Evolutionary relationships of the lungless caecilian Atretochoana eiselti (Amphibia: Gymnophiona: Typhlonectidae)

Appreciation of the diversity of caecilian amphibians has recently been enhanced by the discovery of a radically divergent aquatic caecilian of the Neotropical Typhlonectidae. Atretochoana eiselti is the largest lungless tetrapod and the only lungless caecilian, and it possesses a suite of remarkable cranial modifications that set it apart from all other caecilians. Numerical phylogenetic analyses, using 141 morphological characters, were performed in order to resolve the evolutionary relationships of Atretochoana and representatives of all other typhlonectid genera. These analyses yield a single most parsimonious tree, (Chthonerpeton (Nectocaecilia (Typhlonectes natans, Typhlonectes compressicauda) (Potomotyphlus, Atretochoana)))) , that is both well resolved and, as judged by Bremer support and by bootstrapping, is well supported. This tree is used as a basis for interpreting ecological shifts and associated morphological evolution within the Typhlonectidae. The available data suggest that the rate of morphological evolution in the Atretochoana lineage is significandy greater than that in other typhlonectid lineages.     

Discovery of a new family of amphibians from northeast India with ancient links to Africa

The limbless, primarily soil-dwelling and tropical caecilian amphibians (Gymnophiona) comprise the least known order of tetrapods. On the basis of unprecedented extensive fieldwork, we report the discovery of a previously overlooked, ancient lineage and radiation of caecilians from threatened habitats in the underexplored states of northeast India. Molecular phylogenetic analyses of mitogenomic and nuclear DNA sequences, and comparative cranial anatomy indicate an unexpected sister-group relationship with the exclusively African family Herpelidae. Relaxed molecular clock analyses indicate that these lineages diverged in the Early Cretaceous, about 140 Ma. The discovery adds a major branch to the amphibian tree of life and sheds light on both the evolution and biogeography of caecilians and the biotic history of northeast India-an area generally interpreted as a gateway between biodiversity hotspots rather than a distinct biogeographic unit with its own ancient endemics. Because of its distinctive morphology, inferred age and phylogenetic relationships, we recognize the newly discovered caecilian radiation as a new family of modern amphibians.     

Sexual size dimorphism in caecilian amphibians: analysis, review and directions for future research

Sexual dimorphism, widespread in the animal kingdom, describes differences between the sexes in size, shape and many other traits. Sexual size dimorphism (SSD) plays a significant role in understanding life history evolution and mating systems. The snakelike morphology of limbless caecilian amphibians lacking obvious secondary sexual characters (in contrast to frogs and salamanders) impedes accurate intrasexual comparisons. In this study, sexual size dimorphism in the oviparous caecilian Ichthyophis cf. kohtaoensis, a phylogenetically basal caecilian, was analysed. Females were larger in all body and head characters tested. However, when adjusted to body size (total length), females differed only in their cloacal shape. Clutch volume was positively correlated to female body size, thus female fecundity increased with body size supporting the hypothesis of a fecundity-selected SSD in the oviparous Ichthyophis cf. kohtaoensis. A review of the present SSD data for caecilians shows that many species are monomorphic for body size but show dimorphism in head size, while other species demonstrate female-biased SSD. Male-biased SSD has not been reported for caecilians. To understand life history evolution in caecilians, further studies on the reproductive biology of other taxa are urgently needed, in particular for rhinatrematids and uraeotyphlids. New data will allow phylogenetically controlled comparative analyses to fully explore the pattern of SSD among caecilian lineages.     

