Tesi sul tema "Skinks Phylogeny Molecular aspects"

This thesis concerns the molecular phylogenetics of three tribes of the family Bovidae, the Antilopini, Neotragini, and Tragelaphini. None of these tribes have been studied extensively with molecular techniques. The tribe Antilopini is one of the most speciose tribes (it includes 6 genera with 20 species) and the classification of several species of the genus Gazella is not clear. The tribe Neotragini is thought to be paraphyletic. Mitochondrial sequences of the cytochrome c oxidase ill and cytochrome b genes totalling 1083 base pairs have been determined for 52 taxa and used to determine phylogenetic relationships using cladistic and distance methods. Karyological analysis identified polymorphisms in several species (especially in Gazella saudiya and G. subgutturosa). Karyotypes of G. dorcas pelzelni and an XXY karyotype of a G. dorcas individual are shown for the first time. The main conclusions are that the Antilopini and the Tragelaphini are monophyletic and that the tribe Neotragini is paraphyletic. There is a lack of phylogenetic resolution between tribes which is probably due to the rapid radiation of the different tribes about 20 million years ago. The genus Taurotragus in the tribe Tragelaphini is shown to be paraphyletic and it would be appropriate to incorporate these taxa in the genus Tragelaphus. The genus Gazella could be paraphyletic, due to the position of Antilope cervicapra, in which case the genus needs to be split into two genera or renamed as Antilope. It is also argued that the use of the subgenus Trachelocele should be discontinued and that its only species, G. subgutturosa should be included in the subgenus Gazella. G. rufifrons and G. thomsonii may be more appropriately considered as conspecific. Cytogenetic and sequence data reveal that the herd of G. saudiya in Al Areen Wildlife Park is hybridised with G. bennettii and it is argued that it is important to identify unhybridised G. saudiya in other collections, since this species is on the brink of extinction. This case study demonstrates the need to genetically screen individuals which are part of a captive breeding program, especially if they are intended for reintroduction into the wild.

Thesis (MSc)--Stellenbosch University, 2003.
ENGLISH ABSTRACT: The evolutionary history of the African tribe Tragelaphini (spiral-horn antelope) is controversial. Past phylogenetic relationships among species were based on morphology or limited fossil evidence and are in conflict with mitochondrial DNA sequencing studies that have been conducted more recently. Although the group is distinguished from other African ungulates by the presence of spirally-twisted horns, the nine recognized extant species differ considerably in morphology, feeding habits and their habitat preference. The present study aims to resolve the phylogenetic uncertainties of the Tragelaphini using nuclear DNA sequence data derived from four independent DNA loci (MGF, PRKCl, SPTBN and THY). These data were combined with all previously published DNA sequences to produce a molecular supermatrix comprising approximately 6000 characters. Both parsimony and model based phylogenetic analyses of the nuclear DNA support the associations resulting from the analysis of mitochondrial genes. These findings suggest that the morphological characters previously used to delimit species within the group are subject to convergent evolution. The molecular phylogeny presented herein suggests that early members of Tragelaphini diverged from the other bovids during the mid-Miocene approximately 15.7 million years before present (MYBP). The common nyala (Tragelaphus enqest; and lesser kudu (Tragelaphus imberbis) representing the most basal species, separated from the other tragelaphids approximately 7.1 MYBP. This was subsequently followed by the radiation of those species adapted to a more tropical environment and they include the mountain nyala (Tragelaphus buxtom), bongo (Tragelaphus euryceros), sitatunga (Tragelaphus spekel) and bushbuck (Tragelaphus scriptus), and the arid adapted clade comprising the giant eland (Taurotragus derbianus), common eland (Taurotragus oryx) and greater kudu (Tragelaphus strepsiceros). It is thought that this split occurred at the Miocene-Pliocene boundary approximately 5.4 MYBP. The timing of evolutionary events within the tribe suggests climatic oscillations and subsequent biotic shifts as the major driving forces underpinning speciation in the tribe Tragalaphini.
AFRIKAANSE OPSOMMING; Die evolusionêre geskiedenis van die ras Tragelaphini (spiraalhoringwildsbokke) is kontroversieël. Vorige filogenetiese verwantskappe tussen die spesies is gebaseer op morfologie of beperkte fossiel bewyse. Meer onlangse studies, gebaseer op mitochondriale ONS nukleotieddata, is in teenstryding met baie van die evolusionêre hypotese afkomstig van morfologiese studies. Alhoewel die groep van die ander hoefdiere uitgeken kan word deur die aanwesigheid van spiraalvormige horings, verskil die nege hedendaagse spesies grootliks ten opsigte van morfologie, voedingswyse en habitat. Die hoof doelwit van hierdie studie was om die filogenetise verwantskappe tussen die Tragelaphini spesies te ontleed deur gebruik te maak van nukluêre ONS nukleotieddata afkomstig van vier onafhanklike ONS merkers (MGF, PRKCl, SPTBN en THY). Die data verkry is saamgevoeg by vorige gepubliseerde ONS nukleotidedata om 'n "supermatris" van sowat 6000 karakters te produseer. Parsimonie en modelgebaseerde filogenetise analise van die nukluêre ONS nukleotieddata het ooreengestem met die resultate van vorige mitochondriale studies. Hierdie bevindings dui daarop dat die morfologiese karakters wat voorheen gebruik is om die evolusionêre verwantskappe tussen die Tragelaphini spesies te ontleed onderhewig is aan konvergente evolusie. Die molekulêre filogenie wat hierin beskryf word stel voor dat die ras Tragelaphini gedurende die mid- Miocene, omtrent 15.7 miljoen jaar (MJ) gelede van die ander lede van die subfamilie Bovinae geskei het. Tragelaphus angasi en Tragelaphus imberbis, die mees basale spesies in die filogenie, het omtrent 7.1 MJ gelede van die ander lede van die Tragelaphini geskei. Hierdie skeiding is gevolg deur 'n split tussen die spesies aangepas vir 'n meer tropiese habitat (Tragelaphus buxtoni, Tragelaphus euryceros, Tragelaphus spekei en Tragelaphus scriptus) en die spesies aangepas vir 'n droë habitat (Taurotragus derbianus, Taurotragus oryx en Tragelaphus strepsiceros) Hierdie finale skeiding het gedurende die Miocene-Pliocene oorgang plaasgevind. Die tydsberekening van die evolusionêre gebeurtenisse wat binne die Tragelaphini ras plaasgevind het, gekoppel aan paleoklimaatdata, dui aan dat veranderinge in klimaat en die geassosieerde habitatveranderinge verantwoordelik was vir die spesiasie patroon wat ons vandag in die Tragelaphini ras waarneem.

