Purification and characterization of antimicrobial peptides from the Caribbean frog, Leptodactylus validus (Anura: Leptodactylidae)

Peptidomic analysis of norepinephrine-stimulated skin secretions from the Caribbean frog Leptodactylus validus Garman, 1888 led to the identification of three peptides with previously undescribed sequences that were structurally similar to those of antimicrobial peptides isolated from other species of leptodactylid frogs. These paralogs have been termed ocellatin-V1 (GVVDILKGAGKDLLAHALSKLSEKV.NH(2)), ocellatin-V2 (GVLDILKGAGKDLLAHALSKISEKV.NH(2)), and ocellatin-V3 (GVLDILTGAGKDLLAHALSKLSEKV.NH(2)). The very low antimicrobial potency (MIC>200microM) against Escherichia coli and Staphylococcus aureus associated with the peptides is probably a consequence of their lack of amphipathicity and reduced cationicity compared with active members of the ocellatin family from related species.     

Ontogeny, phylogeny, and morphology in anuran larvae: morphometric analysis of cranial development and evolution in Rana tadpoles (Anura: Ranidae)

Comparative studies of chondrocranial morphology in larval anurans are typically qualitative in nature, focusing primarily on discrete variation or gross differences in the size or shape of individual structures. Detailed data on chondrocranial allometry are currently limited to only two species, Rana sylvatica and Bufo americanus. This study uses geometric morphometric and multivariate statistical analyses to examine interspecific variation in both larval chondrocranial shape and patterns of ontogenetic allometry among six species of Rana. Variation is interpreted within the context of hypothesized phylogenetic relationships among these species. Canonical variates analyses of geometric morphometric datasets indicate that species can be clearly discriminated based on chondrocranial shape, even when whole ontogenies are included in the analysis. Ordinations and cluster analyses based on chondrocranial shape data indicate the presence of three primary groupings (R. sylvatica; R. catesbeiana + R. clamitans; and R. palustris + R. pipiens + R. sphenocephala), and patterns of similarity closely reflect phylogenetic relationships. Analysis of chondrocranial allometry reveals that some patterns are conserved across all species (e.g., most measurements scale with negative allometry, those associated with the posterior palatoquadrate tend to scale with isometry or positive allometry). Ontogenetic scaling along similar allometric trajectories, lateral transpositions of individual trajectories, and variable allometric relationships all contribute to shape differences among species. Overall patterns of similarity among ontogenetic trajectories also strongly reflect phylogenetic relationships. Thus, this study demonstrates a tight link between ontogeny, phylogeny, and morphology, and highlights the importance of including both ontogenetic and phylogenetic data in studies of chondrocranial evolution in larval anurans.     

Cranial ontogeny in Philautus silus (Anura: Ranidae: Rhacophorinae) reveals few similarities with other direct-developing anurans

Direct development has evolved in rhacophorine frogs independently from other anuran lineages, thereby offering an opportunity to assess features associated with this derived life history. Using a developmental series of the direct-developing Philautus silus (Ranidae: Rhacophorinae) from Sri Lanka, we examine features of cranial morphology that are part of a suite of adaptations that facilitate feeding in free-living tadpoles, but have been changed or lost in other direct-developing lineages. Larval-specific upper jaw cartilages, which are absent from many non-rhacophorine direct-developing species (such as Eleutherodactylus coqui), develop in embryos of P. silus. Similarly, lower jaw cartilages initially assume a larval morphology, which is subsequently remodeled into the adult jaw configuration before hatching. However, the cartilaginous jaw suspension and hyobranchial skeleton never assume a typical larval morphology. The palatoquadrate, which suspends the lower jaw, lacks the posterior connections to the braincase found in many metamorphosing species. Unlike in metamorphosing species, bone formation in P. silus begins before hatching. However, the sequence of bone formation resembles that of metamorphosing anurans more than that of other direct developers. In particular, P. silus does not exhibit precocious ossification of the lower jaw, which is characteristic of some frogs and caecilians that lack a free-living tadpole. These data reveal some similarities between Philautus and other direct-developing anurans. However, the departure of Philautus embryos from the generalized tadpole skeletal morphology is less pronounced than that observed in other direct-developing taxa.     

