Two eucalypt clines were examined using morphological, ecophysiological and molecular analyses. The species complexes examined were an ironbark complex (Eucalyptus melanophloia x E. whitei) and a box complex (E. brownii x E. populnea). Both of these complexes demonstrate continuous morphological variation across their clines. The origin of these morphological clines has previously been interpreted as the product of secondary contact between allopatric species. In this study, an analysis of morphological variation across the clines did not identify an increase in trait variance in the intermediate populations, which suggests that previous theories concerning the origin of these clines may not be valid. Genetic structuring in nuclear and chloroplast DNA was examined across the clines to investigate whether the morphological clines were the product of secondary contact between two independent evolutionary lineages, or whether the clines represent a single evolutionary lineage that has undergone primary differentiation. The microsatellite analyses indicated that there was little genetic structuring across either cline, and that there were only low levels of population differentiation. The lack of hierarchical structuring in the distribution of nuclear genetic variation suggests that these clines are unlikely to be the product of recent gene flow between two formerly allopatric species/populations. A nested clade analysis of the JLA+ region of the cpDNA provides additional evidence to reject the null hypothesis that the morphospecies classifications represent distinct evolutionary lineages. Instead the analyses indicate that each cline represents a single cohesion species and a single evolutionary lineage. The phylogeographic distribution of cpDNA haplotypes is likely to have resulted from restricted seed mediated gene flow with isolation by distance. A more cogent explanation for the clines, based on the genetic data, is that they have arisen through the process of continuous morphological diversification that has been promoted by a directional selection gradient. Drought experiments were conducted in the glasshouse to investigate whether differences in physiological performance under water stress helps to explain the maintenance of the ironbark cline. Under increasing water stress, the morphotypes showed differences in their ability to maintain water status and photosynthetic rates, yet there was no obvious pattern to these differences across the cline. Physiological differences are therefore inadequate to explain the maintenance of the ironbark cline and highlight the compensatory role that morphological variation may play in alleviating water stress. The value of adopting the cohesion species concept and a hypothesis-testing framework to assess species status is demonstrated in this study. This framework provided a statistical approach to distinguish independent evolutionary lineages from interspecific populations and provides evidence to refute the current species status of the species complexes studied. Eucalypt classification is predominantly based on morphology, which results in taxonomic classification that may not reflect genealogical relationships. This is due to the disparity between morphological and phylogenetic relationships. I therefore suggest that current presumptions regarding the prevalence and importance of hybridisation within the genus may reflect taxonomic classification. An accurate assessment of the prevalence and importance of hybridisation requires species classification to be based on genealogical relationships.

<p> This work incorporates information from two sources in order to examine the nature of natural selection acting on phenotypic characters in Drosophila melanogaster along a North South cline. Isofemale lines were established from flies collected along a North South cline extending from Winnipeg, Manitoba to Tampa Bay, Florida. Offspring from different lines within each position were then cultured under standardized conditions and used to examine phenotypic variation in 10 morphological characters along the cline. In a separate set of experiments, flies from Vineland, Ontario were mated in a half-sib design in order to estimate the genetic covariance of the set of 10 characters. The results from the clinal and heritability experiments were then combined using Lande's (1979) equation,&.= Gp-ls, to estimate the net selective differentials and net selection gradients for each adjacent set of populations. The study concluded that: 1) Clinal variation is non-linear, with larger flies in the middle latitudes and smaller flies in the north and south. 2) Selection appears to act primarily on body characters in the north (wing width and femur length) and head characters in the south (eye and face width). 3) Scutellum width and wing length generally moderate the prevalent trends in directional selection on the other characters through antagonistic correlated responses. 4) Clinal patterns of variation may not be at equilibrium, but instead dominated by seasonal responses to selection pressures. </p> / Thesis / Master of Science (MSc)