Thesis (MSc)--University of Stellenbosch, 2006.
ENGLISH ABSTRACT: The present study aims to determine the phylogenetic relationships among the sand lizards, Pedioplanis. In addition, a single mitochondrial gene is used to investigate the geographic genetic structure of the widey distributed P. burchelli. With 11 species, Pedioplanis is the most speciose genus among the southern African genera of the family Lacertidae. All the species are restricted to the subcontinent with the exception of three (P. namaquensis, P. undata and P. benguellensis), which extend their range northwards into Angola. A total of 2200 nucleotide positions derived from two mitochondrial markers (ND2 and 16S rRNA) and one nuclear gene (RAG-1) are used to determine the phylogenetic relationships among ten of the eleven Pedioplanis species. The first well resolved gene tree for the genus, drawn from 100 individuals, is presented and this is largely congruent with a phylogeny derived from morphology. Contrary to some previous suggestions, Pedioplanis forms a monophyletic assemblage with Heliobolus and Nucras. The genus Pedioplanis is monophyletic with P. burchelli/P. laticeps forming a sister clade to all the remaining congeners. Two distinct geographic lineages can be identified within the widespread P. namaquensis; one occurs in Namibia, while the other occurs in South Africa. The “P. undata” species complex is monophyletic, but one of its constituent species, P. inornata, is paraphyletic. Relationships among the subspecies of P. lineoocellata are much more complex than previously documented. An isolated population previously assigned to P. l. pulchella is paraphyletic and sister to the three named subspecies. The phylogeny identifies two biogeographical groupings that probably diverged during the mid-Miocene. The development of the Benguella Current could have initiated isolation mechanisms associated with changes in habitat that could have generated barriers and played a role in the evolution of this group. At the lower taxonomic level, the mtDNA phylogeographic structure of the wide spread P. burchelli in South Africa reveal at least six distinct clades that are geographically partitioned. The first one is restricted to the eastern mountains along the Great Escarpment (GE). The next three are found along the Cape Fold Mountains (CFM): the north-west CFM, central CFM and eastern CFM. The fifth one shares samples from central CFM and GE. The last clade is restricted to the eastern central mountains of the GE. These six geographic groupings are genetically divergent from each other and they started separating in the early Pliocene period. Phylogeographic studies on other taxa in the region have found different levels of genetic structuring among or within taxa. The fact that P. burchelli is restricted to high altitude areas could have resulted in limited dispersal and consequently contributed to its geographic structure. However, the exact cause of the pattern obtained is not readily apparent. Habitat fragmentation in the past is probably one of the most influential factors shaping the genetic distribution of the species across South Africa. The inclusion of nuclear markers will shed more light on the evolutionary history of P. burchelli in South Africa.
AFRIKAANSE OPSOMMING: Die huidige studie stel ten doel om ‘n filogenie daar te stel vir die Sand akkedisse, Pedioplanis. ‘n Enkele mitochondriale geen is ook gebruik om die geografiese genetiese struktuur van die wydverspreide P. burchelli vas te stel. Met 11 spesies is Pedioplanis die mees spesieryke genus onder die suidelike Afrika genera wat aan die Lacertidae familie behoort. Al die spesies is beperk tot die subkontinent met die uitsondering van drie (P. namaquensis, P. undata en P. benguellensis), wat ‘n uitgebreide verspreiding het noordwaarts tot in Angola. ‘n Totaal van 2200 nukleotied posisies wat afkomstig is van twee mitochondriale merkers (ND2 en 16S rRNA) en een nukluêre geen (RAG-1) is gebruik om die filogenetiese verwantskappe tussen 10 van die 11 Pedioplanis spesies vas te stel. Die eerste goed geondersteunde geen boom vir die genus, gebasseer op 100 individue, is verkry en dit is meestal ooreenstemmend met ‘n filogenie gebasseer op morfologie. In teenstelling met sekere voorstelle van die verlede vorm Pedioplanis ‘n monofiletiese groep tesame met Heliobolus en Nucras. Die genus Pedioplanis is monofileties met P. burchelli/P. laticeps wat ‘n suster groep vorm van al die oorblywende lede van die genus. Twee herkenbare geografiese lyne kan geidentifiseer word in die wydverspreide P. namaquensis; een kom in Namibia voor, terwyl die ander een in Suid Afrika voorkom. Die “P. undata” spesies kompleks is monofileties, maar een van die spesies wat deel uitmaak van die groep, P. inornata, is parafileties. Verwantskappe tussen die subspesies van P. lineoocellata is meer kompleks as wat aanvanklik aanvaar is. ‘n Geisoleerde bevolkimg wat voorheen toegesê is aan P. l. pulchella is parafileties en verteenwoordig ‘n suster groep van die benaamde subspesies. Die filogenie identifiseer twee biogeografiese groeperings wat moontlik gedivergeer het gedurende die middel-Miocene. Die ontwikkeling van die Benguella stroom het dalk versperrings geinisiëer as gevolg van die gesamentlike veranderinge in habitat wat dalk ook ‘n rol gespeel het in die evolusie van die groep. Op die laer taksonomiese vlak het die mtDNA filogeografiese struktuur van die wydverspreide P. burchelli in Suid Afrika ten minste ses groepe aangetoon wat geografies van mekaar geskei is. Die eerste een is beperk tot die oostelike berge wat aan die Groot Eskarpement (GE) behoort. Die volgende drie word gevind in die Kaapse Vouberge (KVB): die noord-westelike KVB, sentrale KVB en oostelike KVB. Die vyfde een deel eksemplare van beide die GE en die KVB. Die laaste groep is beperk tot die oostelike en sentrale berge van die GE. Hierdie ses geografiese groepe is geneties geskei van mekaar en hulle het begin om apart te ontwikkel gedurende die vroë Pliocene periode. Ander filogeografiese studies in die area het verskillende vlakke van genetiese struktuur vertoon tussen en binne taksa. Die feit dat P. burchelli beperk is tot hoogliggende dele kon moontlik bygedrae het tot die geografiese struktuur. Die presiese oorsaak van die patroon wat verkry is, is nie ooglopend nie. Habitat fragmentasie in die verlede is moontlik een van die mees invloedrykste faktore wat die genetiese verspreiding van die spesie in Suid Afrika beinvloed het. Die insluiting van nukluêre merkers sal meer lig warp op die evolusionêre geskiedenis van P. burchelli in Suid Afrika.

Thesis (DPhil)--University of Stellenbosch, 2008.
ENGLISH ABSTRACT: Sub-Antarctic islands represent the only mid to high latitude terrestrial biomes in the Southern Hemisphere. These islands have various geological origins and histories, well-preserved terrestrial ecosystems and high levels of species endemism. In an attempt to understand the evolution and biogeography of terrestrial taxa in the South Polar Region, the first broad-scale molecular phylogeny was constructed for the unique terrestrial group, the ameronothroid mites (genus Halozetes (Oribatida)), collected from sub-Antarctic and Maritime Antarctic localities. Phylogenetic analyses based on a combined mitochondrial (cytochrome oxidase subunit I (COI)) and nuclear (histone-3 (H3)) sequence dataset indicated that the evolution of these mites were habitat specific (i.e. intertidal, supralittoral and terrestrial). Notwithstanding criticisms levelled against a molecular clock, the mites were evolutionary young (<10myo), contrary to their status as an ancient group predating Gondwana fragmentation. Biogeographic analyses indicated a complex pattern mainly sculpted by multiple independent dispersal events across the Antarctic Polar Frontal Zone similar to previous findings for other marine and terrestrial taxa. Also, the molecular phylogeny displayed considerable discourse with contemporary taxonomy suggesting the need for taxonomic revisions and reassessment of morphological characters. Sub-Antarctic Marion Island, the larger of the two islands comprising the Prince Edward Island archipelago (PEI), has experienced extensive glaciation and volcanism. To assess the impact of historical events (volcanism (including recent eruptions) and glaciation) and contemporary mechanisms (gene flow) on the genetic spatial distribution of species from Marion Island, two mite species namely Eupodes minutus (Prostigmata) and Halozetes fulvus (Oribatida) as well as a single plant species, Azorella selago (Apiaceae), were selected as model organisms. For independent phylogeographic analyses, mitochondrial sequence data (COI) were obtained for both mite species, while chloroplast sequence (trnH-psbA) and amplified fragment length polymorphism (AFLP) data were generated for the cushion plant, A. selago. Since A. selago is typified by two growth forms namely discrete cushions and continuous mats, it was essential to examine the growth dynamics prior to phylogeographic analyses. The sequence and fragment data indicated that both mite and plant species were significantly substructured across Marion Island. Manual comparisons indicated unique populations on the western (Kaalkoppie for H. fulvus, La Grange Kop for E. minutus and Mixed Pickle for A. selago), eastern (Bullard Beach for H. fulvus and Kildalkey Bay for E. minutus), northern (Middelman and Long Ridge for H. fulvus) and southern side (Grey Headed for H. fulvus and Watertunnel for A. selago) of the island. Importantly, the western side had unique localities for all species. Interestingly, based on the H. fulvus data, the western populations were relatively young, characterized by high migration rates, small effective (female) population sizes with no isolation-by-distance. The opposite scenario was found for the eastern populations. This spatial genetic structure described for species on Marion Island can be ascribed to both historical events and environmental conditions. These areas with their unique genetic composition are of special conservational concern; consequently this research will contribute to an active management plan for PEI, South Africa’s only Special Nature Reserve.
AFRIKAANSE OPSOMMING: Sub-Antarktiese eilande verteenwoordig die enigste terrestriële bioom in die middel tot hoër breedtegrades van die Suidelike Halfrond. Hierdie eilande besit ‘n verskeidenheid van geologiese oorspronge en geskiedenisse, goed-bewaarde terrestriële ekosisteme en hoë vlakke van endemisme. In ‘n poging om die evolusie en biogeografie van terrestriële taksa in die Suid Pool Area te verstaan, is die eerste grootskaalse molekulêre filogenie saamgestel vir ‘n unieke terrestriële groep, die ameronothoïed miete (genus Halozetes (Oribatida: Ameronothroidea)), vanaf menigte sub-Antarktiese en Maritime Antarktiese lokaliteite. Filogenetiese analises gebaseer op die saamgestelde mitochondriale (sitokroom oksidase subeenheid I (COI)) en nukluêre (histoon-3 (H3)) basispaarvolgordes het aangedui dat die evolusie van hierdie miete habitat spesifiek is (m.a.w inter-gety, supralitoraal en terrestrieël). Ongeag die kritiek teenoor ‘n molekulêre klok, is hierdie miete evolusionêr jonk (<10mjo), wat teenstrydig is met hulle status as ‘n antieke groep wat terugdateer voor Gondwana fragmentasie. Biogeografiese analises het ‘n komplekse patroon aangedui wat grotendeels gekarakteriseer word deur menigte onafhanklike verspreidingsgebeurtenisse bo-oor die Antarktiese Polêre Frontale Zone, wat ooreenstemmend is met vorige bevindinge vir ander mariene en terrestriële taksa. Die molekulêre filogenie het ook aansienlik verskil van die tradisionele taksonomie, dus is taksonomiese aanpassings en herklassifisering van morfologiese karakters noodsaaklik. Sub-Antarktiese Marion Eiland, die groter eiland van die Prins Edward eilandgroep (PEI), het uitermate glasiasie en vulkanisme ondervind. Om die impak van historiese gebeurtenisse (vulkanisme (insluitend onlangse uitbarstings) en glasiasie) en kontemporêre meganismes (geenvloei) op die genetiesgespasieërde verspreiding van spesies vanaf Marion Eiland te bepaal, was twee mietspesies naamlik Eupodes minutus (Prostigmata) en Halozetes fulvus (Oribatida) asook ‘n enkele plantspesie, Azorella selago (Apiaceae), gekies as model organismes. Vir onafhanklike filogeografiese analises, was die mitochondriale basispaarvolgorde (COI) vir beide mietspesies bepaal, terwyl chloroplast basispaarvolgorde (trnH-psbA) asook geamplifiseerde fragmentlengte polimorfisme (AFLP) data gegenereer was vir die kussingplant, A. selago. Aangesien A. selago gekenmerk word deur twee groeivorme, naamlik diskrete kussings en aaneenlopende matte, was dit noodsaaklik om eers die groeidinamika van die plant te ondersoek alvorens ‘n filogeografiese studie kon geskied. Die basispaarvolgordebepalings en fragmentdata het aangedui dat beide mietspesies sowel as die plantspesie betekenisvolle substruktuur vertoon regoor Marion Eiland. Informele vergelykings het unieke populasies aangedui op die westelike (Kaalkoppie vir H. fulvus, La Grange Kop vir E. minutus en Mixed Pickle vir A. selago), oostelike (Bullardstrand vir H. fulvus en Kildalkeybaai vir E. minutus), noordelike (Middelman en Long Ridge vir H. fulvus) en suidelike kant (Grey Headed vir H. fulvus en Watertunnel vir A. selago) van die eiland. Die westelike kant besit dus unieke lokaliteite vir al die spesies. Interressantheidhalwe het die H. fulvus data getoon dat die westelike populasies relatief jonk is en gekarakteriseer word deur hoë migrasiesyfers en klein effektiewe (vroulike) populasiegroottes met geen isolasie-oor-afstand nie. Die resultate vir die populasies aan die oostelike kant van die Marion Eiland was presies teenoorgesteld. Dié beskryfde substruktuur vir die spesies op Marion Eiland is afkomstig van beide historiese gebeurtenisse asook omgewingstoestande. Hierdie areas met hul unieke genetiese samestelling, is belangrik vir natuurbewaring. Hierdie navorsing sal bydra tot die bestuursriglyne van PEI, Suid Afrika se enigste Spesiale Natuurreservaat.

