Ocean-wide genomic variation in Gray's beaked whales, Mesoplodon grayi.

The deep oceans of the Southern Hemisphere are home to several elusive and poorly studied marine megafauna. In the absence of robust observational data for these species, genetic data can aid inferences on population connectivity, demography and ecology. A previous investigation of genetic diversity and population structure in Gray's beaked whale (Mesoplodon grayi) from Western Australia and New Zealand found high levels of mtDNA diversity, no geographic structure and stable demographic history. To further investigate phylogeographic and demographic patterns across their range, we generated complete mitochondrial and partial nuclear genomes of 16 of the individuals previously analysed and included additional samples from South Africa (n = 2) and South Australia (n = 4), greatly expanding the spatial range of genomic data for the species. Gray's beaked whales are highly elusive and rarely observed, and our data represents a unique and geographically broad dataset. We find relatively high levels of diversity in the mitochondrial genome, despite an absence of population structure at the mitochondrial and nuclear level. Demographic analyses suggest these whales existed at stable levels over at least the past 1.1 million years, with an approximately twofold increase in female effective population size approximately 250 thousand years ago, coinciding with a period of increased Southern Ocean productivity, sea surface temperature and a potential expansion of suitable habitat. Our results suggest that Gray's beaked whales are likely to be resilient to near-future ecosystem changes, facilitating their conservation. Our study demonstrates the utility of low-effort shotgun sequencing in providing ecological information on highly elusive species.

Ecological and geographical speciation in Lucilia bufonivora: The evolution of amphibian obligate parasitism.

Lucilia (Diptera: Calliphoridae) is a genus of blowflies comprised largely of saprophagous and facultative parasites of livestock. Lucilia bufonivora, however, exhibits a unique form of obligate parasitism of amphibians, typically affecting wild hosts. The evolutionary route by which amphibian myiasis arose, however, is not well understood due to the low phylogenetic resolution in existing nuclear DNA phylogenies. Furthermore, the timing of when specificity for amphibian hosts arose in L. bufonivora is also unknown. In addition, this species was recently reported for the first time in North America (Canada) and, to date, no molecular studies have analysed the evolutionary relationships between individuals from Eastern and Western hemispheres. To provide broader insights into the evolution of the amphibian parasitic life history trait and to estimate when the trait first arose, a time-scaled phylogeny was inferred from a concatenated data set comprising mtDNA, nDNA and non-coding rDNA (COX1, per and ITS2 respectively). Specimens from Canada, the UK, Poland, Switzerland, the Netherlands and Germany were analysed, as well as individuals from its sister taxa, the saprophage Lucilia silvarum and a Nearctic species also implicated in amphibian myiasis, Lucilia elongata. Obligate amphibian parasitism appears to have arisen ~4 mya, likely as a result of niche displacement of a saprophagous/facultative parasite ancestor. Consistent paraphyly of L. bufonivora with respect to L. elongata across single-gene phylogenies and high mtDNA genetic distances between Nearctic and Palearctic individuals suggest on-going cryptic speciation facilitated by geographical isolation. These findings suggest that recent reports of L. bufonivora in the Nearctic do not constitute a recent introduction, but instead suggest that it remained unrecorded due to taxonomic confusion and low abundance. This is the first study to confirm the involvement of L. bufonivora in amphibian myiasis in Canada using DNA-based identification methods.

The toad fly Lucilia bufonivora: its evolutionary status and molecular identification.

The blow fly genus Lucilia is composed largely of saprophages and facultative myasis agents, including the economically important species Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) and Lucilia sericata (Meigen). Only one species is generally recognized as an obligate agent of myiasis, Lucilia bufonivora Moniez, and this is an obligate parasite of toads. Lucilia silvarum (Meigen), a sister species, behaves mainly as a carrion breeder; however, it has also been reported as a facultative parasite of amphibians. Morphologically, these species are almost identical, and historically this has led to misidentification, taxonomic ambiguity and a paucity of studies of L. bufonivora. In this study, dipterous larvae were analysed from toad myiasis cases from the U.K., The Netherlands and Switzerland, together with adult specimens of fly species implicated in amphibian parasitism: L. bufonivora, L. silvarum and Lucilia elongata Shannon (from North America). Partial sequences of two genes, cox1 and ef1α, were amplified. Seven additional blow fly species were analysed as outgroups. Bayesian inference trees of cox1, ef1α and a combined-gene dataset were constructed. All larvae isolated from toads were identified as L. bufonivora and no specimens of L. silvarum were implicated in amphibian myiasis. This study confirms L. silvarum and L. bufonivora as distinct sister species and provides unambiguous molecular identification of L. bufonivora.