The evolution of parental investment in caecilian amphibians: a comparative approach

Parental care is widespread among vertebrates and the observed patterns of parental care and investment are extremely diverse. Among amphibians, caecilians (Gymnophiona) exhibit considerable variation in reproductive modes, including both oviparity and viviparity, combined with highly unusual investment strategies (e.g. skin‐feeding and intrauterine feeding). In the present study, current knowledge on the reproductive modes is integrated into an analysis of the evolutionary scenario of parental investment of caecilians. Phylogenetically basal caecilians possessing a biphasic life cycle that includes an aquatic larval stage invest in macrolecithal eggs directly corresponding to size at hatching. Some phylogenetically derived caecilians (i.e. the Teresomata) have a smaller clutch size and show a reduction to either medium‐yolked (mesolecithal) or small‐yolked (microlecithal) eggs. Via alternative pathways of parental investment, such as intrauterine feeding in viviparous taxa and maternal dermatotrophy in oviparous taxa, teresomatan caecilians increase both offspring size and quality. However, more data regarding reproductive biology are needed to obtain a fully resolved understanding of the evolution of reproduction in caecilian amphibians.     

The Braincase of Eocaecilia micropodia (Lissamphibia,Gymnophiona) and the Origin of Caecilians

The scant fossil record of caecilians has obscured the origin and evolution of this lissamphibian group. Eocaecilia micropodia from the Lower Jurassic of North America remains the only stem-group caecilian with an almost complete skull preserved. However, this taxon has been controversial, engendering re-evaluation of traits considered to be plesiomorphic for extant caecilians. Both the validity of the placement of E. micropodia as a stem caecilian and estimates of the plesiomorphic condition of extant caecilians have been questioned. In order to address these issues, the braincase of E. micropodia was examined via micro-computed tomography. The braincase is considered to be a more reliable phylogenetic indicator than peripheral regions of the skull. These data reveal significant new information, including the possession of an ossified nasal septum, ossified anterior wall of the sphenethmoid, long anterolateral processes on the sphenethmoid, and paired olfactory nerve foramina, which are known only to occur in extant caecilians; the latter are possibly related to the evolution of the tentacle, a caecilian autapomorphy. A phylogenetic analysis that included 64 non-amniote taxa and 308 characters represents the first extensive test of the phylogenetic affinities of E. micropodia. The results place E. micropodia securely on the stem of extant caecilians, representing a clade within Temnospondyli that is the sister taxon to batrachians plus Gerobatrachus. Ancestral character state reconstruction confirms the braincase of E. micropodia to be largely representative of the plesiomorphic condition of extant caecilians. Additionally, the results refine the context within which the evolution of the caecilian form can be evaluated. The robust construction and pattern of the dermal skull of E. micropodia is interpreted as symplesiomorphic with advanced dissorophoid temnospondyls, rather than being autapomorphic in its robust construction. Together these data increase confidence in incorporating E. micropodia into discussions of caecilian evolution.     

Mitochondrial evidence on the phylogenetic position of caecilians (Amphibia: Gymnophiona)

The complete nucleotide sequence (17,005 bp) of the mitochondrial genome of the caecilian Typhlonectes natans (Gymnophiona, Amphibia) was determined. This molecule is characterized by two distinctive genomic features: there are seven large 109-bp tandem repeats in the control region, and the sequence for the putative origin of replication of the L strand can potentially fold into two alternative secondary structures (one including part of the tRNA(Cys)). The new sequence data were used to assess the phylogenetic position of caecilians and to gain insights into the origin of living amphibians (frogs, salamanders, and caecilians). Phylogenetic analyses of two data sets-one combining protein-coding genes and the other combining tRNA genes-strongly supported a caecilian + frog clade and, hence, monophyly of modern amphibians. These two data sets could not further resolve relationships among the coelacanth, lungfishes, and tetrapods, but strongly supported diapsid affinities of turtles. Phylogenetic relationships among a larger set of species of frogs, salamanders, and caecilians were estimated with a mitochondrial rRNA data set. Maximum parsimony analysis of this latter data set also recovered monophyly of living amphibians and favored a frog + salamander (Batrachia) relationship. However, bootstrap support was only moderate at these nodes. This is likely due to an extensive among-site rate heterogeneity in the rRNA data set and the narrow window of time in which the three main groups of living amphibians were originated.     

