Within- and among-population impact of genetic erosion on adult fitness-related traits in the European tree frog Hyla arborea.

Assessing in wild populations how fitness is impacted by inbreeding and genetic drift is a major goal for conservation biology. An approach to measure the detrimental effects of inbreeding on fitness is to estimate correlations between molecular variation and phenotypic performances within and among populations. Our study investigated the effect of individual multilocus heterozygosity on body size, body condition and reproductive investment of males (that is, chorus attendance) and females (that is, clutch mass and egg size) in both small fragmented and large non-fragmented populations of European tree frog (Hyla arborea). Because adult size and/or condition and reproductive investment are usually related, genetic erosion may have detrimental effects directly on reproductive investment, and also on individual body size and condition that in turn may affect reproductive investment. We confirmed that the reproductive investment was highly size-dependent for both sexes. Larger females invested more in offspring production, and larger males attended the chorus in the pond more often. Our results did not provide evidence for a decline in body size, condition and reproductive effort with decreased multilocus heterozygosity both within and among populations. We showed that the lack of heterozygosity-fitness correlations within populations probably resulted from low inbreeding levels (inferior to ca. 20% full-sib mating rate), even in the small fragmented populations. The detrimental effects of fixation load were either low in adults or hidden by environmental variation among populations. These findings will be useful to design specific management actions to improve population persistence.

Timeframe of speciation inferred from secondary contact zones in the European tree frog radiation (Hyla arborea group).

BACKGROUND: Hybridization between incipient species is expected to become progressively limited as their genetic divergence increases and reproductive isolation proceeds. Amphibian radiations and their secondary contact zones are useful models to infer the timeframes of speciation, but empirical data from natural systems remains extremely scarce. Here we follow this approach in the European radiation of tree frogs (Hyla arborea group). We investigated a natural hybrid zone between two lineages (Hyla arborea and Hyla orientalis) of Mio-Pliocene divergence (~5 My) for comparison with other hybrid systems from this group. RESULTS: We found concordant geographic distributions of nuclear and mitochondrial gene pools, and replicated narrow transitions (~30 km) across two independent transects, indicating an advanced state of reproductive isolation and potential local barriers to dispersal. This result parallels the situation between H. arborea and H. intermedia, which share the same amount of divergence with H. orientalis. In contrast, younger lineages show much stronger admixture at secondary contacts. CONCLUSIONS: Our findings corroborate the negative relationship between hybridizability and divergence time in European tree frogs, where 5 My are necessary to achieve almost complete reproductive isolation. Speciation seems to progress homogeneously in this radiation, and might thus be driven by gradual genome-wide changes rather than single speciation genes. However, the timescale differs greatly from that of other well-studied amphibians. General assumptions on the time necessary for speciation based on evidence from unrelated taxa may thus be unreliable. In contrast, comparative hybrid zone analyses within single radiations such as our case study are useful to appreciate the advance of speciation in space and time.

Introgressive hybridization of threatened European tree frogs (Hyla arborea) by introduced H. intermedia in Western Switzerland

Hybridization by introduced taxa is a major threat to native species. Characterizing human introductions is thus one of the missions of conservation geneticists. Here we survey a declining population of the regionally endangered European tree frog (Hyla arborea) in the Grangettes natural reserve (Rhone valley, Western Switzerland), where previous evidence indicated human introduction of the Italian taxon H. intermedia. We combined fast-evolving mitochondrial and nuclear markers and an extended sampling to conduct population genetic analyses of the Grangettes and putative source areas. We show that the Grangettes population is a hybrid swarm, with all individuals featuring recent nuclear admixture and mitochondrial DNA of introduced H. intermedia, most likely of proximate south Alpine origin. In contrast, H. arborea and H. intermedia hardly introgress in their natural parapatric ranges, consistent with an advanced reproductive isolation. Thus, potential hybrid incompatibilities may account for the strong decline of this population, despite important conservation efforts. Although their hybrid nature makes them a priori unworthy of any protection, we propose specific measures to recover local H. arborea gene pool and preserve tree frogs in the Grangettes, the last population remaining from this heavily impacted part of the Alps.

High-density sex-specific linkage maps of a European tree frog (Hyla arborea) identify the sex chromosome without information on offspring sex.