Evolutionary process can be inferred at three different levels: the species level, the population level and the molecular level. In this thesis, I applied approaches at these three levels and aimed to get a comprehensive picture of conifer evolution, from speciation and demography to geographic variation and local adaptation, and then to the molecular evolution of proteins and small regulatory RNAs. Spruce species have been observed to possess a large number of trans-species shared polymorphisms. Using an “Isolation with migration” model, we found that the large effective population size of spruce retained these shared polymorphisms, inheriting them from the common ancestor. Post-divergence gene flow only existed between Picea abies and P. glauca, and between P. wilsonii and P. schrenkiana. The combination of Tajima’s D and Fay &amp; Wu’s H at most of loci suggested an ancient and severe bottleneck for most species except P. breweriana. Furthermore, I investigated the effect of local selection in two parallel clines, which is one of the major forces that can cause divergence or even speciation. The timing of bud set and growth cessation was found correlated with latitude in populations of P. abies and P. obovata. Using allele frequency spectrum analyses we identified three genes under local selection in both species including two circadian-clock genes GI and PRR7, and one photoperiodic gene FTL2. This indicated that parallel evolution could occur through groups of genes within related pathways. Clinal variation at expression level provided stronger evidence of selection in FTL2, which has previously been associated with bud set in P. abies. Finally we focused on the molecular evolution of mRNA and small regulatory RNAs in P. abies. With the help of Next-Generation sequencing, we have achieved in spruce the first de novel assembly of the needle transcriptome and a preliminary characterization of sRNA populations. Along with features common in plants, spruce also exhibited novelties in many aspects including lower substitution rate and protein evolutionary rate, dominance of 21-nt sRNA, and a large proportion of TIR-NBS-LRR genes as sRNA sources and targets.

The cosmopolitan Pteridium aquilinum (L.) Kuhn is widespread throughout eastern North American, where it is represented primarily by Tryon's (1941) var. latiusculum (Desv.) Underw. and var. pseudocaudatum (Clute) Heller. The taxonomy of Pteridium is controversial. Fourteen isozyme loci and 12 morphological characters were used to assess the taxonomic relationship of these two varieties. Isozyme data indicated a high mean genetic identity (I = 0.976) between eleven bracken populations. Strong patterns of geographic variation for isozyme allele frequencies were also observed. The isozyme results did not separate the two taxa. Numerical analysis of the morphology distinguished the two taxa when the qualitative characters were used alone or in conjunction with some of the quantitative traits. All qualitative characters differed significantly between the two taxa. No perceptible geographic pattern of variation was observed. Morphological distinctiveness was maintained even in those localities where both taxa were present, with few or no intermediates being found. Isozyme evidence suggestive of gene flow between the two varieties was found at Greensboro, NC, where the two morphotypes were easily recognizable. The isozyme evidence strongly indicates conspecificity, while the morphological evidence supports their status at the varietal level. / Master of Science

The postglacial recolonization of northern Europe supposedly originated from Western Europe and the Russian Plain, however, recent molecular and macrofossil-based investigations suggest that the history may be more complex than previously thought. This study aims to investigate the genetic diversity and population structure of Scots pine from Scandinavia to Russia to re-evaluate its recolonization history, and to examine whether the pattern of spatial genetic diversity has any adaptive significance. Populations ranging from Norway to Russia were sampled and genotyped using genotyping-by-sequencing. The seedlings were freeze tested to provide an average degree of hardiness for every population. Eight hundred and thirty-two seedlings were analyzed, and 6,034 SNPs were recovered in these individuals after stringent filtering. Population structure was investigated using fastStructure and differentiation between populations was estimated with pairwise FST and analysis of molecular variance (AMOVA) to assess the genetic variability. Genetic diversity was measured as observed heterozygosity, H0, in populations, clusters and overall. Two genetic clusters were detected in the samples, one in Norway and Sweden and one in Russia. These clusters are weakly differentiated (FST = 0.01202) with only 0.66 % variation between them. Highest variation was found within populations (98.8 %) and the overall genetic diversity for all populations was high (Ho = 0.2573). The weak differentiation and high diversity are indicative of extensive gene flow between populations in this species. The composition of the clusters across the sampled area suggests a westward recolonization from the Russian Plain into Scandinavia, and a possible local origin of another polymorphism in Norway and Sweden. No clear relationship between cold hardiness and genetic variation was detected. The clinal variation in cold hardiness reflects local adaptation, and the difference between genetic and phenotypic variation is likely due to epigenetic regulation or polygenic inheritance. More extensive genome scan is needed to understand the genetic basis of local adaptation.