A mitogenomic perspective on the phylogeny and biogeography of living caecilians (Amphibia: Gymnophiona)

The caecilians, members of the amphibian Order Gymnophiona, are the least known Order of tetrapods, and their intra-relationships, especially within its largest group, the Family Caeciliidae (57% of all caecilian species), remain controversial. We sequenced thirteen complete caecilian mitochondrial genomes, including twelve species of caeciliids, using a universal primer set strategy. These new sequences, together with eight published caecilian mitochondrial genomes, were analyzed by maximum parsimony, partitioned maximum-likelihood and partitioned Bayesian approaches at both nucleotide and amino acid levels, to study the intra-relationships of caecilians. An additional multiple gene dataset including most of the caecilian nucleotide sequences currently available in GenBank produced phylogenetic results that are fully compatible with those based on the mitogenomic data. Our phylogenetic results are summarized as follow. The caecilian family Rhinatrematidae is the sister taxon to all other caecilians. Beyond Rhinatrematidae, a clade comprising the Ichthyophlidae and Uraeotyphlidae is separated from a clade containing all remaining caecilians (Scolecomorphidae, Typhlonectidae and Caeciliidae). Within this large clade, Scolecomorphidae is the sister taxon of Typhlonectidae and Caeciliidae but this placement did not receive strong support in all analyses. Caeciliidae is paraphyletic with regard to Typhlonectidae, and can be divided into three well-supported groups: Caeciliidae group 1 contains the African caeciliids Boulengerula and Herpele; Caeciliidae group 2 contains Caecilia and Oscaecilia and it is the sister taxon of Typhlonectidae; Caeciliidae group 3 comprises the remaining species of caeciliids. The mitochondrial genome data were also used to calculate divergence times for caecilian evolution using the penalized likelihood method implemented in the program R8S. The newly obtained dating results are compatible with (but a little older than) previous time estimates mainly based on nuclear gene data. The mitogenomic time tree of caecilians suggests that the initial diversification of extant caecilians most probably took place in Late Triassic about 228 (195–260) Ma. Caeciliids currently distributed in India and the Seychelles diverged from their African and American relatives most probably in Late Jurassic about 138 (112–165) Ma, fairly close to the time (130 Ma) when Madagascar–India–Seychelles separated from Africa and South America. The split between the Indian caeciliid Gegeneophis and Seychellean caeciliids occurred about 103 (78–125) Ma, predated the rifting of India and the Seychelles (65 Ma).     

Phylogenetic relationships of Indian caecilians (Amphibia: Gymnophiona) inferred from mitochondrial rRNA gene sequences

India has a diverse caecilian fauna, including representatives of three of the six currently recognized families, the Caeciliidae, Ichthyophiidae, the endemic Uraeotyphlidae, but previous molecular phylogenetic studies of caecilians have not included sequences for any Indian caecilians. Partial 12S and 16S mitochondrial gene sequences were obtained for a single representative of each of the caecilian families found in India and aligned against previously reported sequences for 13 caecilian species. The resulting alignment (16 taxa, 1200 sites, of which 288 cannot be aligned unambiguously) was analyzed using parsimony, maximum-likelihood, and distance methods. As judged by bootstrap proportions, decay indices, and leaf stabilities, well-supported relationships of the Indian caecilians are recovered from the alignment. The data (1) corroborate the hypothesis, based on morphology, that the Uraeotyphlidae and Ichthyophiidae are sister taxa, (2) recover a monophyletic Ichthyophiidae, including Indian and South East Asian representatives, and (3) place the Indian caeciliid Gegeneophis ramaswamii as the sister group of the caeciliid caecilians of the Seychelles. Rough estimates of divergence times suggest an origin of the Uraeotyphlidae and Ichthyophiidae while India was isolated from Laurasia and Africa and are most consistent with an Indian origin of these families and subsequent dispersal of ichthyophiids into South East Asia.     