Identifying homology between sex chromosomes of different species is essential to understanding the evolution of sex determination. Here, we show that the identity of a homomorphic sex chromosome pair can be established using a linkage map, without information on offspring sex. By comparing sex-specific maps of the European tree frog Hyla arborea, we find that the sex chromosome (linkage group 1) shows a threefold difference in marker number between the male and female maps. In contrast, the number of markers on each autosome is similar between the two maps. We also find strongly conserved synteny between H. arborea and Xenopus tropicalis across 200 million years of evolution, suggesting that the rate of chromosomal rearrangement in anurans is low. Finally, we show that recombination in males is greatly reduced at the centers of large chromosomes, consistent with previous cytogenetic findings. Our research shows the importance of high-density linkage maps for studies of recombination, chromosomal rearrangement and the genetic architecture of ecologically or economically important traits.

Unpalatability of Hyla semilineata tadpoles (Anura) to captive and free-ranging vertebrate predators

This study examines the unpalatability of Hyla semilineata tadpoles, relating this possible defence mechanism to their black, presumably aposematic, colouration. Bullfrog tadpoles (Rana catesbeiana), similar in size to the H. semilineata larvae, were used as controls in the experiments. The palatability of H. semilineata tadpoles was tested by offering the tadpoles to Fish (Hoplias malabaricus), free-ranging passerine birds (Pitangus sulphuratus) and hawks (Buteo magnirostris), and domestic chickens (Gallus domesticus). All predators showed a significant preference towards the control R. catesbeiana tadpoles. However, in experiments with fish, this preference was not significant for tadpole capture, only for their ingestion, suggesting that the fish could not distinguish between the two species before tasting them. Although great kiskadees (P. sulphuratus) preferred the control R. catesbeiana tadpoles, they promptly ingested more than half of the test H. semilineata tadpoles when these were offered alone. The chickens, used as naive predators, clearly learned to avoid the black H. semilineata tadpoles after a few trials. The conspicuous colouration and unpalatability of H. semilineata tadpoles may benefit the individual as well as the group, depending on the predator involved.

Evaluating the karyotypic diversity in species of Hyla (Anura; Hylidae) with 2n=30 chromosomes based on the analysis of ten species

Ten species of Hyla with 2n = 30 from Brazilian fauna were analysed cytogenetically. Hyla minuta is the unique presenting all bi-armed metacentric or submetacentric chromosomes in the karyotype, therefore, with the highest FN = 60. The remaining species have a variable number of uni-armed telocentric or subtelo-centric chromosomes: H. cruzi, H. elianeae, and H. rubicundula with three pairs (FN = 54), H. berthalutzae, H. elegans, H. microps, and H. nana with four pairs (FN = 52), and H. nahdereri and H. sanborni with five pairs (FN = 50). The uni-armed elements are among pairs 5, 6, 7, 11, 14, and 15, which also appeared with metacentric or submetacentric morphology. The remaining chromosome pairs 1, 2, 3, 4, 8, 9,10, 12, and 13 were never found to be telocentric or subtelocentric. AgNOR patterns are species-specific, the majority of the species exhibiting a single pair with AgNORs, with the exception of H. elegans and H. nana with more than one chromosome pair bearing this cytological marker. C banding was obtained in H. berthalutzae, H. cruzi, H. elegans, H. elianeae, H. microps, H. minuta, H. nahdereri, and H. nana, which showed positively stained centromeric heterochromatin. Our analysis confirms the great karyotypic diversity in the species of Hyla with 2n = 30, with no species sharing identical karyotypes.

On the identity of Hyla strigilata Spix, 1824 (Anura : Hylidae): redescription and neotype designation for a ghost taxon

Recent field work in Atlantic Rain Forest patches in the southern region of the State of Bahia, Brazil, resulted in the discovery of some populations of an unidentified species of the Scinax catharinae group. An extensive literature review, along with the examination of specimens and distribution patterns of all known species of this group, showed that Hyla strigilata Spix, 1824, a long confused species with lost type material, is an available name for the specimens from Bahia. In order to clarify the taxonomic problems surrounding this taxon, the nomenclatural history of Hyla strigilata is reviewed and a neotype is designated, described, and figured. The association of this name to extant populations from southern Bahia and its consequent stabilization is considered important since it is the type species of the genus Ololygon, a name available for the clade of Scinax catharinae. Data on habits, habitat, and geographic distribution are also presented.

The taxonomic status of Hyla pulchella joaquini (Anura : Hylidae) with description of its tadpole and vocalization

We redescribe Hyla pulchella joaquini and describe its tadpole and vocalization. The taxonomic status of this subspecies is reevaluated; and on the basis of morphology, geographic distribution, and vocalization, we propose the elevation of this subspecies to specific level with the name Hyla joaquini B. Lutz 1968. We also discuss the relationship of H, joaquini within the species groups of H. pulchella Dumeril and Bibron 1841 and H. circumdata (Cope 1871).

