Chez les insectes phytophages, l'adaptation à la plante hôte peut correspondre à l'une des premières étapes de la spéciation. Dans ce contexte, ce manuscrit s'intéresse à Spodoptera frugiperda (Lepidoptera : Noctuidae), un ravageur responsable de dommages importants sur de nombreuses cultures en Amérique et dans les Caraïbes. Spodoptera frugiperda présente deux variants, l'un adapté au riz et l'autre au maïs, et constitue un modèle biologique pertinent pour étudier ce mécanisme. En effet, les deux variants sont morphologiquement identiques, mais ils sont génétiquement différenciés et présentent des différences écologiques et comportementales. L'ensemble de ces caractéristiques suggèrent la présence d'un isolement reproducteur entre les deux variants, qui pourraient alors correspondre à deux espèces différentes. Cependant la présence d'hybrides dans la nature ainsi que des résultats controversés sur le succès d'accouplement entre les deux variants, rendent ambiguë le statut d'espèce de S. frugiperda. L'objectif de ce travail est donc de mieux estimer, à travers une étude intégrative, le niveau de différenciation génétique présent entre les deux variants de S. frugiperda. Grâce à un premier niveau d'analyse, menées à partir de population naturelles, il a été possible de mettre en évidence un niveau de différenciation génétique élevé entre les deux variants, qui est compatible à celui attendu entre deux espèces. Le deuxième niveau d'étude réalisé à partir de population de laboratoire nous a permis de mettre en évidence la présence d'un isolement reproducteur entre les deux variants, se traduisant notamment par une distorsion de ségrégation méiotique des marqueurs moléculaires chez les hybrides. L'ensemble de ces résultats supportent l'hypothèse selon laquelle le variant riz et maïs de Spodoptera frugiperda seraient des entités situées à une étape avancée dans le continuum de la spéciation. / In phytophagous insects, adaptation to host plant could be the first step leading to speciation. In this context, this document focus on moth Spodoptera frugiperda (Lepidoptera: Noctuidae), a pest responsible for serious damages in several crops in the Western hemisphere. Spodoptera frugiperda consists of two host-plant strains, one adapted to rice and the other adapted to maize, which made it a relevant model to study this mechanism. Though the two variants are morphologically identical, they are nonetheless genetically distinguishable and present some ecological and behavioral differences. The species status of S. frugiperda is also highly controversial because hybrids naturally occur in the wild, not to mention the fact of the discrepancies among published results concerning mating success between the two strains. The aim of this thesis is thus to better estimate, through an integrative approach, the level of genetic differentiation between the two variants of S. frugiperda. Starting with natural populations, various phylogenetic methods allowed us to highlight a high level of genetic differentiation between the two variants, compatible with what is expected between distinct species. Furthermore, studies on laboratory populations, including crossing experiments, showed a significant unidirectional bias in inter-strain mating success and the presence of meiotic segregation distortion of molecular markers in hybrid progenies. These results as a whole support the assumption that the two strains of S. frugiperda are well-advanced in the continuum of speciation.

The role of divergent climate adaptation in speciation has received surprisingly little scientific attention. My dissertation research focused on how resting metabolic rate (RMR) relates to the build up of prezygotic and postzygotic isolation in a natural Ficedula flycatcher hybrid zone. RMR is the amount of energy an organism needs to run its internal organs. Since RMR is related to life history traits and thermoregulation in other systems, it is likely to affect speciation processes at secondary contact. I found that adult collared flycatchers displace pied flycatchers into increasingly poor habitats (Paper I). Pied nestlings exhibit lower RMR in poor environments (Paper II), which may promote regional coexistence and habitat isolation by making it possible for pied flycatchers to escape competition from collared flycatchers and reduce the risk of hybridization by breeding in the poorer habitats. Further, I found that while collared flycatcher nestling RMR was not environmentally-dependent (Paper II, Paper III), those collared flycatcher nestlings that had a lower RMR in poor environments tended to have higher condition (Paper III). Further, RMR was genetically linked to a sexual ornament in collared males that has previously been shown to be beneficial in poor environments. Lastly, I found that by seven days old, nestlings increase their metabolic rate when listening to song, indicating that they are listening, and by 9 days they can discriminate between songs (Paper IV). Taken together, RMR could affect pre-zygotic isolation via correlations with life history strategies, song and sexual ornaments. RMR is also related to post zygotic isolation in Ficedula flycatchers. I found that flycatcher hybrids tended to have a higher RMR than the parental species (Paper V), and that there were many differentially expressed genes in energetically expensive organs in hybrids that were related to metabolic function (Paper VI). Thus, metabolic dysfunction, possibly caused by genetic incompatibilities, in Ficedula flycatcher hybrids could be a factor leading to infertility and postzygotic isolation between the parental species. Overall, I find that RMR could be a general physiological trait that affects both pre- and postzygotic isolation in hybridizing species at secondary contact, and ought to be more thoroughly considered in speciation research.