Orientador: Renato Goldenberg
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Miconia é um é um dos maiores gêneros de angiospermas. Tradicionalmente, tem sido considerado complexo e mal circumscrito, assim como outros gêneros aparentados. Estudos filogenéticos recentes, baseados em marcadores moleculares, têm atestado a natureza polifilética de Miconia e suas onze seções. Neste trabalho, é apresentado um estudo filogenético-molecular aprofundado de um dos grupos naturais encontrados em estudos anteriores, o "subclado Miconia discolor", do "clado Miconia IV". Para tanto, foram utilizados sequências de quatro marcadores plastidias (psaI-accD; psnK-L; atpF-H; trnS-G) e dois nucleares (ITS e ETS). Os resultados obtidos permitiram esclarecer o relacionamento interno e externo do grupo. Por meio da otimização de caracteres morfológicos e biogeográficos, são sugeridas hipóteses evolutivas, como a evolução de inflorescências glomeruladas e pseudantos a partir de inflorescências com ramos escorpióides amplos, e a irradiação da diversidade na Floresta Atlântica a partir de poucas colonizações. Os resultados também permitiram a primeira proposição moderna de classificação ingra-genérica para Miconia, através da formalização e circunscrição de novos táxons: Miconia sect. Discolor, e subseções Albicans, Chrysophylla, Discolor e Multispicata, subordinadas à nova seção. Adicionalmente, uma revisão taxonômica é apresentada para a recém proposta subseção Discolor, incluindo descrições, ilustrações, imagens de Microscopia Eletrônica de Varredura, comentários, e uma chave dicotômica de identificação para as espécies. A subseção é composta 32 espécies, das quais três são novas. Trinta e dois nomes heterotípicos foram sinonimizados, e 39 lectotipificações foram propostas. A subseção pode ser reconhecida por apresentar as folhas densamente cobertas por tricomas ramificados na face abaxial (nunca formando um indumento aracnóide), inflorescências geralmente glomeruladas, raramente curtamente scorpióides, lobos do cálice caducos no fruto, e uma distribuição restrita ao sudeste da América do Sul, ocorrendo na Floresta Atlântica e no Cerrado
Abstract: Miconia is one of the biggest genera in angiosperms. It has been traditionally considered very complex and badly circumscribed, and other close genera as well. Phylogenetic studies based on molecular markers have been confirming the polyphily of Miconia and its eleven sections. Here I present a deeper molecular and phylogenetic study of a natural group found out in older studies, the "Miconia discolor subclade", from the "Miconia IV clade". I used sequences of four plastid (psaI-accD; psnK-L; atpF-H; trnS-G) and two nuclear markers (ITS and ETS). The results clarified the inner and outer relationship of that group. Evolutionary hypotheses are suggested by the optimization of morphological and biogeographical charcaters. For example, the evolution of glomerulate inflorescences and pseudanthia from inflorescences of wide, scorpioid branches, and the diversification of the genus in the Atlantic Forest from few colonizations. Additionally, the results allowed the first modern infra-generic rearrangement by the proposition and circumscription of new taxa: Miconia sect. Discolor, with the subsections Albicans, Chrysophylla, Discolor and Multispicata. A revision of the new Miconia subsect. Discolor is presented, with descriptions, illustrations, Scanning Electron Microscopy images, comments, and a taxonomic key for species. The subsection has 32 species and three of them are new. I synonymized 32 heterotypic names proposed 39 lectotipifications. The subsection can be recognized by the abaxial leaf surface usually densely covered by branched trichomes (never forming a cobweb indument), inflorescences usually glomerulate, rarely shortly scorpioid, calyx lobes caducous in fruit, and a distribution restricted to the southeast of South America, in the Atlantic Forest and Cerrado
Doutorado
Biologia Vegetal
Doutora em Biologia Vegetal

Both insect parasitic/entomopathogenic nematodes and plant parasitic nematodes are of great economic importance. Insect parasitic/entomopathogenic nematodes provide an environmentally safe and effective method to control numerous insect pests worldwide. Alternatively, plant parasitic nematodes cause billions of dollars in crop loss worldwide. Because of these impacts, it is important to understand how these nematodes evolve, and, in the case of entomopathogenic nematodes, how their bacterial symbionts evolve. This dissertation contains six chapters. Chapter one is a review of DNA markers and their use in the phylogenetic systematics of entomopathogenic and insect-parasitic nematodes as well as a review of phylogenetic, co-phylogenetic, and population genetic methodologies. Chapter two characterizes positive destabilizing selection on the luxA gene of bioluminescent bacteria. Our data suggests that bacterial ecology and environmental osmolarity are likely driving the evolution of the luxA gene in bioluminescent bacteria. Chapter 3 examines relationships among bacteria within the genus Photorhabdus. Our analyses produced the most robust phylogenetic hypothesis to date for the genus Photorhabdus. Additionally, we show that glnA is particularly useful in resolving specific and intra-specific relationships poorly resolved in other studies. We conclude that P. asymbiotica is the sister group to P. luminescens and that the new strains HIT and JUN should be given a new group designation within P. asymbiotica. Chapter 4 characterizes the morphology of the head and feeding apparatus of fungal feeding and insect infective female morphs of the nematode Deladenus siricidicola using scanning electron microscopy. Results showed dramatic differences in head, face, and stylet morphology between the two D. siricidicola female morphs that were not detected in previous studies using only light microscopy. Chapter five utilizes comparative transciptomics to identify putative plant and insect parasitism genes in the nematode Deladenus siricidicola. Results from this study provide the first transcriptomic characterization for the nematode Deladenus siricidicola and for an insect parasitic member of the nematode infraorder Tylenchomorpha. Additionally, numerous plant parasitism gene homologues were discovered in both D. siricidicola libraries suggesting that this nematode has co-opted these plant parasitism genes for other functions. Chapter six utilizes a phylogenomic approach to estimate the phylogeny of the nematode infraorder Tylenchomorpha.

Orientador: Shirlei Maira Recco Pimentel
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Dendropsophus minutus está amplamente distribuído ao Leste dos Andes, na América do Sul e possui uma grande diversidade acústica e morfológica, o que sugere que possa haver mais de uma espécie sob esse nome. Sua coloração dorsal pode ser classificada em dois padrões principais, hourglass e bivittata. No presente estudo, 14 parâmetros morfométricos e seqüências de DNA com 357 pares de bases do gene citocromo b mitocondrial foram analisados objetivando um melhor entendimento acerca da variação de D. minutus no Brasil. Tanto os resultados moleculares quanto os fenotípicos revelaram a presença de uma alta estruturação da diversidade dessa espécie, mostrando que a divergência entre populações é, geralmente, proporcional à distância geográfica, exceto no estado de São Paulo, sudeste do Brasil. Nessa região, a Serra do Mar está aparentemente agindo como uma barreira geográfica para o fluxo gênico, isolando duas linhagens. A primeira, formada pelas populações da Mata Atlântica, tem padrão hourglass de coloração dorsal. A segunda, do interior de São Paulo, assim como a população do Rio Grande do Sul, possui padrão bivittata de coloração dorsal. Esses resultados corroboram a hipótese de que o táxon D. minutus contém duas linhagens crípticas. Apesar disso, uma amostragem maior se faz necessária, bem como um melhor estudo de caracteres para defini-las como espécies ou não.
Abstract: In despite of its complex reproductive behavior, Dendropsophus minutus has a large distribution at East of Andes, South America and show high acoustic and morphologic diversity, suggesting that possibly more than one species may exist under this name. Its dorsum coloration has basically two main patterns, hourglass or bivittata. Here, 14 morphometric parameters and partial mitochondrial cytochrome b gene sequences (357 base pairs) were analyzed aiming to understand more about Brazilian D. minutus variation. Both molecular and morphologic results agree with a high structuration of this species diversity, showing population divergence generally proportional to their geographic distance, except in São Paulo State, southeastern Brazil. At this region, Serra do Mar high mountains are apparently acting as a barrier for dispersion, isolating two lineages. The first of them, formed by populations from Atlantic Rainforest domain, has an hourglass dorsum pattern, whereas the second, comprising inner São Paulo State populations gathered with D. minutus from Rio Grande do Sul (South Brazil), shows bivittata dorsum coloration pattern. These results corroborate the hypothesis that D. minutus could comprise more than one species, revealing two cryptic lineages. However, these lineages should not be defined as different species before sampling enlargement to the present study.
Mestrado
Biologia Celular
Mestre em Biologia Celular e Estrutural