厄瓜多尔西部新热带蛙(两栖纲: 无尾目:细趾蟾科)的分布特征及生态习性

Leptodactylus labrosus属于陆地坐-等型的捕食者,其食物包括地表的、掘地的及飞行昆虫,其中蚂蚁数量最多。曾有报道Bothrops asper是其天敌。L.labrosus主要栖息于落叶及半落叶林潮湿的环境中,偶尔也栖息于常绿林中。L.labrosus分布于厄瓜多尔西海岸的北部、中部、南部;秘鲁西海岸的北部、中部,分布地海拔可达600 m;及位于南厄瓜多尔和北秘鲁海拔高达1 300 m的干燥的安第斯山谷。其分布地区包括了厄瓜多尔和秘鲁海岸的潮湿的、季节性干燥的森林。在安第斯山脉斜坡的潮湿地带,L.labrosus与其他3种细趾蟾科的分布区重叠,但表现为生境分离。其分布特征与其他几种活动区域受限的两栖动物相似。Tumbesian地区应被认为是地方特有两栖动物的分布区。Choco和Tumbesian的中间区域是L.labrosus与其他细趾蟾科的生境分布重叠区,其生态及气候特征使其形成了独特的动物区系,包括几种地方特有物种。因此该地区应被划为西厄瓜多尔地方种区域。     

Chondrocranial development in larval Rana sylvatica (Anura: Ranidae): morphometric analysis of cranial allometry and ontogenetic shape change

This study provides baseline quantitative data on the morphological development of the chondrocranium in a larval anuran. Both linear and geometric morphometric methods are used to quantitatively analyze size-related shape change in a complete developmental series of larvae of the wood frog, Rana sylvatica. The null hypothesis of isometry was rejected in all geometric morphometric and most linear morphometric analyses. Reduced major axis regressions of 11 linear chondrocranial measurements on size indicate a mixture of allometric and isometric scaling. Measurements in the otic and oral regions tend to scale with negative allometry and those associated with the palatoquadrate and muscular process scale with isometry or positive allometry. Geometric morphometric analyses, based on a set of 11 chondrocranial landmarks, include linear regression of relative warp scores and multivariate regression of partial warp scores and uniform components on log centroid size. Body size explains about one-quarter to one-third of the total shape variation found in the sample. Areas of regional shape transformation (e.g., palatoquadrate, otic region, trabecular horns) are identified by thin-plate spline deformation grids and are concordant with linear morphometric results. Thus, the anuran chondrocranium is not a static structure during premetamorphic stages and allometric patterns generally follow scaling predictions for tetrapod cranial development. Potential implications regarding larval functional morphology, cranial development, and chondrocranial evolution in anurans are discussed.     

Postmetamorphic ontogenetic allometry and the evolution of skull shape in Nest‐building frogs Leptodactylus (Anura: Leptodactylidae)

Allometry constitutes an important source of morphological variation. However, its influence in head development in anurans has been poorly explored. By using geometric morphometrics followed by statistical and comparative methods we analyzed patterns of allometric change during cranial postmetamorphic ontogeny in species of Nest‐building frogs Leptodactylus (Leptodactylidae). We found that the anuran skull is not a static structure, and allometry plays an important role in defining its shape in this group. Similar to other groups with biphasic life‐cycle, and following a general trend in vertebrates, ontogenetic changes mostly involve rearrangement in rostral, otoccipital, and suspensorium regions. Ontogenetic transformations are paralleled by shape changes associated with evolutionary change in size, such that the skulls of species of different intrageneric groups are scaled to each other, and small and large species show patterns of paedomorphic/peramorphic features, respectively. Allometric trajectories producing those phenotypes are highly evolvable though, with shape change direction and magnitude varying widely among clades, and irrespective of changes in absolute body size. These results reinforce the importance of large‐scale comparisons of growth patterns to understand the plasticity, evolution, and polarity of morphological changes in different clades.     