Três tipos de sons de cortejo foram descritos para machos de tefritídeos: de chamada, na ausência de fêmeas; de aproximação, na proximidade de fêmeas; e de pré-cópula, após a monta das fêmeas pelos machos. Nas espécies do gênero Anastrepha em que foram registrados sons de acasalamento, somente dois tipos estão presentes: de chamada e de précópula. As emissões acústicas de acasalamento de algumas espécies foram caracterizadas quanto a parâmetros como duração e freqüência dos sons, incluindo a espécie nominal Anastrepha fraterculus - importante praga da fruticultura nacional. No entanto, até o presente momento, nenhum estudo teve como foco a diferenciação dentro do complexo A. fraterculus, constituído por diversas espécies crípticas - das quais, três caracterizadas para o Brasil: A. sp. 1 aff. fraterculus, A. sp. 2 aff. fraterculus e A. sp. 3 aff. fraterculus. Este trabalho procurou caracterizar o padrão de emissão acústica de chamada de populações do complexo A. fraterculus (duas de A. sp. 1, uma de A. sp 2 e uma de A. sp. 3) e de A. obliqua, em relação à duração de pulso (PD), intervalo entre pulsos (IPI) e freqüência fundamental (FF), comparando com dados disponíveis na literatura para outras espécies. Os parâmetros acústicos de A. sp. 1 de Serra Negra - SP - não se diferenciaram estatisticamente dos parâmetros de A. sp. 2 de São Sebastião - SP. O padrão de emissão de A. sp. 3 de São Sebastião - SP -, difere do de A. sp. 1 de Serra Negra - SP - e de A. sp. 2 de São Sebastião - SP - no parâmetro PD. Os parâmetros acústicos de A. sp. 1 de São Paulo - SP - foram completamente distintos dos parâmetros das demais populações, inclusive dos de A. sp. 1 de Serra Negra - SP. O padrão de emissão acústica A. obliqua de Águas da Prata - SP - também diferencia-se dos das demais espécies/populações, exceto em relação ao PD de A.. sp. 1 de São Paulo - SP. A relação entre picos de distribuição de FF e de harmônicos, bem como a correlação entre parâmetros acústicos e o comprimento das asas sugerem uma relação entre características físicas das asas e os sons emitidos. Porém, outros fatores também devem estar em atuação, uma vez que a relação entre picos e harmônicos não é observada para todas as populações estudadas e a correlação entre tamanho de asa e valores de parâmetros acústicos é baseada em número limitado de espécies. Não há uma relação entre o valores dos parâmetros acústicos e as relações filogenéticas entre as espécies de moscas-das-frutas. Assim, embora os padrões de emissões acústicas permitam diferenciar as espécies entre si, não é possível utilizá-los para agrupar as espécies quanto a afinidades evolutivas. Dentro dos tefritídeos, a evolução dos tipos de emissão acústica do acasalamento provavelmente envolve uma história bastante complexa com várias homoplasias. / Three courtship songs have been described for tephritid males: calling song, when female flies are absent; approach song, near females; and precopulatory song, after male mounting female. For species of Anastrepha in which mating songs were recorded, only two types are present: calling and precopulatory songs. In these cases mating accoustic emissions were characterized regarding parameters such as duration and sound frequency; including for the nominal species Anastrepha fraterculus - a major fruit crop pest in Brazil. However, no study have focused on differences amid A. fraterculus criptic species complex - including the three species that occur in Brazil: A.. sp. 1 aff. fraterculus, A. sp. 2 aff. fraterculus e A. sp. 3 aff. fraterculus. In this work, the acoustic emission patterns for A. sp. 1, A. sp. 2 A. sp. 3, and for A. obliqua population samples were characterized by comparing pulse duration (PD), interpulse interval (IPI) and fundamental frequency (FF), with data available in literature for other tephritid species. The acoustic parameters of A sp. 1 Serra Negra (SP) do not differ statistically from parameters of A. sp. 2 (São Sebastião, SP). The PD parameter of A. sp. 3 (São Sebastião, SP) differs from A. sp. 1 (Serra Negra, SP) and from A. sp. 2 from São Sebastião (SP). The acoustic parameters of A. sp. 1 from São Paulo (SP) were completely different from parameters of other populations, including A. sp. 1 from Serra Negra (SP). The acoustic emission pattern of A. obliqua from Águas da Prata (SP) is also different from other species/populations, except by the PD of A. sp. 1 from São Paulo (SP). The relationship between FF distribution peaks and sound harmonics as well as the correlation between acoustic parameters and wing length, suggest that the emitted sounds are dependent on the wing physical properties. However, other factors might also influence the sound pattern, because the relation between FF peaks and harmonics is not observed for all population samples, and the correlation between wing length and acoustic parameters were evaluated on few species. There is no relation between acoust parameters values and fruit flies phylogeny. Therefore, it is possible to distinguish between species on the basis of sound parameters, but it is not possible to group them by evolutionary affinities. Among the tephritids, mating acoustic emission patterns probably involves a complex evolutionary history with many homoplasies.