Orientadores: Irani Quagio Grassiotto, Daniela Calcagnotto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A superfamília Erythrinoidea é uma das oito famílias que compõem a ordem Characiformes e foi proposta como um grupo monofilético para abrigar as famílias neotropicais Ctenoluciidae, Erythrinidae e Lebiasinidae, e a família africana Hepsetidae. Após a proposição da superfamília, estudos filogenéticos recentes com base em caracteres morfológicos, como osteologia e morfologia de partes moles, e caracteres moleculares tem refutado a ideia de monofiletismo do grupo. Além disso, diferentes autores divergem quanto às relações suprafamiliares de Erythrinoidea. Além dos caracteres tradicionais, o emprego dos caracteres oriundos da análise da ultraestrutura da ontogenia dos espermatozoides e sua forma final tem sido uma importante fonte na complementação dos estudos filogenéticos. O estudo apresentado teve como objetivo realizar uma análise filogenética para testar a monofiletismo de Erythrinoidea e verificar os padrões de relacionamento suprafamiliares em Characiformes, e testar o posicionamento filogenético da família Crenuchidae, supostamente relacionada à superfamília. As análises filogenéticas foram realizadas com base em dados moleculares e dados morfológicos independentemente, e de maneira conjunta (evidência total) através do método de Parcimônia. A ultraestrutura da espermiogênese e dos espermatozoides dos gêneros incluídos em Erythrinoidea, mais a família Crenuchidae e outras possivelmente relacionadas, foram descritos e codificados em uma matriz de dados. Nas três análises realizadas a superfamília Erythrinoidea não é recuperada como monofilética. A família Crenuchidae posiciona-se na base da subordem Characoidei e não está estreitamente relacionada à Erythrinoidea. As análises com dados moleculares e de evidência total apresentaram uma topologia muito semelhante ao nível mais inclusivo e os valores de suporte dos principais clados apresentaram-se maiores na evidência total, mostrando a influência positiva da inserção dos caracteres da morfologia das células espermáticas nas análises filogenéticas
Abstract: The superfamily Erythrinoidea is one of eight superfamilies within the order Characiformes and was proposed to include a monophyletic group composed of the Neotropical fish families Ctenoluciidae, Erythrinidae and Lebiasinidae, and African family Hepsetidae. Current phylogenetic studies based on morphological data, such as osteology and morphology of soft structures, and molecular data have refuted the hypothesis of monophyly of this group. In addition, the suprafamilial relationships in Erythrinoidea have diverged among the authors. In combination with more traditional characters, the analyses of the ultrastructure of sperm ontogeny and its final shape have become an interesting source of characters to complement phylogenetic studies. The goal of this study was to perform a phylogenetic analysis to test the monophyly of the Erythrinoidea and to assess the phylogenetic position of the family Crenuchidae, a group supposed to be closely related to the superfamily. The phylogenetic analyses were performed using molecular and morphological data separately, and in a combined approach (total evidence) through the Parsimony method. The ultrastructure of the spermiogenesis and the sperm shape of Erythrinoidea genera, plus the family Crenuchidae and other species possibly related to the superfamily were described and coded in a matrix data. In all three analyses Erythrinoidea was not recovered as monophyletic. The family Crenuchidae was placed in a basal position within the suborder Characoidei in molecular and total evidence analysis, and was not closely related to the Erythrinoidea. The analyses using only molecular data and total evidence showed a very similar topology for the more inclusive relationship levels. The support values were higher for the total evidence tree when compared to the one resulting of only molecular data possibly pointing to a positive influence of the combination of the spermatic cell data in phylogenetic analyses
Doutorado
Biologia Celular
Doutor em Biologia Celular e Estrutural

Cette thèse a pour but de contribuer à une meilleure compréhension de la tuberculose (TB) et desmycobactéries non-tuberculeuses dans la Caraïbe. La tuberculose a hanté l’humanité depuisplusieurs millénaires et reste de nos jours une des maladies infectieuses faisant le plus de victimeschaque année (1,5 millions de décès en 2013). La connaissance de l’épidémiologie de la tuberculoseest essentielle afin de concevoir des programmes de lutte anti-TB adaptés aux spécificitésrégionales et donc plus efficaces. Dans cette optique, la première partie de ce travail fourni un suivià long-terme de la résistance aux antituberculeux observée en Guadeloupe, Martinique et Guyanefrançaise ainsi qu’un aperçu de la diversité génétique et de la résistance aux antituberculeux dansla Caraïbe. Les données montrent une baisse graduelle de la fréquence des infections causées pardes souches résistantes parmi les nouveaux cas de TB dans les départements français d’Amérique.En ce qui concerne la Caraïbe, des différences marquées ont été observées entre les différentsterritoires, ce qui semble refléter le passé historique de cette région.La deuxième partie est consacrée à la phylogénie et l’évolution de M. tuberculosis, étudié à l’aide dedivers marqueurs génétiques comme spoligotypes, LSP, SNP et MIRU-VNTR. Les profils MIRUVNTR(format 12-loci) ont été étudiés afin de déterminer leur utilité comme marqueurphylogénétique. Il a été montré que ce marqueur est adapté pour retracer la phylogénie ducomplexe M. tuberculosis et que la précision du classement basé sur les MIRUs est supérieure àcelle du classement basé sur les spoligotypes. De plus, la technique MIRU-VNTR permetégalement d’observer la diversification évolutive d’une souche de M. tuberculosis au cours del’infection ou alors d’identifier des patients infectés par plusieurs souches de M. tuberculosis enmême temps. Les deux phénomènes ont été observés au cours de ce travail de thèse et les casconcernés sont décrits dans ce deuxième chapitre.Enfin, un premier aperçu de la diversité des mycobactéries non-tuberculeuses isolées desprélèvements cliniques en Guadeloupe, Martinique et Guyane est présenté dans la troisième partiede ce travail. Des différences marquées dans la fréquence d’isolement de certaines espèces ont puêtre observées entre les trois départements français d’Amérique. M. intracellulare par exemple étaitsignificativement plus abondant en Guadeloupe. Cependant l’existence d’une niche écologiquespécifique à cette île n’a pas pu être mise en évidence. La problématique de l’identification desmycobactéries non-tuberculeuses est abordée également à travers une étude rétrospective del’utilisation de hsp65-PRA pour l’identification des mycobactéries dans un laboratoire de routinemais aussi sous forme d’un travail prospectif visant à la mise en place d’un protocoled’identification de mycobactéries non-tuberculeuses avec MALDI-TOF MS
This thesis aims at providing a better understanding of tuberculosis (TB) and non-tuberculousmycobacteria (NTM) in the Caribbean. TB is an ancient scourge of humanity and remains one ofthe deadliest infectious diseases today having claimed around 1.5 million lives in 2013.Understanding the epidemiology of TB is essential for optimizing regional TB control programs. Inthis context, the first part of this work provides long-term data on drug-resistance in Guadeloupe,Martinique and French Guiana as well as an insight in the genetic diversity and drug-resistance ofM. tuberculosis in twelve Caribbean territories. Encouragingly, the results show a gradual decreaseof drug-resistant TB in newly infected patients in Guadeloupe, Martinique and French Guiana. Onthe Caribbean level, distinct differences were observed from one territory to the next and thecurrent epidemiological landscape seems to reflect the historical past of the region.The second part addresses the phylogeny and evolution of M. tuberculosis using various geneticmarkers such as spoligotyping, large sequence polymorphism (LSP), single nucleotidepolymorphism (SNP), and MIRU-VNTRs. The suitability of 12-loci MIRU-VNTR profiles for use inphylogenetic studies was evaluated and it was found that this marker is not only able to resolvethe evolutionary relationships within the M. tuberculosis complex but also allows to achieve ahigher phylogenetic precision than spoligotyping. MIRU-VNTR also permits the identification ofon-going evolution in TB patients (in-patient microevolution) as well as mixed strain infections.Both phenomena were observed in our setting and the respective cases are described herein.Finally, a first insight in the diversity of NTM isolated from clinical specimen in Guadeloupe,Martinique and French Guiana is provided. The isolation frequency of some NTM species variedconsiderably between the three departments, the most striking example being the relativeabundance of M. intracellulare in Guadeloupe. However, no evidence of a privilegedenvironmental niche/infection source on this island could be found. Last but not least, the subjectof NTM identification is addressed in the form of a retrospective evaluation of hsp65-PRA basedidentification in a routine laboratory and in the form of a prospective study towards theimplementation of a MALDI-TOF MS based identification of NTM at the Pasteur Institute ofGuadeloupe

Un des buts principaux de la biologie évolutive et de la médecine moléculaire consiste à reconstruire les relations phylogénétiques entre organismes à partir de leurs séquences moléculaires. En littérature, cette question est connue sous le nom d’inférence phylogénétique et a d'importantes applications dans la recherche médicale et pharmaceutique, ainsi que dans l’immunologie, l’épidémiologie, et la dynamique des populations. L’accumulation récente de données de séquences d’ADN dans les bases de données publiques, ainsi que la facilité relative avec laquelle des données nouvelles peuvent être obtenues, rend l’inférence phylogénétique particulièrement difficile (l'inférence phylogénétique est un problème NP-Hard sous tous les critères d’optimalité connus), de telle manière que des nouveaux critères et des algorithmes efficaces doivent être développés. Cette thèse a pour but: (i) d’analyser les limites mathématiques et biologiques des critères utilisés en inférence phylogénétique, (ii) de développer de nouveaux algorithmes efficaces permettant d’analyser de plus grands jeux de données.
Doctorat en Sciences
info:eu-repo/semantics/nonPublished

Parmi les paramètres influençant l'inférence d'arbres phylogénétiques, nous nous sommes penchés d'une part sur (i) l'utilisation et l'efficacité de différents marqueurs et (ii) l'influence de la radiation évolutive (la succession rapide d'événements de spéciation) dans la construction d'arbres phylogénétiques et, d'autre part, sur l'applicabilité du modèle de substitution nucléotidique GTR (General Time Reversible).

