Pattern and rate of ovary differentiation with reference to somatic development in anuran amphibians

The rate of somatic development of anuran amphibians is only roughly correlated with the rate of gonad differentiation and varies among species. The somatic stage of a tadpole often does not reflect its age, which seems to be crucial for gonad differentiation rate. We compared the morphology and differentiation of developing ovaries at the light and electron microscopy level, with reference to somatic growth and age of a female. Our observations were performed on 12 species of six families (Rana lessonae, R. ridibunda, R. temporaria, R. arvalis, R. pipiens, R. catesbeiana, Bombina bombina, Hyla arborea, Bufo bufo. B. viridis, Xenopus laevis, Pelobates fuscus) and compared with the results obtained by other authors. This allowed us to describe the unified pattern of anuran female gonad differentiation. Ovary differentiation was divided into 10 stages: I-III, undifferentiated gonad; IV, sexual differentiation; V, first nests of meiocytes; VI, first diplotene oocytes; VII-IX, increasing number of diplotene oocytes and decreasing number of oogonia and nests; X, fully developed ovary composed of diplotene oocytes with rudimental patches of oogonia. We distinguished three types of ovary differentiation rate: basic (most species), retarded (genus Bufo), and accelerated (green frogs of the subgenus Pelophylax genus Rana).     

Variability in timing of larval season in an amphibian community in SW Spain

Timing of larval periods of 9 species of amphibians (Hyla meridionalis, Pelobates cultripes, Discoglossus galganot, Bufo calamita, Bufo bufo, Rana perezi, Triturus marmoratus, Triturus boscat, Pleurodeles waltl) was followed in 7 different years Total larval season may last from autumn to summer, whereas in low rainy years it may last from late winter to summer In rainy years, two species are early breeders, starting to breed m autumn, six species commonly start in mid-winter and one species always breeds later, in spring The widest interannual variability m the onset and duration of larval season occurs in the earliest breeders In general, the more rainy the autumn and the higher temperatures were reached, the earlier the season started In contrast, duration of the season was not correlated with rainfall and temperature Interannual variability is interpreted as the possibility of adjusting the reproduction of the species to the availability of breeding habitats, constrained by physiological features of the species and interspecific relationships     

Infectivity of cultured Trypanosoma fallisi (Kinetoplastida) to various anuran species and its evolutionary implications

Trypanosoma fallisi, a hemoflagellate infecting Bufo americanus from Ontario, was grown in vitro, and metatrypanosomes from the primary culture were inoculated into 4 uninfected test groups from 3 anuran families: Bufonidae, Hylidae, and Ranidae. In vitro-cultured T. fallisi was found to infect B. americanus and to induce transient infections in Bufo valliceps and Hyla versicolor. The flagellate was not infective to Rana clamitans. Trypanosoma ranarum was uninfective to the bufonids and hylids tested. These data suggest that the potential for host-switching decreases with increased evolutionary distance of the potential anuran host.     

Morphological and life history responses to predators in larvae of seven anurans

Amphibian larvae often face two major sources of mortality: pond desiccation and predation. Tadpoles of seven anuran species with different preferences for type of breeding habitat, on a hydroperiod scale, were tested for responses to the presence of predators by raising them experimentally in the presence and absence of a separately caged invertebrate predator that was fed on conspecific tadpoles. The species typically breeding in temporary or semi-permanent ponds ( Rana temporaria , Rana arvalis , Rana dalmatina and Hyla arborea ) – where invertebrate predator populations are predicted to vary considerably spatiotemporally – all showed marked induced increases in tail fin depth in response to predator presence. These species also tended to respond by reduced growth rates. The representative of the most ephemeral habitats, Bufo calamita , did not respond in any of these traits. Its congeneric, Bufo bufo , a toxic inhabitant of permanent ponds and lakes, tended to respond to predator presence by reducing its growth rate, though not by a tail depth increase. I argue that the rather poor swimming performance in Bufo tadpoles may opt for defences other than locomotor ability. The palatable, permanent pond species Pelobates fuscus did not alter either its growth rate or tail morphology. Possible explanations for this result are discussed.     

