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Researchers find missing evolutionary link between frogs and salamanders

Researchers from the University of Calgary, the University of Toronto and McGill University have solved one of the greatest mysteries in vertebrate evolution.

Dr. Jason Anderson of the University of Calgary Faculty of Veterinary Medicine, Dr. Robert Reisz of the University of Toronto and Nadia Fröbisch, a PhD candidate at McGill University's Redpath Museum and colleagues, have proven conclusively that all modern frogs, toads and salamanders descend from a common amphibian ancestor that existed over 200 million years ago. Their results were published May 21 in the journal Nature.

Their findings derive from exhaustive research conducted on Gerobatrachus hottoni (or "Hotton's elder frog,") an ancient fossil discovered in Texas in 1995 by the late Smithsonian Institution researcher Nicholas Hotton. According to the University of Calgary, the fossil went unstudied until it was later "rediscovered" by Dr. Anderson's team.

Gerobachtrachus exhibits a mixture of salamander and frog features in its skull, backbone and teeth.

Moreover, Fröbisch discovered that it had two fused bones in its ankle. "This is a feature usually uniquely seen in salamanders and no other group," she explained.

"The question of whether frogs, toads and salamanders had a common ancestor has been plaguing science for a very long time," Fröbisch said. "There have actually been papers on this topic for 120 years or more."

"The dispute arose because of a lack of transitional forms. This fossil seals the gap," said Anderson, lead author of the study.

"It's very exciting to be doing this kind of breakthrough research," added Fröbisch. "It's actually kind of surreal."

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Frogs and salamanders and a common ancestor, oh my!

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Researchers find missing evolutionary link between frogs and salamanders Researchers from the University of Calgary, the University of Toronto and McGill University have solved one of the greatest mysteries in vertebrate evolution.

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The genus Rhacophorus is unique to house some of the biggest species of flying frogs.

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Episode 2 of David Attenborough's Conquest of the Skies was on TV the other day, and I watched it (I livetweeted throughout, mostly because I wanted to talk about their portrayal of pterosaurs and Mesozoic theropods). And hence I have rhacophorid frogs on my mind - the mostly tropical Afro-Asian group that includes the famous Rhacophorus flying frogs, the best known member of which is Wallace's flying frog R. nigropalmatus from Indonesia, Thailand and adjacent countries. As usual, flying frog were used by Sir David to help illustrate the diversity of animals that have evolved a gliding ability.

Rhacophorids are sometimes called flying frogs, shrub frogs, bush frogs, moss frogs, Old World treefrogs, or Afro-Asian treefrogs, and occur across sub-Saharan Africa, China, much of tropical Asia, Japan, the Philippines and Sulawesi. About 380 species are recognised as of early 2015. The last time I wrote about this group - December 2008 - this number was more like 290, so the rate at which new species are discovered and named is pretty impressive.

A nice illustration of this is provided by Meegaskumbura et al.'s (2002) documentation of more than 100 new rhacophorid species on Sri Lanka alone (just 18 Sri Lankan rhacophorid species were known prior to their work), a discovery that makes Sri Lanka on par with Madagascar, New Guinea and Borneo in terms of anuran diversity.

And, yes, more than 100 new species announced in a single paper. If we look at the discovery record of various of the rhacophorid lineages, we see that - for example - 43 new species of Raorchestes, 30 new species of Rhacophorus, and 51 new species of Pseudophilautus have been named since 2000... 9 new Raorchestes species were named in 2014 alone (Frost 2014). As should be well known, the number of recognised amphibian species has sky-rocketed in recent years, and this really is because of newly discovered species, not just the result of splitting, taxonomic elevation of subspecies, or the recognition of cryptic species that can only be distinguished genetically.

The great paradox is that amphibians are in chronic global decline at the same time, and many species can no longer be located at all. Despite Meegaskumbura et al.'s (2002) 100+ new rhacophorid species, they were unable to find many that had been described in the 19th century, a discovery which implies that the species concerned have gone extinct. As you should also know, amphibian species are currently being 'lost' on a regular and worrying basis - we don't talk of a 'global amphibian crisis' for nothing.

