Estimating the Density of Honeybee Colonies across Their Natural Range to Fill the Gap in Pollinator Decline Censuses

Although pollinator declines are a global biodiversity threat, the demography of the western honeybee (Apis mellifera) has not been considered by conservationists because it is biased by the activity of beekeepers. To fill this gap in pollinator decline censuses and to provide a broad picture of the current status of honeybees across their natural range, we used microsatellite genetic markers to estimate colony densities and genetic diversity at different locations in Europe, Africa, and central Asia that had different patterns of land use. Genetic diversity and colony densities were highest in South Africa and lowest in Northern Europe and were correlated with mean annual temperature. Confounding factors not related to climate, however, are also likely to influence genetic diversity and colony densities in honeybee populations. Land use showed a significantly negative influence over genetic diversity and the density of honeybee colonies over all sampling locations. In Europe honeybees sampled in nature reserves had genetic diversity and colony densities similar to those sampled in agricultural landscapes, which suggests that the former are not wild but may have come from managed hives. Other results also support this idea: putative wild bees were rare in our European samples, and the mean estimated density of honeybee colonies on the continent closely resembled the reported mean number of managed hives. Current densities of European honeybee populations are in the same range as those found in the adverse climatic conditions of the Kalahari and Saharan deserts, which suggests that beekeeping activities do not compensate for the loss of wild colonies. Our findings highlight the importance of reconsidering the conservation status of honeybees in Europe and of regarding beekeeping not only as a profitable business for producing honey, but also as an essential component of biodiversity conservation.

Pathogenicity and molecular genetics of O-specific side-chain lipopolysaccharides of Escherichia coli

Lipopolysaccharide (LPS), a glycolipid molecule found on the outer leaflet of outer membranes of gram-negative bacteria, consists of three moieties: lipid A, core oligosaccharide, and the O-specific polysaccharide chain. The O-specific side chain, which extends to the extracellular milieu, plays an important role in pathogenicity, especially during the initial stages of infection, because of its ability to interact with serum complement. In recent years, several laboratories have used recombinant DNA tools to determine, at the molecular level, the organization, expression, and regulation of genes involved in LPS biosynthesis in Salmonella and Escherichia coli. An increased understanding of the molecular aspects of the O-specific side-chain genes will shed light on the intimate details related with the formation of the O-specific side chain, its assembly onto the lipid A--core, and the translocation and insertion of the complete LPS molecule into the outer membrane. It will also contribute to the understanding of the evolution of these genes and the correlation of chemical diversity of O-specific side chains with the genetic diversity of O-specific side-chain genes. In addition, since the O-specific side chains are involved in the pathogenicity of medically important gram-negative bacteria, a basic understanding of the regulation and expression of O-specific side chain LPS genes will contribute to the field of molecular pathogenesis. This article provides an overview of the role of O-specific side chains in septicemic infections and also discusses the current status of molecular genetic studies on O-specific side-chain genes from E. coli.

Variation in grain arsenic assessed in a diverse panel of rice (Oryza sativa) grown in multiple sites

• Inorganic arsenic (As(i) ) in rice (Oryza sativa) grains is a possible threat to human health, with risk being strongly linked to total dietary rice consumption and consumed rice As(i) content. This study aimed to identify the range and stability of genetic variation in grain arsenic (As) in rice. • Six field trials were conducted (one each in Bangladesh and China, two in Arkansas, USA over 2 yr, and two in Texas, USA comparing flooded and nonflood treatments) on a large number of common rice cultivars (c. 300) representing genetic diversity among international rice cultivars. • Within each field there was a 3-34 fold range in grain As concentration which varied between rice subpopulations. Importantly, As(i) correlated strongly with total As among a subset of 40 cultivars harvested in Bangladesh and China. • Genetic variation at all field sites was a large determining factor for grain As concentration, indicating that cultivars low in grain As could be developed through breeding. The temperate japonicas exhibited lower grain As compared with other subpopulations. Effects for year, location and flooding management were also statistically significant, suggesting that breeding strategies must take into account environmental factors.