Anuran amphibia which are not acclimable to high salt,tolerate high plasma urea

• 1.1. The capacity of five anuran Amphibians (Bufo viridis B. regularis, Rana ridibunda, Hyla arborea and Pelobates syriacus) to acclimate to NaCl and urea solutions was investigated.
• 2.2. All species could be acclimated to relatively high concentrations of urea solutions, while only Bufo viridis and Hyla arborea could be acclimated to 500 mOsm/kg or higher NaCl solutions.
• 3.3. The plasma urea concentration in B. viridis and H. arborea was elevated to levels over 140 mmol/1.
• 4.4. The sum of plasma sodium and chloride concentrations did not increase over 400 mmol/l in any species.
• 5.5. Urine osmolality, which was normally low, increased, but never exceeded the plasma osmolality.
• 6.6. In the urea acclimation conditions, urine electrolytes diminished, similarly in all species in this study.
• 7.7. It is concluded that anuran Amphibians can tolerate high plasma urea concentrations, but only those species which can elevate it, either through retention or net synthesis, can be acclimated to high salt solutions.

     

Comparative studies on the vascular organization of carotid labyrinths of anurans and caudates

The three-dimensional structures of the carotid labyrinth in five species of anurans representing four families (Rana nigromaculata, Rana catesbeiana, Bufo japonicus, Hyla arborea, and Xenopus laevis), and three species of caudates representing three families (Cynops pyrrhogaster, Hynobius nebulosus, Ambystoma mexicanum) were compared using vascular corrosion castings and scanning electron microscopy (SEM). Anuran carotid labyrinths are spherical in shape and are classified into two groups according to the origin of the external and internal carotid arteries. One group, which included Rana, Hyla, and Bufo, is characterized by the presence of a vascular ring at the proximal end and some vascular routes at the distal end of the labyrinth. The external and internal carotid arteries originate from these structures. The other group, which includes only Xenopus, is characterized by the external carotid artery opening directly from the central chamber or the common carotid artery, and by the internal carotid artery originating from within the vascular maze. The vascular maze is most complex in Xenopus, less so in Rana and Bufo, and simplest in Hyla. The carotid labyrinths in Cynops and Hynobius are oblong in shape. The fundamental organization in salamanders is similar to that in anurans. The vascular maze, however, is much simpler than in Hyla. There is no specialized swelling in Ambystoma mexicanum. The present findings suggest that most amphibian carotid labyrinths have the appropriate architecture for controlling vascular tone.     

How can ontogeny help us to understand the morphology of anuran pectoral girdle?

The ontogenetic development of the pectoral girdle in seven anuran species (Xenopus laevis, Discoglossus pictus, Bombina bombina, Bombina variegata, Pelobates fuscus, Bufo bufo and Rana dalmatina) was studied using cleared and stained specimens. The epicoracoid cartilage was found to develop in two different ways resulting in an arciferal or firmisternal type of the pectoral girdle. In the arciferal one, the epicoracoid originates from a medial prolongation of the procoracoid cartilage and broadly overlaps its counter part during further development. In the firmisternal pectoral girdle, the epicoracoids are formed by the widened cartilaginous medial edges of the coracoids that fuse together along the midline. Polarization of ontogenetic characters shows, that omosternum evolves inside Anura, and the type of sternum occurring in basal Anura seems to be an apomorphy of all Batrachia. The sternal elements have a single or paired rudiment, their development is connected with M. rectus abdominis or a zonal area and they remain cartilaginous or ossify during postmetamorphic development. An occurrence of omosternum in Barbourula busuangensis was described for the first time.     

Phylogenetic relationships of Pelobatoidea re-examined using mtDNA

Pelobatoidea is a clade of ancient anurans with obscure relationships to the remaining clades of frogs. We used partial sequences of two mitochondrial genes (cytochrome b and 16S RNA) from all Pelobatoidea subclades, including all species of Pelobatidae and Pelodytidae and four outgroup taxa (Xenopus, Ascaphus, Discoglossus, and Rana), to propose a phylogenetic hypothesis for relationships within Pelobatoidea. Maximum likelihood and Bayesian analyses support the monophyly of Pelobatoidea, but our hypothesis of internal relationships differs substantially from all previous hypotheses. Megophryidae is sister to Pelobates, and this clade is sister to Pelodytes. The most basal clade within Pelobatoidea is formed by Scaphiopus and Spea. The family Pelobatidae, as previously defined is not monophyletic, and it is split into Eurasian spadefoot toads Pelobates which retain the name Pelobatidae and North American spadefoot toads Scaphiopus and Spea which comprise the revived taxon Scaphiopodidae. Our analysis uncovers the existence of morphologically cryptic taxa within previously recognized species of the genus Spea and reveals marked genetic differentiation within Iberian Pelodytes. We discuss biogeographic implications and the evolution of fossoriality in the light of the new phylogenetic hypothesis.     

