Unravelling the taxonomy and identification of a problematic group of benthic fishes from tropical rivers (Gobiidae: Glossogobius).

Flathead gobies (genus Glossogobius) include c. 40 small- to medium-sized benthic fishes found primarily in freshwater habitats across the Indo-Pacific, having biodiversity value as well as cultural and economic value as food fishes, especially in developing countries. To help resolve considerable confusion regarding the identification of some of the larger-growing Glossogobius species, a systematic framework was established using nuclear genetic markers, mitochondrial DNA barcoding and phenotypic evidence for a geographically widespread collection of individuals from the waterways of tropical northern Australia. Species boundaries and distribution patterns were discordant with those previously reported, most notably for the tank goby Glossogobius giuris, which included a cryptic species. Genetic divergence was matched with accompanying unique visual characters that aid field identification. Additional taxonomic complexity was also evident, by comparison with DNA barcodes from international locations, suggesting that the specific names applicable for two of the candidate species in Australia remain unresolved due to confusion surrounding type specimens. Although flathead gobies are assumed to be widespread and common, this study demonstrates that unrealised taxonomic and ecological complexity is evident, and this will influence assessments of tropical biodiversity and species conservation. This study supports the need for taxonomic studies of freshwater fishes to underpin management in areas subject to significant environmental change.

New Sicydiinae phylogeny (Teleostei: Gobioidei) inferred from mitochondrial and nuclear genes: insights on systematics and ancestral areas.

The Sicydiinae subfamily (Teleostei: Gobioidei) is the biggest contributor to the diversity of fish communities in river systems of tropical islands. These species are found in the Indo-Pacific area, the Caribbean region and West Africa. They spawn in freshwater, their planktotrophic larvae drift downstream to the sea where they develop, before returning to the rivers to grow and reproduce. Hence, they are called amphidromous. Their phylogeny has been explored using a total of 3545 sites from 5 molecular markers (mitochondrial DNA: 16S rDNA, cytochrome oxidase I, cytochrome b; nuclear DNA: rhodopsin gene and a nuclear marker specially developed for this study, the interferon regulatory factor 2 binding protein 1-IRF2PB1). Sequences were obtained for 59 Sicydiinae specimens of 9 known genera. The Bayesian and maximum likelihood analyses support the monophyly of the subfamily as well as the monophylyof all genera except Sicydium, which is paraphyletic. Five major clades were identified within this subfamily. One clade contained the genus Stiphodon. Another clade contained Sicyopterus, Sicydium and Parasicydium with Sicyopterus as sister genus of Sicydium. The non-monophyly of Sicydium subclade, because it includes the monotypic genus Parasicydium, challenged the validity of Parasicydium genus. Ancestral area reconstruction showed that the subfamily emerged in the Central West Pacific region implying that previous hypotheses proposing a dispersal route for Sicydiinae into the Atlantic Ocean are unsupported by the present analysis. Our results suggest that the hypotheses for the dispersal route of the genus Sicydium should be reconsidered.

Variable outcomes of hybridization between declining Alosa alosa and Alosa fallax.

Hybridization dynamics between co-occurring species in environments where human-mediated changes take place are important to quantify for furthering our understanding of human impacts on species evolution and for informing management. The allis shad Alosa alosa (Linnaeus, 1758) and twaite shad Alosa fallax (Lacépède, 1803), two clupeids sister species, have been severely impacted by human activities across Europe. The shrinkage of A. alosa distribution range along with the decline of the remaining populations' abundance threatens its persistence. The main objective was to evaluate the extent of hybridization and introgression between those interacting species. We developed a set of 77 species-specific SNP loci that allowed a better resolution than morphological traits as they enabled the detection of hybrids up to the third generation. Variable rates of contemporary hybridization and introgression patterns were detected in 12 studied sites across the French Atlantic coast. Mitochondrial markers revealed a cyto-nuclear discordance almost invariably involving A. alosa individuals with an A. fallax mitochondrial DNA and provided evidence of historical asymmetric introgression. Overall, contemporary and historical introgression revealed by nuclear and mitochondrial markers strongly suggests that a transfer of genes occurs from A. fallax toward A. alosa genome since at least four generations. Moreover, the outcomes of introgression greatly depend on the catchments where local processes are thought to occur. Undoubtedly, interspecific interaction and gene flow should not be overlooked when considering the management of those species.

Fast sequence-based microsatellite genotyping development workflow.

Application of high-throughput sequencing technologies to microsatellite genotyping (SSRseq) has been shown to remove many of the limitations of electrophoresis-based methods and to refine inference of population genetic diversity and structure. We present here a streamlined SSRseq development workflow that includes microsatellite development, multiplexed marker amplification and sequencing, and automated bioinformatics data analysis. We illustrate its application to five groups of species across phyla (fungi, plant, insect and fish) with different levels of genomic resource availability. We found that relying on previously developed microsatellite assay is not optimal and leads to a resulting low number of reliable locus being genotyped. In contrast, de novo ad hoc primer designs gives highly multiplexed microsatellite assays that can be sequenced to produce high quality genotypes for 20-40 loci. We highlight critical upfront development factors to consider for effective SSRseq setup in a wide range of situations. Sequence analysis accounting for all linked polymorphisms along the sequence quickly generates a powerful multi-allelic haplotype-based genotypic dataset, calling to new theoretical and analytical frameworks to extract more information from multi-nucleotide polymorphism marker systems.

Strong population structure deduced from genetics, otolith chemistry and parasite abundances explains vulnerability to localized fishery collapse in a large Sciaenid fish, Protonibea diacanthus.