Most rhacophorids are arboreal or semi-arboreal, living in shrubs, trees and bushes from close to ground level to way up in the forest canopy. There's some uncertainty over how high up in the canopy these frogs actually occur. Nature documentaries (like the aforementioned Attenbourough-led projects) create the impression that they really live tens of metres up in the high canopy but this is hard to confirm and has been doubted on occasion. Recently, however, individuals of some species (like Rhacophorus belalongensis on Borneo, named in 2008) have been recorded from tree-tops 10 m high. Members of some groups are associated with primary forests, but others inhabit agricultural fields, roadsides, cleared forest and villages. [Images below by Σ64 and Alpsdake.]
Such reproductive diversity
Specialised reproductive strategies are widespread across these frogs, and some of the techniques they use mean that they don't have to come down to the ground to breed. Some (like some Philautus species) stick their eggs to the undersides of leaves above the ground and some Philautus species (like P. mjobergi) have been reported to be nepenthiphilous - that is, to lay their eggs inside pitcher plants. While some frogs definitely are nepenthiphilous, the only alleged rhacophorid eggs discovered inside a pitcher plant and subjected to molecular testing turned out to be from the microhylid species Microhyla borneensis (Hertwig et al. 2012). [Photo below by Katja Rembold.]

Those Philautus eggs, by the way, don't produce free-swimming tadpoles: Philautus species are direct developers, which means that the embryos change to froglets within the eggs, a free-living tadpole phase being absent (the developing embryos are lecithotrophic or endotrophic, meaning that they depend on a yolk store). Direct development is also the case in Pseudophilautus and Raorchestes.

While (as just mentioned) some of these direct developers stick their eggs to leaves that are alive and well above ground-level, others come down to the ground and lay their eggs beneath dead leaves. Meegaskumbara et al. (2007) said that these ground-breeding species "deposit their eggs in nests excavated on the forest floor" (p. 9). Waitaminute - frogs excavating nests? Really? I have to look into this...
The foam-nesters
Then there are those rhacophorids that manufacture arboreal foam nests [adjacent nest photos by Alpsdake and Brian Gratwicke]. The females exude a secretion that they (and their male partners) whip up with their legs to form a foam clump that's attached to leaves, branches or aerial roots. It seems that the production of this secretion is quite costly and that a female needs to take a break and re-hydrate herself by soaking up standing water before she can complete a single nest.

This strategy is present in the Afro-Asian foam-nest frogs Chiromantis, most Rhacophorus species, and members of Polypedates. In some species - most famously the Grey foam nest treefrog C. xerampelina of south-eastern Africa - large numbers of these frogs sometimes choose to nest in the same place, meaning that branches or aerial roots can be festooned with whole lines of dripping foam nests. Actually, it isn't just that the frogs 'choose' to nest in the same place - males will deliberately get in on the action if they see a pair working to make a nest, and the result is that single egg clutches are invariably fertilised by more than one male. Byrne & Whiting (2011) showed that this multiple paternity - technically, it's simultaneous polyandry - assists in the survival of the resulting offspring, so it's certainly in the interests of females to solicit as much male attention as possible during these breeding events. [Photos below by Brian Gratwicke, Kapenta and Chintan Sheth.]

The outsides of these foam nests dry to form a hard crust, thereby protecting the eggs within. However, monkeys, snakes and other predators will break into the nests and eat the eggs if they can. Most surprisingly, Fornasini's spiny reed frog Afrixalus fornasini (a member of Hyperoliidae) is a documented foam nest predator, though it can only eat from the nest before the foam has dried (Channing 2001).

The eggs hatch inside the clump, the tadpoles dropping into the stream or pool (sometimes originally formed by rhinos or pigs) below after several days. Polypedates leucomystax bucks the trend by sometimes making foam nests on the ground (Inger & Stuebing 2005). [Adjacent photo by W.A. Djatmiko]. It seems that foam-nesting evolved just once within rhacophorids, since all foam-nesters belong to a single clade ( Frost et al. 2006, Grosjean et al. 2008, Pyron & Wiens 2011).

Foam-nester tadpoles are ectotrophic: free-swimming and completing development outside the egg, and often with a schooling habit. Some live in muddy pools and are of typical, non-specialised morphology. Others (like those of Rhacophorus penanorum) are specialised stream-dwellers with streamlined bodies, sucker-like mouths and elongate, muscular tails. These tadpoles are rheophilous (associated with fast-flowing streams) and inhabit rocky pools that are sometimes also home to Megophrys/ Xenophrys spadefoot tadpoles and Ansonia toad tadpoles (Haas et al. 2012).