Postglacial dispersal, rather than in situ glacial survival, best explains the disjunct distribution of the Lusitanian plant species Daboecia cantabrica (Ericaceae)

Aim:

The distribution of the Lusitanian flora and fauna, species which are found only in southern and western Ireland and in northern Spain and Portugal but which are absent from intervening countries, represents one of the classic conundrums of biogeography. The aim of the present study was to determine whether the distribution of the Lusitanian plant species Daboecia cantabrica was due to persistence in separate Irish and Iberian refugia, or has resulted from post-glacial recolonization followed by subsequent extinction of intervening populations.

Location:

Northern Spain and Co. Galway, western Ireland.

Methods:

Palaeodistribution modelling using Maxent was employed to identify putative refugial areas for D. cantabrica at the Last Glacial Maximum (LGM). Phylogeographical analysis of samples from 64 locations in Ireland and Spain were carried out using a chloroplast marker (atpB–rbcL), the nuclear ITS region, and an anonymous nuclear single-copy locus.

Results:

The palaeodistribution model indicated areas with a high probability of survival for D. cantabrica at the LGM off the western coast of Galicia in Spain, and in the Bay of Biscay. Spanish populations exhibited substantially higher genetic diversity than Irish populations at all three loci, as well as geographical structuring of haplotypes within Spain consistent with divergence in separate refugia. Spanish populations also exhibited far more endemic haplotypes. Divergence time between Irish and Spanish populations associated with the putative Biscay refugium was estimated as 3.333–32 ka.

Main conclusions:

Our data indicate persistence by D. cantabrica throughout the LGM in two separate southern refugia: one in western Galicia and one in the area off the coast of western France which now lies in the Bay of Biscay. Spain was recolonized from both refugia, whilst Ireland was most likely recolonized from the Biscay refugium. On the balance of evidence across the three marker types and the palaeodistribution modelling, our findings do not support the idea of in situ survival of D. cantabrica in Ireland, contrary to earlier suggestions. The fact that we cannot conclusively rule out the existence of a small, more northerly refugium, however, highlights the need for further analysis of Lusitanian plant species.

Conservation genetics of Ireland's sole population of the River water crowfoot (Ranunculus fluitans Lam.)

Populations of many freshwater species are becoming increasingly threatened as a result of a wide range of anthropogenically mediated factors. In the present study, we wanted to assess levels and patterns of genetic diversity in Ireland's sole population of the River water crowfoot (Ranunculus fluitans), which is restricted to a 12 km stretch of a single river, to assist the formation of conservation strategies. Analysis using amplified fragment length polymorphism (AFLP) indicated comparable levels of genetic diversity to those exhibited by a more extensive population of the species in England, and revealed no evidence of clonal reproduction. Allele-specific PCR analysis of five nuclear single nucleotide polymorphisms (SNPs) indicated no evidence of hybridization with its more abundant congener Ranunculus penicillatus, despite previous anecdotal reports of the occurrence of hybrids. Although the population currently exhibits healthy levels of genetic diversity and is not at risk of genetic assimilation via hybridization with R. penicillatus, it still remains vulnerable to other factors such as stochastic events and invasive species. © 2013 Elsevier B.V. All rights reserved.

Neutral Genomic Microevolution of a Recently Emerged Pathogen, Salmonella enterica Serovar Agona

Salmonella enterica serovar Agona has caused multiple food-borne outbreaks of gastroenteritis since it was first isolated in 1952. We analyzed the genomes of 73 isolates from global sources, comparing five distinct outbreaks with sporadic infections as well as food contamination and the environment. Agona consists of three lineages with minimal mutational diversity: only 846 single nucleotide polymorphisms (SNPs) have accumulated in the non-repetitive, core genome since Agona evolved in 1932 and subsequently underwent a major population expansion in the 1960s. Homologous recombination with other serovars of S. enterica imported 42 recombinational tracts (360 kb) in 5/143 nodes within the genealogy, which resulted in 3,164 additional SNPs. In contrast to this paucity of genetic diversity, Agona is highly diverse according to pulsed-field gel electrophoresis (PFGE), which is used to assign isolates to outbreaks. PFGE diversity reflects a highly dynamic accessory genome associated with the gain or loss (indels) of 51 bacteriophages, 10 plasmids, and 6 integrative conjugational elements (ICE/IMEs), but did not correlate uniquely with outbreaks. Unlike the core genome, indels occurred repeatedly in independent nodes (homoplasies), resulting in inaccurate PFGE genealogies. The accessory genome contained only few cargo genes relevant to infection, other than antibiotic resistance. Thus, most of the genetic diversity within this recently emerged pathogen reflects changes in the accessory genome, or is due to recombination, but these changes seemed to reflect neutral processes rather than Darwinian selection. Each outbreak was caused by an independent clade, without universal, outbreak-associated genomic features, and none of the variable genes in the pan-genome seemed to be associated with an ability to cause outbreaks. © 2013 Achtman et al

