Paul E. Moler

Is The Amphibian Tree of Life really fatally flawed

Wiens (2007 , Q. Rev. Biol. 82, 55–56) recently published a severe critique of Frost et al.s (2006, Bull. Am. Mus. Nat. Hist. 297, 1–370) monographic study of amphibian systematics, concluding that it is “a disaster” and recommending that readers “simply ignore this study”. Beyond the hyperbole, Wiens raised four general objections that he regarded as “fatal flaws”: (1) the sampling design was insufficient for the generic changes made and taxonomic changes were made without including all type species; (2) the nuclear gene most commonly used in amphibian phylogenetics, RAG‐1, was not included, nor were the morphological characters that had justified the older taxonomy; (3) the analytical method employed is questionable because equally weighted parsimony “assumes that all characters are evolving at equal rates”; and (4) the results were at times “clearly erroneous”, as evidenced by the inferred non‐monophyly of marsupial frogs. In this paper we respond to these criticisms. In brief: (1) the study of Frost et al. did not exist in a vacuum and we discussed our evidence and evidence previously obtained by others that documented the non‐monophyletic taxa that we corrected. Beyond that, we agree that all type species should ideally be included, but inclusion of all potentially relevant type species is not feasible in a study of the magnitude of Frost et al. and we contend that this should not prevent progress in the formulation of phylogenetic hypotheses or their application outside of systematics. (2) Rhodopsin, a gene included by Frost et al. is the nuclear gene that is most commonly used in amphibian systematics, not RAG‐1. Regardless, ignoring a study because of the absence of a single locus strikes us as unsound practice. With respect to previously hypothesized morphological synapomorphies, Frost et al. provided a lengthy review of the published evidence for all groups, and this was used to inform taxonomic decisions. We noted that confirming and reconciling all morphological transformation series published among previous studies needed to be done, and we included evidence from the only published data set at that time to explicitly code morphological characters (including a number of traditionally applied synapomorphies from adult morphology) across the bulk of the diversity of amphibians (Haas, 2003, Cladistics 19, 23–90). Moreover, the phylogenetic results of the Frost et al. study were largely consistent with previous morphological and molecular studies and where they differed, this was discussed with reference to the weight of evidence. (3) The claim that equally weighted parsimony assumes that all characters are evolving at equal rates has been shown to be false in both analytical and simulation studies. (4) The claimed “strong support” for marsupial frog monophyly is questionable. Several studies have also found marsupial frogs to be non‐monophyletic. Wiens et al. (2005, Syst. Biol. 54, 719–748) recovered marsupial frogs as monophyletic, but that result was strongly supported only by Bayesian clade confidence values (which are known to overestimate support) and bootstrap support in his parsimony analysis was < 50%. Further, in a more recent parsimony analysis of an expanded data set that included RAG‐1 and the three traditional morphological synapomorphies of marsupial frogs, Wiens et al. (2006, Am. Nat. 168, 579–596) also found them to be non‐monophyletic. Although we attempted to apply the rule of monophyly to the naming of taxonomic groups, our phylogenetic results are largely consistent with conventional views even if not with the taxonomy current at the time of our writing. Most of our taxonomic changes addressed examples of non‐monophyly that had previously been known or suspected (e.g., the non‐monophyly of traditional Hyperoliidae, Microhylidae, Hemiphractinae, Leptodactylidae, Phrynobatrachus, Ranidae, Rana, Bufo; and the placement of Brachycephalus within “Eleutherodactylus”, and Lineatriton within “Pseudoeurycea”), and it is troubling that Wiens and others, as evidenced by recent publications, continue to perpetuate recognition of non‐monophyletic taxonomic groups that so profoundly misrepresent what is known about amphibian phylogeny.

American Crocodile (Crocodylus Acutus) in Florida: Recommendations for Endangered Species Recovery and Ecosystem Restoration

Abstract When the American Crocodile (Crocodylus acutus) was declared endangered in 1975, scant data were available for making management decisions. Results of intensive studies conducted during the late 1970s and early 1980s by the National Park Service, Florida Game and Fresh Water Fish Commission, and Florida Power and Light Company resulted in an optimistic outlook for crocodiles. However, new issues face crocodiles today. Florida and Biscayne bays have undergone changes that have caused concern for the health of these ecosystems. The purpose of this paper is to review results of monitoring programs for C. acutus that have been used as a basis for consideration of reclassification of this endangered species and for restoration of its endangered ecosystem. More crocodiles and nests occur in more places today than in 1975. The maximum number of nesting females in Florida has increased from 20 in 1975 to 85 in 2004, and the number of concentrations of nesting effort from two to four. This evidence supports the proposed reclassification of the American Crocodile from endangered to threatened. However, crocodiles are still threatened by modification of habitat because of development adjacent to crocodile habitat and will benefit from restored freshwater flow into estuaries. As crocodiles continue to increase in number and expand into new areas, interactions with humans will occur more frequently. The challenge of integrating a recovering population of the American Crocodile with an ever-increasing use of coastal areas by humans will be the final challenge in successful recovery of this once critically endangered species.