A comparative study of locomotion in the caecilians Dermophis mexicanus and Typhlonectes natans (Amphibia: Gymnophiona)

We compared locomotion of two species of caecilian using x-ray videography of the animals traversing smooth-sided channels and a pegboard. Two channel widths were used, a body width channel and a body width + 20% channel. The terrestrial caecilian, Dermophis mexicanus , used internal concertina locomotion in both channels and lateral undulation on the pegboard. The aquatic caecilian, Typhlonectes natans , was not able to move at all in the body width channel. In the wider channel Typhlonectes proceeded at the same speed as Dermophis while using normal, rather than internal, concertina locomotion. On the pegboard, Typhlonectes used lateral undulation and achieved 2.5 times the speed managed by Dermophis. A phylogenetic analysis of this, and other, evidence shows that (1) internal concertina evolved in the ancestor to extant caecilians and (2) internal concertina locomotion was secondarily lost in the aquatic caecilians.     

Morphometric analysis of the skull of Dermophis mexicanus (Amphibia: Gymnophiona)

The levels of integration, patterns of allometric growth and size-related differences in skull dimensions of the caecilian Dermophis mexicanus were investigated by means of univariate, bivariate and multivariate statistics. The analyses indicate that (1) most but not all measurements of the skull arc more variable in adults than in juveniles; (2) growth is allometric and results in changes in shape between juveniles and adults; (3) there is a high level of integration of all variables by general size; and (4) homologous paired measurements show concordant allometric trends and are integrated beyond the sole effect of general size. These analyses provide baseline data, a set of hypotheses and a methodological framework for comparative studies of patterns of variation and integration among caecilians and among other vertebrates.     

Phylogenetic implications of the morphology of the braincase of caecilian amphibians (Gymnophiona)

Caecilian morphology is strongly modified in association with their fossorial mode of life. Currently phylogenetic analyses of characters drawn from the morphology of caecilians lack resolution, as well as complementarity, with results of phylogenetic analyses that employ molecular data. Stemming from the hypothesis derived from the mammal literature that the braincase has the greatest potential (in comparison to other cranial units) to yield phylogenetic information, the braincase and intimately associated stapes of 27 species (23 genera) of extant caecilians were examined using images assembled via microcomputed tomography. Thirty‐four new morphological characters pertaining to the braincase and stapes were identified and tested for congruence with previously recognized morphological characters. The results reveal that when added to previous character matrices, characters of the braincase and stapes resolve generic‐level relationships in a way that is largely congruent with the results of molecular analyses. Analysis of a combined data set of molecular and morphological data provides a framework for conducting ancestral character state reconstructions, which resulted in the identification of 95 new synapomorphies for various clades and taxa, 27 of which appear to be unique for the taxa that possess them. Together these data demonstrate the utility of the application of characters of the braincase and stapes for resolving phylogenetic relationships for a group whose morphology is largely confounded by functional modifications. In addition this study provides evidence of the utility of the braincase in resolving problematic morphology‐based phylogeny outside of Amniota, in an amphibian group. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 166 , 160–201.     

Cellular migration and morphological complexity in the caecilian brain

The morphology of the tectum mesencephali and the medial pallium is studied in species representing the six families of caecilians (Amphibia: Gymnophiona) in order to determine whether differences in brain morphology are related to function, phylogenetic history, or life history strategies. In general, the caecilian tectum is characterized by simplification in having little to no lamination and few migrated cells. The degree of morphological complexity differs between species and between brain regions. Our data suggest that changes in brain morphology are due to a mosaic of different influences. We did not find a strict correlation between visual system reduction and tectal morphologies. However, phylogenetic effects exist. The greatest degree of morphological complexity is found in members of the Rhinatrematidae, a family that is considered basal to the lineage. Thus, simplification of brain morphology in caecilians must be considered a secondary or derived rather than a primitive feature. Direct development and miniaturization are correlated with the greatest simplification in the tectum mesencephali and medial pallium. There is a relationship between differences in brain morphology and heterochrony in caecilians, as in other amphibians. J. Morphol. 231:11–27, 1997. © 1997 Wiley-Liss, Inc.     