Phylogeographical analysis of two cold-tolerant plants with disjunct Lusitanian distributions does not support in situ survival during the last glaciation

Aim: We used a combination of modelling and genetic approaches to investigate whether Pinguicula grandiflora and Saxifraga spathularis, two species that exhibit disjunct Lusitanian distributions, may have persisted through the Last Glacial Maximum (LGM, c. 21 ka) in separate northern and southern refugia.

Location: Northern and eastern Spain and south-western Ireland.

Methods: Palaeodistribution modelling using maxent was used to identify putative refugial areas for both species at the LGM, as well as to estimate their distributions during the Last Interglacial (LIG, c. 120 ka). Phylogeographical analysis of samples from across both species' ranges was carried out using one chloroplast and three nuclear loci for each species.

Results: The palaeodistribution models identified very limited suitable habitat for either species during the LIG, followed by expansion during the LGM. A single, large refugium across northern Spain and southern France was postulated for P. grandiflora. Two suitable regions were identified for S. spathularis: one in northern Spain, corresponding to the eastern part of the species' present-day distribution in Iberia, and the other on the continental shelf off the west coast of Brittany, south of the limit of the British–Irish ice sheet. Phylogeographical analyses indicated extremely reduced levels of genetic diversity in Irish populations of P. grandiflora relative to those in mainland Europe, but comparable levels of diversity between Irish and mainland European populations of S. spathularis, including the occurrence of private hapotypes in both regions.

Main conclusions: Modelling and phylogeographical analyses indicate that P. grandiflora persisted through the LGM in a southern refugium, and achieved its current Irish distribution via northward dispersal after the retreat of the ice sheets. Although the results for S. spathularis are more equivocal, a similar recolonization scenario also seems the most likely explanation for the species' current distribution.

Origin of British and Irish mammals: disparate post-glacial colonisation and species introductions

Global climate changes during the Quaternary reveal much about broader evolutionary effects of environmental change. Detailed regional studies reveal how evolutionary lineages and novel communities and ecosystems, emerge through glacial bottlenecks or from refugia. There have been significant advances in benthic imaging and dating, particularly with respect to the movements of the British (Scottish) and Irish ice sheets and associated changes in sea level during and after the Last Glacial Maximum (LGM). Ireland has been isolated as an island for approximately twice as long as Britain with no evidence of any substantial, enduring land bridge between these islands after ca 15 kya. Recent biogeographical studies show that Britain's mammal community is akin to those of southern parts of Scandinavia, The Netherlands and Belgium, but the much lower mammal species richness of Ireland is unique and needs explanation. Here, we consider physiographic, archaeological, phylogeographical i.e. molecular genetic, and biological evidence comprising ecological, behavioural and morphological data, to review how mammal species recolonized western Europe after the LGM with emphasis on Britain and, in particular, Ireland. We focus on why these close neighbours had such different mammal fauna in the early Holocene, the stability of ecosystems after LGM subject to climate change and later species introductions.