Spatial and temporal habitat partitioning by calliphorid blowflies.

Calliphorid blowflies perform an essential ecosystem service in the consumption, recycling and dispersion of carrion nutrients and are considered amongst the most important functional groups in an ecosystem. Some species are of economic importance as facultative agents of livestock myiasis. The interspecific ecological differences that facilitate coexistence within the blowfly community are not fully understood. The aim of this work was to quantify differences in habitat use by calliphorid species. Thirty traps were distributed among three habitats at two sites in southwest England for collections made during March-August 2016. A total of 17 246 specimens were caught, of which 2427 were Lucilia sericata, 51 Lucilia richardsi, 6580 Lucilia caesar, 307 Lucilia ampullacea, 4881 Calliphora vicina and 2959 Calliphora vomitoria (all: Diptera: Calliphoridae). Lucilia sericata was the dominant species in open habitats, whereas L. caesar was the most abundant species in shaded habitats. Calliphora specimens were more abundant in the cooler months. These findings suggest that Calliphora and Lucilia species show strong temporal segregation mediated by temperature, and that species of the genus Lucilia show differences in the use of habitats that are likely to be driven by differences in humidity tolerance and light intensity. These factors in combination result in effective niche partitioning.

Mesotocin influences pinyon jay prosociality.

Many species exhibit prosocial behaviour, in which one individual's actions benefit another individual, often without an immediate benefit to itself. The neuropeptide oxytocin is an important hormonal mechanism influencing prosociality in mammals, but it is unclear whether the avian homologue mesotocin plays a similar functional role in birds. Here, we experimentally tested prosociality in pinyon jays (Gymnorhinus cyanocephalus), a highly social corvid species that spontaneously shares food with others. First, we measured prosocial preferences in a prosocial choice task with two different pay-off distributions: Prosocial trials delivered food to both the subject and either an empty cage or a partner bird, whereas Altruism trials delivered food only to an empty cage or a partner bird (none to subject). In a second experiment, we examined whether administering mesotocin influenced prosocial preferences. Compared to choices in a control condition, we show that subjects voluntarily delivered food rewards to partners, but only when also receiving food for themselves (Prosocial trials), and administration of high levels of mesotocin increased these behaviours. Thus, in birds, mesotocin seems to play a similar functional role in facilitating prosocial behaviours as oxytocin does in mammals, suggesting an evolutionarily conserved hormonal mechanism for prosociality.

Understanding and managing fish populations: keeping the toolbox fit for purpose.

Wild fish populations are currently experiencing unprecedented pressures, which are projected to intensify in the coming decades. Developing a thorough understanding of the influences of both biotic and abiotic factors on fish populations is a salient issue in contemporary fish conservation and management. During the 50th Anniversary Symposium of The Fisheries Society of the British Isles at the University of Exeter, UK, in July 2017, scientists from diverse research backgrounds gathered to discuss key topics under the broad umbrella of 'Understanding Fish Populations'. Below, the output of one such discussion group is detailed, focusing on tools used to investigate natural fish populations. Five main groups of approaches were identified: tagging and telemetry; molecular tools; survey tools; statistical and modelling tools; tissue analyses. The appraisal covered current challenges and potential solutions for each of these topics. In addition, three key themes were identified as applicable across all tool-based applications. These included data management, public engagement, and fisheries policy and governance. The continued innovation of tools and capacity to integrate interdisciplinary approaches into the future assessment and management of fish populations is highlighted as an important focus for the next 50 years of fisheries research.

Atlantic salmon Salmo salar in the chalk streams of England are genetically unique.