A really interesting thing that's been noted for rheophilous tadpoles is that their limb development seems to be offset, time-wise, relative to the condition in related, non-rheophilous species. This is presumably an evolutionary response to the fact that developing hindlimbs might affect their streamlining and ability to cling to rocks in fast-flowing water. They also keep sucker-like mouths and other features for longer than do other tadpoles (Nodzenski & Inger 1990). Accordingly, it can be difficult to say reliable things about their age and estimated metamorphic stage.

Finally, there are yet other rhacophorids where egg clumps are laid in arboreal settings, but not in foam nests. In some Theloderma species, egg clumps are laid in water-filled tree hollows, and the tadpoles complete their development here. In captivity, these frogs will lay their egg clumps attached to the bark, just above the hollow, the hatching tadpoles then dropping into the water (Tapley 2009). Oh, there are also a few foam-nesting Rhacophorus species that lay their eggs in tree hollows, the most famous of which is R. vampyrus from Vietnam (after hatching inside a foam nest, the tadpoles drop into a water-filled tree hollow). This species saw international stardom a couple of years ago when it was revealed that the tadpoles have black, hooked fangs on the lower jaw that -it's presumed - are used when feeding on unfertilised eggs provided by their mother: these tadpoles, it seems, practise obligate oophagy, eating R. vampyrus eggs and nothing else ( Vassilieva et al. 2013).

Finally finally, the possibility exists that a completely unique reproductive strategy was present in a species that now seems to be extinct. The holotype female specimen of Philautus maia, collected on Sri Lanka prior to 1876, had a disc-shaped mass of eggs attached to its belly, raising the possibility that members of this species carried their eggs around with them (Günther 1876). Alas, Meegaskumbura et al. (2007) discussed how unlikely this proposal was, concluding that a more plausible possibility is that the individual concerned was collected while in the process of laying and positioning her egg clutch on a leaf. Alas, only further observations can establish the 'truth' and... sadly, P. maia is one of those many, many frog species that seems to have become extinct between its 19th century discovery and the present. Don't forget: Global. Amphibian. Crisis.
Mossy-skinned, warty-skinned, smooth-skinned, and with winglets
External appearance is variable in rhacophorids. For all their fame as 'flying frogs', it has to be said that the vast majority look - to those unenlightened in anuran diversity - like standard 'treefrogs'. They're generally small (SVLs of 40 mm or less), wide-headed, big-eyed frogs with expanded digit-tips and a (normally) prominent tympanum. Many are smooth-skinned but spiny tubercles cover the skin in some taxa, and others are notably warty, with a rough, bumpy skin that aids camouflage.

The latter is most developed in the grotesque Rough treefrog Theloderma horridum of Thailand, peninsula Malaysia and Borneo. Indeed, this is one of several species (most of which belong to Theloderma) that resemble moss or bark in external texture and colour [adjacent photos of T. corticale by Steven G. Johnson and Václav Gvoždík]. T. asperum - patterned in brownish and pale blotches - superficially resembles a bird dropping and is sometimes called the Bird poop frog. Vocal sacs are absent in some taxa (like Nyctixalus) but big and obvious in others.

Wallace's flying frog and the other gliding Rhacophorus species are pretty remarkable. They're very big compared to most other members of the group, SVLs being 90-100 mm in females and 80-90 mm in males. Their fully webbed hands and feet are enormous, but they also have flaps of skin - winglets, if you like - on the arms and legs, and sometimes on the body. Glides of more than 15 m have been recorded. Exactly how many Rhacophorus species are true gliders is uncertain: the ability is confirmed for just a handful of species but more may have it (Inger & Stuebing 2005). Several smaller ones (including R. angulirostris, R. cyanopunctatus and R. gauni) have only partial digital webbing and either lack those skin flaps or only have small versions.

Finally, where do rhacophorids fit within the anuran radiation? Molecular studies find them to be close to Ranidae, the familiar neobatrachian clade that includes European water frogs, brown frogs, the American bullfrog, leopard frogs and so many others ( Frost et al. 2006, Pyron & Wiens 2011). They're clearly not at all close to hylid treefrogs (hylids are part of the same clade as glassfrogs, toads and kin). I also need to say that a huge amount of work - scarcely any of which is cited in the article you're reading now - has recently been devoted to the in-group relationships of Rhacophoridae, several conventional 'genera' being the subject of substantial disagreement due to proposals that they might be paraphyletic or polyphyletic. At the risk of elaborating further, I must stop here. Oh, I seem to have blogged about anurans again.