La première partie de ce travail étudie l'évolution des cétacés en se basant sur les séquences des génomes mitochondriaux, sur le motif d'insertion de rétroposons SINEs (short interspersed elements) nouvellement isolés et les loci nucléaires de ces derniers. Le choix des cétacés est motivé par la présence, durant leur évolution, de radiations évolutives, qui sont propices au tri différentiel de lignées généalogiques: si des séquences de gènes ou des allèles restent polymorphes entre des événements de spéciations, il est possible, et même probable, d'observer une incompatibilité entre les histoires évolutives de ces marqueurs, malgré que celles-ci soient bien correctes. Nous abordons l'étude du tri différentiel des lignées généalogiques par le biais des SINEs, dont l'insertion aléatoire et irréversible confère à ces marqueurs un risque de convergence particulièrement faible.

Notre approche multi-marqueur nous permet de reconstruire un arbre robuste à partir duquel nous analysons ces différents marqueurs à l'aide des rapports signal/bruit (la qualité du contenu informatif du marqueur) et effort/signal (les efforts à mettre en oeuvre pour obtenir du signal phylogénétique). Nous discutons également les relations conflictuelles/incorrectes obtenues à partir des différents marqueurs, notamment des motifs d'insertion de SINEs pour lesquels nous décrivons un test objectif nous permettant de différencier le tri différentiel de lignées généalogiques et la convergence.

Les modèles de substitutions nucléotidiques sont à la base de nombreuses méthodes d'inférence phylogénétiques. Parmi ces modèles, le modèle GTR est un des plus complets et des plus utilisés. Waddell and Steel [1997] ont décrit une procédure qui permet d'estimer les distances et les taux instantanés de substitution pour des séquences évoluant selon les hypothèses du modèle GTR. Il existe néanmoins des conditions qui rendent cette procédure, et donc l'utilisation du modèle GTR, inapplicables.

Nous avons simulé l'évolution de séquences d'ADN le long de 12 arbres caractérisés par un ensemble de conditions biologiquement plausibles (différentes longueurs de branches, des conditions de (non-)homogénéité de la matrice de taux instantanés de substitution et différentes longueurs de séquences). Pour chaque ensemble de conditions, nous avons évalué (i) l'applicabilité du modèle GTR et (ii) la qualité des alignements obtenus à partir des données simulées.

Nos résultats indiquent que l'inapplicabilité de la procédure de Waddell and Steel [1997] peut effectivement être considérée comme un problème pratique car elle apparaît avant les difficultés d'alignement (étape nécessaire et préalable à toute inférence phylogénétique). La probabilité de cette inapplicabilité dépend du taux de substitution et de la taille des données.

Doctorat en sciences, Spécialisation biologie moléculaire
info:eu-repo/semantics/nonPublished

Orientadores: Ana Maria Lima de Azeredo-Espin, Nilson Ivo Tonin Zanchin
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A superfamília Oestroidea (Diptera:Brachycera:Calyptratae), com +13.000 espécies descritas, compreende um dos grupos mais numerosos e ecologicamente diversos da ordem Diptera. O grupo possui grande interesse para atividades humanas por englobar espécies de importância médica, veterinária e forense, muitas das quais compõem a família Calliphoridae. Apesar do grande número de estudos disponíveis, as relações evolutivas no grupo, o qual é composto predominantemente por linhagens de rápida diversificação e radiação, ainda são controversas e pouco compreendidas, encorajando a caracterização de novos marcadores moleculares para análises de filogenia molecular. Neste contexto, esta tese foi desenvolvida e organizada em três capítulos descrevendo estudos genéticoevolutivos em espécies da superfamília Oestroidea, com ênfase em Calliphoridae. O primeiro capítulo trata da caracterização e avaliação do segundo espaçador transcrito interno (ITS2) do DNA ribossomal como um marcador molecular para análises filogenéticas em Calliphoridae, incorporando informações tanto da sequência primária quanto da estrutura secundária adquirida pela região. A análise do ITS2 revelou um padrão hierarquicamente organizado das distâncias genéticas nos níveis de espécies, gêneros e subfamílias, enquanto pouca variação intra-específica foi encontrada. As árvores inferidas recuperaram muitas das relações comumente aceitas entre os táxons amostrados, sendo que a inclusão da informação estrutural nas análises resultou na recuperação de topologias mais confiáveis. Sendo assim, o potencial da região ITS2 como um marcador molecular para análises evolutivas na família Calliphoridae foi confirmado e seu uso em análises de maior escala, incluindo marcadores de diferentes naturezas de evolução, encorajado. O segundo capítulo da tese descreve a caracterização in vitro da estrutura secundária adquirida pelo ITS2, através de padrões de digestão enzimática e análise dos fragmentos gerados, em espécies representantes das três superfamílias de Calyptratae: Glossina morsitans, Musca domestica e Cochliomyia hominivorax. A análise do padrão de fragmentos gerados pelas enzimas RNAse I, A, T1 e V1, quando mapeados na estrutura secundária predita in silico, corroborou muitos dos domínios inicialmente preditos pelo método computacional, ressaltando a importância e confiabilidade desses métodos na predição de estruturas secundárias. O terceiro capítulo da tese descreve análises de filogenia molecular na superfamília Oestroidea, com ênfase na amostragem de espécies de Calliphoridae, utilizando quatro marcadores moleculares, dois nucleares (ITS2 e 28S) e dois mitocondriais (COI e 16). As análises, que incluíram uma extensa avaliação dos efeitos de diferentes estratégias de particionamento dos dados em análises de inferência Bayesiana (por conformação estrutural e posição no códon), revelaram a existência de dois clados principais em Oestroidea: Tachinidae + Mesembrinellinae e Oestridae + Rhiniinae + Sarcophagidae + Calliphoridae (definida em senso estrito). O status de família recentemente atribuído à Rhiniinae foi encontrado, enquanto há também evidências para sugerir o mesmo para a subfamília Mesembrinellinae, como proposto anteriormente por outros autores. As diferentes estratégias de particionamento do conjunto de dados amostrados resultaram em diferenças discretas em termos de topologia, comprimentos de ramo e suporte geral das filogenias inferidas. Embora o resultado geral indique uma melhor resolução das análises quando do uso de combinações de partições e modelos mais complexas, as mesmas podem ocasionar também um aumento considerável na incerteza associada às análises
Abstract: The Oestroidea superfamily (Diptera: Brachycera: Calyptratae), with +13,000 described species, comprises one of the most numerous and ecologically diverse groups in the Diptera order. The group is actually of great interest for human activities since it includes species of medical, veterinary and forensic importance, most of them included in the Calliphoridae family. Despite the existence of several studies addressing the issue, evolutionary relationships in Oestroidea, a group mainly composed of rapidly diverged lineages, remains contentious and poorly understood, encouraging the characterization of new molecular markers for phylogenetic inference analyses. In this context, this thesis was developed and organized in three chapters describing genetic and evolutionary studies in species of the Oestroidea superfamily, with emphasis in Calliphoridae. The first chapter deals with the characterization and evaluation of the second internal transcribed spacer region (ITS2) of the ribosomal DNA cluster as a molecular marker for phylogenetic inference in Calliphoridae, including information of both primary sequence and secondary structure. The analyses revealed an hierarchically organized pattern of genetic distances in the specific, generic and subfamilial level, while little intraspecific variation was detected. Inferred trees were able to recover most of the commonly accepted relationships among the sampled taxa, with the consideration of structural information resulting in better supported topologies. Thereby, the potential of the ITS2 region as a molecular marker for phylogenetic inference in the Calliphoridae family was corroborated and its use in larger scale analyses, including other markers with different evolutionary patterns, encouraged. Chapter II describes the in vitro characterization of the secondary structure of the ITS2 region, through patterns of enzymatic digestion and analysis of the generated fragments, in representative species of the three superfamilies of the Calyptratae clade: Glossina morsitans, Musca domestica and Cochliomyia hominivorax. Analyses of the patterns of the fragments generated by enzymatic digestions with the RNAses I, A, T1 and V1, when mapped in the in silico predicted secondary structure, corroborated the folding of most of the domains predicted by computational methods, highlighting the importance and reliability of these methods in secondary structure prediction. Chapter III describes molecular phylogenetic analyses in the Oestroidea superfamily, with emphasis on the Calliphoridae family, using four different molecular markers, two nuclear (ITS2 and 28S) and two mitochondrial (COI and 16S) regions. The analyses, which included a comprehensive evaluation of the effects of different data partitioning strategies in a Bayesian framework (by structural conformation and codon position), revealed the existence of two main clades in Oestroidea: Tachinidae+Mesembrinellinae and Oestridae+Rhiniinae+Sarcophagidae+Calliphoridae (defined in a strict sense). The recently attributed family status to Rhiniinae was confirmed, and there are evidence to also suggest the same for Mesembrinellinae, as previously pointed out by other studies. The different data partitioning strategies used in the sampled dataset resulted in small differences in terms of inferred topologies, estimated branch lengths and average support. Although the overall results indicate a significant increase in phylogeny resolution when more complex and parameter-rich models / partitions combinations are used, they can also lead to an increased uncertainty in the phylogenetic estimation process
Doutorado
Genetica Animal e Evolução
Soutor em Genética e Biologia Molecular