Mitochondrial phylogeny of Anura (Amphibia): a case study of congruent phylogenetic reconstruction using amino acid and nucleotide characters

We explore whether phylogenetic analyses of the same sequence data set at the amino acid and nucleotide level are able to recover congruent topologies, as well as the advantages and limitations of both alternative approaches. As a case study, mitochondrial protein-coding genes were used to discern among competing hypotheses on the phylogenetic relationships of major anuran amphibian lineages. To properly address this phylogenetic question, the complete nucleotide sequences of the mitochondrial genomes of two archaeobatrachian species, Ascaphus truei and Pelobates cultripes, were determined anew. Bayesian and maximum likelihood phylogenetic inferences of the same sequence data set were performed based on both amino acid and nucleotide characters, with the latter analysed either as codons or as a reduced data set of first+second (P12) codon positions. In addition, likelihood-based ratio tests were performed to evaluate the support of alternative topologies. The different data sets arrived at congruent and highly supported topologies, suggesting a similar phylogenetic resolving power of the two character types provided that correctly selected sites and appropriate evolutionary models are used. The reconstructed anuran mitochondrial phylogeny supports the paraphyly of Archaeobatrachia, with Ascaphus as sister group to all the remaining anurans, and Pelobates as sister group of Neobatrachia. However, the employed tree reconstruction methods and likelihood-based ratio tests seemed to be negatively affected by the fast evolving sequences of neobatrachians, suggesting that the phylogeny of Anura here presented is not definitive, and needs further investigation using an extended taxon sampling.     

Electrotonic transmission between primary afferent fibers and the motor neurons of the isolated spinal cord of various representative amphibians

Studies have been made of monosynaptic excitatory postsynaptic potentials elicited by stimulation of the posterior root in motoneurones of the isolated spinal cord of the frog Rana ridibunda, toad Bufo bufo, and clawed toad Xenopus laevis. In all the amphibians studied, the early component of monosynaptic EPSP was not blocked in Ca-free medium containing 2 mM Mn2+. It is suggested that electrical coupling in anuran amphibians reflects certain stage of evolution of the synaptic transmission in vertebrates.     

The development and fine structure of ultimobranchial glands in larval anurans

A comparative optical microscopic and ultrastructural study on the ultimobranchial (UB) glands of three common species of Israeli anurans: Bufo viridis, Hyla arborea, and Rana ridibunda during metamorphosis is presented. The UB glands typically consist of a single follicle with a central lumen, though occasionally secondary follicles are present in Hyla and Rana. A single UB cell type is found which appears either in a very electron-dense "dark" form or as a less dense "light" form, though the ratio of dark: light cells from gland to gland at any one stage of metamorphic development is quite variable. By the end of metamorphosis in Bufo and Hyla all the UB cells are usually of the light variety, whereas in Rana the dark cells persist. The organelles of these secretory cells including secretory granules, granular endoplasmic reticulum, free ribosomes, tonofilaments, microtubules, Golgi bodies, and lipid droplets, their distribution, abundance, and possible functions in relation to metamorphosis are described. Apocrine secretion into the central lumen of the gland is also described and discussed.     

