Cryptic diversity within endemic Italian barbels: revalidation and description of new Barbus species (Teleostei: Cyprinidae).

Two fluviolacustrine lineages (SI1 Barbus and SI2 Barbus) of the Barbus genus have been recently detected in the Apulia-Campania ichthyogeographic district (southern Italy). The aim of this study was to determine the taxonomic status of these lineages by comparing them with the two already-established Italian fluviolacustrine species Barbus plebejus and Barbus tyberinus through a more in-depth molecular and morphological investigation. Genetic analyses were performed on both mitochondrial (cytochrome b) and nuclear (growth hormone paralog 2) DNA markers, and morphological analyses were carried out on specific age classes and purebred populations. Molecular analyses detected four evolutionary lineages at the mitochondrial level, whereas the nuclear data set highlighted the strict evolutionary relation between B. plebejus sensu stricto and the new lineages, converged in the B. plebejus complex clade. The morphological analyses allowed us to discriminate SI1 Barbus and SI2 Barbus from both B. plebejus and B. tyberinus. The new taxa could be discriminated by the greatest maximum body height and the longest pre-orbital distance, respectively. Both the new lineages have longer ventral and pectoral fins than B. plebejus and B. tyberinus, a larger caudal fin than B. tyberinus and a lower number of scales along the lateral line than B. plebejus. Both molecular and morphological results suggested the two southern Italian lineages could be considered as distinct endemic species: the formal description of the new species Barbus samniticus sp. nov. (i.e., SI1 Barbus) and the revalidation of Barbus fucini Costa, 1853 (i.e., SI2 Barbus) were thus proposed, and, for both species, molecular and morphological diagnosis were provided.

Intrinsic and extrinsic factors act at different spatial and temporal scales to shape population structure, distribution and speciation in Italian Barbus (Osteichthyes: Cyprinidae).

Previous studies have given substantial attention to external factors that affect the distribution and diversification of freshwater fish in Europe and North America, in particular Pleistocene and Holocene glacial cycles. In the present paper we examine sequence variation at one mitochondrial and four nuclear loci (over 3 kbp) from populations sampled across several drainages of all species of Barbus known to inhabit Italian freshwaters (introduced B. barbus and native B. balcanicus, B. caninus, B. plebejus and B. tyberinus). By comparing species with distinct ecological preferences (rheophilic and fluvio-lacustrine) and using a fossil-calibrated phylogeny we gained considerable insight about the intrinsic and extrinsic processes shaping barbel distribution, population structure and speciation. We found that timescales of Italian barbel diversification are older than previously thought, starting in the Early Miocene, and involving local and regional tectonism and basin paleo-evolution rather than Pleistocene glacial cycles. Conversely, more recent environmental factors associated with glaciation-deglaciation cycles have influenced species distributions. These events had a more marked impact on fluvio-lacustrine than on rheophilic species by means of river confluence at low sea levels. We show that genetic structure is influenced by species ecology: populations of small rheophilic species inhabiting upper river stretches of large basins are less connected and more differentiated than large fluvio-lacustrine species that inhabit lower river courses. We report the existence of both natural and human-induced interspecific gene flow, which could have great impacts on the evolution and persistence of species involved. In addition, we provide evidence that B. tyberinus is genetically distinguishable from all other Italian taxa and that its morphological similarity to B. plebejus and intermediacy with B. caninus are best explained by recent common ancestry and similar ecology with the former, rather than by hybrid origin involving these two species as previously hypothesized.

Morphological divergence of lake and stream Phoxinus of Northern Italy and the Danube basin based on geometric morphometric analysis.

Minnows of the genus Phoxinus are promising candidates to investigate adaptive divergence, as they inhabit both still and running waters of a variety of altitudes and climatic zones in Europe. We used landmark-based geometric morphometric methods to quantify the level of morphological variability in Phoxinus populations from streams and lakes of Northern Italy and the Danube basin. We analyzed body shape differences of populations in the dorsal, lateral, and ventral planes, using a large array of landmarks and semilandmarks. As the species identification of Phoxinus on morphological characters is ambiguous, we used two mitochondrial genes to determine the genetic background of the samples and to ensure we are comparing homogenous groups. We have found significant body shape differences between habitats: Minnow populations inhabiting streams had a deeper body and caudal peduncle and more laterally inserted pectoral fins than minnows inhabiting lakes. We have also found significant body shape differences between genetic groups: Italian minnows had deeper bodies, deeper and shorter caudal peduncles, and a shorter and wider gape than both groups from the Danube. Our results show that the morphology of Phoxinus is highly influenced by habitat and that body shape variation between habitats was within the same range as between genetic groups. These morphological differences are possibly linked to different modes of swimming and foraging in the respective habitats and are likely results of phenotypic plasticity. However, differences in shape and interlandmark distances between the groups suggest that some (though few) morphometric characters might be useful for separating Phoxinus species.

The Alps as barrier to dispersal in cold-adapted freshwater fishes? Phylogeographic history and taxonomic status of the bullhead in the Adriatic freshwater drainage.

The freshwater faunas of the Italian peninsula are isolated from the rest of Europe by the geographic barrier of the Alps and consequently have developed many endemic forms and contain few non-endemic species. However, some 'non-endemics' may either represent recent invaders of the Adriatic basin or cryptic endemic species. To test these two hypotheses against each other, we studied the origin and phylogenetic relationships of bullheads, cold adapted freshwater fishes of the genus Cottus, from both sides of the Alps and Dinaric Mountains. From the Adriatic basin, Cottus ferrugineus () was described as an endemic species, but the present analyses of sequences of the complete mitochondrial control region of 146 individuals from 43 localities showed no major differentiation between bullheads from both sides of the Alps. The very low diversification between representatives across the Alps suggests active transfers of haplotypes across this geographic barrier from the glacial cycles up to recent times. The transfers are most likely based on stream capture, since the cold-adapted bullhead is able to colonise the highest stretches of the water courses. No other freshwater fish in Europe is known to have experienced such an extensive gene flow across the highest European Mountains. In contrast, the Dinaric Mountains seem to have been a much more effective barrier between the Danube and the Adriatics. Our data reject the hypothesis of C. ferrugineus as an endemic species in the whole Adriatic drainage.