middle East

Background  

To help conservation programs of the endangered spur-thighed tortoise and to gain better insight into its systematics, genetic variation and evolution in the tortoise species Testudo graeca (Testudines: Testudinidae) was investigated by sequence analysis of a 394-nucleotide fragment of the mitochondrial 12S rRNA gene for 158 tortoise specimens belonging to the subspecies Testudo graeca graeca, Testudo graeca ibera, Testudo graeca terrestris, and a newly recognized subspecies Testudo graeca whitei. A 411-nucleotide fragment of the mitochondrial D-loop was additionally sequenced for a subset of 22 T. graeca, chosen because of their 12S gene haplotype and/or geographical origin.     

Bioregion heterogeneity correlates with extensive mitochondrial DNA diversity in the Namaqua rock mouse, <Emphasis Type="Italic">Micaelamys namaquensis</Emphasis>(Rodentia: Muridae) from southern Africa - evidence for a species complex

Background  

Intraspecific variation within the diverse southern African murine rodents has not been extensively investigated, yet cryptic diversity is evident in several taxa studied to date. The Namaqua rock mouse, Micaelamys namaquensis Smith, 1834 is a widespread endemic murine rodent from the subregion. Currently, a single species with four subspecies is recognised, but in the past up to 16 subspecies were described. Thus, this species is a good candidate for the investigation of patterns and processes of diversification in a diverse but under-studied mammalian subfamily and geographic region. Here, we report genetic differentiation based on mitochondrial DNA (mtDNA) cytochrome b (cyt b) sequences among samples collected over an extensive coverage of the species' range.     

Phylogeography of the California sheephead,Semicossyphus pulcher: the role of deep reefs as stepping stones and pathways to antitropicality

In the past decade, the study of dispersal of marine organisms has shifted from focusing predominantly on the larval stage to a recent interest in adult movement. Antitropical distributions provide a unique system to assess vagility and dispersal. In this study, we have focused on an antitropical wrasse genus, Semicossyphus, which includes the California sheephead, S. pulcher, and Darwin's sheephead, S. darwini. Using a phylogenetic approach based on mitochondrial and nuclear markers, and a population genetic approach based on mitochondrial control region sequences and 10 microsatellite loci, we compared the phylogenetic relationships of these two species, as well as the population genetic characteristics within S. pulcher. While S. pulcher and S. darwini are found in the temperate eastern Pacific regions of the northern and southern hemispheres, respectively, their genetic divergence was very small (estimated to have occurred between 200 and 600 kya). Within S. pulcher, genetic structuring was generally weak, especially along mainland California, but showed weak differentiation between Sea of Cortez and California, and between mainland California and Channel Islands. We highlight the congruence of weak genetic differentiation both within and between species and discuss possible causes for maintenance of high gene flow. In particular, we argue that deep and cooler water refugia are used as stepping stones to connect distant populations, resulting in low levels of genetic differentiation.     

Comparison of the feeding apparatus and diet of European sardines Sardina pilchardus of Atlantic and Mediterranean waters: ecological implications

In this study, the feeding apparatus (gill rakers, GR) and the diet composition of European sardine Sardina pilchardus populations living in two contrasting environments were compared: the upwelling area off western Iberia and the comparatively less productive region of the north‐western Mediterranean Sea. The importance of local adaptations in the trophic ecology of this species was estimated. Sardina pilchardus from the Atlantic Iberian coast and from the north‐western Mediterranean Sea have clear differences in the feeding apparatus and diet compositions. Those from the Atlantic Iberian coast have significantly more GRs than S. pilchardus of the same size range in the Mediterranean Sea. While S. pilchardus from the Mediterranean Sea mostly depend on prey ranging between 750–1500 and 3000–4000 µm, corresponding mostly to cladocerans, decapods and copepods, those from the Atlantic depend on smaller prey (50–500 and 1000–1500 µm) that include phytoplankton and copepods, particularly during summer months, and S. pilchardus eggs during the winter. The marked difference between the trophic ecology of S. pilchardus in the two areas studied appears to have originated from different dietary strategies that the two populations have adopted in contrasting feeding environments. These differences are shown to profoundly affect the size and quality of prey consumed, and the effect of cannibalism on the populations.     

