Genome scans reveal signals of selection associated with pollution in fish populations of Basilichthys microlepidotus, an endemic species of Chile.

The Maipo River catchment is one of Chile's most polluted basins. In recent decades, discharges of untreated sewage and organic matter have caused eutrophication and water quality degradation. We employed the indigenous silverfish species Basilichthys microlepidotus as a model organism to investigate the process of adaptation and selection on genes influenced by pollution. Using variation at single nucleotide polymorphisms (SNPs), we determined the temporal stability of the population structure patterns previously identified in this species by varying SNPs. We also examined local adaptation to pollution-selected genes. Using the genotypes of 7684 loci in 180 individuals, we identified 429 and 700 loci that may be undergoing selection. We detected these loci using the FSTHET and ARLEQUIN outlier detection software, respectively. Both software packages simultaneously identified a total of 250 loci. B. microlepidotus' population structure did not change over time at contaminated or unpolluted sites. In addition, our analysis found: (i) selection of genes associated with pollution, consistent with observations in other organisms; (ii) identification of candidate genes that are functionally linked to the same biological processes, molecular functions and/or cellular components that previously showed differential expression in the same populations; and (iii) a candidate gene with differential expression and a non-synonymous substitution.

Presence of microplastics in water, sediments and fish species in an urban coastal environment of Fiji, a Pacific small island developing state.

Microplastics (MP) in the marine environment are widely reported. However, MP occurrence in some geographical areas such as from Small Island Developing States (SIDS) is missing. The main aim of this study was to assess MP levels in the urban coastal environment of Suva, the capital of the Republic of Fiji. Microplastics were measured from surface water, sediments and fish gastrointestinal tracts (GI) from three sites adjacent to Suva. In addition, an evaluation on the contribution from the local sewage treatment plant to MP levels was undertaken. In general, low levels of MP in water and sediments were detected, but significant differences were observed among sites. The sewage treatment plant was shown to contribute to MP levels in sediment but not in water. Species ingestion of MP was high and associated with MP in sediments. Fibres and fragments were the predominant type of MP, and fifteen different polymers were detected, with higher percentages of polyethylene, latex and polypropylene. In conclusion, MP were found in Suva coastal environment and these MP were associated to land based human activities.

Indications of strong adaptive population genetic structure in albacore tuna (Thunnus alalunga) in the southwest and central Pacific Ocean.

Albacore tuna (Thunnus alalunga) has a distinctly complex life history in which juveniles and adults separate geographically but at times inhabit the same spaces sequentially. The species also migrates long distances and presumably experiences varied regimes of physical stress over a lifetime. There are, therefore, many opportunities for population structure to arise based on stochastic differences or environmental factors that promote local adaptation. However, with the extent of mobility consistently demonstrated by tagged individuals, there is also a strong argument for panmixia within an ocean basin. It is important to confirm such assumptions from a population genetics standpoint for this species in particular because albacore is one of the principal market tuna species that sustains massive global fisheries and yet is also a slow-growing temperate tuna. Consequently, we used 1,837 neutral SNP loci and 89 loci under potential selection to analyze population genetic structure among five sample groups collected from the western and central South Pacific. We found no evidence to challenge panmixia at neutral loci, but strong indications of structuring at adaptive loci. One population sample, from French Polynesia in 2004, was particularly differentiated. Unfortunately, the current study cannot infer whether the divergence is geographic or temporal, or possibly caused by sample distribution. We encourage future studies to include potentially adaptive loci and to continue fine-scale observations within an ocean basin, and not to assume genome-wide panmixia.

Essential waters: Young bull sharks in Fiji's largest riverine system.