For previous Tet Zoo articles on frogs and toads, see...

Refs - -
Byrne, P. G. & Whiting, M. J. 2011. Effects of simultaneous polyandry on offspring fitness in an African tree frog. Behavioral Ecology 22, 385-391.
Channing, A. 2001. Amphibians of Central and Southern Africa. Cornell University Press, Ithaca and London.

Frost, D. R. 2014. Amphibian Species of the World: an Online Reference. Version 6.0 (30th December 2014). Electronic Database accessible at http://research.amnh.org/herpetology/amphibia/index.html. American Museum of Natural History, New York, USA.
- ., Grant, T., Faivovich, J., Bain, R. H., Haas, A., Haddad, C. F. B., De Sá, R. O., Channing, A., Wilkinson, M., Donnellan, S. C., Raxworthy, C. J., Campbell, J. A., Blotto, B. L., Moler, P., Drewes, R. C., Nussbaum, R. A., Lynch, J. D., Green, D. M. & Wheeler, W. C. 2006. The amphibian tree of life. Bulletin of the American Museum of Natural History 297, 1-370.
Grosjean, S., Delorme, M., Dubois, A. & Ohler, A. 2008. Evolution of reproduction in the Rhacophoridae (Amphibia, Anura). Journal of Zoological Systematics and Evolutionary Research 462, 169-176.

Günther, A. 1876. Note on the mode of propagation of some Ceylonese tree-frogs, with description of two new species. Annals and Magazine of Natural History (4) 17, 377-380.

Haas, A., Hertwig, S. T., Krings, W., Braskamp, E., Dehling, J. M., Min, P. Y., Jankowski, A., Schweizer, M. & Das, I. 2012. Description of three Rhacophorus tadpoles (Lissamphibia: Anura: Rhacophoridae) from Sarawak, Malaysia (Borneo). Zootaxa 3328, 1-19.

Hertwig, S. T., Lilje, K. E., Min, P. Y., Haas, A. & Das, I. 2012. Molecular evidence for direct development in the rhacophorid frog, Philautus acutus (Rhacophoridae, Anura) from Borneo. The Raffles Bulletin of Zoology 60, 559-567.

Inger, R. F. & Stuebing, R. B. 2005. A Field Guide of the Frogs of Borneo. Natural History Publications (Borneo), Kota Kinabalu.
Meegaskumbura, M., Bossuyt, F., Pethiyagoda, R., Manamendra-Arachchi, K., Bahir, M., Milinkovitch, M. C. & Schneider, C. J. 2002. Sri Lanka: an amphibian hotspot. Science 298, 379.
- ., Manamendra-Arachchi, K., Schneider, C. J. & Pethiyagoda, R. 2007. New species amongst Sri Lanka's extinct shrub frogs (Amphibia: Rhacophoridae: Philautus). Zootaxa 1397, 1-15.

Nodzenski, E. & Inger, R. F. 1990. Uncoupling of related structural changes in metamorphosing torrent-dwelling tadpoles. Copeia 1990, 1047-1054.
Pyron, A. R. & Wiens, J. J. 2011. A large-scale phylogeny of Amphibia including over 2800 species, and a revised classification of extant frogs, salamanders, and caecilians. Molecular Phylogenetics and Evolution 61, 543-583.
Tapley, B. 2009. Aspects of captive husbandry of Taylor's Bug-eyed Frog, Theloderma stellatum (Taylor, 1962). Herpetological Bulletin 108, 31-33.
Vassilieva, A., Galoyan, E. & Poyarkov, N. 2013. Rhacophorus vampyrus (Anura:Rhacophoridae) reproductive biology: a new type of oophagous tadpole in Asian treefrogs. Journal of Herpetology 47, 607-614.

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There Is So Much More to Flying Frogs Than Flying

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Episode 2 of David Attenborough's Conquest of the Skies was on TV the other day, and I watched it (I livetweeted throughout, mostly because I wanted to talk about their portrayal of pterosaurs and Mesozoic theropods).

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Polypedate
- Common tree frog
- Spotted Leg Tree frog
- File-Eared Tree frog
- Bongao Tree frog
- Black Spotted Tree frog
Chiromantis

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Pacific Tree Frog, Pseudacris P. regilla
A total group of amphibians that include the order anura, frogs and toads.