Peptidomic analysis of skin secretions from Rana heckscheri and Rana okaloosae provides insight into phylogenetic relationships among frogs of the Aquarana species group.

The members of the Aquarana (or Rana catesbeiana species group) form a monophyletic group comprising seven species: R. catesbeiana, Rana clamitans, Rana grylio, Rana virgatipes, Rana septentrionalis, Rana heckscheri and Rana okaloosae. Previous work has led to structural characterization of the antimicrobial peptides present in electrically-stimulated skin secretions from the first five species listed and this study presents the primary structures of orthologs from the river frog R. heckscheri and the Florida bog frog R. okaloosae. Peptidomic analysis of R. heckscheri and R. okaloosae skin secretions led to the identification of peptides with antimicrobial activity belonging to the ranalexin, ranatuerin-2, and temporin families. In addition, a peptide (GFLDIIKDTGKDFAVKILNNLKCKLAGGCPR) was isolated from R. okaloosae whose primary structure identified it as a member of the palustrin-2 family. Consistent with previous data based upon morphological analysis and comparisons of the nucleotide sequences of mitochondrial and ribosomal genes, cladistic analysis based upon a comparison of the amino acid sequences of antimicrobial peptides indicates a sister-group relationship between R. heckscheri and R. grylio and a close, but less well defined, phylogenetic relationship between R. okaloosae and R. clamitans.

HEALTH ASSESSMENT OF FREE-RANGING ALLIGATOR SNAPPING TURTLES (MACROCHELYS TEMMINCKII) IN GEORGIA AND FLORIDA

The Alligator Snapping Turtle (Macrochelys temminckii) is a large freshwater turtle endemic to river systems that drain into the Gulf of Mexico. Turtle populations were sharply reduced by commercial harvest in the 1970s and 1980s; however, the species has yet to be protected under the Endangered Species Act. While anthropogenic stressors such as habitat fragmentation and degradation and illegal capture continue to threaten populations, the degree to which disease may be contributing to any decline of the Alligator Snapping Turtle is unknown. Data were collected from 97 free-ranging Alligator Snapping Turtles in nine waterways in Florida and Georgia from 2001 to 2006. Eleven turtles were captured more than once, resulting in a total sample pool of 123. Reference ranges were established for complete blood count, plasma biochemistry values, trace metals (mercury, zinc, copper, lead, and arsenic), and nutrient parameters (vitamins A, E, D, and selenium). Variations by capture location, sex, and season were detected and likely resulted from external factors such as habitat and diet. Turtles sampled in one location were positive for tortoise herpesviral antibodies. Blood mercury values also differed among populations. This study provides justification for the use of these long-lived aquatic turtles as biologic monitors of the health of local freshwater ecosystems.

The amphibian chytrid fungus, Batrachochytrium dendrobatidis, in fully aquatic salamanders from Southeastern North America.

Little is known about the impact that the pathogenic amphibian chytrid fungus, Batrachochytrium dendrobatidis (Bd), has on fully aquatic salamander species of the eastern United States. As a first step in determining the impacts of Bd on these species, we aimed to determine the prevalence of Bd in wild populations of fully aquatic salamanders in the genera Amphiuma, Necturus, Pseudobranchus, and Siren. We sampled a total of 98 salamanders, representing nine species from sites in Florida, Mississippi, and Louisiana. Overall, infection prevalence was found to be 0.34, with significant differences among genera but no clear geographic pattern. We also found evidence for seasonal variation, but additional sampling throughout the year is needed to clarify this pattern. The high rate of infection discovered in this study is consistent with studies of other amphibians from the southeastern United States. Coupled with previously published data on life histories and population densities, the results presented here suggest that fully aquatic salamanders may be serving as important vectors of Bd and the interaction between these species and Bd warrants additional research.

Evolution of Gigantism in Amphiumid Salamanders

The Amphiumidae contains three species of elongate, permanently aquatic salamanders with four diminutive limbs that append one, two, or three toes. Two of the species, Amphiuma means and A. tridactylum, are among the largest salamanders in the world, reaching lengths of more than one meter, whereas the third species (A. pholeter), extinct amphiumids, and closely related salamander families are relatively small. Amphiuma means and A. tridactylum are widespread species and live in a wide range of lowland aquatic habitats on the Coastal Plain of the southeastern United States, whereas A. pholeter is restricted to very specialized organic muck habitats and is syntopic with A. means. Here we present analyses of sequences of mitochondrial and nuclear loci from across the distribution of the three taxa to assess lineage diversity, relationships, and relative timing of divergence in amphiumid salamanders. In addition we analyze the evolution of gigantism in the clade. Our analyses indicate three lineages that have diverged since the late Miocene, that correspond to the three currently recognized species, but the two gigantic species are not each others closest relatives. Given that the most closely related salamander families and fossil amphiumids from the Upper Cretaceous and Paleocene are relatively small, our results suggest at least two extreme changes in body size within the Amphuimidae. Gigantic body size either evolved once as the ancestral condition of modern amphiumas, with a subsequent strong size reduction in A. pholeter, or gigantism independently evolved twice in the modern species, A. means and A. tridactylum. These patterns are concordant with differences in habitat breadth and range size among lineages, and have implications for reproductive isolation and diversification of amphiumid salamanders.