Recent models argue that thermal environments are the major cause of ectotherm life-history clines. However, elevational clines in body size in the mountain dusky salamander Desmognathus ocoee (family Plethodontidae) shift from positive at hatching, to negative at metamorphosis to positive again as adults, and so are not consistent with this explanation. The clinal shift from hatching to metamorphosis was investigated by examining the clinal and seasonal feeding patterns of larval salamanders at high and low elevation sites in rockface and woodland habitats. Repeated cohort sampling was also used to examine clinal and seasonal patterns in body size and to estimate average growth rates. Larval growth in both rockface and woodland habitats was tightly correlated with feeding activity. Although temperature was found to vary between high and low elevation sites, the greatest growth occurred in a cold woodland habitat with a high elevation, and the lowest growth occurred in an adjacent rockface habitat. Because this difference in growth cannot be attributed to thermal differences, we conclude that local food resource levels are the predominant source of local differences in growth. These findings, clinal patterns of variation in other predatory salamanders, and experimental analyses in which both food and temperature are orthogonally manipulated, indicate that general models that single out temperature as the principle cause of ectotherm life-history clines should be viewed with caution.

<p>In this thesis, quantitative genetic approaches, common-garden experiments, and field studies were combined to examine patterns of population differentiation and the genetic architecture of characters of putative adaptive significance in the widely distributed perennial herb Lythrum salicaria. In this work, I (1) documented patterns of population differentiation in phenology, life-history, and morphology along latitudinal gradients at different geographical scales, (2) investigated the genetic architecture of phenology, flower morphology, and inflorescence size, and (3) combined estimates of phenotypic selection in the field with information on the genetic variance-covariance matrix (G) to examine potential constraints to adaptive evolution. </p><p>A common-garden experiment demonstrated latitudinal variation in life-history, and phenology of growth and reproduction among L. salicaria populations sampled across Sweden. Flower morphology varied significantly among populations, but was, with the exception of calyx length, not related to latitude of origin. A second experiment, which included two Swedish, two Dutch, and two Spanish populations, indicated that the latitudinal gradient in reproductive and vegetative phenology might extend throughout Europe.</p><p>A quantitative-genetic study of two Swedish populations revealed significant additive genetic variation for all phenological and morphological traits investigated. The G matrices of the populations differed significantly according to common principal component analysis, and genetic correlations within the study populations did not strictly correspond to trait correlations observed among populations.</p><p>In a field study, I detected directional selection through female function for larger inflorescences in two consecutive years. Relative fitness increased disproportionately with inflorescence size in the year when supplemental hand-pollination indicated that pollen limitation was severe. Genetic correlations with inflorescence size considerably influenced predicted response to selection in other characters.</p><p>Taken together, the results suggest that among-population differences in phenology and life-history in L. salicaria have evolved in response to latitudinal variation in length of the growing season. They demonstrate that the evolutionary potential of local populations may be considerable. The genetic covariance structure substantially influences predicted short-term evolutionary trajectories. However, the weak correspondence between genetic correlations documented within populations and trait correlations among populations, suggest that the G matrix has not imposed strict constraints on patterns of among-population differentiation.</p>

In this thesis, quantitative genetic approaches, common-garden experiments, and field studies were combined to examine patterns of population differentiation and the genetic architecture of characters of putative adaptive significance in the widely distributed perennial herb Lythrum salicaria. In this work, I (1) documented patterns of population differentiation in phenology, life-history, and morphology along latitudinal gradients at different geographical scales, (2) investigated the genetic architecture of phenology, flower morphology, and inflorescence size, and (3) combined estimates of phenotypic selection in the field with information on the genetic variance-covariance matrix (G) to examine potential constraints to adaptive evolution. A common-garden experiment demonstrated latitudinal variation in life-history, and phenology of growth and reproduction among L. salicaria populations sampled across Sweden. Flower morphology varied significantly among populations, but was, with the exception of calyx length, not related to latitude of origin. A second experiment, which included two Swedish, two Dutch, and two Spanish populations, indicated that the latitudinal gradient in reproductive and vegetative phenology might extend throughout Europe. A quantitative-genetic study of two Swedish populations revealed significant additive genetic variation for all phenological and morphological traits investigated. The G matrices of the populations differed significantly according to common principal component analysis, and genetic correlations within the study populations did not strictly correspond to trait correlations observed among populations. In a field study, I detected directional selection through female function for larger inflorescences in two consecutive years. Relative fitness increased disproportionately with inflorescence size in the year when supplemental hand-pollination indicated that pollen limitation was severe. Genetic correlations with inflorescence size considerably influenced predicted response to selection in other characters. Taken together, the results suggest that among-population differences in phenology and life-history in L. salicaria have evolved in response to latitudinal variation in length of the growing season. They demonstrate that the evolutionary potential of local populations may be considerable. The genetic covariance structure substantially influences predicted short-term evolutionary trajectories. However, the weak correspondence between genetic correlations documented within populations and trait correlations among populations, suggest that the G matrix has not imposed strict constraints on patterns of among-population differentiation.