Development of the nasal chemosensory organs in two terrestrial anurans: the directly developing frog, Eleutherodactylus coqui (Anura: Leptodactylidae), and the metamorphosing toad, Bufo americanus (Anura: Bufonidae)

Nearly all vertebrates possess an olfactory organ but the vomeronasal organ is a synapomorphy for tetrapods. Nevertheless, it has been lost in several groups of tetrapods, including aquatic and marine animals. The present study examines the development of the olfactory and vomeronasal organs in two terrestrial anurans that exhibit different developmental modes. This study compares the development of the olfactory and vomeronasal organs in metamorphic anurans that exhibit an aquatic larva (Bufo americanus) and directly developing anurans that have eliminated the tadpole (Eleutherodactylus coqui). The olfactory epithelium in larval B. americanus is divided into dorsal and ventral branches in the rostral and mid-nasal regions. The larval olfactory pattern in E. coqui has been eliminated. Ontogeny of the olfactory system in E. coqui embryos starts to vary substantially from the larval pattern around the time of operculum development, the temporal period when the larval stage is hypothesized to have been eliminated. The nasal anatomy of the two frogs does not appear morphologically similar until the late stages of embryogenesis in E. coqui and the terminal portion of metamorphosis in B. americanus. Both species and their respective developing offspring, aquatic tadpoles and terrestrial egg/embryos, possess a vomeronasal organ. The vomeronasal organ develops at mid-embryogenesis in E. coqui and during the middle of the larval period in B. americanus, which is relatively late for neobatrachians. Development of the vomeronasal organ in both frogs is linked to the developmental pattern of the olfactory system. This study supports the hypothesis that the most recent common ancestor of tetrapods possessed a vomeronasal organ and was aquatic, and that the vomeronasal organ was retained in the Amphibia, but lost in some other groups of tetrapods, including aquatic and marine animals.     

Cladistic analysis and osteological descriptions of the frog species in the Leptodactylus fuscus species group (Anura, Leptodactylidae)

The genus Leptodactylus is predominantly Neotropical (a few species have colonized the southern Neartic region) and is distributed from Texas to Argentina and on certain Caribbean islands. Leptodactylus was divided into five groups of species: Leptodactylus melanonotus , Leptodactylus ocellatus, Leptodactylus fuscus , Leptodactylus pentadactylus and Leptodactylus marmoratus . Among these, the L. fuscus group is the one with most species, with 27 taxa. Characters unverified in most of the species are used to define the L. fuscus group. However, the monophyly of the group has never been tested rigorously in a quantitative phylogenetic context. Thus, the main goal of this study was to test such monophyly and to construct a phylogeny of the L. fuscus group. A matrix of 114 characters scored across 43 taxa was constructed, with 31 characters taken from external morphology, 58 from adult skeletons, 16 from larval chondrocranium, 5 from ethology and 4 from morphometric data were included. Out of all the species examined, 23 belonged to the ingroup and 20 to the outgroup. The data set was analysed with implied weights, by using TNT software. The monophyly of the group was strongly supported in the fittest cladogram obtained. The optimizations of some characters on this hypothesis support traditional evolutionary hypotheses. The optimizations also suggest the presence of paedomorphic character states in some species, which is also discussed.     