Morphological, mitochondrial DNA and allozyme evolution in representative amphibians and reptiles inhabiting each side of the Strait of Gibraltar

Patterns of differentiation in morphology, mitochondrial DNA and allozymes in amphibians and reptiles inhabiting northern and southern shores of the Strait of Gibraltar are not concordant, suggesting that each taxon was affected differently by events preceding or following the formation of the Strait of Gibraltar. Mitochondrial DNA and allozyme differentiation between Discoglossus jeanneae and Discoglossus scovazzi (Anura, Discoglossidae), Rana perezi and Rana saharica (Anura, Ranidae), and Blanus cinereus and Blanus tingitanus (Squamata, Amphisbaenia, Amphisbaenidae) is substantial, whereas morphological differentiation is moderate in Rana and Blanus , but is substantial in Discoglossus . Differentiation in mitochondrial DNA and morphology between Timon ( Lacerta ) lepidus and Timon ( Lacerta ) tangitanus (Squamata, Lacertoidea, Lacertidae) is considerable, but allozyme differentiation is low. In members of type-I and -II Podarcis vaucheri (Squamata, Lacertoidea, Lacertidae), morphology and mitochondrial DNA are moderately differentiated, but allozyme differentiation is low. Spanish and Moroccan populations of Hyla meridionalis (Anura, Hylidae), Mauremys leprosa (Testudines, Geoemydidae), and Macroprotodon brevis (Squamata, Serpentes, Colubridae) demonstrate little allozyme and mitochondrial DNA differentiation, but whereas morphological differentiation between Mauremys and Macroprotodon populations is moderate, Hyla demonstrate substantial morphological differentiation between continental populations. These data suggest that sex-limited mitochondrial markers are reflective of ancient phylogenetic history, whereas biparentally inherited allozyme markers and morphological characteristics reflect more recent population structure and movement.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 445–461.     

Variations in anuran embryogenesis: yolk-rich embryos of Hyperolius puncticulatus (Hyperoliidae)

Anuran development is usually described using model species, most notably Xenopus laevis and Rana pipiens. We describe the development of the East African Reed Frog, Hyperolius puncticulatus, a species displaying development that is highly divergent from the "classic" anuran developmental pattern. Although having small eggs, the eggs of H. puncticulatus are characterized by a large amount of yolk, and embryonic development is reminiscent of species with large eggs. The eggs are teleolecithal and the cleavage is holoblastic, with a "pseudo-meroblastic" pattern. Gastrulation proceeds primarily at the dorsal lip and is characterized by reduced embryonic cavities. Gastrulation ends with a thickened "embryonic mantle" that sits upon a large yolk mass and forms most of the tissues of the embryo. The embryonic axis curves across the yolk mass, instead of the typical lengthening of anuran embryos. The tadpole hatches with a large ventral yolk mass which is gradually absorbed. We hypothesize about the developmental mechanisms that underlie this unusual development, based on comparisons with other anuran and fish species. We suggest that this type of development is not unique to this species, but can be found in many species of different anuran taxonomic groups. Comparing the development of H. puncticulatus and similar species to what is known about the development of model species, such as X. laevis, shows us the variation in early anuran embryogenesis. Knowing the existing diversity is a prerequisite to understanding the evolution of early anuran development and the changes in patterning mechanisms in different lineages.     

Histochemical demonstration of 5-hydroxytryptamine in poison glands of amphibian skin

Summary The aim of this project was to see whether 5-hydroxytryptamine (5-HT) believed to be present in the skin glands of anuran amphibians can be demonstrated histochemically. The Periodic acid-Schiff technique (P.A.S.) and three histochemical methods known to demonstrate 5-HT in enterochromaffin cells (Pontana's, diazonium and Schmorl's) were applied to the dorsal skin of three species—Xenopus laevis, Rana angolensis and Bufo regularis.Mucous glands were identified by their P.A.S. reactivity in all three species. Poison glands were identifiable in Bufo regularis and Xenopus laevis only. The secretory granules of the latter glands have strong positive reactions with all three histochemical techniques used.It is concluded that the poison glands of anuran amphibians contain 5-HT demonstrable by histochemical means.     