There is general concordance of archaeological and molecular genetic evidence where data allow some insight into history after the LGM. Phylogeography reveals the process of recolonization, e.g. with respect to source of colonizers and anthropogenic influence, whilst archaeological data reveal timing more precisely through carbon dating and stratigraphy. More representative samples and improved calibration of the ‘molecular clock’ will lead to further insights with regards to the influence of successive glaciations. Species showing greatest morphological, behavioural and ecological divergence in Ireland in comparison to Britain and continental Europe, were also those which arrived in Ireland very early in the Holocene either with or without the assistance of people. Cold tolerant mammal species recolonized quickly after LGM but disappeared, potentially as a result of a short period of rapid warming. Other early arrivals were less cold tolerant and succumbed to the colder conditions during the Younger Dryas or shortly after the start of the Holocene (11.5 kya), or the area of suitable habitat was insufficient to sustain a viable population especially in larger species. Late Pleistocene mammals in Ireland were restricted to those able to colonize up to ca 15 kya, probably originating from adjacent areas of unglaciated Britain and land now below sea level, to the south and west (of Ireland). These few, early colonizers retain genetic diversity which dates from before the LGM. Late Pleistocene Ireland, therefore, had a much depleted complement of mammal species in comparison to Britain.

Mammal species, colonising predominantly from southeast and east Europe occupied west Europe only as far as Britain between ca 15 and 8 kya, were excluded from Ireland by the Irish and Celtic Seas. Smaller species in particular failed to colonise Ireland. Britain being isolated as an island from ca. 8 kya has similar species richness and composition to adjacent lowland areas of northwest continental Europe and its mammals almost all show strongest genetic affinity to populations in neighbouring continental Europe with a few retaining genotypes associated with earlier, western lineages.

The role of people in the deliberate introduction of mammal species and distinct genotypes is much more significant with regards to Ireland than Britain reflecting the larger species richness of the latter and its more enduring land link with continental Europe. The prime motivation of early people in moving mammals was likely to be resource driven but also potentially cultural; as elsewhere, people exploring uninhabited places introduced species for food and the materials they required to survive. It is possible that the process of introduction of mammals to Ireland commenced during the Mesolithic and accelerated with Neolithic people. Irish populations of these long established, introduced species show some unique genetic variation whilst retaining traces of their origins principally from Britain but in some cases, Scandinavia and Iberia. It is of particular interest that they may retain genetic forms now absent from their source populations. Further species introductions, during the Bronze and late Iron Ages, and Viking and Norman invasions, follow the same pattern but lack the time for genetic divergence from their source populations. Accidental introductions of commensal species show considerable genetic diversity based on numerous translocations along the eastern Atlantic coastline. More recent accidental and deliberate introductions are characterised by a lack of genetic diversity other than that explicable by more than one introduction.

The substantial advances in understanding the postglacial origins and genetic diversity of British and Irish mammals, the role of early people in species translocations, and determination of species that are more recently introduced, should inform policy decisions with regards to species and genetic conservation. Conservation should prioritise early, naturally recolonizing species and those brought in by early people reflecting their long association with these islands. These early arrivals in Britain and Ireland and associated islands show genetic diversity that may be of value in mitigating anthropogenic climate change across Europe. In contrast, more recent introductions are likely to disturb ecosystems greatly, lead to loss of diversity and should be controlled. This challenge is more severe in Ireland where the number and proportion of invasive species from the 19th century to the present has been greater than in Britain.

Pseudomonas aeruginosa exhibits frequent recombination, but only a limited association between genotype and ecological setting

Pseudomonas aeruginosa is an opportunistic pathogen and an important cause of infection, particularly amongst cystic fibrosis (CF) patients. While specific strains capable of patient-to-patient transmission are known, many infections appear to be caused by unique and unrelated strains. There is a need to understand the relationship between strains capable of colonising the CF lung and the broader set of P. aeruginosa isolates found in natural environments. Here we report the results of a multilocus sequence typing (MLST)-based study designed to understand the genetic diversity and population structure of an extensive regional sample of P. aeruginosa isolates from South East Queensland, Australia. The analysis is based on 501 P. aeruginosa isolates obtained from environmental, animal and human (CF and non-CF) sources with particular emphasis on isolates from the Lower Brisbane River and isolates from CF patients obtained from the same geographical region. Overall, MLST identified 274 different sequence types, of which 53 were shared between one or more ecological settings. Our analysis revealed a limited association between genotype and environment and evidence of frequent recombination. We also found that genetic diversity of P. aeruginosa in Queensland, Australia was indistinguishable from that of the global P. aeruginosa population. Several CF strains were encountered frequently in multiple ecological settings; however, the most frequently encountered CF strains were confined to CF patients. Overall, our data confirm a non-clonal epidemic structure and indicate that most CF strains are a random sample of the broader P. aeruginosa population. The increased abundance of some CF strains in different geographical regions is a likely product of chance colonisation events followed by adaptation to the CF lung and horizontal transmission among patients.