The Rufous-naped lark Mirafra africana complex consists of 22 subspecies spread across the African continent. Several of the subspecies have recently been suggested to potentially be treated as separate species. In this study a comparative analysis was done on the song from seven of the subspecies: M. a. africana, M. a. athi, M. a. grisescens, M. a. kabalii, M. a. nyikae, M. a. transvaalensis and M. a. tropicalis. The results showed that M. a. athi, M. a. kabalii and M. a. nyikae are all very divergent from each other as well as from the other four subspecies. In contrast, M. a. tropicalis, M. a. grisescens, M. a. africana and M. a. transvaalensis are not clearly separable from each other. Based on the results, I suggest that M. a. athi, M. a. kabalii and M. a. nyikae can be classified as separate species, with M. a. africana, M. a. tropicalis, M. a grisescens and M. a. transvaalensis forming a fourth species (M. africana sensu stricto). Finally, I conclude that this study shows that more studies need to be done on the subspecies of the Mirafra africana complex.

We reviewed the taxonomy of the Trogon rufus species-complex under the premises of the Biological Species Concept. Putative taxonomic units, breaks and transition zones, were visualised by heatmaps and isophenes (phenotypic contour lines) of the colour, barring, morphometric and song characters and tested by discriminant function analyses. Colourmetric data were obtained via spectrometry and barring patterns analysed via high quality digital photographs. We found four distinct biological species. Trogon chrysochloros Pelzeln 1856 from the Atlantic Forest with its denser and blacker undertail and wing covert barring, larger size and faster, generally higher song with more notes. Its upperparts vary from bluer to more coppery-green with increasing altitude. The bill is also relatively smaller and more serrated, linked to a diet that consists almost exclusively of large arthropods, making it the most insectivorous new world Trogon species yet known, which may account for its relative rarity compared to other Trogonids with which it is sympatric. Trogon tenellus Cabanis 1862, from Central America, and Trogon cupreicauda Chapman 1914 from the Chocó-Magdalena provide a classic case of typical biological species, coming into contact in the extreme NW Chocó Province, Colombia, but without intermediate forms. T. tenellus is identified by its blue to blue-green uppertail, blue or grey eye-rings, grey tarsi and song with 2-4 notes, longer note duration and greater change in peak and high frequencies between the intro note and loudsong. This contrasts with the shiny olive-green to coppery green uppertails, yellow eye-ring, usually olive tarsi, brown wash on the undertail of females and song with 6-8 notes of shorter duration and little change in frequency between the intro note and loudsong of T. cupreicauda. T. cupreicauda varies clinally from generally bluer- to more coppery-green plumage and from thicker to thinner black bars in a gradient from the Pacific coast on the border with Ecuador to the Magdalena Valley. The greater difference in colour and barring relative to T. tenellus in the region they come into contact provides possible evidence of character displacement as a result of the competitive exclusion between these two species, maintaining their parapatric distributions. The Amazonian population belongs to a single species, Trogon rufus Gmelin 1788, but with two highly distinct forms that we designate as Trogon rufus rufus in the Guiana Shield and Trogon rufus sulphureus in S & W Amazonia, for which Todd\'s amazonicus is synonymised. They are morphologically and, to a lesser extent, vocally distinct across the lower Rio Negro and matrix of highland and open habitats of the Rio Branco basin but show limited character exchange between the 52-58th parallels west on the southern bank of the Amazon, centred around the Rio Arapiuns on the left bank of mouth of the Tapajos. We postulate that this is the result of secondary contact as a consequence of shifts in the course of the main channel of the Amazon River at times of lower sea levels during the Plio-Pleistocene. T .r. sulphureus is identified by a typically coppery uppertail with subterminal tailband of greener hue, yellow eye-ring, low barring density and broad black bars of