I used mitochondrial COI sequence data from forty one individuals to investigate phylogenetic relationships among populations of two morphologically similar species of the pseudoscorpion genus Apochthonius, A. minimus and A. occidentalis, in western Washington, Oregon, and northern California. My goal was to assess whether genetic structure in the two species was congruent with geography. Many plant and animal species in the Pacific Northwestern United States have shown patterns of genetic differentiation that follow both north-south and east-west trends, indicating that geologic and climatic events in the past separated populations to the extent that they became genetically differentiated. A distinct geographic pattern emerged within A. occidentalis, with at least one northern and two southern populations. A clade containing all A. minimus sequences was recovered. However, this clade falls within the larger clade of A. occidentalis, rendering A. occidentalis paraphyletic. Furthermore, the A. minimus sequences showed north-south geographic structuring within the clade. Population genetic analyses were performed based on geographic location within the Pacific Northwest. I found high genetic differentiation coupled with low gene flow between most populations, with the exception of the Portland and North Coast Range populations. These data suggest the presence of more than two species of Apochthonius in the Pacific Northwest.

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Population structure, biogeography and phylogenetic relationships of the fruit bat genus Rousettus have been studied in Africa and adjacent regions. The current population patterns of rousettine fruit bats in the Old World are influenced by several environmental attributes, namely the topography, climate and land cover. These variables are mirrored in fruit bat plesiomorphies related to the ecological niche of tropical flying frugivore, as well as apomorphies of rousettines including echolocation ability, roosting in caves and dispersal capacity in open landscapes with discontinuous tree cover. Phylogenetic relationships among species and subspecies of the genus have been indicated and confronted with the existing colonization scenarios. Insular populations (including habitat islands within desert oases) show frequent genetic differentiation from their mainland relatives suggesting successful founder events after traversing stretches of unsuitable habitats. Genetic differentiation evolving in less distant islands suggests involving behavioural mechanisms maintaining cohesion of isolated demes as site fidelity and natal habitat-biased dispersal. In sub-Saharan mainland Africa within the large range reaching from the southern border of Sahara to Cape Peninsula, Rousettus populations share a...

The mountain whitefish (Prosopium williamsoni) has been largely overlooked in population genetic analyses despite its wide distribution in discrete drainage basins in western North America for over four million years. Its closest sister taxa the Bear Lake whitefish (P. abyssicola), Bonneville cisco (P. gemmifer), and Bonneville whitefish (P. spilonotus) are found only in Bear Lake Idaho-Utah and were also included in the analyses. A total of 1,334 cytochrome b and 1,371 NADH dehydrogenase subunit 2 sequences from the Bonneville Basin, the Columbia River Sub-basin, the lower Snake River Sub-basin, the upper Snake River Sub-basin, the Green River Basin, the Lahontan Basin, and the Missouri Basin were examined to test for geographically based genetic differentiation between drainage basins and sub-basins and phylogeographic relationships to determine the invasion route of Prosopium into western North America and to aid in understanding current relationships. Prosopium entered the region via the Missouri River connection to Hudson Bay and moved in two waves: one colonized the lower Snake River Sub-basin, Columbia River Sub-basin, and the Lahontan Basin; the second wave colonized the upper Snake River Sub-basin, Bonneville Basin, Green River Basin, and established the Bear Lake Prosopium. Mountain whitefish exhibit a large amount of geographical genetic differentiation based on drainage basin except between the upper Snake River and the Bonneville Basin while the Bear Lake Prosopium show large amounts of gene flow between the three species. The apparent paraphyly of the mountain whitefish and the limited genetic structure of the Bear Lake Prosopium warrant recognition in the management of Prosopium and raise questions regarding species definitions in the group.

Phylogeography and molecular phylogenetics have proven remarkably useful for understanding the patterns and processes influencing historical diversification of biotic lineages at and below the species level, as well as delimiting morphologically cryptic species. In this dissertation, I used an integrative approach coupling comparative phylogeography and coalescent-based species delimitation to improve our understanding of the biogeography and species limits of Central American freshwater fishes. In Chapter 1, I conducted a literature review of the contributions of phylogeography to understanding the origins and maintenance of lower Central American biodiversity, in light of the geological and ecological setting. I highlighted emerging phylogeographic patterns, along with the need for improving regional historical biogeographical inference and conservation efforts through statistical and comparative phylogeographic studies. In Chapter 2, I compared mitochondrial phylogeographic patterns among three species of livebearing fishes (Poeciliidae) codistributed in the lower Nicaraguan depression and proximate uplands. I found evidence for mixed spatial and temporal divergences, indicating phylogeographic “pseudocongruence” suggesting that multiple evolutionary responses to historical processes have shaped population structuring of regional freshwater biota, possibly linked to recent community assembly and/or the effects of ecological differences among species on their responses to late Cenozoic environmental events. In Chapter 3, I used coalescent-based species tree and species delimitation analyses of a multilocus dataset to delimit species and infer their evolutionary relationships in the Poecilia sphenops species complex (Poeciliidae), a widespread but morphologically conserved group of fishes. Results indicated that diversity is underestimated and overestimated in different clades by c. ±15% (including candidate species); that lineages diversified since the Miocene; and that some evidence exists for a more probable role of hybridization, rather than incomplete lineage sorting, in shaping observed gene tree discordances. Last, in Chapter 4, I used a comparative phylogeographical analysis of eight codistributed species/genera of freshwater fishes to test for shared evolutionary responses predicted by four drainage-based hypotheses of Neotropical fish diversification. Integrating phylogeographic analyses with paleodistribution modeling revealed incongruent genetic structuring among lineages despite overlapping ancestral Pleistocene distributions, suggesting multiple routes to community assembly. Hypotheses tests using the latest approximate Bayesian computation model averaging methods also supported one pulse of diversification in two lineages diverged in the San Carlos River, but multiple divergences of three lineages across the Sixaola River basin, Costa Rica, correlated to Neogene sea level events and continental shelf width. Results supported complex biogeographical patterns illustrating how species responses to historical drainage-controlling processes have influenced Neotropical fish diversification.