Recent research has identified genetic groups of Atlantic salmon Salmo salar that show association with geological and environmental boundaries. This study focuses on one particular subgroup of the species inhabiting the chalk streams of southern England, U.K. These fish are genetically distinct from other British and European S. salar populations and have previously demonstrated markedly low admixture with populations in neighbouring regions. The genetic population structure of S. salar occupying five chalk streams was explored using 16 microsatellite loci. The analysis provides evidence of the genetic distinctiveness of chalk-stream S. salar in southern England, in comparison with populations from non-chalk regions elsewhere in western Europe. Little genetic differentiation exists between the chalk-stream populations and a pattern of isolation by distance was evident. Furthermore, evidence of temporal stability of S. salar populations across the five chalk streams was found. This work provides new insights into the temporal stability and lack of genetic population sub-structuring within a unique component of the species' range of S. salar.

Contrasting patterns of population structure and gene flow facilitate exploration of connectivity in two widely distributed temperate octocorals.

Connectivity is an important component of metapopulation dynamics in marine systems and can influence population persistence, migration rates and conservation decisions associated with Marine Protected Areas (MPAs). In this study, we compared the genetic diversity, gene flow and population structure of two octocoral species, Eunicella verrucosa and Alcyonium digitatum, in the northeast Atlantic (ranging from the northwest of Ireland and the southern North Sea, to southern Portugal), using two panels of 13 and 8 microsatellite loci, respectively. Our results identified regional genetic structure in E. verrucosa partitioned between populations from southern Portugal, northwest Ireland and Britain/France; subsequent hierarchical analysis of population structure also indicated reduced gene flow between southwest Britain and northwest France. However, over a similar geographical area, A. digitatum showed little evidence of population structure, suggesting high gene flow and/or a large effective population size; indeed, the only significant genetic differentiation detected in A. digitatum occurred between North Sea samples and those from the English Channel/northeast Atlantic. In both species the vast majority of gene flow originated from sample sites within regions, with populations in southwest Britain being the predominant source of contemporary exogenous genetic variants for the populations studied. Overall, historical patterns of gene flow appeared more complex, though again southwest Britain appeared to be an important source of genetic variation for both species. Our findings have major conservation implications, particularly for E. verrucosa, a protected species in UK waters and listed by the IUCN as 'Vulnerable', and for the designation and management of European MPAs.

Symbiodinium thermophilum sp. nov., a thermotolerant symbiotic alga prevalent in corals of the world's hottest sea, the Persian/Arabian Gulf.

Coral reefs are in rapid decline on a global scale due to human activities and a changing climate. Shallow water reefs depend on the obligatory symbiosis between the habitat forming coral host and its algal symbiont from the genus Symbiodinium (zooxanthellae). This association is highly sensitive to thermal perturbations and temperatures as little as 1°C above the average summer maxima can cause the breakdown of this symbiosis, termed coral bleaching. Predicting the capacity of corals to survive the expected increase in seawater temperatures depends strongly on our understanding of the thermal tolerance of the symbiotic algae. Here we use molecular phylogenetic analysis of four genetic markers to describe Symbiodinium thermophilum, sp. nov. from the Persian/Arabian Gulf, a thermally tolerant coral symbiont. Phylogenetic inference using the non-coding region of the chloroplast psbA gene resolves S. thermophilum as a monophyletic lineage with large genetic distances from any other ITS2 C3 type found outside the Gulf. Through the characterisation of Symbiodinium associations of 6 species (5 genera) of Gulf corals, we demonstrate that S. thermophilum is the prevalent symbiont all year round in the world's hottest sea, the southern Persian/Arabian Gulf.

Long-term effects of stock transfers: synergistic introgression of allochthonous genomes in salmonids.

The genus Salmo was employed as a model to study introgression of genes between species due to secondary contacts. Seven microsatellite loci, the LDH-C1* locus and the 5S ribosomal DNA were studied. Results showed the mutually enhanced introgression of allochthonous genomes into southern European salmonids. This phenomenon appears to go beyond a simple consequence of the altered behaviour of domestic individuals. Invasions of autochthonous genomes by allochthonous genes would be enhanced by human activities such as stock transfers, which would simultaneously promote allochthonous and allospecific (from other species) introgressions in a synergistic process in Atlantic salmon Salmo salar and brown trout Salmo trutta. As a minor result, the data do not support the value of the microsatellite locus SsaD486 as a species-specific marker.