Ma, Ka Yan.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2009.
Includes bibliographical references (leaves 88-103).
Abstracts in English and Chinese.
ABSTRACT --- p.i
ACKNOWLEDGEMENTS --- p.vii
CONTENTS --- p.ix
LIST OF TABLES --- p.xi
LIST OF FIGURES --- p.xii
Chapter Chapter 1 --- Introduction
Chapter 1.1 --- Molecular phylogenetics --- p.1
Chapter 1.2 --- Phylogeny of the penaeoid shrimps --- p.2
Chapter 1.2.1 --- Interfamilial relationships of Penaeoidea --- p.3
Chapter 1.2.2 --- Ingergeneric relationships of Penaeidae --- p.8
Chapter 1.2.3 --- Interspecific relationships of Penaeus s.l --- p.11
Chapter 1.3 --- Molecular markers for decapods phylogenetics studies --- p.14
Chapter 1.3.1 --- Mitochondrial markers --- p.14
Chapter 1.3.2 --- Nuclear markers --- p.16
Chapter Chapter 2 --- Molecular phylogeny of superfamily Penaeoidea
Chapter 2.1 --- Introduction --- p.19
Chapter 2.2 --- Materials and methods --- p.21
Chapter 2.3 --- Results --- p.28
Chapter 2.4 --- Discussion --- p.40
Chapter 2.5 --- Conclusions --- p.48
Chapter Chapter 3 --- Molecular phylogeny of genus Penaeus sensu lato
Chapter 3.1 --- Introduction --- p.50
Chapter 3.2 --- Materials and methods --- p.50
Chapter 3.3 --- Results --- p.56
Chapter 3.4 --- Discussion --- p.74
Chapter 3.5 --- Conclusions --- p.84
Chapter Chapter 4 --- General conclusions --- p.85
References --- p.88

In eukaryotes, the defined loci on each chromosome, the centromeres, accomplish the critical task of correct cell division. In some organisms, centromeres are composed of a euchromatic central core region embedded in a stretch of heterochromatin and the inheritance and maintenance of centromeres are controlled by dynamic epigenetic phenomena. Although the size of centromeres differs between organisms, its organization, and the placement of euchromatic and heterochromatic regions is conserved from the fission yeast, Schizosaccharomyces pombe, to humans, Homo sapiens. However, relatively little is known about centromeres in the filamentous fungi from the Ascomycota, representing the largest group of fungi and fungal pathogens. Further, studies from humans, flies, yeast and plants have shown that the inheritance of centromeres is not strictly guided by centromeric DNA content, which is highly AT-rich, repetitive and constantly evolving. Therefore, it is difficult to align ans assemble the sequenced contigs of centromeric regions of higher eukaryotes, including most filamentous fungi. A genetic technique, tetrad (or octad) analysis has helped to map the centromeres of the filamentous fungus Neurospora crassa early on. The research presented in this dissertation used N. crassa as a model to focus on characterizing different features of centromeres with an emphasis on the centromere-specific histone H3 (CenH3) protein. Data included here represent the first study on centromere-specific proteins in Neurospora, and demonstrate that the central core of the centromeres are heterochromatic, showing enrichment of silent histone marks, which is in contrast to the centromere arrangement in fission yeast. The CenH3 protein, whose deposition on the genome licenses formation or maintenance of centromeres, shows highly divergent N-terminal regions and a conserved histone fold domain (HFD) in all eukaryotes. This bipartite nature of CenH3 is also observed in the Ascomycota, which provides an opportunity for functional complementation assays by replacing Neurospora CenH3 (NcCenH3) with CenH3 genes from other species within the Ascomycota. The results from this experimental approach provide good measures for (1) determining the specific regions of CenH3 required for the assembly of centromeres during meiotic and mitotic cell divisions and (2) analyzing the resistance to changes in the organization of centromeres in N. crassa. The genetic analysis showed that the divergent N-terminal region is essential for the proper assembly of centromeres, and that the conserved carboxy-terminus of CenH3 is important for the process of meiosis but not mitotic cell division. ChIP-seq analyses suggest that the observed loss of Podospora anserina CenH3 (PaCenH3- GFP) from certain N. crassa centromeres does not result in obvious phenotypic defects, e.g. diminished growth or evidence for aneuploidy. Further, the low enrichment of PaCenH3-GFP at certain centromeres is possibly predetermined during meiosis, which results in irreversible and progressive decreases in enrichment. It remains to be determined if this process is random as far as selection of centromeres is concerned. Together the results presented here suggest that during meiosis more stringent structural requirements for centromere assembly apply and that these are dependent on CenH3, and that depletion of CenH3 from centromeres does not critically affect mitosis in the asynchronously dividing nuclei of Neurospora hyphae.
Graduation date: 2013

Li, Chi Pang.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2010.
Includes bibliographical references (leaves 127-141).
Abstracts in English and Chinese.
Abstract --- p.i
Abstract (Chinese) --- p.iii
Acknowledgements --- p.v
Contents --- p.vi
List of Tables --- p.ix
List of Figures --- p.x
Chapter Chapter 1 --- General Introduction --- p.1
Chapter Chapter 2 --- Literature Review --- p.3
Chapter 2.1 --- Caridean phylogeny --- p.3
Chapter 2.1.1 --- Informative morphological characters in Caridean shrimps --- p.3
Chapter 2.1.2 --- Brief history of Caridean classifications --- p.4
Chapter 2.1.3 --- Natantia/Reptantia scheme vs. Dendrobranchiata/Pleocyemata scheme --- p.8
Chapter 2.2 --- Phylogney of the family Hippolytidae --- p.10
Chapter 2.3 --- Molecular approach to phylogeny --- p.11
Chapter 2.3.1 --- Use of molecular data --- p.11
Chapter 2.3.2 --- Use of mitochondrial gene markers in crustaceans --- p.12
Chapter 2.3.3 --- Use of nuclear gene markers in crustaceans --- p.14
Chapter Chapter 3 --- Phylogeny of the Infraorder Caridea Based on five Nuclear Genes --- p.27
Chapter 3.1 --- Introduction --- p.27
Chapter 3.2 --- Materials and Methods --- p.28
Chapter 3.2.1 --- Sample Collection --- p.28
Chapter 3.2.2 --- DNA extraction and PCR amplification --- p.28
Chapter 3.2.3 --- DNA sequencing --- p.29
Chapter 3.2.4 --- Phylogenetic analysis --- p.30
Chapter 3.3 --- Results --- p.34
Chapter 3.3.1 --- Enolase --- p.34
Chapter 3.3.2 --- NaK --- p.35
Chapter 3.3.3 --- PEPCK --- p.37
Chapter 3.3.4 --- Histone --- p.38
Chapter 3.3.5 --- 18S rRNA --- p.39
Chapter 3.3.6 --- Combined dataset --- p.41
Chapter 3.3.7 --- Substitution saturation analysis --- p.43
Chapter 3.4 --- Discussion --- p.44
Chapter 3.4.1 --- Evaluation of the five nuclear gene markers --- p.44
Chapter 3.4.1.1 --- Nuclear protein coding genes --- p.44
Chapter 3.4.1.2 --- 18S rRNA --- p.81
Chapter 3.4.2 --- Superfamilies and families --- p.82
Chapter 3.4.2.1 --- Superfamilies --- p.82
Chapter 3.4.2.2 --- Families --- p.86
Chapter 3.4.3 --- Basal groups --- p.86
Chapter 3.4.4 --- Procarididae --- p.88
Chapter Chapter 4 --- Phylogeny of the family Hippolytidae --- p.90
Chapter 4.1 --- Introduction --- p.90
Chapter 4.2 --- Materials and Methods --- p.91
Chapter 4.2.1 --- Sample Collection --- p.91
Chapter 4.2.2 --- DNA extraction and PCR amplification --- p.91
Chapter 4.2.3 --- DNA sequencing --- p.95
Chapter 4.2.4 --- Phylogenetic analysis --- p.95
Chapter 4.3 --- Results --- p.95
Chapter 4.3.1 --- Enolase --- p.95
Chapter 4.3.2 --- NaK --- p.98
Chapter 4.3.3 --- 16S rRNA --- p.99
Chapter 4.3.4 --- Combined dataset --- p.100
Chapter 4.4 --- Discussion --- p.118
Chapter 4.4.1 --- "Resurrection of family Lysmatidae Dana,1852" --- p.118
Chapter 4.4.2 --- Other hippolytid clades --- p.120
Chapter 4.4.2.1 --- """Hippolytidae""" --- p.120
Chapter 4.4.2.2 --- Bythocarididae --- p.121
Chapter 4.4.3 --- Superfamily Alpheoidea --- p.122
Chapter Chapter 5 --- General Conclusion --- p.125
References --- p.127

