Contrasted influences of moon phases on the reproduction and movement patterns of four amphibian species inhabiting different habitats in central Italy

Many studies have provided evidence that prey adjust their behaviour to adaptively balance the fitness effects of reproduction and predation risk. Nocturnal terrestrial animals should deal with a range of environmental conditions during the reproductive season at the breeding sites, including a variable amount of natural ambient light. High degrees of illumination are expected to minimize those behaviours that might increase the animal detection by predators. Therefore, under habitat variable brightness conditions and in different ecosystems, the above mentioned behaviours are expected to depend on the variation in predation risk. Although moon effects on amphibian biology have been recognized, the direction of this influence is rather controversial with evidences of both increased and depressed activity under full moon. We tested in four nocturnal amphibian species (Hyla intermedia, Rana dalmatina, Rana italica, Salamandrina perspicillata) the effects of different (i) light conditions and (ii) habitats (open land vs. dense forest) on the reproductive phenology. Our results showed that the effects of the lunar cycle on the study species are associated with the change in luminosity, and there is no evidence of an endogenous rhythm controlled by biological clocks. The habitat type conditioned the amphibian reproductive strategy in relation to moon phases. Open habitat breeders (e. g., ponds with no canopy cover) strongly avoided conditions with high brightness, whereas forest habitat breeders were apparently unaffected by the different moon phases. Indeed, for all the studied species no effects of the moon phase itself on the considered metrics were found. Rather, the considered amphibian species seem to be conditioned mainly by moonlight irrespective of the moon phase. The two anurans spawning in open habitat apparently adjust their oviposition timing by balancing the fitness effects of the risk to be detected by predators and the reproduction.

Inferring the degree of incipient speciation in secondary contact zones of closely related lineages of Palearctic green toads (Bufo viridis subgroup).

Reproductive isolation between lineages is expected to accumulate with divergence time, but the time taken to speciate may strongly vary between different groups of organisms. In anuran amphibians, laboratory crosses can still produce viable hybrid offspring >20 My after separation, but the speed of speciation in closely related anuran lineages under natural conditions is poorly studied. Palearctic green toads (Bufo viridis subgroup) offer an excellent system to address this question, comprising several lineages that arose at different times and form secondary contact zones. Using mitochondrial and nuclear markers, we previously demonstrated that in Sicily, B. siculus and B. balearicus developed advanced reproductive isolation after Plio-Pleistocene divergence (2.6 My, 3.3-1.9), with limited historic mtDNA introgression, scarce nuclear admixture, but low, if any, current gene flow. Here, we study genetic interactions between younger lineages of early Pleistocene divergence (1.9 My, 2.5-1.3) in northeastern Italy (B. balearicus, B. viridis). We find significantly more, asymmetric nuclear and wider, differential mtDNA introgression. The population structure seems to be molded by geographic distance and barriers (rivers), much more than by intrinsic genomic incompatibilities. These differences of hybridization between zones may be partly explained by differences in the duration of previous isolation. Scattered research on other anurans suggests that wide hybrid zones with strong introgression may develop when secondary contacts occur <2 My after divergence, whereas narrower zones with restricted gene flow form when divergence exceeds 3 My. Our study strengthens support for this rule of thumb by comparing lineages with different divergence times within the same radiation.