Reproductive isolation can be achieved through multiple types of barriers and is essential for speciation. In flowering plants, pre-pollination barriers (e.g. differentiation in pollination niches) are believed to be the most efficient at preventing gene flow across species boundaries. In closely related species that come into secondary contact, such barriers can evolve to prevent competition for pollinator service and/or interspecific pollen transfer, which can have fitness costs. Hence pollination niche differentiation should be stronger in sympatric populations than in allopatric populations (i.e. character displacement). To investigate the differences in pollination niches and to see if it is consistent with a hypothesis of character displacement, I used the two closely related and phenotypically similar orchid species, Gymnadenia conopsea and G. densiflora. I sampled mixed and pure populations of G. conopsea and G. densiflora on Öland during the summer of 2020. In these populations, I used video cameras and pollinator catches to record pollinator activity and characterize the composition of pollinator communities. Estimation of pollinator efficiency was also assessed by analyzing the number of pollinia carried by each pollinator. Contrary to my expectations, I found that both orchids had their visitation peak during the night and that the most frequent and efficient pollinators were Autographa gamma or Deilephila porcellus for both of them. Furthermore, no increased differentiation between the two species was found in mixed compared to pure populations. My results suggest that plant-pollinator interactions do not act as efficient pre-pollination barriers in these two orchid species, and that competition for pollinator service and through interspecific pollen transfer seem to be too weak to drive pollination niche partitioning.

