Phylogeography of Italian endemic Flea Beetles (Chrysomelidae, Galerucinae, Alticini): biogeographic, systematic, and conservation implications

biodiversity hotspot within the Mediterranean global biodiversity hotspot. However, most of the knowledge we have on phylogeographic patterns of Italian endemic species come from studies that have used vertebrates as model species, although they represent only a small part of the global biodiversity of the species (<1.5%). This makes our appraisal of the ecological response of species to past climatic changes far from complete. In my PhD project, I used the highly diverse group of flea beetles endemic of the central Apennines (Coleoptera Chrysomelidae, Galerucinae, Alticini), to investigate the ecological, evolutionary and biogeographic processes primed by Quaternary glaciations cycles on temperate and high-altitude biota. These insects show distribution patterns from micro to macro scale, being present in most mountain ecosystems, and have restrictive and well-known ecological requirements, thus making them ideal phylogeographic models. The main aim of the thesis project is to study the historical processes that have shaped current patterns of genetic diversity within different endemic Alticni species of Apennines and to compare how species with different ecological requirements have responded to Quaternary climate change. To achieve these main aim, I have adopted a multi-taxa approach including (i) an integrative taxonomic approach to delimit the phylogenetic and taxonomic units, and (ii) a multilocus phylogeographic approach to identify the main Pleistocene refugia within the Apennine system, and to infer demographic and biogeographic processes underlying the formation of the observed intraspecific diversity patterns. Six target species were selected based on their distribution and ecology: Longitarsus springeri, Psylliodes biondii and Psylliodes springeri linked to high mountain environments; Longitarsus laureolae and Longitarsus zangherii linked to temperate environments and Psylliodes ruffoi linked to xerophilous environments. Three large sampling campaigns were performed to sample the populations of target species. A first integrative morphological and molecular delimitation of the target species was done as a fundamental and preliminary step to the phylogeographic study. In this framework, three main issues related to the mismatch between molecular and morphological identification were pointed out and resolved. First, within the genus Longitarsus, we observed several instances of species misidentification and taxonomic incongruences within the public databases of DNA sequences (GenBank and BOLD). This makes difficult to implement a molecular identification of species by comparing the DNA sequence of a target gene (cox1) of a sampled individual with deposited sequences in public database through a DNA barcoding approach. To overcome this drawback, we implemented a methodical pipeline with a posteriori taxonomic revision that allow improving the quality of the reference-sequence library (from GenBank and BOLD) and the effectiveness of the DNA barcoding tool. Second, we observed that some samples, morphological identified as P. ruffoi, had molecular characters (i.e. DNA sequences) typical of the related species P. kiesenwetteri. The use of an integrative taxonomy approach within a phylogeographic framework allowed to delimit the two species P. ruffoi and P. kiesenwetteri from a molecular and morphological point of view, clarifying their phylogenetic and taxonomic relationship. Third, following the molecular assessment of the Maiella population of P. biondii, we observed that this population show a remarkable genetic distance relative to other populations of P. biondii, suggesting they are possibly two distinct species. Thanks to an integrative taxonomy approach, we were able to solve these taxonomic and systematic problems, to identify a new lineage (possibly a new species) endemic to the Maiella mountains, and thus to identify the evolutionary units on which to deepen the phylogeographical study. [...]