Evolution of Y chromosomes : lessons from mammals and amphibians

RÉSUMÉ : Le sexe des individus peut être déterminé par l'environnement ou la génétique. Lorsque la détermination du sexe est génétique, il y a dans le génome, la présence de chromosomes spécifiques qui détermineront le sexe. Dans cette thèse, j'ai étudié l'évolution des chromosomes sexuels et dans quel contexte des marqueurs sur ces chromosomes peuvent être utilisés. Pour explorer la formation du chromosome Y, nous avons étudié les caractéristiques des chromosomes sexuels chez la rainette verte, Hyla arborea. Dans un premier temps, nous avons utilisé un marqueur situé sur les chromosomes sexuels X et Y chez plusieurs espèces appartenant au groupe de la rainette verte. Cela nous a permis de révéler chez toutes ces espèces une hétérogamétie mâle. Dans un deuxième temps, nous avons tiré profit de deux autres marqueurs situés sur les chromosomes sexuels pour montrer que la recombinaison est supprimée chez les mâles mais pas chez les femelles. Pour expliquer la réduction de la variabilité sur le chromosome Y, il n'est pas nécessaire d'invoquer le balayage sélectif ou la sélection d'arrière-plan : le nombre de copies plus petit du chromosome Y dans le génome et l'absence de recombinaison suffisent à l'expliquer. Nous avons également analysé plus en détail la suppression de la recombinaison chez les mâles de H. arborea. Les modèles classiques de l'évolution des chromosomes sexuels supposent que la taille de la région non-recombinante augmente progressivement pendant l'évolution du chromosome Y, due à l'accumulation de changements structuraux. Dans cette étude, nous montrons un modèle différent, à savoir que la recombinaison est supprimée ou diminuée non seulement sur les chromosomes sexuels mais aussi sur les autosomes chez les mâles, dû à l'action de modificateurs généraux. En utilisant des marqueurs localisés sur le chromosome Y, ainsi que sur l'ADN mitochondrial et le chromosome X, nous avons étudié l'histoire évolutive de la musaraigne musette, Crocidura russula. Cette étude illustre que les analyses génétiques avec plusieurs types de marqueurs génétiques peuvent faciliter l'interprétation de l'histoire évolutive des espèces, mais que l'utilisation des marqueurs sur les chromosomes X et Y pour des études phylogéographiques est limitée par le peu de polymorphisme observé sur ces deux types de marqueurs. Le même jeu de données combiné avec des simulations a été employé pour comprendre les facteurs responsables de la faible variabilité sur le chromosome Y qui peut être expliqué, dans notre étude, par la démographie et les traits d'histoire de vie de C. russula. SUMMARY The sex of an individual is determined either by its environment or its genetics. Genetic sex determination relies on the presence of specific chromosomes that will determine the sex of their bearer. In this thesis, I studied the evolution of the sex chromosomes and the context in which markers on this type of chromosomes can be used. To explore the evolution of a Y chromosome, we studied the nascent sex chromosomes in the European tree frog Hyla arborea. First; we amplified a sex specific marker in several related species of European tree frog and found a homogeneous pattern of male heterogamety. Secondly, we used two additional sex-specific markers to show that recombination is suppressed in males but not in females. There is, therefore, no need to invoke background selection or selective sweeps to explain the reduced genetic variability on the Y chromosome, because the lower number of copies of the Y chromosomes per breeding pair and the absence of recombination are sufficient. To further analyze the suppression of recombination in male European. tree frogs, we constructed a microsatellite linkage map for this species. Classical models of sex-chromosome evolution assume that the non-recombining region expands progressively during the long-term evolution of the Y chromosome, owing to the accumulation of structural changes. Here we show a strikingly different pattern: recombination is suppressed or depressed both on sex chromosomes and autosomes in the heterogametic sex, presumably due to the action of general modifiers. We investigated the evolutionary history of the greater white-toothed shrew, Crocidura russula, using markers on both sex chromosomes and mtDNA. This study illustrates that multilocus genetic analyses facilitates the interpretation of a species' evolutionary history. It also demonstrates that phylogeographic inferences from X and Y chromosomal markers are restricted by the low levels of observed polymorphism. Combining this genetic study with simulations, we determined that the demography and the life-history traits of this species can alone be responsible for the low Y diversity. In conclusion, this thesis shows that sex chromosomes, in combination with autosomes or mtDNA, are necessary to understand the evolution of sex chromosomes and to precisely infer the population history of a species.

Integrating detection probabilities in species distribution models of amphibians breeding in Mediterranean temporary ponds