Distribution and reproductive strategies of Gyrinicola batrachiensis (Oxyuroidea: Pharyngodonidae) in larvae of eight species of amphibians from Nebraska

In total, 462 tadpoles and salamander larvae of 8 species were examined for the presence of Gyrinicola batrachiensis from 5 locations in Nebraska. Infection by G. batrachiensis occurred in tadpoles of Rana blairi , Rana catesbeiana, Rana pipiens, and Bufo woodhousii. Tadpoles of Hyla chrysoscelis , Spea bombifrons, and Pseudacris maculata and larvae of Ambystoma mavortium were not infected with G. batrachiensis. Population structure, defined as prevalence, mean abundance, and mean intensity of G. batrachiensis, varied among tadpoles of different amphibian species and was determined by collection locality, developmental period of tadpole hosts, amphibian species co-occurrence, and different reproductive strategies of G. batrachiensis , or a combination. Gyrinicola batrachiensis observed in all ranid tadpoles and B. woodhousii tadpoles from where bufonids were the only anuran species present, confirmed to the didelphic haplodiploidy and monodelphic parthenogenetic reproductive strategies, respectively. However, tadpoles of B. woodhousii that co-occurred with tadpoles of R. pipiens at Cedar Creek were inconsistent with these predictions and contained both male and didelphic female nematodes, but at a low mean intensity (1.61 ± 0.70). Didelphic female nematodes from B. woodhousii tadpoles at Cedar Creek only produced thick-shelled eggs, whereas nematodes in R. pipiens tadpoles had a high mean intensity (14.88 ± 23.83) from this location and contained both thick-shelled and thin-shelled eggs in their respective uteri. More importantly, adult female nematodes from tadpoles of R. pipiens and B. woodhousii from Cedar Creek were morphologically more similar to each other than to female nematodes recovered from tadpoles of other anuran species, other locations, or both. These data suggest that when strains of G. batrachiensis are shared by tadpoles of different amphibian species that differ in developmental period, the nematodes have an intermediate reproductive strategy in amphibian species, with tadpoles having short development.     

黑眶蟾蜍、黑斑蛙、中国雨蛙不同地理居群的染色体多样性

本文研究温州地区的黑眶蟾蜍、黑斑蛙、中国雨蛙的核型,分析了三个地理居群的黑星期五蟾蜍、四个地理居群的黑斑蛙、三个地理居群的中国雨蛙核型。结果表明不同地理居群的同种蛙有相同的染色体数和核型模式。黑星期五蟾蜍为2n＝22,NF＝44,核模式6＋5;黑斑蛙为2n=26,NF=52,核模式5＋8;中国雨蛙为2n＝24,NF＝48,核模式6＋6。但同一种蛙的不同地理居群之间在SM数目和顺序、次缢痕或随体的位置等有所不同。说明不同地理居群的同种蛙的染色体具有丰富的多样性。     

Laich-Fressen durch Kaulquappen als mögliche Ursache spezifischer Biotoppräferenzen und kurzer Laichzeiten bei europäischen Froschlurchen (Amphibia,Anura)

Zusammenfassung In einem Kunstweiher aufgezogene Kaulquappen von Rana temporaria fressen in der Versuchsanordnung von Abb. I den Laich folgender sympatrischer Anuren-Arten: Rana temporaria (Kannibalismus), Rana esculenta, Rana ridibunda, Bufo bufo, Bufo calamita, Bombina variegata und Hyla arborea (Laichräubern). — Daß die Kaulquappen der früh im Jahr laichenden Rana temporaria später gelegten arteigenen und artfremden Laich fressen, wirkt als Selektionsdruck in folgende Richtungen: Rana temporaria züchtet sich selbst eine kurze Laichzeit. Spät in seichtem Wasser laichende Arten werden im Biotop von Rana temporaria unterdrückt (Bombina variegata, Bufo calamita, Hyla arborea). Rana esculenta, Rana ridibunda und Bufo bufo sind dem Laichräubern durch Kaulquappen wenig ausgesetzt, weil sie in größeren Wassertiefen laichen. Zudem ist Bufo bufo ebenfalls ein Frühlaicher. Alytes obstetricans steht außer Konkurrenz, weil er die Eier nicht ins Wasser setzt. — Untersuchungen über die Biotoppräferenzen und Laichzeiten der betroffenen Arten bestätigen diese Interpretation.