Conserved karyotypes in the Hyla pukhella species group (Anura, Hylidae)

Cytogenetic analyses were done on specimens of Hyla marginata and on three populations of H. semiguttata differing in morphology and in the physical parameters of their advertisement call, as well as in individuals of Hyla sp. (aff. semiguttata). All specimens had 2n = 24 chromosomes with a morphology very similar to that of other 24-chromosome Hyla species. Hyla semiguttata and H. marginata showed the same C-banding pattern but were distinguished by the location of the NOR on pair 1 in H. semiguttata (in the three populations) and Hyla sp. (aff. semiguttata), and on pair 10 in H. marginata. The H. semiguttata populations did not differ cytogenetically, despite variations in their morphology and advertisement calls. Similarly, H. semiguttata and H. p. joaquini studied previously had identical C-banding patterns and NOR locations, suggesting that they are very closely related.

Bioacoustic analysis of advertisement call in Hyla nana and Hyla sanborni (Anura, Hylidae) in Botucatu, São Paulo, Brazil

As características relacionadas à comunicação sonora em duas espécies aparentadas, Hyla nana e H. sanborni, pertencentes ao grupo nana de espécies, foram estudadas, entre agosto de 1997 e junho de 1999, em duas poças permanentes de grande porte em área aberta na região de Botucatu, Estado de São Paulo. Foram obtidas gravações de 131 exemplares, 71 indivíduos de H. nana e 58 de H. sanborni, durante início de atividade de vocalização e atividade de vocalização em coro. Houve diferença nos ritmos de emissão de notas dos cantos entre o início das atividades e durante os coros. O canto de anúncio das espécies consiste na emissão de séries consecutivas de notas simples, pulsadas, com taxa de repetição rápida. Hyla nana e H. sanborni apresentam dois tipos de notas em seu canto de anúncio, denominados aqui como tipos A e B. Notas do tipo A, introdutórias, apresentam maior duração e número de pulsos, e suas emissões foram mais freqüentes durante o início das atividades de vocalização. As notas introdutórias são as primeiras da série emitida em atividade de coro. As notas do tipo B, secundárias, são curtas e com menor número de pulsos, sendo emitidas durante as vocalizações em coro. Os dois tipos de notas encontrados diferem significativamente em sua estrutura temporal. As duas espécies apresentaram segregação acústica tanto na estrutura espectral como na temporal de seus cantos de anúncio.

Schooling and swimming behaviors of Hyla semilineata tadpoles (Anura, Hylidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

O status taxonômico de Hyla cochranae Mertens e recaracterização de Aplastodiscus A. Lutz (Anura, Hylidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

A molecular perspective on the phylogeny of the Hyla pulchella species group (Anura, Hylidae)

A molecular phylogenetic analysis of the Hyla pulchella species group was performed to test its monophyly, explore the interrelationships of its species, and evaluate the validity of the taxa that were considered subspecies of H. pulchella. Approximately 2.8 kb from the mitochondrial genes 12s, tRNA valine, 16s, and Cytochrome b were sequenced. The analysis included 50 terminals representing 10 of the 14-15 species currently recognized in the H. pulchella group, including samples from several localities for some taxa, several outgroups, as well as two species previously suspected to be related with the group (Hyla guentheri and Hyla hischoffi). The results show that the H. pulchella and Hyla circumdata groups are distantly related, and, therefore, should be recognized as separate groups. As currently defined, the H. pulchella group is paraphyletic with respect to the Hyla polytaenia group; therefore, we recognize the Hyla polytaenia clade in the H. pulchella group. Two subspecies of H. pulchella recognized by some authors are considered full species including Hyla pulchella riojana because it is only distantly related to H. pulchella, and Hyla pulchella cordobae because molecular and non-molecular evidence suggests that it is specifically distinct. With the inclusion of the H. polytaenia clade, H. guentheri, and H. bischoffi, and the recognition of the two former subspecies of H. pulchella as distinct species, the H. pulchella group now comprises 25 described species. All representatives of the H. pulchella group with an Andean distribution are monophyletic and nested within a clade from the Atlantic forest from south-southeastern Brazil/northeastern Argentina, and Cerrado gallery forest from central Brazil. (C) 2004 Elsevier B.V. All rights reserved.