Coastal and estuarine systems provide critical shark habitats due to their relatively high productivity and shallow, protected waters. The young (neonates, young-of-the-year, and juveniles) of many coastal shark species occupy a diverse range of habitats and areas where they experience environmental variability, including acute and seasonal shifts in local salinities and temperatures. Although the location and functioning of essential shark habitats has been a focus in recent shark research, there is a paucity of data from the South Pacific. In this study, we document the temporal and spatial distribution, age class composition, and environmental parameters of young bull sharks (Carcharhinus leucas) in the Rewa, Sigatoka, and Navua Rivers, Fiji's three largest riverine systems. One hundred and seventy-two young bull sharks were captured in fisheries-independent surveys from January 2016 to April 2018. The vast majority of the captures were neonates. Seasonality in patterns of occurrence of neonate individuals suggests a defined parturition period during summer. Environmental parameters between the Rewa and the Sigatoka River differed significantly, as did the recorded young bull sharks abundance. According to the surveys, young bull sharks occur in all three rivers with the Rewa River likely representing essential habitat for newly born bull sharks. These results enhance the understanding of bull shark ecology in Fiji and provide a scientific basis for the implementation of local conservation strategies that contribute to the protection of critical habitats.

Community profiling of the intestinal microbial community of juvenile Hammerhead Sharks (Sphyrna lewini) from the Rewa Delta, Fiji.

Fourteen juvenile scalloped hammerhead sharks (Sphyrna lewini; SHS) were captured between November and December 2014 in the Rewa Delta in Fiji, and assessed for intestinal microflora characterisation using 16S rRNA amplicon sequencing by Illumina Miseq. The microbial population revealed a fluctuating dominance between the Enterobacteriaceae and Vibrionaceae families, namely Citrobacter and Photobacterium spp. Other related marine operational taxonomic units were closely related to Afipia felis, Chloroflexus aggregans, Psychrobacter oceani, Pontibacter actiniarum and Shigella sonnei. Two sharks had distinctive profiles that were dominated by known pathogens, namely Aeromonas salmonicida and Klebsiella pneumonia. The presence of a Methanosaeta species, and of Shigella and Psychrobacter, would suggest sewage contamination because of a spill that occurred on the 6th of December 2014. This study successfully establishes a baseline for future research.

The population genomics of yellowfin tuna (Thunnus albacares) at global geographic scale challenges current stock delineation.

Yellowfin tuna, Thunnus albacares, is one of the most important seafood commodities in the world. Despite its great biological and economic importance, conflicting evidence arises from classical genetic and tagging studies concerning the yellowfin tuna population structure at local and global oceanic scales. Access to more powerful and cost effective genetic tools would represent the first step towards resolving the population structure of yellowfin tuna across its distribution range. Using a panel of 939 neutral Single Nucleotide Polymorphisms (SNPs), and the most comprehensive data set of yellowfin samples available so far, we found genetic differentiation among the Atlantic, Indian and Pacific oceans. The genetic stock structure analysis carried out with 33 outlier SNPs, putatively under selection, identified discrete populations within the Pacific Ocean and, for the first time, also within the Atlantic Ocean. Stock assessment approaches that consider genetic differences at neutral and adaptive genomic loci should be routinely implemented to check the status of the yellowfin tuna, prevent illegal trade, and develop more sustainable management measures.

Discovery of a multispecies shark aggregation and parturition area in the Ba Estuary, Fiji Islands.

Population declines in shark species have been reported on local and global scales, with overfishing, habitat destruction and climate change posing severe threats. The lack of species-specific baseline data on ecology and distribution of many sharks, however, makes conservation measures challenging. Here, we present a fisheries-independent shark survey from the Fiji Islands, where scientific knowledge on locally occurring elasmobranchs is largely still lacking despite the location's role as a shark hotspot in the Pacific. Juvenile shark abundance in the fishing grounds of the Ba Estuary (north-western Viti Levu) was assessed with a gillnet- and longline-based survey from December 2015 to April 2016. A total of 103 juvenile sharks identified as blacktip Carcharhinus limbatus (n = 57), scalloped hammerhead Sphyrna lewini (n = 35), and great hammerhead Sphyrna mokarran (n = 11) sharks were captured, tagged, and released. The condition of umbilical scars (68% open or semihealed), mean sizes of individuals (±SD) (C. limbatus: 66.5 ± 3.8 cm, S. lewini: 51.8 ± 4.8 cm, S. mokarran 77.4 ± 2.8 cm), and the presence of these species over recent years (based on fishermen interviews), suggest that the Ba Estuary area is a critical habitat for multiple species that are classified as "Near Threatened" or "Endangered." Specifically, the area likely acts as a parturition ground over the studied period, and potentially as a subsequent nursery area. We identified subareas of high abundance and found that temperature, salinity and depth acted as small-scale environmental drivers of shark abundance. The data suggests a tendency for species-specific spatial use, both horizontally (i.e., between sampling areas) and vertically (i.e., across the water column). These results enhance the understanding of shark ecology in Fiji and provide a scientific basis for the implementation of local conservation strategies that contribute to the protection of these threatened species.