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Summary Tadpoles of Rana temporaria which have been raised in an artificial pond prey in the experimental situation shown in Fig. 1 upon the spawn of the following sympatric European Anura species: Rana temporaria (cannibalism), Rana esculenta, Rana ridibunda, Bufo bufo, Bufo calamita, Bombina variegata and Hyla arborea (predation). — Rana temporaria is an early breeder. Thus selection pressure acts in the following ways: Rana temporaria establishes for itself an explosive breeding pattern. Species which breed later in the year in shallow water are suppressed in the biotope of Rana temporaria and develop other biotope preferences (Bombina variegata, Bufo calamita, Hyla arborea). Rana esculenta, Rana ridibunda and Bufo bufo spawn in deeper water. Therefore they are scarcely exposed to the tadpoles of Rana temporaria. Moreover Bufo bufo, too, is an early breeder. Alytes obstetricans laying no spawn in the water is not endangered. — An analysis of biotope preferences and breeding times of the species involved confirms these interpretations.

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Mit Unterstützung des Schweizerischen Nationalfonds zur Förderung der wissenschaftlichen Forschung.     

Mitochondria-rich cells in anuran amphibia: chloride conductance and regional distribution over the body surface

The distribution and density (D(mrc)) of mitochondria-rich cells (MR cells) in skin epithelium, were determined over the whole body surface in nine species of anuran Amphibia that live in a variety of habitats. It was found that the more terrestrial species (beginning with Hyla arborea) have a higher density of MR cells in their pelvic region. In the skin of aquatic (Xenopus laevis) or fossorial (Pelobates syriacus) species, D(mrc) is evenly distributed over the whole body surface. In dorsal skin pieces of H. arborea that lack detectable MR cells, transepithelial voltage activation did not induce Cl(-) conductance as it did in ventral pieces. Skins from Bufo viridis and X. laevis, both have MR cells in their skin, differ markedly in their biophysical properties: a Cl(-) specific current conductance is predominant in the skin epithelium of B. viridis, and is absent in X. laevis. In the latter, anionic conductance is due to glandular secretion. The biophysical properties cannot therefore be related solely to the presence or density of MR cells. Mitochondria-rich cells are sites of Cl(-) conductance across the skin of those amphibians that show this property, but must have different function(s) in other species. It is suggested that the specific zonal distribution of MR cells in the species that were examined in this study could be due to ion exchange activity and water conservation in more terrestrial environments.     

西藏蟾蜍线粒体COI和Cyt b基因序列的分析研究

采用聚合酶链式反应克隆西藏蟾蜍Bufo tibetanus线粒体COI和cyt b基因,首次报道该物种这两个基因的全序列,利用分子生物学软件结合比较与其他7种两栖动物的同源序列.结果显示:西藏蟾蜍两个基因序列中碱基G含量明显低于其它三种碱基,密码子第三位碱基G含量在4种蟾蜍中是最低的;碱基替换主要发生在第三位,属内转换率大于颠换率;相比核苷酸数据,氨基酸序列显示的遗传距离表明氨基酸序列更加保守,遗传距离显示西藏蟾蜍与中华大蟾蜍B.gargarizans的遗传距离最小;构建系统进化树,显示西藏蟾蜍和中华大蟾蜍的亲缘关系最近.     

Using heterochrony plots to detect the dissociated coevolution of characters

The comparison of developmental sequences among species is notoriously difficult. Here, heterochrony plots are introduced as a new graphic method to detect temporal shifts in the development of characters in pair-wise species comparisons. Plotting the timing of character development in one species against the timing of character development in another species allows us to compare a principally unlimited number of characters simultaneously and can detect whether suites of characters are dissociated from one another or not. Such heterochrony plots can be embedded into a comparative phylogenetic analysis in order to establish whether observed patterns of character codissociation are indeed due to their dissociated coevolution. Comparative phylogenetic analysis may also reveal multiple independent events of dissociated coevolution of the same suite of characters in a certain lineage, suggesting that the characters of this suite reciprocally constrain their evolutionary modifiability, thereby forming a unit of evolution. This ability to identify units of evolution is a prerequisite for assessing the validity of recently proposed scenarios, suggesting that modules of development and/or function tend to act as units of evolution. Starting from a detailed heterochrony plot comparing development in the direct developing frog Eleutherodactylus coqui and in the biphasically developing frog Discoglossus pictus, this comparative approach is illustrated focusing on the evolution of development of limbs, the nervous system and the pharyngeal arches in amphibians.