Ultrastructure of poison glands of South American frogs: a comparison between Physalaemus albonotatus and Leptodactylus chaquensis (Anura: Leptodactylidae)

Serous (poison) cutaneous glands of the leptodactylid species Physalaemus albonotatus and Leptodactylus chaquensis were compared using light and transmission electron microscopy. Glands in the two species share structural traits common in anurans, including the peripheral contractile sheath (myoepithelium) and the syncytial secretory unit that produces, stores, and modifies the poison. At the ultrastructural level, early steps of poison production are also similar and fit the usual path of proteosynthesis, involving rough endoplasmic reticulum (RER) and Golgi stacks (dictyosomes) in the peripheral syncytial cytoplasm. However, several differences are obvious during the maturational processes that lead post-Golgian products to their ultimate ultrastructural traits. In P. albonotatus, the dense product released from the dictyosomes acquires a thick repeating substructure, which, however, becomes looser in the inner portion of the syncytium. In L. chaquensis, serous maturation involves gradual condensation, and opaque, somewhat "vacuolized" granules are formed. These different maturational paths expressed during poison manufacturing in the two species agree with the polyphyletic origin of the family Leptodactylidae. On the other hand, data collected for P. albonotatus fit previous findings from P. biligonigerus and stress the view that poisons produced by congeneric species share similar (or identical) ultrastructural features.     

Species affinity and infracommunity ordination of helminths of Leptodactylus chaquensis (Anura: Leptodactylidae) in two contrasting environments from northeastern Argentina

One hundred seventy-two frogs (Leptodactylus chaquensis) were collected from November 2002 to November 2003, in agricultural (n = 132) and nonagricultural (n = 40) areas. Both sites are near the city of Corrientes, Argentina. The main goals of this study were as follows: (1) to determine the helminth parasite community in agricultural and nonagricultural habitats; (2) to analyze the relationships between helminth parasites and site of infection, frog body size, and gender; (3) to identify and examine covariation and association of helminth communities; and (4) to determine the mean richness and diversity of parasite communities. The helminth compound community of this amphibian species consisted of 24 species: 19 in agricultural habitats and 18 in nonagricultural habitats. The mean richness, mean diversity, and evenness of helminths were significantly different between the habitats (P < 0.05). The body size of the host was the important factor in determining parasite richness. Both habitats exhibited differences in community ordination. The helminth species in the 2 habitats exhibited the same interspecific relationships, although differences were observed in the intensity of infection.     

Cryptic species complexes,widespread species and conservation: lessons from Amazonian frogs of the Leptodactylus marmoratus group (Anura: Leptodactylidae)

Morphologically cryptic species act as a wild card when it comes to biodiversity assessments and conservation, with the capacity to dramatically alter our understanding of the biological landscape at the taxonomic, ecological, biogeographic, evolutionary, and conservation levels. We discuss the potential effects that cryptic species may have on biodiversity assessments and conservation, as well as some of the current issues involving the treatment of cryptic species both at taxonomic and conservation levels. In addition, using a combination of advertisement call and morphological data, we describe a new species of the Leptodactylus marmoratus group from the upper Amazon basin, and we assess how cryptic species can affect conservation assessments of species in the Leptodactylus marmoratus group by examining how recent findings affect our understanding of the distribution of what is assumed to be a widespread Amazonian species, Leptodactylus andreae.     

Hyperossification in miniaturized toadlets of the genus Brachycephalus (Amphibia: Anura: Brachycephalidae): Microscopic structure and macroscopic patterns of variation

Species of the genus Brachycephalus, have a snout‐vent length of less than 18 mm and are believed to have evolved through miniaturization. Brachycephalus ephippium, is particularly interesting; because its entire skull is hyperossified, and the presacral vertebrae and transverse processes are covered by a dorsal shield. We demonstrate in this paper that, at the macroscopic level, a completely hyperossified skull and dorsal shield occur only in B. ephippium, but not in B. ferruginus, B. izechsohni, B. pernix, B. pombali, B. brunneus, B. didactylus, and B. hermogenesi. An intermediate condition, in which the skull is hyperossified but a dorsal shield is absent, occurs in B. vertebralis, B. nodoterga, B. pitanga, and B. alipioi. The microscopic structure of hyperossification was examined in skulls of B. ephippium and B. pitanga, revealing a complex organization involving the presence of Sharpey fibers, which in humans are characteristic of periodontal connections. J. Morphol., 2009. © 2009 Wiley‐Liss, Inc.     