The effects of a stressed inshore urban reef on coral recruitment in Suva Harbour, Fiji.

A relic inshore reef ecosystem adjacent to the Fijian capital of Suva and another remote inshore reef were monitored monthly from July 2014 to July 2015 for coral recruitment, sedimentation rates, coral cover, temperature, and light intensity. Despite a major sewage spill in Suva Harbour in December 2014, the municipal inshore site exposed to constant anthropogenic activity, recorded no significant differences in coral spat abundance (except for the family Poritidae) on artificial substrata compared to the remote inshore site. Total yearly spat abundance was 106 on municipal reef and 132 on remote reef, while average daily sediment trap collection rates (g cm2/day) were significantly higher in the municipal site for the entire duration of monitoring. Total annual particulate organic matter content in sediment was also significantly higher in the municipal site (107.51 g cm2), compared to the remote site (43.37 g cm2). Mean light intensity was significantly lower for the municipal site (69.81 lum/ft2) compared to the remote site (239.26 lum/ft2), with Photosynthetically Active Radiation also lower for the former (800-1,066.66 µmol m-2 s-1) compared to the latter (3,266.66-3,600 µmol m-2 s-1). The lack of significant differences in coral spat recruitment rates suggests that settling larvae may be unable to distinguish between sub-optimal and optimal sites probably as a consequence of interference with coral settlement cues arising from anthropogenic development.

Fisheries-independent surveys identify critical habitats for young scalloped hammerhead sharks (Sphyrna lewini) in the Rewa Delta, Fiji.

Sharp declines in numerous shark populations around the world have generated considerable interest in better understanding and characterising their biology, ecology and critical habitats. The scalloped hammerhead shark (SHS, Sphyrna lewini) is subject to a multitude of natural and anthropogenic threats that are often exacerbated within the coastal embayments and estuaries used during SHS early life stages. In this study, we describe the temporal and spatial distribution, age class composition, and reproductive biology of SHS in the Rewa Delta (RD), Fiji. A total of 1054 SHS (including 796 tagged individuals; 101 of which were recaptured) were captured from September 2014 to March 2016 in the RD. A majority of the captures in this area were neonates and young-of-the-year (YOY) (99.8%). Significant seasonality in patterns of occurrence of both neonates and YOY individuals suggests a defined parturition period during the austral summer. Between the seven sampling sites in the RD we also found significant differences in SHS neonate catch per unit of effort, and average total length of individuals. According to the data, the RD is likely to represent an important nursery area for SHS up to one year of age.

Transcriptomic response to thermal and salinity stress in introduced and native sympatric Palaemon caridean shrimps.

Organisms develop local adaptations to cope with spatially and temporally variable environments such as estuarine habitats, where abiotic parameters such as salinity and temperature fluctuate continuously. Studying the regulation of gene expression in a variable environment allows us to understand the underlying molecular mechanisms of these adaptations and the relative roles of the genetic and plastic response. The transcriptomes of the European native Palaemon longirostris (PL) and the introduced P. macrodactylus (PM) shrimps are described and compared after an experiment simulating summer conditions in the Guadalquivir Estuary, Spain. Specimens, collected in the Guadalquivir Estuary, were maintained at a temperature and salinity of 20 °C and 5 ppt for the control, and 30 °C and 15 ppt for the stress treatment. A large amount of differential gene expression was observed: 16,013 and 2,594 for PL and PM respectively. Functionally annotated unigenes revealed some differences, with PL seemingly having to face stronger physiological stress than PM. Thus, PM seems to have greater resistance than PL under conditions of high temperature and salinity. These results constitute a step forward in the understanding of the underlying molecular mechanisms of genetic adaptation of native invertebrates, and alien taxa that have successfully invaded estuaries in temperate regions around the world.