Three‐dimensional post‐glacial expansion and diversification of an exploited oceanic fish

Vertical divergence in marine organisms is being increasingly documented, yet much remains to be carried out to understand the role of depth in the context of phylogeographic reconstruction and the identification of management units. An ideal study system to address this issue is the beaked redfish, Sebastes mentella – one of four species of ‘redfish’ occurring in the North Atlantic – which is known for a widely distributed ‘shallow‐pelagic’ oceanic type inhabiting waters between 250 and 550 m, and a more localized ‘deep‐pelagic’ population dwelling between 550 and 800 m, in the oceanic habitat of the Irminger Sea. Here, we investigate the extent of population structure in relation to both depth and geographic spread of oceanic beaked redfish throughout most of its distribution range. By sequencing the mitochondrial control region of 261 redfish collected over a decadal interval, and combining 160 rhodopsin coding nuclear sequences and previously genotyped microsatellite data, we map the existence of two strongly divergent evolutionary lineages with significantly different distribution patterns and historical demography, and whose genetic variance is mostly explained by depth. Combined genetic data, analysed via independent approaches, are consistent with a Late Pleistocene lineage split, where segregation by depth probably resulted from the interplay of climatic and oceanographic processes with life history and behavioural traits. The ongoing process of diversification in North Atlantic S. mentella may serve as an ‘hourglass’ to understand speciation and adaptive radiation in Sebastes and in other marine taxa distributed across a depth gradient.     

Evolutionary diversification of cryophilic <Emphasis Type="Italic">Grylloblatta</Emphasis>species (Grylloblattodea: Grylloblattidae) in alpine habitats of California

Background  

Climate in alpine habitats has undergone extreme variation during Pliocene and Pleistocene epochs, resulting in repeated expansion and contraction of alpine glaciers. Many cold-adapted alpine species have responded to these climatic changes with long-distance range shifts. These species typically exhibit shallow genetic differentiation over a large geographical area. In contrast, poorly dispersing organisms often form species complexes within mountain ranges, such as the California endemic ice-crawlers (Grylloblattodea: Grylloblattidae: Grylloblatta). The diversification pattern of poorly dispersing species might provide more information on the localized effects of historical climate change, the importance of particular climatic events, as well as the history of dispersal. Here we use multi-locus genetic data to examine the phylogenetic relationships and geographic pattern of diversification in California Grylloblatta.     

Scale morphometry allows discrimination of European sardine Sardina pilchardus and round sardinella Sardinella aurita and among their local populations

Landmark‐based geometric morphometric analysis revealed differences in scale shape between European sardine Sardina pilchardus and round sardinella Sardinella aurita as well as among the local populations of each species. Fish scale measurements from four different areas in the central and eastern Mediterranean Sea showed that the mean scale shape of the two species using landmark data could be differentiated with high certainty. Populations of S. aurita from the central and eastern Mediterranean Sea could be separated reliably (P < 0·001) with an average discrimination rate of 91%, whereas the average discrimination of the S. pilchardus populations was lower (80%), albeit still high.     

18S rRNA V9 metabarcoding for diet characterization: a critical evaluation with two sympatric zooplanktivorous fish species

The potential of the 18S rRNA V9 metabarcoding approach for diet assessment was explored using MiSeq paired‐end (PE; 2 × 150 bp) technology. To critically evaluate the method′s performance with degraded/digested DNA, the diets of two zooplanktivorous fish species from the Bay of Biscay, European sardine (Sardina pilchardus) and European sprat (Sprattus sprattus), were analysed. The taxonomic resolution and quantitative potential of the 18S V9 metabarcoding was first assessed both in silico and with mock and field plankton samples. Our method was capable of discriminating species within the reference database in a reliable way providing there was at least one variable position in the 18S V9 region. Furthermore, it successfully discriminated diet between both fish species, including habitat and diel differences among sardines, overcoming some of the limitations of traditional visual‐based diet analysis methods. The high sensitivity and semi‐quantitative nature of the 18S V9 metabarcoding approach was supported by both visual microscopy and qPCR‐based results. This molecular approach provides an alternative cost and time effective tool for food‐web analysis.