As pressure on coastal marine resources is increasing globally, the need to quantitatively assess vulnerable fish stocks is crucial in order to avoid the ecological consequences of stock depletions. Species of Sciaenidae (croakers, drums) are important components of tropical and temperate fisheries and are especially vulnerable to exploitation. The black-spotted croaker, Protonibea diacanthus, is the only large sciaenid in coastal waters of northern Australia where it is targeted by commercial, recreational and indigenous fishers due to its food value and predictable aggregating behaviour. Localized declines in the abundance of this species have been observed, highlighting the urgent requirement by managers for information on fine- and broad-scale population connectivity. This study examined the population structure of P. diacanthus across north-western Australia using three complementary methods: genetic variation in microsatellite markers, otolith elemental composition and parasite assemblage composition. The genetic analyses demonstrated that there were at least five genetically distinct populations across the study region, with gene flow most likely restricted by inshore biogeographic barriers such as the Dampier Peninsula. The otolith chemistry and parasite analyses also revealed strong spatial variation among locations within broad-scale regions, suggesting fine-scale location fidelity within the lifetimes of individual fish. The complementarity of the three techniques elucidated patterns of connectivity over a range of spatial and temporal scales. We conclude that fisheries stock assessments and management are required at fine scales (100 s of km) to account for the restricted exchange among populations (stocks) and to prevent localized extirpations of this species. Realistic management arrangements may involve the successive closure and opening of fishing areas to reduce fishing pressure.

The complete mitochondrial genome of the black jewfish Protonibea diacanthus (Perciformes: Sciaenidae).

We describe the complete mitochondrial genome of the black Jewfish Protonibea diacanthus. It was assembled from approximately 1.6 million reads produced by Ion Torrent next generation sequencing. The complete genome was 16,521 bp in length consisting of 13 protein-coding regions, 22 tRNA, 12S and 16S rRNA as well as two non-coding regions. The A+T base content (52.8%) is similar to other teleosts.

The complete mitochondrial genome of the golden snapper Lutjanus johnii (Perciformes: Lutjanidae).

We describe the complete mitochondrial genome of the golden snapper Lutjanus johnii. It was assembled from approximately 1.4 million reads produced by Ion Torrent next generation sequencing. The complete genome was 16,596 bp in length consisting of 13 protein-coding regions, 22 tRNA, 12S and 16S rRNA as well as two non-coding regions. The A+T base content (52.8%) is similar to other teleosts.

The complete mitochondrial genome of the grass emperor, Lethrinus laticaudis (Perciformes: Lethrinidae).

The grass emperor Lethrinus laticaudis is a coral reef fish that has high value to fisheries and is vulnerable to overharvesting. The complete mitochondrial genome was assembled from approximately 5.5 million reads produced by Illumina MiSeq. The 16,758 bp consisted of 13 protein-coding genes, 22 transfer RNA genes and two ribosomal RNA genes (12S and 16S). The genes and RNAs order and orientation on as well as the A + T base content (50.7%) was similar to what is found in other Teleosts. A phylogenetic tree with the most closely related species available in GenBank was built to validate L. laticaudis mitogenome.

Characterization, development and multiplexing of microsatellite markers in three commercially exploited reef fish and their application for stock identification.

Thirty-four microsatellite loci were isolated from three reef fish species; golden snapper Lutjanus johnii, blackspotted croaker Protonibea diacanthus and grass emperor Lethrinus laticaudis using a next generation sequencing approach. Both IonTorrent single reads and Illumina MiSeq paired-end reads were used, with the latter demonstrating a higher quality of reads than the IonTorrent. From the 1-1.5 million raw reads per species, we successfully obtained 10-13 polymorphic loci for each species, which satisfied stringent design criteria. We developed multiplex panels for the amplification of the golden snapper and the blackspotted croaker loci, as well as post-amplification pooling panels for the grass emperor loci. The microsatellites characterized in this work were tested across three locations of northern Australia. The microsatellites we developed can detect population differentiation across northern Australia and may be used for genetic structure studies and stock identification.

Contrasting genetic structure among populations of two amphidromous fish species (Sicydiinae) in the Central West Pacific.

Both present-day and past processes can shape connectivity of populations. Pleistocene vicariant events and dispersal have shaped the present distribution and connectivity patterns of aquatic species in the Indo-Pacific region. In particular, the processes that have shaped distribution of amphidromous goby species still remain unknown. Previous studies show that phylogeographic breaks are observed between populations in the Indian and Pacific Oceans where the shallow Sunda shelf constituted a geographical barrier to dispersal, or that the large spans of open ocean that isolate the Hawaiian or Polynesian Islands are also barriers for amphidromous species even though they have great dispersal capacity. Here we assess past and present genetic structure of populations of two amphidromous fish (gobies of the Sicydiinae) that are widely distributed in the Central West Pacific and which have similar pelagic larval durations. We analysed sections of mitochondrial COI, Cytb and nuclear Rhodospine genes in individuals sampled from different locations across their entire known range. Similar to other Sicydiinae fish, intraspecific mtDNA genetic diversity was high for all species (haplotype diversity between 0.9-0.96). Spatial analyses of genetic variation in Sicyopus zosterophorum demonstrated strong isolation across the Torres Strait, which was a geologically intermittent land barrier linking Australia to Papua New Guinea. There was a clear genetic break between the northwestern and the southwestern clusters in Si. zosterophorum (φST = 0.67502 for COI) and coalescent analyses revealed that the two populations split at 306 Kyr BP (95% HPD 79-625 Kyr BP), which is consistent with a Pleistocene separation caused by the Torres Strait barrier. However, this geographical barrier did not seem to affect Sm. fehlmanni. Historical and demographic hypotheses are raised to explain the different patterns of population structure and distribution between these species. Strategies aiming to conserve amphidromous fish should consider the presence of cryptic evolutionary lineages to prevent stock depletion.