Sex Chromosome Evolution, Heterochiasmy, and Physiological QTL in the Salmonid Brook Charr Salvelinus fontinalis.

Whole-genome duplication (WGD) can have large impacts on genome evolution, and much remains unknown about these impacts. This includes the mechanisms of coping with a duplicated sex determination system and whether this has an impact on increasing the diversity of sex determination mechanisms. Other impacts include sexual conflict, where alleles having different optimums in each sex can result in sequestration of genes into nonrecombining sex chromosomes. Sex chromosome development itself may involve sex-specific recombination rate (i.e., heterochiasmy), which is also poorly understood. The family Salmonidae is a model system for these phenomena, having undergone autotetraploidization and subsequent rediploidization in most of the genome at the base of the lineage. The salmonid master sex determining gene is known, and many species have nonhomologous sex chromosomes, putatively due to transposition of this gene. In this study, we identify the sex chromosome of Brook Charr Salvelinus fontinalis and compare sex chromosome identities across the lineage (eight species and four genera). Although nonhomology is frequent, homologous sex chromosomes and other consistencies are present in distantly related species, indicating probable convergence on specific sex and neo-sex chromosomes. We also characterize strong heterochiasmy with 2.7-fold more crossovers in maternal than paternal haplotypes with paternal crossovers biased to chromosome ends. When considering only rediploidized chromosomes, the overall heterochiasmy trend remains, although with only 1.9-fold more recombination in the female than the male. Y chromosome crossovers are restricted to a single end of the chromosome, and this chromosome contains a large interspecific inversion, although its status between males and females remains unknown. Finally, we identify quantitative trait loci (QTL) for 21 unique growth, reproductive, and stress-related phenotypes to improve knowledge of the genetic architecture of these traits important to aquaculture and evolution.

Null alleles are ubiquitous at microsatellite loci in the Wedge Clam (Donax trunculus).

Recent studies have reported an unusually high frequency of nonamplifying alleles at microsatellite loci in bivalves. Null alleles have been associated with heterozygous deficits in many studies. While several studies have tested for its presence using different analytical tools, few have empirically tested for its consequences in estimating population structure and differentiation. We characterised 16 newly developed microsatellite loci and show that null alleles are ubiquitous in the wedge clam, Donax trunculus. We carried out several tests to demonstrate that the large heterozygous deficits observed in the newly characterised loci were most likely due to null alleles. We tested the robustness of microsatellite genotyping for population assignment by showing that well-recognised biogeographic regions of the south Atlantic and south Mediterranean coast of Spain harbour genetically different populations.

Isolation mediates persistent founder effects on zooplankton colonisation in new temporary ponds.

Understanding the colonisation process in zooplankton is crucial for successful restoration of aquatic ecosystems. Here, we analyzed the clonal and genetic structure of the cyclical parthenogenetic rotifer Brachionus plicatilis by following populations established in new temporary ponds during the first three hydroperiods. Rotifer populations established rapidly after first flooding, although colonisation was ongoing throughout the study. Multilocus genotypes from 7 microsatellite loci suggested that most populations (10 of 14) were founded by few clones. The exception was one of the four populations that persisted throughout the studied hydroperiods, where high genetic diversity in the first hydroperiod suggested colonisation from a historical egg bank, and no increase in allelic diversity was detected with time. In contrast, in another of these four populations, we observed a progressive increase of allelic diversity. This population became less differentiated from the other populations suggesting effective gene flow soon after its foundation. Allelic diversity and richness remained low in the remaining two, more isolated, populations, suggesting little gene flow. Our results highlight the complexity of colonisation dynamics, with evidence for persistent founder effects in some ponds, but not in others, and with early immigration both from external source populations, and from residual, historical diapausing egg banks.