Morphology of the cranial skeleton and musculature in the obligate carnivorous tadpole of Lepidobatrachus laevis (Anura: Ceratophryidae)

Ziermann, J.M., Infante, C., Hanken, J. and Olsson, L. 2011. Morphology of the cranial skeleton and musculature in the obligate carnivorous tadpole of Lepidobatrachus laevis (Anura: Ceratophryidae). —Acta Zoologica (Stockholm) 00 :1–12. Lepidobatrachus laevis (Ceratophryidae: Ceratophryinae) is a bizarre frog endemic to the Chacoan desert of central South America. Its tadpole is an obligate carnivore that can catch and consume live prey nearly its own size. Morphological adaptations associated with this unique feeding mode, including the larval skull anatomy and associated cranial musculature, have only been partly described. We studied the head of Stages 26–27 larvae using gross dissection, immunohistochemistry, and standard histology. Derived features of this tadpole compared to the microphagous, herbivorous larvae of most other anurans include simplified chondrocranial cartilages and very robust jaw muscles. The mm. suspensorio‐ et quadratoangularis do not take their origin from the processus muscularis of the palatoquadrate, as in most other tadpoles, but instead originate from the corpus of the palatoquadrate caudal to this process. The jaw levators are unusually large. The tadpole of Ceratophrys, another member of the ceratophryine clade, also consumes large animal prey, but its morphology is very different. It probably has evolved independently from a generalized, mainly herbivorous tadpole similar to the larva of Chacophrys, the third ceratophryine genus. Most specialized features of the larval head of Lepidobatrachus laevis are adaptations for ‘megalophagy’—ingestion of whole, very large animal prey.     

Morphology and metamorphosis of Eupsophus calcaratus tadpoles (Anura: Leptodactylidae)

Eupsophus calcaratus, a leptodactyloid frog from the austral Andean forests of Argentina and Chile, has endotrophic, nidicolous tadpoles. We studied a metamorphic series from Stages 31 to 46 of Gosner's developmental table (1960). Other than the scarce pigmentation, proportionately large eyes, and massive developing hindlimbs, the remaining external characters are similar to those of generalized, exotrophic larvae. At the same time, internal morphology does not reveal any character state attributable to the endotrophic-nidicolous way of life; conversely, structures such as the hyobranchial skeleton and the mandibular cartilages are similar to those of exotrophic-macrophagous tadpoles. The metamorphic process is characterized by the delayed development of diverse structures (e.g., ethmoid region, palatoquadrate, and hyobranchial apparatus), and the retention of some larval characters (e.g., parietal fenestrae, overall absence of ossification) with the absence of development of some "juvenile" characters (e.g., adult otic process, several bones) in metamorphosed individuals. These heterochronic processes and truncation of larval development are related to a shorter larval life (when compared to other species of the austral Andean region) and to the small size at metamorphosis.     

Morphology and feeding in tadpoles of Ceratophrys cranwelli (Anura: Leptodactylidae)

This paper provides data on the skeleton, musculature, buccal apparatus, buccopharyngeal cavity and diet of Ceratophrys cranwelli tadpoles, and attempts to contribute to the knowledge of relations between morphology and ecology in anuran larvae. Both in morphological characters and feeding habits, these tadpoles are very similar to other species within the genus. They possess many of the structural features usually found in predaceous tadpoles: strong, keratinized jaw sheaths and keratodonts, reduced buccal papillation, high values of in‐lever arm proportion and buccal floor area, well‐developed ceratohyals, and hypertrophied jaw muscles. Food sources consist of other tadpoles, microcrustaceans, larvae of insects, plant fragments, as well as rotifers and microalgae. As facultative carnivores, they are likely to play an important role in regulating the aquatic communities of the ephemeral ponds where they develop.