the undertail and wing-coverts barring with and lack of a pectoral band. They are also sometimes distinguishable in song by a higher frequency introduction note and/or more pronounced descending modulation across the loudsong. This varies clinally on a west-east gradient, from strong-coppery to shiny olive-green uppertails with more to less distinct subterminal tailbands, diminishing black bar widths with corresponding increasing density and decreasing intro note low frequency. T. r. rufus have green uppertails, blue eye-rings, presence or absence of a white pectoral band and denser undertail and wing panel barring with thinner black bars. These characters were shown to change as a function of geographic distance between specimens of sulphureus and rufus, connected via the \'Arapiuns contact zone\', suggesting isolation by distance. This is reminiscent of a ring species pattern and two specimens with a possible mixture of characters were indeed found from the upper Rio Negro and in Pantepui, where T. r. rufus and T. r. sulphureus would be expected to come into contact, effectively \'closing the ring\'. Whether Trogon rufus constitutes a valid ring species requires further testing, preferably including molecular characters, but this clearly illustrates that the distinction between clinal variation and ring-species is a matter of degree, not kind, with the formation of the ring-species necessarily passing through a clinal stage with no overlap between terminal taxa. We therefore propose the concept of a loop species, where the terminal forms do not overlap but are connected via a series of intergrading populations. It seems likely that such patterns are more widespread in Amazonia than presently known due to the propensity for clinal variation and parapatric speciation lended by its massive geographical extent and abundance of biogeographical semi-permeable barriers. With regards to the population from the Pernambuco Center of Endemism, the few records suggest that it is a valid taxonomic unit. It has the unique combination of a song very similar to T. r. sulphureus due to the high introduction note frequency and pronounced descent in frequencies across the loudsong, with a corresponding widening range but moderately large size, serrated bill and blue eye-ring but this certainly requires confirmation. This requires urgent attention, as the remnant population is very small and localised, recorded only from the Murici municipality, Alagoas. / Nós revisamos a taxonomia do complexo Trogon rufus sob o conceito Biológico de Espécies. Unidades taxonômicas possíveis, quebras e zonas de transição taxonômicas foram definidas usando mapas de calor e isofenas (linhas de contorno de fenótípo) baseados em caráteres de cor, barramento e morfometria. Esses possíveis táxons foram testados pelas análises de discriminantes. Dados de coloração foram obtidos por meio de espectrometria, e os padrões de barramento por meio de fotos de alta qualidade. Nós encontramos quatro espécies biológicas distintas Trogon chrysochloros Pelzeln 1856, da Mata Atlântica, diagnosticável pelo barramento mais escuro e denso na face inferior da cauda e coberteiras da asa, maior tamanho corpóreo, canto mais rápido, com mais notas e de frequência mais alta. Suas partes superiores (cabeça, dorso e cauda) variam de azul a verde acobreado com o aumento da altitude. O bico também é relativamente menor e mais serrilhado, o que está ligado a uma dieta que consiste quase exclusivamente de grandes artrópodes, fazendo desta espécie o Trogon mais insetívoro do mundo, o que deve ser a razão de sua relativa raridade quando comparado a outros Trogonidae com os quais vive em simpatia. Trogon tenellus Cabanis 1862, da América Central, e Trogon cupreicauda Chapman 1914, do Chocó-Magdalena, formam um caso clássico de espécies biológicas, entrando em contato no extremo noroeste da província de Chocó, na Colômbia, sem a presença de formas intermediárias. T. tenellus é identificável pela face superior da cauda azul ou azul esverdeado, anel perioftálmico azul ou cinza, tarso cinza, e voz com de 2 a 4 notas, maior duração das notas e maior mudança entre frequência de pico e frequência alta entre a nota introdutória e nota principal. Essas características contrastam com a cor verde-oliva brilhante da face superior da cauda, anel perioftámico amarelo, tarso