Use of multiple markers demonstrates a cryptic western refugium and postglacial colonisation routes of Atlantic salmon (Salmo salar L.) in Northwest Europe.

Glacial and postglacial processes are known to be important determinants of contemporary population structuring for many species. In Europe, refugia in the Italian, Balkan and Iberian peninsulas are believed to be the main sources of species colonising northern Europe after the glacial retreat; however, there is increasing evidence of small, cryptic refugia existing north of these for many cold-tolerant species. This study examined the glacial history of Atlantic salmon in western Europe using two independent classes of molecular markers, microsatellites (nuclear) and mitochondrial DNA variation. Alongside the well-documented refuge in the Iberian Peninsula, evidence for a cryptic refuge in northwest France is also presented. Critically, methods utilised to estimate divergence times between the refugia indicated that salmon in these two regions had diverged a long time before the last glacial maximum; coalescence analysis (as implemented in the program IMa2) estimated divergence times at around 60 000 years before present. Through the examination of haplotype frequencies, previously glaciated areas of northwest Europe, that is, Britain and Ireland, appear to have been colonised from salmon expanding out of both refugia, with the southwest of England being the primary contact zone and exhibiting the highest genetic diversity.

Parasite epidemiology in a changing world: can molecular phylogeography help us tell the wood from the trees?

SUMMARY Molecular phylogeography has revolutionised our ability to infer past biogeographic events from cross-sectional data on current parasite populations. In ecological parasitology, this approach has been used to address fundamental questions concerning host-parasite co-evolution and geographic patterns of spread, and has raised many technical issues and problems of interpretation. For applied parasitologists, the added complexity inherent in adding population genetic structure to perceived parasite distributions can sometimes seem to cloud rather than clarify approaches to control. In this paper, we use case studies firstly to illustrate the potential extent of cryptic diversity in parasite and parasitoid populations, secondly to consider how anthropogenic influences including movement of domestic animals affect the geographic distribution and host associations of parasite genotypes, and thirdly to explore the applied relevance of these processes to parasites of socio-economic importance. The contribution of phylogeographic approaches to deeper understanding of parasite biology in these cases is assessed. Thus, molecular data on the emerging parasites Angiostrongylus vasorum in dogs and wild canids, and the myiasis-causing flies Lucilia spp. in sheep and Cochliomyia hominovorax in humans, lead to clear implications for control efforts to limit global spread. Broader applications of molecular phylogeography to understanding parasite distributions in an era of rapid global change are also discussed.

Microsatellite standardization and evaluation of genotyping error in a large multi-partner research programme for conservation of Atlantic salmon (Salmo salar L.).

Microsatellite genotyping is a common DNA characterization technique in population, ecological and evolutionary genetics research. Since different alleles are sized relative to internal size-standards, different laboratories must calibrate and standardize allelic designations when exchanging data. This interchange of microsatellite data can often prove problematic. Here, 16 microsatellite loci were calibrated and standardized for the Atlantic salmon, Salmo salar, across 12 laboratories. Although inconsistencies were observed, particularly due to differences between migration of DNA fragments and actual allelic size ('size shifts'), inter-laboratory calibration was successful. Standardization also allowed an assessment of the degree and partitioning of genotyping error. Notably, the global allelic error rate was reduced from 0.05 ± 0.01 prior to calibration to 0.01 ± 0.002 post-calibration. Most errors were found to occur during analysis (i.e. when size-calling alleles; the mean proportion of all errors that were analytical errors across loci was 0.58 after calibration). No evidence was found of an association between the degree of error and allelic size range of a locus, number of alleles, nor repeat type, nor was there evidence that genotyping errors were more prevalent when a laboratory analyzed samples outside of the usual geographic area they encounter. The microsatellite calibration between laboratories presented here will be especially important for genetic assignment of marine-caught Atlantic salmon, enabling analysis of marine mortality, a major factor in the observed declines of this highly valued species.

Identification and lineage genotyping of South American trypanosomes using fluorescent fragment length barcoding.