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Hippopotamus, Hippopatmaus H. amphibus
Western Green Mamba, H. Mambas
Giant Golden-Crowned flying fox, Acereodon A. jubatus
Three-toed sloth, Bradypus Linnaeus
Sea Otter, Enhydra E. lutris
Crab-Eating Racoon, Pyrocon P. cancrivrous

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Closest Living relatives: The salamander

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Closest Living relatives: The salamander

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Sequence Alignment and Phylogenetic Reconstruction.

A total of 12 complete mitochondrial genomes representing the major groups of tetrapods were analyzed (GenBank accession nos. for African lungfish, Protopterus dolloi, L42813; coelacanth, Latimeria chalumnae, U82228; clawed frog, Xenopus laevis, M10217; caecilian, Typhlonectes natans, AF154051; Lusehan's salamander, Mertensiella luschani, AF154053; skink, Eumeces egregius, AB016606; painted turtle, Chrysemys picta, AF069423; alligator, Alligator mississippiensis, Y13113; chicken, Gallus gallus, X52392; opossum, Didelphis virginiana, Z29573; blue whale, Balaenoptera musculus, X72204; human, Homo sapiens, D38112). Tetrapod mtDNAs were selected so that their molecular evolutionary rates were not statistically different and so that long-branch attraction effects could be avoided (e.g., the snake mtDNA shows an unusually accelerated evolutionary rate and was omitted; ref. 28). Nucleotide sequences were aligned by using clustal x ( 29) and refined by eye. Gaps resulting from the alignment were treated as missing data. Ambiguous alignments, mainly in 5′ and 3′ ends of protein coding genes in the DHU and TψC arms of the tRNAs and in several highly variable regions of the rRNA genes were excluded from the phylogenetic analyses. Maximum parsimony (MP) analyses were performed by using heuristic searches [tree bisection and reconnection (TBR) branch swapping; multiple trees (MULTREE) option in effect] with 10 random stepwise additions of taxa. Transitions (Ti) and transversions (Tv) were given equal weight according to the empirical Ti:Tv ratio (1:1). The neighbor-joining (NJ) ( 30) analysis of the alignment was based on an HKY85 ( 31) distance matrix. In the maximum likelihood (ML) analyses [Hasegawa, Kishino, Yano (HKY)85 model], Ti:Tv ratios were optimized to maximize the likelihood analysis; empirical base frequencies were used. Robustness of the phylogenetic results was tested either by bootstrap analyses ( 32) with 500 pseudoreplications (MP and NJ) or by the quartet puzzling method ( 33) with 1,000 puzzling steps (ML). All phylogenetic analyses were performed by using paup* Version 4.0b4a ( 34).

Complete Mitochondrial DNA Evidence Supports the Batrachia Hypothesis.

Mitochondrial protein-coding, tRNA, and rRNA gene sequences were combined into a single data set that produced an alignment of 15,686 positions; of those positions, 2,635 were excluded because of ambiguity, 4,825 were constant, and 6,472 were parsimony-informative. All three commonly used methods of phylogenetic inference (MP, NJ, and ML) arrived at the same tree topology (Fig. 2). This fully resolved tree supports the monophyly of living amphibians with respect to other living tetrapods ( 1, 8- 16) and favors a sister group relationship for salamanders and frogs (the Batrachia hypothesis; refs. 1, 10, 11, 13, and 22). Robustness of our results was confirmed by high bootstrap support of all nodes in the tree (Fig. 2). This result contradicts the hypothesis of a salamander + caecilian grouping ( 12) that was suggested by previous molecular work based on rRNA data sets ( 15, 23, 24, 26).

Several recent studies have pointed out the risk of recovering misleading phylogenetic relationships among distantly related taxa when only short sequence data sets are analyzed ( 35- 38). Larger or relatively heterogenous data sets, i.e., complete mitochondrial genomes with many phylogenetically informative sites, are required to confidently resolve most questions dealing with major phylogenetic events ( 35- 38). Our results would seem to support the conclusions of these studies.

