Drift rather than selection dominates MHC class II allelic diversity patterns at the biogeographical range scale in natterjack toads Bufo calamita.

Study of major histocompatibility complex (MHC) loci has gained great popularity in recent years, partly due to their function in protecting vertebrates from infections. This is of particular interest in amphibians on account of major threats many species face from emergent diseases such as chytridiomycosis. In this study we compare levels of diversity in an expressed MHC class II locus with neutral genetic diversity at microsatellite loci in natterjack toad (Bufo (Epidalea) calamita) populations across the whole of the species' biogeographical range. Variation at both classes of loci was high in the glacial refugium areas (REF) and much lower in postglacial expansion areas (PGE), especially in range edge populations. Although there was clear evidence that the MHC locus was influenced by positive selection in the past, congruence with the neutral markers suggested that historical demographic events were the main force shaping MHC variation in the PGE area. Both neutral and adaptive genetic variation declined with distance from glacial refugia. Nevertheless, there were also some indications from differential isolation by distance and allele abundance patterns that weak effects of selection have been superimposed on the main drift effect in the PGE zone.

Effects of road mortality and mitigation measures on amphibian populations.

Road mortality is a widely recognized but rarely quantified threat to the viability of amphibian populations. The global extent of the problem is substantial and factors affecting the number of animals killed on highways include life-history traits and landscape features. Secondary effects include genetic isolation due to roads acting as barriers to migration. Long-term effects of roads on population dynamics are often severe and mitigation methods include volunteer rescues and under-road tunnels. Despite the development of methods that reduce road kill in specific locations, especially under-road tunnels and culverts, there is scant evidence that such measures will protect populations over the long term. There also seems little likelihood that funding will be forthcoming to ameliorate the problem at the scale necessary to prevent further population declines.

When is a species declining? Optimizing survey effort to detect population changes in reptiles.

Biodiversity monitoring programs need to be designed so that population changes can be detected reliably. This can be problematical for species that are cryptic and have imperfect detection. We used occupancy modeling and power analysis to optimize the survey design for reptile monitoring programs in the UK. Surveys were carried out six times a year in 2009-2010 at multiple sites. Four out of the six species--grass snake, adder, common lizard, slow-worm -were encountered during every survey from March-September. The exceptions were the two rarest species--sand lizard and smooth snake--which were not encountered in July 2009 and March 2010 respectively. The most frequently encountered and most easily detected species was the slow-worm. For the four widespread reptile species in the UK, three to four survey visits that used a combination of directed transect walks and artificial cover objects resulted in 95% certainty that a species would be detected if present. Using artificial cover objects was an effective detection method for most species, considerably increased the detection rate of some, and reduced misidentifications. To achieve an 85% power to detect a decline in any of the four widespread species when the true decline is 15%, three surveys at a total of 886 sampling sites, or four surveys at a total of 688 sites would be required. The sampling effort needed reduces to 212 sites surveyed three times, or 167 sites surveyed four times, if the target is to detect a true decline of 30% with the same power. The results obtained can be used to refine reptile survey protocols in the UK and elsewhere. On a wider scale, the occupancy study design approach can be used to optimize survey effort and help set targets for conservation outcomes for regional or national biodiversity assessments.

Lusitania revisited: a phylogeographic analysis of the natterjack toad Bufo calamita across its entire biogeographical range.

Attempts to understand the current distributions of plants and animals require both historical and ecological information. Phylogeography has proved highly effective in elucidating historical events such as postglacial colonisations in north temperate zones. However, interesting questions still await resolution. Lusitanian distributions of fauna and flora in western Europe, for example, have puzzled biogeographers for more than 150 years. Lusitanian species have highly disjunct distributions in Ireland and in Iberia, often with few or no other populations inbetween. Despite much debate, no agreed explanation for Lusitanian distributions has yet emerged. We investigated the phylogeographic structure of one Lusitanian species, the natterjack toad Bufo calamita, using mitochondrial DNA control region sequences and allelic variation at eight microsatellite loci. Our results show that this amphibian must have survived in north European refugia, as well as in Iberia, during and since the last (Weichselian) glacial maximum around 20,000 years before present (BP). Subsequent local recolonisation after the Younger Dryas cooling around 11,000 years BP best explains the Lusitanian aspect of natterjack toad distribution.

Population genetics of a successful invader: the marsh frog Rana ridibunda in Britain.