Comparative morphology of caecilian sperm (Amphibia: Gymnophiona)

The morphology of mature sperm from the testes of 22 genera and 29 species representing all five families of caecilians (Amphibia: Gymnophiona) was examined at the light microscope level in order to: (1) determine the effectiveness of silver-staining techniques on long-preserved, rare material, (2) assess the comparative morphology of sperm quantitatively, (3) compare patterns of caecilian sperm morphology with that of other amphibians, and (4) determine if sperm morphology presents any characters useful for systematic analysis. Although patterns of sperm morphology are quite consistent intragenerically and intrafamilially, there are inconsistencies as well. Two major types of sperm occur among caecilians: those with very long heads and pointed acrosomes, and those with shorter, wider heads and blunt acrosomes. Several taxa have sperm with undulating membranes on the flagella, but limitations of the technique likely prevented full determination of tail morphology among all taxa. Cluster analysis is more appropriate for these data than is phylogenetic analysis. © 1994 Wiley-Liss, Inc.     

A field guide to the caecilians of the Western Ghats, India

Caecilians are legless amphibians quite characteristic of the Western Ghats. Fourteen out of 16 Indian species occur in the Western Ghats and all are endemic. The present paper deals with the biology of caecilians with reference to external morphology and general breeding behaviour. It consolidates information on 26 morphological parameters which are used in caecilian identification. Metric multidimensional scaling of 16 species of caecilians using pair-wise euclidian distances calculated on the basis of 11 important morphometric parameters clearly depicts morphological distances between different species and more so the genera, thereby validating the classification. A field guide has been developed for the identification of caecilians based on a survey made all over the Western Ghats, observation of holotypes at the Natural History Museum, London and review of the literature. The study also reveals the microhabitat requirements of the caecilians. Further, the localities of caecilian distribution in the Western Ghats are mapped. The taxonomy of Indian caecilians is discussed.     

Development of the tectum in Gymnophiones,with comparison to other amphibians

Tectal development in a number of caecilian (Gymnophiona: Amphibia) species was examined and compared with that in frogs and salamanders. The caecilian optic tectum develops along the same rostrocaudal and lateromedial gradients as those of frogs and salamanders. However, differences exist in the time course of development. Our data suggest that, as in salamanders, simplification of morphological complexity in caecilians is due to a retardation or loss of late developmental stages. Differences in the time course of development (heterochrony) among different caecilian species are correlated with phylogenetic history as well as with variation in life histories. The most pronounced differences in development occur between the directly developing Hypogeophis rostratus and all other species examined. In this species, the increase in the degree of morphological complexity is greatly accelerated. J. Morphol. 236:233–246, 1998. © 1998 Wiley-Liss, Inc.     

A stem-group caecilian (Lissamphibia: Gymnophiona) from the Lower Cretaceous of North Africa

Of living amphibian groups, the limbless burrowing caecilians are amongst the most highly specialised, but are the least known. Their fossil record is extremely poor, leaving unresolved questions as to their origins, relationships and early distribution. We describe here caecilian remains from a Lower Cretaceous (Berriasian) microfossil locality near Anoual, Morocco. This material represents the second oldest record for the group, after the Jurassic Eocaecilia of North America, and the earliest caecilian record for Gondwana. It forms the basis of a new genus, Rubricacaecilia , which appears slightly more derived than Eocaecilia , but lacks major features of crown-group taxa. We support the use of Apoda Oppel, 1811 for the crown-group alone, and Gymnophiona Rafinesque 1814 for the clade comprising stem-group taxa + Apoda.     