geralmente oliva, presença de marrom claro na face inferior da cauda das fêmeas e canto com entre 6 e 8 notas, de menor duração, pouca mudança na frequência entre a primeira nota e o canto principal de T. cupreicauda. Este varia clinalmente de azul para uma plumagem mais verde acobreada e de barramento preto mais espesso para mais fino em uma gradiente da costa do Pacífico, do noroeste do Equador até o Vale do Magdalena. A maior distinção de estados de caráter relativa a T. tenellus na região onde os dois grupos se encontram provê uma possível evidência de deslocamento de caracteres como resultado de exclusão competitiva entre estas duas espécies, mantendo suas distribuições parapátricas. A população amazônica pertence a única espécie biológica Trogon rufus Gmelin 1788, mas com duas formas altamente distintas que designamos como Trogon rufus rufus do Escudo Guianense, e Trogon rufus sulphureus no sul e oeste da Amazônia, com a qual amazonicus de Todd é sinonimizada. Estas são morfologicamente, e em menor escala, vocalmente distintas nas duas margens do baixo Rio Negro e áreas abertas e/ou montanhosas da bacia do Rio Branco, mas apresentam troca de caracteres limitada nas longitudes entre 52 e 58 Oeste na margem sul do Rio Amazonas, centrado nos arredores do Rio Arapiuns, na margem esquerda da foz do Tapajós. Nós postulamos que isto é um resultado de contato secundário, como consequência de mudanças no curso principal do Rio Amazonas em tempos de níveis mais baixos do mar durante o Plio-Pleistoceno. T. r. sulphureus é identificado pela coloração tipicamente acobreada da face superior da cauda com uma banda sub-terminal de tonalidade esverdeada, anel perioftálmico amarelo, barras negras espessas e de baixa densidade na face inferior da cauda e nas coberteiras das asas e pela ausência de uma faixa peitoral. Em certos casos eles também podem ser distinguíveis pela voz com uma nota introdutória de maior frequência e/ou uma modulação descendente mais pronunciada no canto principal. Este grupo varia clinalmente em gradiente de oeste para leste, de uma coloração cobre forte até verde oliva na face superior da cauda com faixa sub-terminal mais ou menos distinta, anel perioftálmico azul, presença ou ausência de uma faixa peitoral branca e barramento na face inferior da cauda e coberteiras de menor espessura e, consequentemente, maior densidade. Nossos dados apontam que estes caracteres mudam linearmente entre sulphureus e rufus de acordo com a distância ao longo de uma gradiente clinal em forma de laço do Oeste Amazônico até o Escudo das Guianas, conectado pela zona de contato de Arapiuns, sugerindo isolamento por distância. Isto sugere um remanescente de um padrão de espécie em anel. Além disso, dois espécimes com possível mistura de caracteres foram de fato encontrados no alto Rio Negro e Pantepui, onde se espera que T. r. rufus e T. r. sulphureus entrem em contato, fechando o anel efetivamente. Ainda é necessário testar se Trogon rufus constitui uma espécie em anel válida, preferencialmente usando dados moleculares, mas este caso ilustra claramente que a distinção entre variação clinal e espécie em anel é uma questão de grau e não de tipo, com a formação de espécie em anel necessariamente passando por um estágio clinal sem sobreposição entre os grupos terminais. Nós assim sugerimos o conceito de espécie em laço, onde as formas terminais não se sobrepõem, mas são ligadas através de uma série de populações onde há fluxo gênico. Parece provável que estes padrões são mais amplamente distribuídos na Amazônia do que é sabido atualmente devido a uma propensão à variação clinal e especiação parapátrica causada por sua grande extensão geográfica e abundância de barreiras geográficas semipermeáveis. Em relação a população do Centro de Endemismo Pernambuco, os poucos registros sugerem que esta é uma unidade taxonômica válida. Este grupo apresenta uma combinação única de canto muito similar ao de T. r. sulphureus, devido à alta frequência da nota introdutória e pronunciada modulação descendente ao longo do canto principal, com uma maior amplitude da frequência, combinada a maior tamanho, bico serrilhado, e anel perioftálmico azul, mas isso requer confirmação. Este caso demanda atenção urgente, já que a população remanescente é muito pequena e de distribuição restrita ao município de Murici, em Alagoas.