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      Morphological and ecological convergence at the lower size limit for vertebrates highlighted by five new miniaturised microhylid frog species from three different Madagascan genera

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          Abstract

          Miniaturised frogs form a fascinating but poorly understood amphibian ecomorph and have been exceptionally prone to taxonomic underestimation. The subfamily Cophylinae (family Microhylidae), endemic to Madagascar, has a particularly large diversity of miniaturised species which have historically been attributed to the single genus Stumpffia largely based on their small size. Recent phylogenetic work has revealed that several independent lineages of cophyline microhylids evolved towards highly miniaturised body sizes, achieving adult snout–vent lengths under 16 mm. Here, we describe five new species belonging to three clades that independently miniaturised and that are all genetically highly divergent from their relatives: (i) a new genus (Mini gen. nov.) with three new species from southern Madagascar, (ii) one species of Rhombophryne, and (iii) one species of Anodonthyla. Mini mum sp. nov. from Manombo in eastern Madagascar is one of the smallest frogs in the world, reaching an adult body size of 9.7 mm in males and 11.3 mm in females. Mini scule sp. nov. from Sainte Luce in southeastern Madagascar is slightly larger and has maxillary teeth. Mini ature sp. nov. from Andohahela in southeast Madagascar is larger than its congeners but is similar in build. Rhombophryne proportionalis sp. nov. from Tsaratanana in northern Madagascar is unique among Madagascar’s miniaturised frogs in being a proportional dwarf, exhibiting far less advanced signs of paedomorphism than other species of similar size. Anodonthyla eximia sp. nov. from Ranomafana in eastern Madagascar is distinctly smaller than any of its congeners and is secondarily terrestrial, providing evidence that miniaturisation and terrestriality may be evolutionarily linked. The evolution of body size in Madagascar’s microhylids has been more dynamic than previously understood, and future studies will hopefully shed light on the interplay between ecology and evolution of these remarkably diverse frogs.

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          Size and shape in ontogeny and phylogeny

          We present a quantitative method for describing how heterochronic changes in ontogeny relate to phyletic trends. This is a step towards creating a unified view of developmental biology and evolutionary ecology in the study of morphological evolution. Using this representation, we obtain a greatly simplified and logical scheme of classification. We believe that this scheme will be particularly useful in studying the data of paleontology and comparative morphology and in the analysis of processes leading to adaptive radiation. We illustrate this scheme by examples drawn from the literature and our own work.
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            Vast underestimation of Madagascar's biodiversity evidenced by an integrative amphibian inventory.

            Amphibians are in decline worldwide. However, their patterns of diversity, especially in the tropics, are not well understood, mainly because of incomplete information on taxonomy and distribution. We assess morphological, bioacoustic, and genetic variation of Madagascar's amphibians, one of the first near-complete taxon samplings from a biodiversity hotspot. Based on DNA sequences of 2,850 specimens sampled from over 170 localities, our analyses reveal an extreme proportion of amphibian diversity, projecting an almost 2-fold increase in species numbers from the currently described 244 species to a minimum of 373 and up to 465. This diversity is widespread geographically and across most major phylogenetic lineages except in a few previously well-studied genera, and is not restricted to morphologically cryptic clades. We classify the genealogical lineages in confirmed and unconfirmed candidate species or deeply divergent conspecific lineages based on concordance of genetic divergences with other characters. This integrative approach may be widely applicable to improve estimates of organismal diversity. Our results suggest that in Madagascar the spatial pattern of amphibian richness and endemism must be revisited, and current habitat destruction may be affecting more species than previously thought, in amphibians as well as in other animal groups. This case study suggests that worldwide tropical amphibian diversity is probably underestimated at an unprecedented level and stresses the need for integrated taxonomic surveys as a basis for prioritizing conservation efforts within biodiversity hotspots.
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              The use of bioacoustics in anuran taxonomy: theory, terminology, methods and recommendations for best practice.

              Vocalizations of anuran amphibians have received much attention in studies of behavioral ecology and physiology, but also provide informative characters for identifying and delimiting species. We here review the terminology and variation of frog calls from a perspective of integrative taxonomy, and provide hands-on protocols for recording, analyzing, comparing, interpreting and describing these sounds. Our focus is on advertisement calls, which serve as premating isolation mechanisms and, therefore, convey important taxonomic information. We provide recommendations for terminology of frog vocalizations, with call, note and pulse being the fundamental subunits to be used in descriptions and comparisons. However, due to the complexity and diversity of these signals, an unequivocal application of the terms call and note can be challenging. We therefore provide two coherent concepts that either follow a note-centered approach (defining uninterrupted units of sound as notes, and their entirety as call) or a call-centered approach (defining uninterrupted units as call whenever they are separated by long silent intervals) in terminology. Based on surveys of literature, we show that numerous call traits can be highly variable within and between individuals of one species. Despite idiosyncrasies of species and higher taxa, the duration of calls or notes, pulse rate within notes, and number of pulses per note appear to be more static within individuals and somewhat less affected by temperature. Therefore, these variables might often be preferable as taxonomic characters over call rate or note rate, which are heavily influenced by various factors. Dominant frequency is also comparatively static and only weakly affected by temperature, but depends strongly on body size. As with other taxonomic characters, strong call divergence is typically indicative of species-level differences, whereas call similarities of two populations are no evidence for them being conspecific. Taxonomic conclusions can especially be drawn when the general advertisement call structure of two candidate species is radically different and qualitative call differences are thus observed. On the other hand, quantitative differences in call traits might substantially vary within and among conspecific populations, and require careful evaluation and analysis. We provide guidelines for the taxonomic interpretation of advertisement call differences in sympatric and allopatric situations, and emphasize the need for an integrative use of multiple datasets (bio-acoustics, morphology, genetics), particularly for allopatric scenarios. We show that small-sized frogs often emit calls with frequency components in the ultrasound spectrum, although it is unlikely that these high frequencies are of biological relevance for the majority of them, and we illustrate that detection of upper harmonics depends also on recording distance because higher frequencies are attenuated more strongly. Bioacoustics remains a prime approach in integrative taxonomy of anurans if uncertainty due to possible intraspecific variation and technical artifacts is adequately considered and acknowledged.
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                Author and article information

                Journal
                PLOS ONE
                PLoS ONE
                Public Library of Science (PLoS)
                1932-6203
                March 27 2019
                March 27 2019
                : 14
                : 3
                : e0213314
                Article
                10.1371/journal.pone.0213314
                e0d7d245-53c2-4e0a-b89b-4768a9f18fb3
                © 2019

                http://creativecommons.org/licenses/by/4.0/

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