Finally, the gene tree of the true crabs, Brachyura, confirms that the basal "Podotremata" is paraphyletic, with the Raninoidea and Cyclodorippoidea more closely related to Eubrachyura than to the other podotremes. Within the monophyletic Eubrachyura, the analysis supports the reciprical monophyly of the two subsections, Heterotremata and Thoracotremata. All of the Old World freshwater crabs cluster together, representing an early diverged lineage in the Heterotremata.
From the inferred phylogeny, we have obtained new insights on the evolution of decapods. First, the spiny lobster from the family Palinuridae is found to be paraphyletic with the polyphyletic Synaxidae nested within it. The Stridentes forms a monophyletic assemblage, indicating that the stridulating sound producing organ evolved only once in the spiny lobsters. Moreover, the spiny lobsters originated in the shallower water rocky reefs of the Southern Hemisphere and then invaded deep sea habitats and diversified.
In sum, I demonstrate the utility of the nuclear protein-coding gene markers in decapod phylogeny and they are informative across a wide range of taxonomic levels. I propose that nuclear protein-coding genes should constitute core markers for future phylogenetic studies of decapods, especially for higher systematics.
Second, we show that hermit crabs have a single origin, but surprisingly, that almost all other major clades and body forms within the Anomura, are derived from within the hermit crabs. The crab-like form and squat lobster form have each evolved at least twice from separate symmetrical hermit crab ancestors. These remarkable cases of multiple parallelism suggest considerable phenotypic flexibility within the hermit crab ground plan, with a general tendency towards carcinization. Rather than having a separate origin from other major clades, hermit crabs have given rise to most other major anomuran body types.
The high diversity of decapods has attracted the interest of carcinologists but there is no consensus on decapod phylogeny in spite of the endeavors using both morphological and molecular approaches. New sources of information are necessary to elucidate the phylogenetic relationships among decapods. In the present study, I attempted to develop and apply the nuclear protein-coding gene markers on decapod phylogeny. Using only two protein-coding genes, we have successfully resolved most of the infraordinal relationships with good statistical support, indicating the superior efficiency of these markers compared to nuclear ribosomal RNA and mitochondrial genes commonly used in phylogenetic reconstruction of decapods. Apparently these two types of markers suffer from the problems of alignment ambiguities and rapid saturation, respectively. Subsequently, I tried to apply the nuclear protein-coding genes in revealing interfamilial and intergeneric evolutionary history in three selected decapod groups, the spiny lobster (family Palinuridae), the infraorder Anomura and the true crabs of the infraorder Brachyura to further evaluate the utility of these markers and reconstruct the evolutionary history the groups. Trees with robust support can be obtained using sequences of three to five genes for the infraorders and families tested including the most speciose Brachyura. The genes are shown to be informative in elucidating interspecific phylogeny as well.
Tsang, Ling Ming.
Adviser: Ka Hou Chu.
Source: Dissertation Abstracts International, Volume: 73-02, Section: B, page: .
Thesis (Ph.D.)--Chinese University of Hong Kong, 2010.
Includes bibliographical references (leaves 127-153).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstract also in Chinese.

Ho, Ka Chai.
Thesis submitted in: November 2006.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2007.
Includes bibliographical references (leaves 127-145).
Abstracts in English and Chinese.
Abstract --- p.i
Abstract (Chinese) --- p.iii
Acknowledgements --- p.v
Table of Contents --- p.vi
List of Tables --- p.ix
List of Figures --- p.x
Chapter Chapter 1 --- General Introduction --- p.1
Chapter 1.1 --- Molecular phylogeny of Metanephrops --- p.1
Chapter 1.2 --- Identification of Nephropidae using DNA barcodes --- p.3
Chapter Chapter 2 --- Literature Review --- p.5
Chapter 2.1 --- Molecular phylogenetic studies of crustaceans --- p.5
Chapter 2.1.1 --- Molecular phylogeny and reasons of using molecular markers in phylogenetic studies --- p.5
Chapter 2.1.2 --- Characteristics of animal mitochondrial genome --- p.7
Chapter 2.1.3 --- Examples of crustacean phylogenetic studies derived from mitochondrial DNA --- p.8
Chapter 2.2 --- Identification of species based on DNA barcode --- p.17
Chapter 2.2.1 --- Traditional taxonomy and its current practice --- p.17
Chapter 2.2.2 --- Needs for DNA barcode --- p.18
Chapter 2.2.3 --- Molecular identification based on DNA barcodes --- p.21
Chapter 2.3 --- Taxonomy of Nephropidae --- p.28
Chapter 2.3.1 --- Classification and phylogenetic relationship of Nephropidae --- p.28
Chapter 2.3.2 --- Classification and distribution of Metanephrops --- p.31
Chapter 2.3.3 --- Evolutionary history of Metanephrops --- p.36
Chapter Chapter 3 --- Molecular Phylogeny of Metanephrops --- p.38
Chapter 3.1 --- Introduction --- p.38
Chapter 3.2.1 --- Species studied and sample collection --- p.41
Chapter 3.2.2 --- DNA extraction --- p.43
Chapter 3.2.3 --- Amplification of mitochondrial genes --- p.43
Chapter 3.2.4 --- Nucleotide sequencing --- p.46
Chapter 3.2.4.1 --- Asymmetric PCR --- p.46
Chapter 3.2.4.2 --- Purification of asymmetric PCR products --- p.47
Chapter 3.2.5 --- Sequence alignment --- p.47
Chapter 3.2.6 --- Phylogenetic analyses --- p.48
Chapter 3.3 --- Results --- p.50
Chapter 3.3.1 --- PCR products of 16S rRNA and COI genes --- p.50
Chapter 3.3.2 --- Nucleotide composition of 16S rRNA gene alignments --- p.52
Chapter 3.3.3 --- Nucleotide composition of COI gene alignments --- p.54
Chapter 3.3.4 --- Intraspecific and interspecific genetic variation --- p.56
Chapter 3.3.5 --- Phylogenetic analysis based on 16S rRNA gene sequences --- p.61
Chapter 3.3.6 --- Phylogenetic analysis based on COI gene sequences --- p.68
Chapter 3.3.7 --- Phylogenetic analysis based on combined data set --- p.74
Chapter 3.4 --- Discussion --- p.80
Chapter 3.4.1 --- Interspecific genetic divergence --- p.80
Chapter 3.4.2 --- Monophyly of the four species groups --- p.81
Chapter 3.4.3 --- Phylogenetic relationship in Metanephrops --- p.84
Chapter 3.4.4 --- Evolutionary history of Metanephrops --- p.90
Chapter Chapter 4 --- Molecular Identification of Nephropidae --- p.92
Chapter 4.1 --- Introduction --- p.92
Chapter 4.2 --- Materials and methods --- p.93
Chapter 4.2.1 --- Species studied and sample collection --- p.93
Chapter 4.2.2 --- DNA extraction --- p.95
Chapter 4.2.3 --- Amplification of genes --- p.95
Chapter 4.2.4 --- PCR profiles for mitochondrial genes --- p.97
Chapter 4.2.5 --- Nucleotide sequencing --- p.97
Chapter 4.2.6 --- Purification of asymmetric PCR products --- p.97
Chapter 4.2.7 --- Sequence alignment --- p.97
Chapter 4.2.8 --- Cluster analysis --- p.97
Chapter 4.2.9 --- Graphical summary of species similarity --- p.98
Chapter 4.2.10 --- Testing of molecular identification system in Nephropidae --- p.98
Chapter 4.3 --- Results --- p.100
Chapter 4.3.1 --- PCR products and sequence alignments of 16S rRNA and COI genes --- p.100
Chapter 4.3.2 --- Species identification for clawed lobsters --- p.100
Chapter 4.3.2.1 --- 16S rRNA profile --- p.100
Chapter 4.3.2.2 --- COI profile --- p.108
Chapter 4.4 --- Discussion --- p.116
General Conclusion --- p.124
Literature Cited --- p.127
Appendices --- p.146

Pearl oysters of the genus Pinctada include some economically important species. The taxonomy of some of the species is problematic. Phylogenetic relationship of the species in the genus is also poorly studied. In the present study, phylogenetic relationships of P. chemnitzi, P. fucata, P. margaritifera, P. maxima, P. nigra, P. radiata (from China), P. fucata martensii (from Japan), P. albina and P. imbricata (from Australia) were studied with Pteria penguin as an outgroup, and genetic variation of Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata populations were investigated (1) to address the taxonomic confusion and phylogeny of pearl oysters, (2) to understand the genetic connections between the Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata in west Pacific and (3) to provide information for the genetic improvement program initiated in China.
Since P. fucata, P. fucata martensii and P. imbricata are synonymous, to study the genetic differentiation and genetic variation of such widely distributed populations is helpful in understanding their genetic connections. For this purpose, five populations, three from China (Daya Bay, Sanya Bay and Beibu Bay), one from Japan (Mie Prefecture) and one from Australia (Port Stephens) were studied using AFLP technique. Three primer pairs generated 184 loci among which 91.8-97.3% is polymorphic. An overall genetic among populations and an average of 0.37 within populations (ranging from 0.35 in Japanese population to 0.39 in Beibu Bay population) were observed. Genetic differentiation among the five populations is low but significant as indicated by pairwise GST (0.0079-0.0404). AMOVA further shows that differentiation is significant among the five populations but is not significant at a broader geographical scale, among the three groups of Chinese. Japanese and Australian populations or among the two groups of Australian and north Pacific populations. The low level of genetic differentiation indicated that P. fucata populations in the west Pacific are genetically linked. Among the five populations, the Australian one is more differentiated from the others, based on both pairwise AMOVA and GST analyses, and is genetically isolated by distance as indicated by Mantel test. However, genetic differences among the three Chinese populations are not correlated with the geographic distances, suggesting that Hainan Island and Leizhou Peninsula may act as barriers blocking gene flow.
The above three wild Chinese populations in southern China were compared with the three adjacent cultured populations using AFLP markers. Three pairs of primers generated 184 loci among 179 individuals in populations from Beibu Bay, Daya Bay and Sanya Bay. A high level of genetic diversity, ranging from 0.363 in a wild population in Sanya Bay to 0.388 in a wild population in Beibu Bay, was observed within both wild and cultured populations, indicating an absence of strong bottleneck effects in the history of cultured P. fucata populations. Yet cultured populations in Sanya Bay and Beibu Bay had more fixed loci than the corresponding wild populations. Genetic differentiation in most pairwise comparisons of populations was significant. AMOVA indicated that genetic variation among populations were very low (1.77%) though significant, while more than 98% variation resided among individuals within population. These findings provide no evidence to show that hatchery practice of pearl oyster in China to date has significantly affected the genetic diversity of the cultured populations, and suggest that all populations are competent for selection. Yet the significant genetic differentiation among populations implies that any translocation of individuals for genetic improvement program should be managed with caution for the preservation of genetic diversity in natural populations.
The internal transcribed spacers (ITS1 and ITS2) of nuclear ribosomal DNA were compared among the above nine taxa, based on sequences determined by the present study and those available from Genl3ank. The phylogenetic analysis indicates that the pearl oysters studied constitute three clades: clade I with the small oysters P. fucata, P. fucata martensii and P. imbricata, clade II with P. albina, P. nigra, P. chemnitzi and P. radiata, and clade III and clade III with the big pearl oysters P. margaritifera and P. maxima forming the basal clade. Clade II is made up two subclades: clade IIA consisting of P. albina and P. nigra and clade IIB consisting of P. chemnitzi and P. radiata. The topology of the phylogenetic tree and substitution pattern of ITS sequences suggest that P. margaritifera and P. maxima are primitive species and P. chemnitzi is a recent species. The genetic divergences between clades ranged from 28% to 76.5%, and between subclades, 8.7-10.2%. In clade I, the interspecific genetic divergences ranged from 0.6% to 1.4%, and overlapped with interspecific divergences (0.6-1.1%), indicating that P. fucata, P. fucata martensii and P. imbricata may be conspecific. Based on amplified fragment length polymorphism (AFLP) markers and ITS sequences from more individuals, analyses of the populations of these three taxa also support the conclusion that Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata are the same species, with P. fucata being the correct name. The genetic divergence between P. albina and P. nigra was also very low (1.2%), suggesting that they may represent two subspecies that can only be distinguished by shell color. The genetic divergences between P. maxima and P. margaritifera, and between clade IIA and clade IIB ranged from 8.3% to 10.2%, suggesting that they are closely related, respectively. The ITS1 sequence of P. radiata from GenBank is almost identical to that of P. chemnitzi determined in the present study, suggesting that the specimen used for the P. radiata sequence was possibly misidentified.
Yu Dahui.
"August 2005."
Adviser: Ka Hou Chu.
Source: Dissertation Abstracts International, Volume: 67-11, Section: B, page: 6125.
Thesis (Ph.D.)--Chinese University of Hong Kong, 2005.
Includes bibliographical references (p. 100-124).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstracts in English and Chinese.
School code: 1307.