Unique mitochondrial DNA lineages in Irish stickleback populations: cryptic refugium or rapid recolonization?

Repeated recolonization of freshwater environments following Pleistocene glaciations has played a major role in the evolution and adaptation of anadromous taxa. Located at the western fringe of Europe, Ireland and Britain were likely recolonized rapidly by anadromous fishes from the North Atlantic following the last glacial maximum (LGM). While the presence of unique mitochondrial haplotypes in Ireland suggests that a cryptic northern refugium may have played a role in recolonization, no explicit test of this hypothesis has been conducted. The three-spined stickleback is native and ubiquitous to aquatic ecosystems throughout Ireland, making it an excellent model species with which to examine the biogeographical history of anadromous fishes in the region. We used mitochondrial and microsatellite markers to examine the presence of divergent evolutionary lineages and to assess broad-scale patterns of geographical clustering among postglacially isolated populations. Our results confirm that Ireland is a region of secondary contact for divergent mitochondrial lineages and that endemic haplotypes occur in populations in Central and Southern Ireland. To test whether a putative Irish lineage arose from a cryptic Irish refugium, we used approximate Bayesian computation (ABC). However, we found no support for this hypothesis. Instead, the Irish lineage likely diverged from the European lineage as a result of postglacial isolation of freshwater populations by rising sea levels. These findings emphasize the need to rigorously test biogeographical hypothesis and contribute further evidence that postglacial processes may have shaped genetic diversity in temperate fauna.

The heart of a dragon: 3D anatomical reconstruction of the 'scaly-foot gastropod' (Mollusca: Gastropoda: Neomphalina) reveals its extraordinary circulatory system

Introduction: The 'scaly-foot gastropod' (Chrysomallon squamiferum Chen et al., 2015) from deep-sea hydrothermal vent ecosystems of the Indian Ocean is an active mobile gastropod occurring in locally high densities, and it is distinctive for the dermal scales covering the exterior surface of its foot. These iron-sulfide coated sclerites, and its nutritional dependence on endosymbiotic bacteria, are both noted as adaptations to the extreme environment in the flow of hydrogen sulfide. We present evidence for other adaptations of the 'scaly-foot gastropod' to life in an extreme environment, investigated through dissection and 3D tomographic reconstruction of the internal anatomy.

Results: Our anatomical investigations of juvenile and adult specimens reveal a large unganglionated nervous system, a simple and reduced digestive system, and that the animal is a simultaneous hermaphrodite. We show that Chrysomallon squamiferum relies on endosymbiotic bacteria throughout post-larval life. Of particular interest is the circulatory system: Chrysomallon has a very large ctenidium supported by extensive blood sinuses filled with haemocoel. The ctenidium provides oxygen for the host but the circulatory system is enlarged beyond the scope of other similar vent gastropods. At the posterior of the ctenidium is a remarkably large and well-developed heart. Based on the volume of the auricle and ventricle, the heart complex represents approximately 4 % of the body volume. This proportionally giant heart primarily sucks blood through the ctenidium and supplies the highly vascularised oesophageal gland. Thus we infer the elaborate cardiovascular system most likely evolved to oxygenate the endosymbionts in an oxygen poor environment and/or to supply hydrogen sulfide to the endosymbionts.

Conclusions: This study exemplifies how understanding the autecology of an organism can be enhanced by detailed investigation of internal anatomy. This gastropod is a large and active species that is abundant in its hydrothermal vent field ecosystem. Yet all of its remarkable features-protective dermal sclerites, circulatory system, high fecundity-can be viewed as adaptations beneficial to its endosymbiont microbes. We interpret these results to show that, as a result of specialisation to resolve energetic needs in an extreme chemosynthetic environment, this dramatic dragon-like species has become a carrying vessel for its bacteria.