Trophic ecology of East African caecilians (Amphibia: Gymnophiona), and their impact on forest soil invertebrates

Diets of the syntopic caecilian amphibians Boulengerula boulengeri and Scolecomorphus vittatus were studied from gut content analyses of 108 specimens collected from Nilo Forest Reserve, East Usambara Mountains, Tanzania. Head length and width relative to body length is greater in male B. boulengeri than in females, but no such dimorphism is apparent in S. vittatus . No differences, other than females consuming disproportionately more ants and invertebrate eggs and fewer termites than males, occur in the diets of B. boulengeri . The gut contents of S. vittatus are dominated by large, surface-active earthworms, in contrast to those of B. boulengeri which contained smaller, endogeic earthworms and a much greater number and diversity of soil arthropods (particularly termites, ants and Diptera larvae). These dietary characteristics underline differences in microhabitat use between these caecilian species and thus corroborate a previously proposed hypothesis of niche separation. The ecological impact of caecilians and other limbless endogeic vertebrates is little studied and poorly known. Approximations based on current knowledge are not clear, but are interpreted as indicating that predation by caecilians is unlikely to be among the most important factors influencing population densities of soil-dwelling termites and perhaps also ants: two groups recognized as soil ecosystem engineers.     

Bayesian and Maximum Likelihood Analyses of Rotifer–Acanthocephalan Relationships

Rotifera is composed of groups with unusual ultrastructural, physiological, and reproductive characters. Our ability to understand the evolution of these features is complicated by the fact that the phylogenetic relationships among the three traditional rotifer groups (Seisonidea, Monogononta, and Bdelloidea) and Acanthocephala remain unresolved. Here, I present maximum likelihood and Bayesian analyses of rotifer–acanthocephalan relationships using both the protein-coding gene hsp82 and a combined data set of hsp82 and ribosomal small subunit (SSU) DNA sequences, using nucleotide and codon based models of evolution. Statistical analysis of the phylogenetic support for any of the likely relationships among rotifer groups suggests that more than a combined hsp82 + SSU data set will be needed to resolve rotifer–acanthocephalan phylogeny with any degree of certainty.     

Phylogeny of caecilian amphibians (Gymnophiona) based on complete mitochondrial genomes and nuclear RAG1

We determined the complete nucleotide sequence of the mitochondrial (mt) genome of five individual caecilians (Amphibia: Gymnophiona) representing five of the six recognized families: Rhinatrema bivittatum (Rhinatrematidae), Ichthyophis glutinosus (Ichthyophiidae), Uraeotyphlus cf. oxyurus (Uraeotyphlidae), Scolecomorphus vittatus (Scolecomorphidae), and Gegeneophis ramaswamii (Caeciliidae). The organization and size of these newly determined mitogenomes are similar to those previously reported for the caecilian Typhlonectes natans (Typhlonectidae), and for other vertebrates. Nucleotide sequences of the nuclear RAG1 gene were also determined for these six species of caecilians, and the salamander Mertensiella luschani atifi. RAG1 (both at the amino acid and nucleotide level) shows slower rates of evolution than almost all mt protein-coding genes (at the amino acid level). The new mt and nuclear sequences were compared with data for other amphibians and subjected to separate and combined phylogenetic analyses (Maximum Parsimony, Minimum Evolution, Maximum Likelihood, and Bayesian Inference). All analyses strongly support the monophyly of the three amphibian Orders. The Batrachia hypothesis (Gymnophiona, (Anura, Caudata) receives moderate or good support depending on the method of analysis. Within Gymnophiona, the optimal tree (Rhinatrema, (Ichthyophis, Uraeotyphlus), (Scolecomorphus, (Gegeneophis Typhlonectes) agrees with the most recent morphological and molecular studies. The sister group relationship between Rhinatrematidae and all other caecilians, that between Ichthyophiidae and Uraeotyphlidae, and the monophyly of the higher caecilians Scolecomorphidae+Caeciliidae+Typhlonectidae, are strongly supported, whereas the relationships among the higher caecilians are less unambiguously resolved. Analysis of RAG1 is affected by a spurious local rooting problem and associated low support that is ameliorated when outgroups are excluded. Comparisons of trees using the non-parametric Templeton, Kishino-Hasegawa, Approximately Unbiased, and Shimodaira-Hasegawa tests suggest that the latter may be too conservative.