The origin and maintenance of much of natural biodiversity can likely be explained by theecological speciation theory. Ecological speciation is defined as the process whenreproductive isolation arises through divergent selection caused by ecological factors, such asbiotic interactions and abiotic conditions. Many populations of European whitefish(Coregonus lavaretus) are currently going through sympatric speciation from a single littoralecotype to two or more ecologically separate ecotypes. It has been hypothesized that northernpike (Esox lucius) is promoting speciation in whitefish by exposing it to a trade-off betweenpredation risk and resource availability causing an initial divergence in size eventuallyleading to two or more ecologically and genetically distinct ecotypes. After initial divergencesmall whitefish ecotypes often abandon ancestral spawning sites to spawn in streams or deepwater suggesting that the ecotypes are at least partially reproductively isolated. The driver ofthis reproductive isolation could be predation risk as pike mainly forage in the littoral. If thisis true, we would expect to find significantly higher predation risk for small whitefish at thelittoral spawning sites than on alternative spawning sites. To test this idea, hooks baited withsmall whitefish was placed at different spawning sites during the fall spawning season andchecked every 12 hour for 36 hours. Relative predation risk for small whitefish wassignificantly higher at the ancestral littoral spawning sites than at stream spawning sites. Inaddition, there was a significant decrease in predation risk later in the spawning season. Thisindicates that predation is promoting reproductive isolation in European whitefish byselecting against small individuals spawning at littoral spawning sites, but potentially also byselecting for delaying spawning in small individuals in the littoral. To fully understand thespeciation process in this system more studies must be done considering the complexity ofthe speciation process. Because of the lack of studies investigating predation as a cause ofbehavioral changes, that in turn promotes reproductive isolation, future studies should putmore emphasis on predation as a possible driver of reproductive isolation.

The goal of this study was to examine the taxonomy, ecology, molecular phylogenetics and biosystematics of two apparently closely related species and to elucidate congruent patterns from these often divergent sub-disciplines of systematics. Phylogenetic analysis of DNA sequences showed that Phacelia fimbriata and Phacelia purshii are sister species that demonstrate species-specific differences in germination but little morphologic or genetic differentiation. While a geographic region of sympatry exists, elevation differences preclude co-occurrence. The widespread P. purshii supports higher levels of morphological and genetic diversity compared to the narrow endemic, P. fimbriata. Gene flow between members of this closely related species pair is blocked by hybrid inviability whose expression occurs following normal pollen tube growth and apparent fertilization. Hybrid seeds are inviable, an effect likely caused by an embryo-endosperm incompatibility that, following fertilization, permits development of maternal, but not hybrid fruit and seed tissues. This reproductive barrier adds to our knowledge of an already diverse suite of isolating mechanisms in Phacelia subgenus Cosmanthus.

Collectively, populations of Acanthocyclops vernalis, a species complex of freshwater copepods, are remarkably similar as to morphology and DNA content, despite variability in chromosome number. Reproductive isolation had been reported among some populations, but with each new investigation the species boundaries and factors that may influence them appeared less clear. To clarify the pattern of biological species within this group of populations, we adopted a comprehensive approach and examined patterns of reproductive isolation in populations for which morphology, chromosome number, DNA content, and 18S rDNA sequences are known. In this study we established nine isofemale lines from four sites in Wisconsin and performed 266 crosses. Crosses within and among these lines were used to relate the degree of reproductive isolation to chromosome differences and to construct a model to explain the origin and maintenance of chromosome number variability. Different gametic and somatic chromosome numbers were observed among specimens within some isofemale lines. In a few cases, gametes with different haploid numbers were produced by a single female. Matings within isofemale lines always produced at least some reproductively successful replicate crosses (produced viable, fertile offspring). Crosses between lines from the same site showed reduced success relative to within-line crosses. Crosses between populations from distant sites showed limited genetic compatibility, producing viable, fertile F1 offspring but infertile F2 adults. One cross between lines with different chromosome numbers (one with 2n = 8 and one with 2n = 10) produced fertile viable offspring, which reproduced for at least 60 generations. These hybrids had either eight or nine chromosomes in the third generation of inbreeding, and eight chromosomes after 20 generations. These hybrids also had reduced nuclear DNA contents at the third generation, a level that persisted through the 20th generation. Successful backcrosses between some hybrids and their parental lines further demonstrated the potential for genetic compatibility among forms with different chromosome numbers. We propose a model in which alterations due to Robertsonian fusions, translocations, and/or loss of chromosomal fragments generate heritable variation, only some of which leads to reproductive isolation. Hence, some of the criteria traditionally used to recognize species boundaries in animals (morphology, DNA content, chromosome number) may not apply to this species complex.