Aim  The imperfect detection of species may lead to erroneous conclusions about species-environment relationships. Accuracy in species detection usually requires temporal replication at sampling sites, a time-consuming and costly monitoring scheme. Here, we applied a lower-cost alternative based on a double-sampling approach to incorporate the reliability of species detection into regression-based species distribution modelling.Location  Doñana National Park (south-western Spain).Methods  Using species-specific monthly detection probabilities, we estimated the detection reliability as the probability of having detected the species given the species-specific survey time. Such reliability estimates were used to account explicitly for data uncertainty by weighting each absence. We illustrated how this novel framework can be used to evaluate four competing hypotheses as to what constitutes primary environmental control of amphibian distribution: breeding habitat, aestivating habitat, spatial distribution of surrounding habitats and/or major ecosystems zonation. The study was conducted on six pond-breeding amphibian species during a 4-year period.Results  Non-detections should not be considered equivalent to real absences, as their reliability varied considerably. The occurrence of Hyla meridionalis and Triturus pygmaeus was related to a particular major ecosystem of the study area, where suitable habitat for these species seemed to be widely available. Characteristics of the breeding habitat (area and hydroperiod) were of high importance for the occurrence of Pelobates cultripes and Pleurodeles waltl. Terrestrial characteristics were the most important predictors of the occurrence of Discoglossus galganoi and Lissotriton boscai, along with spatial distribution of breeding habitats for the last species.Main conclusions  We did not find a single best supported hypothesis valid for all species, which stresses the importance of multiscale and multifactor approaches. More importantly, this study shows that estimating the reliability of non-detection records, an exercise that had been previously seen as a naïve goal in species distribution modelling, is feasible and could be promoted in future studies, at least in comparable systems.

Amphibians in the diet of European Barn Owls

We present a review of the propensity to eat amphibians in the Barn Owl Tyto alba in Europe. Based on the analysis of 596 published studies reporting 3.32 million prey items identified in pellets, 17 869 amphibians (0.54%) were found. An analysis of 9036 amphibians identified to the species level showed that Barn Owls avoid consuming toxic species, and they are able to capture tree frogs (Hylidae) only rarely. The true frogs (Ranidae) are by far the most frequently captured amphibians followed by spadefoot toads (Pelobatidae) and Parsley frogs (Pelodytidae).

Iowa's frog and toad survey, 1995-2003.

This survey began in response to widespread interest of declines in amphibians. More recently, a comprehensive statewide planning group discovered 44% of Iowa’s herpetofauna (amphibians and reptiles) to be of special concern. In response to these concerns, the Iowa Department of Natural Resources Wildlife Diversity Program (WDP) initiated an auditory survey for calling anurans to determine geographic distributions within the state. This survey has established itself as an extensive, long term monitoring program. This 2005 report is the second edition since the first report of this survey was shared in 1998 by then program biologist Lisa Hemesath. The goals of the survey are to: (1) determine the distributions of Iowa’s anuran species, (2) determine population trends for each species, and (3) promote education about aquatic life by using volunteers to conduct the survey. In addition to Iowa, volunteer-based auditory surveys for frogs and toads are currently being used in the Midwest by Wisconsin, Minnesota, Missouri, and Illinois.

Buccal swabs allow efficient and reliable microsatellite genotyping in amphibians

Buccal swabs have recently been used as a minimally invasive sampling method in genetic studies of wild populations, including amphibian species. Yet it is not known to date what is the level of reliability for microsatellite genotypes obtained using such samples. Allelic dropout and false alleles may affect the genotyping derived from buccal samples. Here we quantified the success of microsatellite amplification and the rates of genotyping errors using buccal swabs in two amphibian species, the Alpine newt Triturus alpestris and the Green tree frog Hyla arborea, and we estimated two important parameters for downstream analyses, namely the number of repetitions required to achieve typing reliability and the probability of identity among genotypes. Amplification success was high, and only one locus tested required two to three repetitions to achieve reliable genotypes, showing that buccal swabbing is a very efficient approach allowing good quality DNA retrieval. This sampling method which allows avoiding the controversial toe-clipping will likely prove very useful in the context of amphibian conservation.

Effect of biogeographic history on population vulnerability in European amphibians.

The genetic diversity of populations, which contributes greatly to their adaptive potential, is negatively affected by anthropogenic habitat fragmentation and destruction. However, continental-scale losses of genetic diversity also resulted from the population expansions that followed the end of the last glaciation, an element that is rarely considered in a conservation context. We addressed this issue in a meta-analysis in which we compared the spatial patterns of vulnerability of 18 widespread European amphibians in light of phylogeographic histories (glacial refugia and postglacial routes) and anthropogenic disturbances. Conservation statuses significantly worsened with distances from refugia, particularly in the context of industrial agriculture; human population density also had a negative effect. These findings suggest that features associated with the loss of genetic diversity in post-glacial amphibian populations (such as enhanced fixation load or depressed adaptive potential) may increase their susceptibility to current threats (e.g., habitat fragmentation and pesticide use). We propose that the phylogeographic status of populations (i.e., refugial vs. post-glacial) should be considered in conservation assessments for regional and national red lists.

