Heterochrony and limb form variation in crested newts N. Tomaševic ´ Kolarov Department of Evolutionary Biology, Institute for Biological Research ‘Siniša Stankovic ´ ’, University of Belgrade, Belgrade, Serbia Keywords limbs; sequence heterochrony; allometric heterochrony; crested newts. Correspondence Nataša Tomaševic ´ Kolarov, Department of Evolutionary Biology, Institute for Biological Research ‘Siniša Stankovic ´ ’, University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia. Tel: +381 11 2078378; Fax: +381 11 2761433 Email: natasha@ibiss.bg.ac.rs; t.k.natasa@gmail.com Editor: Mark-Oliver Rödel Received 15 August 2012; revised 28 December 2012; accepted 29 December 2012 doi:10.1111/jzo.12016 Abstract The crested newt superspecies includes four morphotypes, each characterized by a specific body-to-limb conformation associated with their respective ecologies: newts with large, stocky bodies and robust limbs are more terrestrial, whereas newts with small, elongate bodies and small limbs are more aquatic. This study investigated whether heterochronic transformations can account for limb form variation in light of phylogenetic history and ecology. Ossification sequence analy- ses revealed some synapomorphic heterochronic shifts specific to crested newts, including delay of the ossification in the second finger and accelerations in meta- carpal III and metatarsal V. These shifts involve a change from pre-axial to post-axial dominance in a developmental sequence uncommon to caudate sala- manders. No adaptive explanation of these shifts is apparent. The allometric trajectories of crested newt species were similar after metamorphosis; however, pre-metamorphic growth showed species differences, potentially reflecting differ- ences among species in ecological or functional demands. Introduction The crested newt superspecies includes four morphotypes, each characterized by a specific body-to-limb conformation associated with their respective ecologies (Vukov et al., 2011; Wielstra & Arntzen, 2011). Species with large, stocky bodies and robust limbs are largely terrestrial, with an average of 3 (Triturus karelinii group), 4 (Triturus macedonicus and Tritu- rus carnifex) or 5 (Triturus cristatus) months spent in the water. In contrast, the most aquatic species, Triturus dobrogi- cus, has a small, slender, elongate body and small limbs, and spends an average of 6 months in the water (Wielstra & Arntzen, 2011). The correlation between ecology and body shape is further supported by the most robust body build, marbled newt Triturus marmoratus, which spends only 2 months in aquatic habitats, the shortest time of any Triturus newt species (Wielstra & Arntzen, 2011). Of interest in this study is heterochronic perspective of the differences in the ontogenetic development of the limbs. Het- erochrony (i.e. evolutionary changes in the timing and rate of development) lies at the intersection of evolution and devel- opment (McKinney & McNamara, 1991; Smith, 2003). It is considered an important evolutionary mechanism underlying morphological changes, acting across all animal groups and stages of development (Gould, 1977; Alberch et al., 1979; McKinney, 1988; McKinney & McNamara, 1991; Richardson, 1999). The study of heterochrony originally focused on differences among developing organisms in the relative growth rates of selected organs or regions (McKinney, 1988). More recent work on heterochrony has focused on evolutionary changes in the temporal order of developmental events (Smith, 2001). Sequence heterochrony has a number of significant advantages (see Smith, 2003), with great potential in the studies of evolution and developmental time, mostly due to advances in analytic techniques and computation ability (Harrison & Larsson, 2008; Maxwell & Harrison, 2009). Heterochronic modifications are thought to occur fre- quently in the evolution of salamanders and other amphibians (Alberch & Alberch, 1981; Wake, Wake & Wake, 1983; Wake, 1986; Hanken, 1989; Hinchliffe & Vorobyeva, 1999; Buckley et al., 2007). The evolution of various body traits, reproductive modes and complex life cycles are often related to these modifications (Hanken, 1992; Bruce, 2003). Accord- ing to Blanco & Alberch (1992) and Vorobyeva (2009), het- erochrony and larval adaptations have played important roles in the diversification of the distal limb skeleton in caudate Journal of Zoology Journal of Zoology. Print ISSN 0952-8369 Journal of Zoology •• (2013) ••–•• © 2013 The Zoological Society of London 1