Ronald M. Bonett, Paul T. Chippindale, Paul E. Moler, R. Wayne Van Devender, David B. Wake

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 other's 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.