Interestingly, morphology- and molecule-based studies have disagreed profoundly regarding extant amphibian relationships. Most morphological studies have consistently concluded that salamanders and frogs are sister groups ( 1, 10, 11, 13), whereas molecular studies based on nuclear and mitochondrial rRNA data have supported caecilians as the sister taxon of salamanders ( 15, 23, 24, 26). Our analyses based on complete mitochondrial sequence data may contribute to a resolution of the apparent conflict between morphological and molecular evidence. In agreement with previous morphology-based hypotheses, our molecular data overwhelmingly support monophyly of living amphibians and the hypothesis that salamanders and frogs are more closely related to each other than to caecilians (Fig. 2). There are several reasons why we believe that our extensive molecular data set may be more reliable than the smaller rRNA data sets on this question. It is well known that rRNA data sets show a high degree of among-site rate variation, and consequently, few sites are phylogenetically informative at any given level of divergence ( 39). This pattern can lead to the recovery of spurious phylogenies based on homoplasious rather than phylogenetically informative sites, and likely explains why previous molecular studies based on rRNA data were prone to recover a salamander + caecilian clade. In fact, a recent reanalysis of the mitochondrial rRNA evidence was able to moderately support a salamander + frog sister group relationship ( 22).

Identity of the Sister Group of Lissamphibia Can Be Gleaned Only from Paleontological Data.

Of course, molecular data cannot directly assess the monophyly of Lissamphibia because it also includes fossil groups. However, as indicated by our molecular data (Fig. 2), a sister group relationship between salamanders and frogs can indirectly reject those hypotheses based on morphological and paleontological data that suggest nonmonophyly of living amphibians because of an independent origin of salamanders (refs. 18- 21; Fig. 1 d). Moreover, the monophyly of living amphibians with respect to living amniotes (Fig. 2) can indirectly reject those hypotheses that relate caecilians and salamanders to microsaurs, and they suggest a closer relationship of the latter to amniota rather than to Temnospondyli (refs. 17- 21; Fig. 1 d). In this regard, the molecular phylogeny presented further supports the hypothesis that lissamphibians (including fossil groups) constitute a single clade with respect to extinct amphibian groups.

Unfortunately, molecular data cannot provide information on the question of the closest extinct relative of the Lissamphibia. Only paleontological evidence can determine whether the extinct Temnospondyli (e.g., refs. 13 and 40) or the Lepospondyli ( 14, 16, 20) are the closest sister group to modern amphibians (but see below).

Congruent Molecular and Morphological Evidence Provides a Robust Phylogenetic Framework That Is Necessary for Comparative Studies Among Amphibians.

The hypothesis presented here implies that frogs provide the ideal extant outgroup to analyze phylogenetic relationships within salamanders (including, e.g., the open question of the relative positions of the families Sirenidae and Proteidae; ref. 41); inversely, salamanders are likely the best living outgroup to study anuran systematics (including the contentious monophyly of archaeobatrachia; ref. 42). The use of the closest living relative as outgroup is relevant both in morphological (because it could directly affect the polarity of several characters) and molecular (e.g., because it could prevent the attraction of long branches to basal positions) studies. Moreover, our results have implications for the decision to choose between alternative positions of several fossil taxa that are closely related to the origin of the three lineages of living amphibians. For instance, the recent placement of the albanerpetontids (a group of salamander-like fossil amphibians) as sister group of the Batrachia ( 43) is supported by the molecular phylogeny (a putative caecilian + salamander clade would have contradicted it). Furthermore, by providing an independent source of phylogenetic information, our results confirm the authenticity of some of the putative morphological synapomorphies (up to nine unique shared-derived characters; see ref. 13) that diagnose the Batrachia, and might encourage the search for new shared-derived traits between salamanders and frogs. Moreover, our results invite further research on the causes of the now seemingly homoplasious characters that group caecilians with salamanders.

The geographic argument based on the current world distribution of amphibians and rRNA evidence that salamanders and caecilians originated from a common ancestor because of the breakup of the supercontinent Pangea ( 15) is invalidated by our data. Furthermore, the recent discovery of Czatkobatrachus from the early Triassic of Poland ( 44) suggests that, at least, Salientia (the stem-group of anurans) occurred in all of Pangea ( Triadobatrachus is from early Triassic of Madagascar; ref. 45), and that the separation of salamanders and frogs predated the breakup of Pangea.

Our results will also be helpful in polarizing the divergent developmental patterns exhibited by Caudata, Anura, and Gymnophiona ( 46, 47). For instance, vertebral development in frogs is interpreted as being rather similar to that inferred for Temnospondyli ( 21), which may represent the primitive condition for all tetrapods. However, the pattern of vertebral centra ossification in salamanders ( 48) and caecilians ( 49) is more similar to that inferred for Lepospondyli (representing a derived condition). If lissamphibians evolved from lepospondyls ( 14, 16), and accepting that salamanders and frogs are sister group taxa, the most parsimonious explanation to the observed patterns is that the type of vertebral development in salamanders and caecilians is primitive for lissamphibians, and that the ancestor of frogs reelaborated the developmental pattern found in the extinct temnospondyls (this pattern may be a constraint associated with metamorphosis in frogs; ref. 21). Alternatively, whether temnospondyls are the sister group of modern amphibians, the early ossification of vertebral centra evolved independently in caecilians and salamanders, whereas frogs would have retained the primitive condition of all tetrapods.