A new species of frog (Ranidae: Rana) from Northwestern Florida

Pine Barrens treefrog (Hyla andersonii) in Okaloosa County, Florida, 21 July 1982, I heard a distinctive but unfamiliar frog call. Investigation disclosed a small Rana sitting in shallow water along a clear stream. Through the remainder of the evening, similar calls were heard at five additional sites, and five more specimens were collected. It soon became apparent that these frogs represented an undescribed species. A total of 34 adults, three recently transformed juveniles and numerous tadpoles are now available. Specimens have been deposited in the Florida State Museum (UF) and the United States National Museum (USNM).

Taxonomic assessment of Alligator Snapping Turtles (Chelydridae: Macrochelys), with the description of two new species from the southeastern United States.

The Alligator Snapping Turtle, Macrochelys temminckii, is a large, aquatic turtle limited to river systems that drain into the Gulf of Mexico. Previous molecular analyses using both mitochondrial and nuclear DNA suggested that Macrochelys exhibits significant genetic variation across its range that includes three distinct genetic assemblages (western, central, and eastern = Suwannee). However, no taxonomic revision or morphological analyses have been conducted previously. In this study, we test previous hypotheses of distinct geographic assemblages by examining morphology, reanalyzing phylogeographic genetic structure, and estimating divergence dating among lineages in a coalescent framework using Bayesian inference. We reviewed the fossil record and discuss phylogeographic and taxonomic implications of the existence of three distinct evolutionary lineages. We measured cranial (n=145) and post-cranial (n=104) material on field-captured individuals and museum specimens. We analyzed 420 base pairs (bp) of mitochondrial DNA sequence data for 158 Macrochelys. We examined fossil Macrochelys from ca. 15-16 million years ago (Ma) to the present to better assess historical distributions and evaluate named fossil taxa. The morphological and molecular data both indicate significant geographical variation and suggest three species-level breaks among genetic lineages that correspond to previously hypothesized genetic assemblages. The holotype of Macrochelys temminckii is from the western lineage. Therefore, we describe two new species as Macrochelys apalachicolae sp. nov. from the central lineage and Macrochelys suwanniensis sp. nov. from the eastern lineage (Suwannee River drainage). Our estimates of divergence times suggest that the most recent common ancestor (MRCA) of M. temminckii (western) and M. apalachicolae (central) existed 3.2-8.9 Ma during the late Miocene to late Pliocene, whereas M. temminckii-M. apalachicolae and M. suwanniensis last shared a MRCA 5.5-13.4 Ma during the mid-Miocene to early Pliocene. Examination of fossil material revealed that the fossil taxon M. floridana is actually a large Chelydra. Our taxonomic revision of Macrochelys has conservation and management implications in Florida, Georgia, and Alabama.

Allozyme variation and systematics of the Nerodia fasciata-Nerodia clarkii complex of water snakes (Serpentes: Colubridae)

extensive area of the coastal plain of eastern North America. They are parapatric with the closely related N. sipedon along the fall line and hybridization between these two species sometimes occurs in areas of habitat disturbance. Nerodia f. clarkii, N. f. compressicauda, and N. f. taeniata are inhabitants of a narrow coastal saltwater zone and are distributed almost continuously from the middle Atlantic coast of Florida to southern Texas and including the Florida Keys. These snakes are physiologically well adapted to exploit their saline environment. The geographic ranges of the two ecologically distinct groups sometimes overlap in the saltwater-freshwater ecotone, yet, because of their divergent adaptations, they are for the most part microallopatric in these areas. Analyses of allozyme data from 33 protein coding loci by several methods show that, except for areas of considerable habitat disturbance, gene flow between the two groups is very slight or absent and is on the same order as that seen between the freshwater group and N. sipedon. Elevation to species level is recommended for the two groups. Within-group divergence is minimal at the allozyme level, and the question of whether N. f. taeniata arose from ancestral N. f. clarkii or ancestral N. f. compressicauda could not be resolved by analyses of these data.

Effect of human-mediated migration and hybridization on the recovery of the American crocodile in Florida (USA)

The American crocodile (Crocodylus acutus) is a large apex predator with an extensive but fragmented range. Crocodylus acutus suffered a population crash and was almost extirpated from Florida (USA) during the 1970s; however, in 2007 it was federally downlisted from endangered to threatened based on an increase of suitable habitat and a growing population. A genetic assessment of this population has not been performed even though foreign crocodiles have been released into Florida waters. Herein, we use mitochondrial and nuclear DNA markers to characterize relationships among American crocodiles along the southern coast of Florida and reference samples from other countries. We detected inter- and intra-specific hybridization and unexpected population structure attributed to human-mediated migration of crocodiles from Latin America and the Greater Antilles. Our results suggest that the population size of crocodiles actually native to Florida should be reevaluated, particularly in light of ongoing admixture in this population. We reemphasize the utility of genetic markers in conservation and management programs for endangered species; especially those that can hybridize with closely related congeners. The American crocodile in Florida has recovered owing to successful conservation initiatives, but a long-term management protocol that takes genetic data into account is still needed.