The paradox of invasion: Reeves’ muntjac deer invade the British Isles from a limited number of founding females

High levels of genetic diversity and high propagule pressure are favoured by conservation biologists as the basis for successful reintroductions and ensuring the persistence of populations. However, invasion ecologists recognize the ‘paradox of invasion’, as successful species introductions may often be characterized by limited numbers of individuals and associated genetic bottlenecks. In the present study, we used a combination of high-resolution nuclear and mitochondrial genetic markers to investigate the invasion history of Reeves' muntjac deer in the British Isles. This invasion has caused severe economic and ecological damage, with secondary spread currently a concern throughout Europe and potentially globally. Microsatellite analysis based on eight loci grouped all 176 introduced individuals studied from across the species' range in the UK into one genetic cluster, and seven mitochondrial D-loop haplotypes were recovered, two of which were present at very low frequency and were related to more common haplotypes. Our results indicate that the entire invasion can be traced to a single founding event involving a low number of females. These findings highlight the fact that even small releases of species may, if ignored, result in irreversible and costly invasion, regardless of initial genetic diversity or continual genetic influx.

A genomic island linked to ecotype divergence in Atlantic cod

The genomic architecture underlying ecological divergence and ecological speciation with gene flow is still largely unknown for most organisms. One central question is whether divergence is genome-wide or localized in 'genomic mosaics' during early stages when gene flow is still pronounced. Empirical work has so far been limited, and the relative impacts of gene flow and natural selection on genomic patterns have not been fully explored. Here, we use ecotypes of Atlantic cod to investigate genomic patterns of diversity and population differentiation in a natural system characterized by high gene flow and large effective population sizes, properties which theoretically could restrict divergence in local genomic regions. We identify a genomic region of strong population differentiation, extending over approximately 20 cM, between pairs of migratory and stationary ecotypes examined at two different localities. Furthermore, the region is characterized by markedly reduced levels of genetic diversity in migratory ecotype samples. The results highlight the genomic region, or 'genomic island', as potentially associated with ecological divergence and suggest the involvement of a selective sweep. Finally, we also confirm earlier findings of localized genomic differentiation in three other linkage groups associated with divergence among eastern Atlantic populations. Thus, although the underlying mechanisms are still unknown, the results suggest that 'genomic mosaics' of differentiation may even be found under high levels of gene flow and that marine fishes may provide insightful model systems for studying and identifying initial targets of selection during ecological divergence.

Novel polymorphic microsatellite loci for the protogynous hermaphrodite slinger sea bream (Chrysoblephus puniceus, Sparidae)

As a result of the global decline of fish stocks, an increasing number of fish species are becoming targets of heavy exploitation, often concomitantly with a lack of biological knowledge on their structure and demographics. Here we present 11 new polymorphic microsatellite loci, isolated from the slinger sea bream (Chrysoblephus puniceus, Sparidae), a relatively recent target of coastal fisheries in eastern South Africa. Levels of genetic diversity were assessed in 39 individuals collected from the KwaZulu-Natal coast (Park Rynie, South Africa). Observed and expected heterozygosities varied between 0.39 and 0.97 and between 0.53 and 0.96, respectively. One locus (SL35) showed significant heterozygote deficiency and linkage disequilibrium was detected between SL35 and SL1. Importantly, five of these microsatellites cross-amplify in Cheimerius nufar, a sympatric species also subjected to exploitation.

New microsatellite loci for the longnose velvet dogfish Centroselachus crepidater (Squaliformes: Somniosidae) and other deep sea sharks

The continuing over-exploitation of traditional coastal stocks has resulted in the shift of commercial fishing towards deep-sea ecosystems in many parts of the world. The effects on target and non-target species have been dramatic; particularly for the deep-sea sharks. With the aim of providing tools that will allow the assessment of population genetic structure of Centroselachus crepidater, novel microsatellite loci have been developed for this deep-sea elasmobranch. Seven of these markers showed between 3 and 7 alleles per locus in two North Atlantic populations, with observed and expected heterozygosities between 0.18-0.95 and 0.25-0.82, respectively. Additionally, ten loci cross-amplify in other Elasmobranch species.