The efficiency of indicator groups for the conservation of amphibians in the Brazilian Atlantic Forest

The adequate selection of indicator groups of biodiversity is an important aspect of the systematic conservation planning. However, these assessments differ in the spatial scale, in the methods used and in the groups considered to accomplish this task, which generally produces contradictory results. The quantification of the spatial congruence between species richness and complementarity among different taxonomic groups is a fundamental step to identify potential indicator groups. Using a constructive approach, the main purposes of this study were to evaluate the performance and efficiency of eight potential indicator groups representing amphibian diversity in the Brazilian Atlantic Forest. Data on the geographic range of amphibian species that occur in the Brazilian Atlantic Forest was overlapped to the full geographic extent of the biome, which was divided into a regular equal-area grid. Optimization routines based on the concept of complementarily were applied to verify the performance of each indicator group selected in relation to the representativeness of the amphibians in the Brazilian Atlantic Forest as a whole, which were solved by the algorithm"simulated annealing", through the use of the software MARXAN. Some indicator groups were substantially more effective than others in regards to the representation of the taxonomic groups assessed, which was confirmed by the high significance of data (F = 312.76; p < 0.01). Leiuperidae was considered as the best indicator group among the families analyzed, as it showed a good performance, representing 71% of amphibian species in the Brazilian Atlantic Forest (i.e. 290 species), which may be associated with the diffuse geographic distribution of its species. This study promotes understanding of how the diversity standards of amphibians can be informative for systematic conservation planning on a regional scale.

Control of respiration in fish, amphibians and reptiles

Fish and amphibians utilise a suction/force pump to ventilate gills or lungs, with the respiratory muscles innervated by cranial nerves, while reptiles have a thoracic, aspiratory pump innervated by spinal nerves. However, fish can recruit a hypobranchial pump for active jaw occlusion during hypoxia, using feeding muscles innervated by anterior spinal nerves. This same pump is used to ventilate the air-breathing organ in air-breathing fishes. Some reptiles retain a buccal force pump for use during hypoxia or exercise. All vertebrates have respiratory rhythm generators (RRG) located in the brainstem. In cyclostomes and possibly jawed fishes, this may comprise elements of the trigeminal nucleus, though in the latter group RRG neurons have been located in the reticular formation. In air-breathing fishes and amphibians, there may be separate RRG for gill and lung ventilation. There is some evidence for multiple RRG in reptiles. Both amphibians and reptiles show episodic breathing patterns that may be centrally generated, though they do respond to changes in oxygen supply. Fish and larval amphibians have chemoreceptors sensitive to oxygen partial pressure located on the gills. Hypoxia induces increased ventilation and a reflex bradycardia and may trigger aquatic surface respiration or air-breathing, though these latter activities also respond to behavioural cues. Adult amphibians and reptiles have peripheral chemoreceptors located on the carotid arteries and central chemoreceptors sensitive to blood carbon dioxide levels. Lung perfusion may be regulated by cardiac shunting and lung ventilation stimulates lung stretch receptors.

Autonomic control of cardiorespiratory interactions in fish, amphibians and reptiles

Control of the heart rate and cardiorespiratory interactions (CRI) is predominantly parasympathetic in all jawed vertebrates, with the sympathetic nervous system having some influence in tetrapods. Respiratory sinus arrhythmia (RSA) has been described as a solely mammalian phenomenon but respiration-related beat-to-beat control of the heart has been described in fish and reptiles. Though they are both important, the relative roles of feed-forward central control and peripheral reflexes in generating CRI vary between groups of fishes and probably between other vertebrates. CRI may relate to two locations for the vagal preganglionic neurons (VPN) and in particular cardiac VPN in the brainstem. This has been described in representatives from all vertebrate groups, though the proportion in each location is variable. Air-breathing fishes, amphibians and reptiles breathe discontinuously and the onset of a bout of breathing is characteristically accompanied by an immediate increase in heart rate plus, in the latter two groups, a left-right shunting of blood through the pulmonary circuit. Both the increase in heart rate and opening of a sphincter on the pulmonary artery are due to withdrawal of vagal tone. An increase in heart rate following a meal in snakes is related to withdrawal of vagal tone plus a non-adrenergic-non-cholinergic effect that may be due to humoral factors released by the gut. Histamine is one candidate for this role.