Trypanosoma cruzi and Trypanosoma rangeli are human-infective blood parasites, largely restricted to Central and South America. They also infect a wide range of wild and domestic mammals and are transmitted by a numerous species of triatomine bugs. There are significant overlaps in the host and geographical ranges of both species. The two species consist of a number of distinct phylogenetic lineages. A range of PCR-based techniques have been developed to differentiate between these species and to assign their isolates into lineages. However, the existence of at least six and five lineages within T. cruzi and T. rangeli, respectively, makes identification of the full range of isolates difficult and time consuming. Here we have applied fluorescent fragment length barcoding (FFLB) to the problem of identifying and genotyping T. cruzi, T. rangeli and other South American trypanosomes. This technique discriminates species on the basis of length polymorphism of regions of the rDNA locus. FFLB was able to differentiate many trypanosome species known from South American mammals: T. cruzi cruzi, T. cruzi marinkellei, T. dionisii-like, T. evansi, T. lewisi, T. rangeli, T. theileri and T. vivax. Furthermore, all five T. rangeli lineages and many T. cruzi lineages could be identified, except the hybrid lineages TcV and TcVI that could not be distinguished from lineages III and II respectively. This method also allowed identification of mixed infections of T. cruzi and T. rangeli lineages in naturally infected triatomine bugs. The ability of FFLB to genotype multiple lineages of T. cruzi and T. rangeli together with other trypanosome species, using the same primer sets is an advantage over other currently available techniques. Overall, these results demonstrate that FFLB is a useful method for species diagnosis, genotyping and understanding the epidemiology of American trypanosomes.

Characterization and utilization of microsatellite loci in the New World screwworm fly, Cochliomyia hominivorax.

New World screwworm populations in North and Central America have been the targets of virtually continuous eradication attempts by sterile insect technique (SIT) since the 1950s. Nevertheless, in some areas, such as Jamaica, SIT control programmes have failed. Reasons for the failure of SIT-based control programmes in some locations are unknown, but it has been hypothesized that failure may be related to mating incompatibility between sterile and wild fly populations or to the existence of sexually incompatible cryptic species. This paper outlines the development of a suite of four new microsatellite loci which can be used to study intra-specific relationships between populations of Cochliomyia hominivorax from the Caribbean and South America, which represent those populations involved in, or earmarked for, forthcoming SIT control. Cross-amplification with the secondary screwworm, Cochliomyia macellaria, was also successful with three of the new loci. We present results which suggest that populations from Trinidad and Jamaica form distinct groupings of flies and that C. hominivorax from Trinidad appears particularly distinct.

Phylogenetic analysis of New World screwworm fly, Cochliomyia hominivorax, suggests genetic isolation of some Caribbean island populations following colonization from South America.

Larval infestations of the New World screwworm (NWS) fly, Cochliomyia hominivorax, cause considerable economic losses through the direct mortality and reduced production of livestock. Since the 1950s, NWS populations in North and Central America have been the target of virtually continuous eradication attempts by sterile insect technique (SIT). Nevertheless, in some areas, such as Jamaica, SIT-based control programmes have failed. Reasons for the failure of SIT-based programmes in some locations are unknown, but it is hypothesized that failure may be related to the mating incompatibility between sterile and wild flies or to the existence of sexually incompatible cryptic species. Accordingly, the current research investigates intraspecific phylogenetic relationships and associated biogeographic patterns between NWS populations from the Caribbean and South America, which represent those populations involved in, or earmarked for, forthcoming SIT programmes. Uniquely, this study also includes analyses of two North American samples, collected in Texas in 1933 and 1953 prior to initiation of the SIT-based eradication programme. The study utilizes three nucleotide datasets: elongation factor-1alpha (nuclear); cytochrome oxidase subunit 1 (mitochondrial), and 12S rRNA (mitochondrial). Phylogenetic analysis of these data, representing populations from across the Caribbean, South America and Texas, indicates sub-structuring of fly populations on several of the larger Caribbean islands, suggesting a period of isolation and/or founder effects following colonization from South America; significantly, our findings do not support a North American origin for Cuban flies. The importance of these findings in the light of proposed SIT programmes in the region is discussed.

Global gene expression analysis of bovine somatic cell nuclear transfer blastocysts and cotyledons.