Reproductive barriers are vital to generating new species as well as maintaining distinct species. Investigating reproductive barriers between closely related plant taxa helps us to understand how these barriers are maintained, particularly between rare and widespread relatives. Layia jonesii, a rare San Luis Obispo County serpentine endemic, and L. platyglossa, a common coastal species, co-occur on serpentine derived hillsides and are interfertile. At these locations, L. jonesii is isolated to dry soils near serpentine rock outcrops and L. platyglossa is located on slightly deeper grassland soils surrounding the rock outcrops. On hillsides where they co-occur, I observe two morphologically distinct species, therefore the two species must be maintaining reproductive barriers, yet mechanisms that maintain this isolation are unknown. I studied this system to investigate possible mechanisms contributing to the maintenance of reproductive barriers. I hypothesize prezygotic reproductive isolation in this system is due to (1) habitat isolation due to local adaptation to differential edaphic environments on the hillside, (2) flowering time differences, and (3) reduced seed set resulting from hybrid crosses. To investigate the local adaptation of L. jonesii and L. platyglossa, I reciprocally transplanted both species into the center of each species’ distribution. I also conducted a competition experiment to determine if L. jonesii is sensitive to resource competition beyond its natural distribution. To investigate flowering time differences, I tracked flowering time of both wild and reciprocally transplanted populations. I also performed controlled crosses to determine if heterospecific, or hybrid crosses, result in lowered seed set than conspecific crosses. The reciprocal transplants showed L. platyglossa is locally adapted to the grassland habitat. Local adaptation likely prevents L. playtyglossa from dispersing into the rock outcrop habitat. Results of the competition experiment revealed L. jonesii is sensitive to competition and this may contribute to its constrained distribution to shallow soils. Local adaptation and competition likely contribute to habitat isolation between the two species. I also documented stark differences in flowering time between the species which contributes to reproductive isolation by reducing pollen flow. Hybrid crosses also resulted in lowered seed set than conspecific crosses. These results suggest prezygotic barriers to reproduction likely maintain the majority of isolation between the two species. These results provide insight into mechanisms that maintain reproductive barriers between closely related taxa existing in similar habitats. The results also contribute to our understanding of how rare plants preserve genetic integrity near common and interfertile relatives.

In basidiomycetous fungi, the viability of gametes is an important component of sexual fitness and can have implications for speciation events. Prior estimates of basidiospore germination are highly variable and the occurrence of reproduction between these lineages suggests that reproductive isolation is incomplete. Genetic incompatibilities during meiosis have been attributed to much of the offspring inviability. However, the influence of environmental factors on basidiospore germination in Cryptococcus are not well known. In this study, we used human opportunistic yeast pathogens, Cryptococcus neoformans and Cryptococcus deneoformans, as models to investigate the potential effects of selected genetic and environmental factors on basidiospore germination. We evaluated basidiospore germination of six genetic crosses by pairing a total of five strains, three intraspecific crosses and three interspecific crosses, between C. neoformans and C. deneoformans. The offspring of genetic crosses were incubated under multiple media and temperature treatments and scored for their germination ability. In general, spores from intraspecific crosses had greater germination potential than those from interspecific crosses. Growth under high temperatures was the most significant influence on basidiospore germination on these crosses. Furthermore, there were notable interaction effects between environmental factors and parental strains or strain pairs on basidiospore germination. The interaction between the sex-specific genes and environmental pressures impact reproductive barriers and blur species distinctions within the Cryptococcus neoformans species complex. And so, reduced viability of hybrid offspring can have implications for Cryptococcus speciation, ecology, and pathogenesis as hybridization events are an effective method of increasing pathogenicity, expanding species distributions and increasing tolerance to novel environments or hosts. / Thesis / Master of Science (MSc) / In basidiomycetous fungi, the viability of gametes is an important component of sexual fitness and can have implications for speciation events. The Cryptococcus neoformans species complex are a group of opportunistic pathogens, for which hybridization can be facilitated readily under laboratory conditions, creating offspring that are generally completely inviable or have low germination potential. Antagonistic genetic interactions are mostly responsible for offspring inviability, yet the impacts of environmental factors is not known. Multiple genetic crosses between Cryptococcus strains were used as a model to investigate species relationships by examining the impacts of genetic and environmental factors on offspring germination potential.

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