Implications for management and conservation of the population genetic structure of the wedge clam Donax trunculus across two biogeographic boundaries.

In a resource management perspective, the understanding of the relative influence of the physical factors on species connectivity remains a major challenge and is also of great ecological and conservation biology interest. Despite the overfishing threat on the wedge clam Donax trunculus in Europe, relatively little information is known about its population genetic structure and connectivity and their consequences on conservation policies. We employed 16 microsatellite loci to characterise the genetic diversity and population structure of D. trunculus. A total of 514 samples from seven different localities along the Atlantic-Mediterranean transition, from the Atlantic (Gulf of Cádiz) to the north-western Mediterranean were genotyped. The analysis of the population genetic structure displayed a clear distinction along the Atlantic-Mediterranean transition with different clusters in the Atlantic Ocean, the Alboran Sea and the northwestern Mediterranean. Consequently, we recommend that these three areas should be considered as different management units. We showed that all populations seem to be at high long-term risk of extinction with the exception of the protected Doñana National Park population which still seems to have evolutionary potential. Therefore, our results emphasized the necessity of protection of this economic resource and the validity of molecular tools to evaluate the population dynamics.

Effect of the enzyme and PCR conditions on the quality of high-throughput DNA sequencing results.

Library preparation protocols for high-throughput DNA sequencing (HTS) include amplification steps in which errors can build up. In order to have confidence in the sequencing data, it is important to understand the effects of different Taq polymerases and PCR amplification protocols on the DNA molecules sequenced. We compared thirteen enzymes in three different marker systems: simple, single copy nuclear gene and complex multi-gene family. We also tested a modified PCR protocol, which has been suggested to reduce errors associated with amplification steps. We find that enzyme choice has a large impact on the proportion of correct sequences recovered. The most complex marker systems yielded fewer correct reads, and the proportion of correct reads was greatly affected by the enzyme used. Modified cycling conditions did reduce the number of incorrect sequences obtained in some cases, but enzyme had a much greater impact on the number of correct reads. Thus, the coverage required for the safe identification of genotypes using one of the low quality enzymes could be seven times larger than with more efficient enzymes in a biallelic system with equal amplification of the two alleles. Consequently, enzyme selection for downstream HTS has important consequences, especially in complex genetic systems.

High genetic diversity and absence of founder effects in a worldwide aquatic invader.

The introduced oriental shrimp Palaemon macrodactylus has recently become widespread in temperate estuaries worldwide. However, this recent worldwide spread outside of its native range arises after a previous introduction to the US Pacific coast, where it was restricted for more than 30 years. Using a phylogeographic approach, the present work investigates the genetic history of the invasion of this decapod worldwide. Japan acted as the main native source area for worldwide introduced populations, but other native areas (likely South Korea and China) may act as source populations as well. The recently introduced European and NW Atlantic populations result from colonization from both Japan and an unknown area of the native range, although colonization from the NE Pacific could not be ruled out. Most introduced populations had higher haplotypic diversity than most native populations. P. macrodactylus has a strong potential to become one of the most widespread introduced species and may become the dominant estuarine shrimp in Europe. The ecological and economic consequences of this invasion remain to be thoroughly evaluated.

Combining next-generation sequencing and online databases for microsatellite development in non-model organisms.

Next-generation sequencing (NGS) is revolutionising marker development and the rapidly increasing amount of transcriptomes published across a wide variety of taxa is providing valuable sequence databases for the identification of genetic markers without the need to generate new sequences. Microsatellites are still the most important source of polymorphic markers in ecology and evolution. Motivated by our long-term interest in the adaptive radiation of a non-model species complex of whitefishes (Coregonus spp.), in this study, we focus on microsatellite characterisation and multiplex optimisation using transcriptome sequences generated by Illumina® and Roche-454, as well as online databases of Expressed Sequence Tags (EST) for the study of whitefish evolution and demographic history. We identified and optimised 40 polymorphic loci in multiplex PCR reactions and validated the robustness of our analyses by testing several population genetics and phylogeographic predictions using 494 fish from five lakes and 2 distinct ecotypes.