We investigated the genetic outcome of successful invasion by an alien species, the marsh frog Rana ridibunda, in Britain. Twelve adults translocated from Hungary into Kent (Romney) in 1935 resulted rapidly in a large localized population. A further successful translocation in 1973 from Romney to Sussex (Lewes), together with other range extensions, provided an opportunity to test bottleneck effects during colonization events. Romney and Lewes frogs had similar genetic diversities to those in Hungary at 14 random amplified polymorphic DNA marker (RAPD) and five microsatellite loci. The introduced populations were, however, differentiated genetically from each other and from a reference population in Hungary. Fitness assessments (larval growth and survival) revealed no differences between the Lewes and Romney populations. Despite starting with few founders, significant bottleneck effects on R. ridibunda in Britain were therefore undetectable, presumably because population expansions were rapid immediately after the translocations.

Population on the verge of a mutational meltdown? Fitness costs of genetic load for an amphibian in the wild.

The fitness costs of high genetic load in wild populations have rarely been assessed under natural conditions. Such costs are expected to be greatest in small, bottlenecked populations, including those occurring near range edges. Britain is at the northwesterly range limit of the natterjack toad Bufo calamita. We compared fitness attributes in two populations of this amphibian with very different recent histories. Key larval fitness attributes in B. calamita, notably growth rate and metamorph production, were substantially higher in the large outbreeding population (Ainsdale) than in the small and isolated one (Saltfleetby). These differences were manifest under seminatural conditions, when larvae were reared in mesh cages within breeding ponds at the site of the small population, and were exacerbated by high stress treatments. The results indicate that genetic load effects can be sufficiently severe enough to predispose extinction over relatively short time frames, as predicted by extinction vortex models.

Ecological responses to recent climate change.

There is now ample evidence of the ecological impacts of recent climate change, from polar terrestrial to tropical marine environments. The responses of both flora and fauna span an array of ecosystems and organizational hierarchies, from the species to the community levels. Despite continued uncertainty as to community and ecosystem trajectories under global change, our review exposes a coherent pattern of ecological change across systems. Although we are only at an early stage in the projected trends of global warming, ecological responses to recent climate change are already clearly visible.

Prototheca richardsi, a pathogen of anuran larvae, is related to a clade of protistan parasites near the animal-fungal divergence.

Prototheca richardsi is a protist of uncertain taxonomy which mediates growth inhibition in anuran larvae. Cells of P. richardsi were isolated from tadpole faeces and DNA was purified by Qiagen chromatography. Nuclear small-subunit (18S) rDNA (ssu-rDNA) was amplified by PCR using universal primers, cloned, and six clones (two from each of three separate isolates) were sequenced. All clones yielded an essentially identical sequence of 1802 nucleotides. In situ hybridization of fluorescent Prototheca-specific oligonucleotide probes, designed using the derived 18S rDNA sequence, confirmed that the sequence was indeed from P. richardsi cells and not from other components of tadpole faeces. The P. richardsi sequence was aligned with ssu-rDNA from a range of other eukaryotes, and phylogenetic analyses were carried out using several inference methods. P. richardsi consistently and stably grouped within a novel clade that contains rDNAs from an apparently heterodisperse group of parasitic micro-organisms assigned to the class Ichthyosporea. P. richardsi is evidently misplaced in the genus Prototheca, and the authors propose its inclusion in a new genus Anurofeca.

The basis of niche separation during terrestrial life between two species of toad (Bufo bufo and Bufo calamita): competition or specialisation?

Two congeneric anurans with highly overlapping geographical ranges, Bufo bufo and B. calamita, were investigated with respect to isolating mechanisms during the terrestrial phase of life in the three habitat types in which both species occur in Britain. Marked spatial niche separations were observed in all three habitats (coastal dunes, upper saltmarshes and lowland heaths). B. bufo was associated with complex, dense vegetation structures and had a relatively wide niche, while B. calamita had a narrower niche and occurred mostly on open, poorly-vegetated ground. Food niche overlap was high at all three study sites. Animals translocated into atypical habitats returned to their preferred type whenever possible, and suffered increased mortality or loss of condition if prevented from doing so. B. calamita operated at significantly higher body temperatures (by an average of 1.4° C) than B. bufo when hunting at night. Experiments in outdoor vivaria indicated that B. calamita survived in open habitats because individuals escaped desiccation in daytime by burrowing into the substrate, whereas B. bufo lacked this behaviour and perished on the inhospitable surface. By contrast, B. calamita individuals lost mass and became hyperactive in densely-vegetated conditions because their hunting efficiency was selectively reduced relative to that of B. bufo. Taken together, the data indicated that spatial niche separation between these species during the terrestrial phase of life was dictated primarily by behavioural and physiological factors, and not by competition.