Brain development of frogs and salamanders differs considerably. The brains of frogs are morphologically much more complex than those of salamanders ( 50). On the other hand, salamanders and caecilians show similarity in many features of brain development ( 51). These patterns seemingly would contradict the Batrachia hypothesis. However, the simple brain morphology of salamanders may be a secondarily derived condition associated with an increase in genome, cell size, and pedomorphic evolution ( 51, 52). Our phylogeny supports this hypothesis of a secondary simplification in salamanders.

The three orders of living amphibians differ markedly in their limb morphologies ( 6). By providing a robust phylogenetic framework, our results may also aid in polarizing the divergent limb development patterns found in amphibians ( 53). For instance, frogs and salamanders follow divergent pathways of digital formation (e.g., refs. 46 and 54). Frogs develop their digits in a posterior to anterior sequence ( 55). However, salamanders show the opposite pattern ( 46). Based on strict ontogenetic criteria, these patterns were used in the past as evidence of amphibian (and tetrapod) polyphyly (e.g., ref. 56; see ref. 47 for a review). Yet, based on the robust and congruent morphological and molecular evidence that supports the Batrachia hypothesis, the interpretation of these developmental patterns would need to be quite different. Digit formation in frogs is considered to be the ancestral condition for all tetrapods, whereas the salamander pattern represents a derived condition ( 46, 55, 57, 58). Recent molecular developmental studies demonstrate that expression patterns of Hoxa-11 in the limb buds of frogs are similar to those found in amniota ( 59), whereas salamanders show a derived pattern.

The interpretation of many comparative studies among the three orders of living amphibians largely depends on whether the salamanders are the closest living relative of frogs or the living sister group of caecilians ( 60). In contrast to previous molecular analyses based on nuclear and mitochondrial rRNA data, our results, based on the analysis of complete mitochondrial genome data, support the Batrachia (salamander + frog) hypothesis and are thereby in agreement with most morphological evidence. Our analyses demonstrate that some phylogenetic controversies between morphology and molecules that exist in the literature are only apparent rather than real and frequently caused by a lack of phylogenetically informative characters in one of the two (morphological or molecular) relatively small data sets analyzed. General support of the Batrachia hypothesis allows a confident polarization of characters in many comparative studies that explore the astonishing biological diversity within and among extant orders of amphibians.

Received September 22, 2000.

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On the origin of and phylogenetic relationships among living amphibians

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The phylogenetic relationships among the three orders of modern amphibians (Caudata, Gymnophiona, and Anura) have been estimated based on both morphological and molecular evidence. Most morphological and paleontological studies of living and fossil amphibians support the hypothesis that salamanders and frogs are sister lineages (the Batrachia hypothesis) and that caecilians are more distantly related.

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Spotted Salamander, Mole Salamander
Western Toad, North American Toad
Boophis luteus, Boophis
Common Reed Frog, Hyperolius
Mudpuppies, Necturus N. Maculosus

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Hammerhead shark, Sphyrna S. lewini
American Black Bear: Bears, Ursus U. Americanus
Giant Oceanic Manta Ray, Mobula M. birostris
Green Iguana, Iguana I. igunana
Garden snail, Helix Linnaeus
Angler fish, Melanocetus M. johnsonii

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Facts about frogs, specifically Parachute Frogs

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Facts about frogs, specifically Parachute Frogs

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Wallace’s Flying Frog | National Geographic

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Discover a frog that soars through the air with the greatest of ease. Learn how these amphibians have adapted to a life in the trees.

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Blue Poison Dart Frog, Dendrobates D. tinctorius
Bronze Frog, Lithobates L. Clamitans
Northern Green Frog, Lithobates L. Clamitans
Wood Frog- Lithobates L. sylvaticus
Black Rain Frog- Breviceps B. fuscus

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Kingdom: Animalia
Phylum: Chordata
Family: Rhacophoridae
Class: Amphibia
Clade: Salientia
Order: Anura
Genus: Rhacophorus
Species: R. nigropalmatus