Y-chromosome analysis in Retuertas horses.

Several studies based on a variety of genetic markers have attempted to establish the origins of horse domestication. Thus far a discrepancy between the results of mitochondrial DNA analysis, which show high levels of diversity, and results from the Y-chromosome, with almost no genetic variability, has been identified. Most previous work on the horse Y-chromosome has focused on widespread, popular breeds or local Asian breeds. It is possible that these breeds represent a reduced set of the genetic variation present in the species. Additional genetic variation may be present in local breeds and ancient feral populations, such as the Retuertas horse in Spain. In this study we analyzed the Y-chromosome of the Retuertas horse, a feral horse population on the Iberian Peninsula that is at least several hundred years old, and whose genetic diversity and morphology suggests that it has been reproductively isolated for a long time. Data from the Retuertas horse was compared to another 11 breeds from the region (Portugal, Spain and France) or likely of Iberian origin, and then to data from 15 more breeds from around the globe. We sequenced 31 introns, Zinc finger Y-chromosomal protein (ZFY) and anonymous Y-linked fragments and genotyped 6 microsatellite loci found on the Y-chromosome. We found no sequence variation among all individuals and all breeds studied. However, fifteen differences were discovered between our data set and reference sequences in GenBank. We show that these likely represent errors within the deposited sequences, and suggest that they should not be used as comparative data for future projects.

Genetic diversity at neutral and adaptive loci determines individual fitness in a long-lived territorial bird.

There is compelling evidence about the manifest effects of inbreeding depression on individual fitness and populations' risk of extinction. The majority of studies addressing inbreeding depression on wild populations are generally based on indirect measures of inbreeding using neutral markers. However, the study of functional loci, such as genes of the major histocompatibility complex (MHC), is highly recommended. MHC genes constitute an essential component of the immune system of individuals, which is directly related to individual fitness and survival. In this study, we analyse heterozygosity fitness correlations of neutral and adaptive genetic variation (22 microsatellite loci and two loci of the MHC class II, respectively) with the age of recruitment and breeding success of a decimated and geographically isolated population of a long-lived territorial vulture. Our results indicate a negative correlation between neutral genetic diversity and age of recruitment, suggesting that inbreeding may be delaying reproduction. We also found a positive correlation between functional (MHC) genetic diversity and breeding success, together with a specific positive effect of the most frequent pair of cosegregating MHC alleles in the population. Globally, our findings demonstrate that genetic depauperation in small populations has a negative impact on the individual fitness, thus increasing the populations' extinction risk.

Major histocompatibility complex variation in insular populations of the Egyptian vulture: inferences about the roles of genetic drift and selection.

Insular populations have attracted the attention of evolutionary biologists because of their morphological and ecological peculiarities with respect to their mainland counterparts. Founder effects and genetic drift are known to distribute neutral genetic variability in these demes. However, elucidating whether these evolutionary forces have also shaped adaptive variation is crucial to evaluate the real impact of reduced genetic variation in small populations. Genes of the major histocompatibility complex (MHC) are classical examples of evolutionarily relevant loci because of their well-known role in pathogen confrontation and clearance. In this study, we aim to disentangle the partial roles of genetic drift and natural selection in the spatial distribution of MHC variation in insular populations. To this end, we integrate the study of neutral (22 microsatellites and one mtDNA locus) and MHC class II variation in one mainland (Iberia) and two insular populations (Fuerteventura and Menorca) of the endangered Egyptian vulture (Neophron percnopterus). Overall, the distribution of the frequencies of individual MHC alleles (n=17 alleles from two class II B loci) does not significantly depart from neutral expectations, which indicates a prominent role for genetic drift over selection. However, our results point towards an interesting co-evolution of gene duplicates that maintains different pairs of divergent alleles in strong linkage disequilibrium on islands. We hypothesize that the co-evolution of genes may counteract the loss of genetic diversity in insular demes, maximize antigen recognition capabilities when gene diversity is reduced, and promote the co-segregation of the most efficient allele combinations to cope with local pathogen communities.