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Context: West Nile virus (WNV) is an arthropod-borne virus considered a One Health challenge because of its increasing impact on human and animal health. It is one the most widely distributed viruses of the encephalitic Flaviviruses. It may cause severe neurological symptoms in humans and animals and is recognized as a serious public health problem also because of its impact on blood transfusion and organ transplantation. First identified in Africa in 1937, it was later introduced and spread in Italy, where in many regions it is now endemic, due to the increasingly favorable climatic and environmental conditions. Aim: The main objectives of this study, based on an interdisciplinary One Health approach, were: (1) to characterize the geographical distribution within specific host and vector populations in Africa; (2) to describe its phylogeographical patterns between Africa and Europe; (3) to define the genetic structure and epidemiology of Italian WNV strains, giving an insight of the viral circulation dynamics in the Italian territory. Methodology: Ecological and epidemiological studies were combined with molecular and phylogenetic analyses, carrying out field sampling activities, cellular culture, viral infection, immunofluorescent assay, multiplexed RT-PCR, sequencing, data analysis, and novel technique design. These activities were carried out both in Italy and in Senegal. Results: Our study evidences: (i) the circulation of several WNV lineages [Lineage 1 (L1), 2 (L2), 7 (L7), and 8 (L8)] in the African Continent; (ii) the presence of diverse competent mosquito vectors in Africa, mainly belonging to the Culex genus; (iii) the lack of vector competence studies for several other mosquito species found naturally infected with WNV in Africa; (iv) the need of more vector competence studies on ticks; (v) the circulation of WNV among humans, animals and vectors in at least 28 African countries; (vi) the lack of knowledge on the epidemiological situation of WNV for 19 African countries, and (vii) the importance of carrying out specific serological surveys in order to avoid possible bias on WNV circulation in Africa (objective 1). Furthermore, a new set of WNV L1 and L2 genome-specific primers for tiled-amplicon sequencing have been designed and a consistent dataset of 64 WNV L2 and 31 WNV L1 Italian genomes and of 3 WNV L2 and 7 WNV L1 Senegalese genome sequences from samples collected in Italy and Senegal between 2006 and 2022 has been produced. Twenty more WNV L1 and L2 Senegalese sequences obtained from samples collected in Senegal between 1985 and 2018 have been shared by the Institut Pasteur Dakar of Senegal and added to the dataset. This allowed the conduction of phylogenetic and phylogeographic analyses, evidencing: (viii) the presence of a strong viral connection between Africa and Europe, with intercontinental circulation supported by birds crossing international boundaries while migrating through the African-Eurasian flyways; (ix) the WNV L1 Western-Mediterranean cluster probable spread from Senegal, where the virus was first reported in 1979, to Italy, where the lineage first appeared in Europe in 1998, and to France in 2000, and the presence of back re-introductory events from Italy, Spain, and France to North and West Africa from the 2010s; and (x) the first African introduction of WNV L2 in Europe in Hungary in 2004, possibly from South African countries (objective 2). Our study also gives an insight of the dynamics of the viral circulation in Italy, demonstrating: (xi) the endemic presence of WNV L1 and L2 in part of Italy supported by resident wild birds and vector competent mosquitoes mainly belonging to the Culex genus; (xii) the current existence of two diverse WNV L1 strains circulating in Italy, one in the North-East, and one circulating intra-regionally in the Campania region; (xiii) suggested characteristic silent periods observed for WNV L1 in the country, with unnoticed circulation lasting sometimes for more than 10 years; (xiv) the 2022 WNV L1 increasing incidence of neurological disease cases in humans; (xv) the presence of genetically stable WNV L2 strains in Italy with continuous circulation throughout the time; (xvi) the presence of overwintering mechanisms supported by bird-to bird, rodent-to bird, or mosquito-to bird transmission routes; (xvii) the existence of WNV L1 and L2 co-infections in birds and mosquitoes; (xviii) the existence of a continuous transmission of the two strains between Western Mediterranean countries, supported by short distance migratory birds; and (xix) the crucial importance of the surveillance system other than the strategic role of wildlife rescue centers in monitoring both the introduction and circulation of avian emerging zoonotic diseases in Italy (objective 3). Conclusion: Our work points out the existence of high genetic diversity of WNV strains in Africa, the spread of L1 and L2 strains from Africa to Europe, and the existence of continuous transmission episodes among several Western-Mediterranean countries, with few recently suspected back introductory events from Europe to Africa. The progressive increase of the WNV L2 circulation both temporally and spatially in the Mediterranean countries and the WNV L1 re-appearance in Europe, both associated with a significant impact on humans and animal health, other than the strong WNV incidence in Italy and its endemization in part of its territory, evidence a solid WNV epidemic risk for Italy and a persistent threat for WNV spread into new areas. To predict and control future epidemics, it is crucial to constantly monitor the circulation and evolution of WNV in Europe and Africa, and to implement coordinated surveillance plans in both Continents, even in areas which are not currently affected.