Habitat Split as a Cause of Local Population Declines of Amphibians with Aquatic Larvae

Most amphibian species have biphasic life histories and undergo an ontogenetic shift from aquatic to terrestrial habitats. In deforested landscapes, streams and forest fragments are frequently disjunct, jeopardizing the life cycle of forest-associated amphibians with aquatic larvae. We tested the impact of habitat split-defined as human-induced disconnection between habitats used by different life-history stages of a species-on four forest-associated amphibian species in a severely fragmented landscape of the Brazilian Atlantic Forest. We surveyed amphibians in forest fragments with and without streams (referred to as wet and dry fragments, respectively), including the adjacent grass-field matrix. Our comparison of capture rates in dry fragments and nearby streams in the matrix allowed us to evaluate the number of individuals that engaged in high-risk migrations through nonforested habitats. Adult amphibians moved from dry fragments to matrix streams at the beginning of the rainy season, reproduced, and returned at the end of the breeding period. Juveniles of the year moved to dry fragments along with adults. These risky reproductive migrations through nonforested habitats that expose individuals to dehydration, predation, and other hazards may cause population declines in dry fragments. Indeed, capture rates were significantly lower in dry fragments compared with wet fragments. Declining amphibians would strongly benefit from investments in the conservation and restoration of riparian vegetation and corridors linking breeding and nonbreeding areas.

Cutaneous Resistance to Evaporative Water Loss in Brazilian Rhinella (Anura: Bufonidae) from Contrasting Environments

Influenced by taxonomic position. For example, bufonids are regarded as exhibiting a permeable skin that seems typical for terrestrial anurans. However, this assumption is supported by information on only four bufonid species; therefore, the enormous ecological diversity of the family remains poorly Investigated. To assess whether variation in R(s) within related bufonids correlates with environmental aridity, we measured area-specific rates of EWL of two Brazilian populations of Rhinella granulosa (previously Bufo granulosus), one from the Atlantic Forest and other from the semi-arid Caatinga, and compared both with the forest species R. ornato. Rhinella granulosa from the Atlantic Forest had higher cutaneous resistance than conspecifics from Caatinga and R. ornata. Rhinella ornato presented the lowest cutaneous resistance values. However, Rs were very close to zero In all three populations. We conclude that enhanced Rs is not part of the suite of traits allowing R. granulosa to exploit the Caatinga, and that variation in R(s) within bufonids may relate to traits other than water conservation. Some Information on microhabitat occupation and ventral skin morphology supports the idea that exceptional abilities for detecting and taking up water may be the key factors enhancing the survival of R. granulosa, and possibly other bufonids, in xeric environments.

Seasonal metabolic changes in a year-round reproductively active subtropical tree-frog (Hypsiboas prasinus)

Although seasonal metabolic variation in ectothermic tetrapods has been investigated primarily in the context of species showing some level of metabolic depression during winter, but several species of anurans maintain their activity patterns throughout the year in tropical and subtropical areas. The tree-frog Hypsiboas prasinus occurs in the subtropical Atlantic Forest and remains reproductively active during winter, at temperatures below 10 degrees C. We compared males calling in summer and winter, and found that males of H. prasinus exhibit seasonal adjustments in metabolic and morphometric variables. Individuals calling during winter were larger and showed higher resting metabolic rates than those calling during summer. Calling rates were not affected by season. Winter animals showed lower liver and heart activity level of citrate synthase (CS), partially compensated by larger liver mass. Winter individuals also showed higher activity Of pyruvate kinase (PK) and lower activity of CS in trunk muscles, and higher activity of CS in leg muscles. Winter metabolic adjustments seem to be achieved by both compensatory mechanisms to the lower environmental temperature and a seasonally oriented aerobic depression of several organs. The impact of seasonal metabolic changes on calling performance and the capacity of subtropical anurans for metabolic thermal acclimatization are also discussed. (C) 2008 Elsevier Inc. All rights reserved.