Low developmental competence of bovine somatic cell nuclear transfer (SCNT) embryos is a universal problem. Abnormal placentation has been commonly reported in SCNT pregnancies from a number of species. The present study employed Affymetrix bovine expression microarrays to examine global gene expression patterns of SCNT and in vivo produced (AI) blastocysts as well as cotyledons from day-70 SCNT and AI pregnancies. SCNT and AI embryos and cotyledons were analyzed for differential expression. Also in an attempt to establish a link between abnormal gene expression patterns in early embryos and cotyledons, differentially expressed genes were compared between the two studies. Microarray analysis yielded a list of 28 genes differentially expressed between SCNT and AI blastocysts and 19 differentially expressed cotyledon genes. None of the differentially expressed genes were common to both groups, although major histocompatibility complex I (MHCI) was significant in the embryo data and approached significance in the cotyledon data. This is the first study to report global gene expression patterns in bovine AI and SCNT cotyledons. The embryonic gene expression data reported here adds to a growing body of data that indicates the common occurrence of aberrant gene expression in early SCNT embryos.

Complete mitochondrial control region sequences indicate a distinct variety of brown trout Salmo trutta in the Aral Sea.

Complete sequencing of the mtDNA control region (CR) from five specimens of brown trout Salmo trutta from the Amu Darya River identified two novel haplotypes belonging to the Danubian lineage. This finding supports the long-standing hypothesis that brown trout in the Aral Sea represent a distinct genetic stock and also illustrates the benefits that complete sequencing of the CR can provide for elucidating phylogeographic relationships.

Kinetoplastid phylogenetics, with special reference to the evolution of parasitic trypanosomes.

To fully understand the evolutionary history of parasitic kinetoplastids and to understand the context within which the evolution of each parasite group has developed, an understanding not just of the parasites, but of all kinetoplastids is required. Accordingly, this paper provides an overview of kinetoplastid evolution and systematics, including coverage of the proposal by Moreira et al. (2004) to divide kinetoplasts into Prokinetoplastina (Ichthyobodo and Perkinsiella) and Metakinetoplastina (other bodonids and trypanosomatids). The implications of such a revision, with regard to correctly identifying outgroup taxa for studies of evolution within taxa of medical importance, are addressed, together with a more detailed review of the evolution and origins of the trypanosomes in the light of new phylogenies, new approaches and revisions in kinetoplastid systematics.

Molecular characterization and phylogenesis of Steganinae (Diptera, Drosophilidae) inferred by the mitochondrial cytochrome c oxidase subunit 1.

The subfamily Steganinae (Diptera, Drosophilidae) includes flies which display zoophilic feeding behaviour in the larval and/or adult stages, some of which act as vectors of Spirurida eyeworms, which infect both carnivores and humans. To date, the taxonomy and phylogeny of the subfamily Steganinae has been studied only superficially and many aspects of their systematics remain unresolved. Thus, the present study aimed to provide a molecular dataset to facilitate the identification and phylogenetic analysis of Steganinae species based on partial ( approximately 700 basepairs) mitochondrial cytochrome c oxidase subunit 1 (cox1) sequences. A total of 134 flies belonging to 13 species and eight genera of Steganinae were subjected to molecular and phylogenetic analyses. The mean nucleotide variation within the Steganinae subfamily was 8.1%, with a variation within genera for which more than one species was examined ranging from 1.6% (in Phortica spp.) to 21.8% (in Amiota spp.). Interspecific pairwise divergence ranged from 1.6% (Phortica variegata vs. Phortica semivirgo) to 24.8% (Cacoxenus indagator vs. Amiota alboguttata) and intraspecific variation ranged from 0% to 1%. Seventy of the 233 amino acids were variable, including 26 parsimony informative sites and 44 singleton sites, with some highly conserved residues identified within the genera Stegana and Amiota. Parsimony and maximum likelihood-based phylogenetic analyses provided strong support for the genus Phortica, phylogenetically distinct from the genus Amiota. Gitona distigma was placed in an unresolved position adjacent to the outgroup taxa, Drosophila yakuba and Drosophila melanogaster. The molecular data reported here represent the first molecular dataset based on cox1 of Steganinae flies and provide a base for further investigations into the evolutionary relationships among this little-studied subfamily.