Ph.D.
Pimelea Banks & Sol. Ex Gaertn. nom. cons. is a large genus consisting of 110 species, of which 90 species are endemic to Australia, 19 to New Zealand and one to Lord Howe Island. The genus has a great diversity of life forms, breeding systems and habitat. Its closest related genus is Thecanthes Wikstr. Thecanthes comprises five species of annual herbs occurring in the Philippines, New Ireland and northern Australia. Australasian Thecanthes and Pimelea are the only genera within sub-tribe Pimeleinae (angiosperm family Thymelaeaceae) and are characterised by the reduction to two stamens. Here I present the most comprehensive molecular phylogenetic study for Pimelea and Thecanthes. Sequences data from nuclear ITS rDNA and plastid rbcL, rps16, matK and trnL-F intergeneric spacer were used to reconstruct a phylogeny for these genera. I have produced 457 new DNA sequences (five genes and 150 taxa) for the present analyses. The resulting phylogeny was used to assess the taxonomic status of Thecanthes and to evaluate the relationships with Pimeleinae since previous studies indicated a close relationship between Pimelea, Thecanthes and species of Gnidia L. from tropical Africa. The morphological delimitation of sections within Pimelea, the biogeography and the radiation of the genus have been revaluated. Pimelea was found to be monophyletic. It was concluded that Pimelea and Thecanthes are congeneric; consequently a paper has been submitted transferring all species of Thecanthes into Pimelea and making the new combination Pimelea filifolia (Rye) Motsi & Rye. Data analysis revealed very low sequence variation within the subtribe Pimeleinae. This suggested a rapid radiation of the genera, which was confirmed by my molecular dating analyses. Based on molecular clock techniques, I calculated the following ages for the origin of Pimelea: 4.1 mya for New Zealand Pimelea spp. and 13.38 mya for other Pimelea spp. The molecular data also indicated that Pimelea and South Africa Gnidia have a direct common ancestor. I also show that the New Zealand Pimelea are derived and dispersed from Australian

Rabies, one of the oldest recognized viral zoonotic diseases, is a fatal encephalomyelitis transmitted to man via contact with infected animals. Evan today, rabies still is a disease of public health concern with many potentially preventable deaths occurring mainly in Asia, Africa and Latin America. Rabies and rabies-related viruses are members of the Lyssavirus genus, which comprises the rabies virus (genotype 1), Lagos bat virus (genotype 2), Mokola virus (genotype 3), Duvenhage virus (genotype 4), European bat lyssaviruses 1 and 2 (genotypes 5 and 6) and the Australian bat lyssavirus (genotype 7). Antigenic and genetic studies have shown that rabies virus strains circulating in particular host species tend to undergo genetic adaptation and evolve into distinct biotypes that differ in antigenicity and pathogenicity. Two biotypes of rabies virus are recognized in southern Africa. The first called the canid viruses, infect carnivores of the family Canidae (dogs, jackals and bat-eared foxes) and the second, the viverrid viruses, infect carnivores of the family Herpestidae (the yellow mongoose Cynictis penicil!ata and the slender mongoose Galerella sanguinea). In an endeavour to better understand the molecular epidemiology of lyssaviruses in Zimbabwe and South Africa, we analysed nucleotide sequences of the glycoprotein and the G-L intergenic region (rabies viruses) and the nucleoprotein gene (Mokola viruses). The main aim of the studies described in this thesis was to characterise lyssaviruses (genotypes I and 3) from Zimbabwe and compare them to those present in South Africa. In addition, we wanted to establish the role of the various rabies variants in rabies epizootics in the southern African subcontinent. It could be shown from this study that all the southern African canid viruses were closely related, with no general distinction between viruses from any of the canid species. Despite the general overall similarity between the canid viruses, certain phylogenetic groupings were apparent and by association with host species, geography and year of isolation, certain groups could be identified as particular epidemiological cycles. A high genetic diversity was evident amongst viverrid rabies viruses, the opposite of our observation for canid viruses. The viverrid virus groups corresponded to geographical pockets that were independent of host species. Mokola viruses from Zimbabwe were shown to be different from those from South Africa and phylogenetic relationships of these viruses were related to their geographical location of origin. This study has demonstrated the value of multinational surveillance and investigation in understanding the epidemiology of lyssaviruses in southern Africa and elsewhere in Africa. The results presented here will serve as basis for future studies on lyssaviruses in Africa and will contribute to the improved surveillance and control programs of rabies and Mokola viruses in the region.
Thesis (PhD (Microbiology))--University of Pretoria, 2006.
Microbiology and Plant Pathology
unrestricted

Data from nuclear ribosomal internal transcribed spacer regions (nrITS) and chloroplast DNA (cpDNA) have failed to resolve phylogenetic relationships in Pinus. To provide greater interspecific resolution, five low-copy nuclear genes were developed from mapped conifer anchor loci. Four genes were sequenced from species representing all Pinus subsections. Individual loci do not uniformly support the nrITS or cpDNA hypotheses. Combined analysis of low-copy nuclear loci produces a well-supported subsectional topology. The phylogenetic positions of P. nelsonii and P. krempfii are of systematic interest. Results strongly support P. nelsonii as sister to sect. Parrya, and suggest a moderately well-supported position of P. krempfii as sister to the remaining sect. Quinquefoliae. The most informative locus, a Late Embryogenesis Abundant-like gene, was used to explore phylogenetic relationships among closely related species in subg. Strobus. Thirty-nine species were sequenced, with two or more alleles representing 33 species. Nineteen of 33 species exhibited allelic nonmonophyly in the strict consensus tree. Nucleotide diversity was strongly associated (P<0.0001) with the degree of species monophyly. While species nonmonophyly complicates phylogenetic interpretations, this locus offers greater topological support than cpDNA or nrITS. Lacking evidence for hybridization, recombination, or imperfect taxonomy, incomplete lineage sorting remains the best explanation for trans-species polymorphisms. The absence of allelic coalescence is a severe constraint in the application of phylogenetic methods in Pinus, and taxa sharing similar life history traits may show analogous patterns. While lack of coalescence may limit their utility in traditional phylogenetics, nuclear genes remain highly informative in describing speciation events. Pinus chiapensis is a threatened species originally described as a variety of P. strobus. Prior morphological work suggests P. chiapensis is a distinct species, but that taxonomy is not universally accepted. Multiple accessions of three probable progenitors were sequenced at three nuclear loci. No interspecific allele sharing occurs with P. chiapensis, and its alleles are monophyletic at two loci. Results demonstrate that P. chiapensis is a distinct species. However, determination of the sister species is complicated by lack of species monophyly and interlocus variability. Pinus ayacahuite is the least likely progenitor, but the relationship of P. chiapensis to P. monticola or P. strobus is unclear.
Graduation date: 2006

Low copy number anonymous nuclear loci were used to search for species markers in four species of Klamath Basin suckers. We sequenced 28 randomly chosen loci representing 10,421 bp; 21 loci were similar to sequences in GenBank. Eight fixed sequence differences were found among Klamath species. Locus 120 contained rare but diagnostic markers for Deltistes luxatus and for Catostomus rimiculus. Locus 4 also contained three rare but unique sites in Catostomus rimiculus. No sequence differences were found between Chasmistes brevirostris and Catostomus snyderi. Loci 4 and 120 exhibited allele frequency differences between Rogue River C. rimiculus and all Klamath Basin suckers. Genotype BB of locus 4 was a fixed diagnostic marker and genotype BB of locus 120 was a frequency dependent marker for Rogue C. rimiculus. Although Klamath suckers represent three genera, very limited variation was found among 10,431 base pairs. We examined phylogenetic patterns of five loci in eleven catostomid genera and 25 species to determine if the homogeneity in the Upper Klamath Basin was due to massive hybridization and introgression or to retention of ancestral sequences. Two loci with no similarity to GenBank sequences (non-coding loci) and three loci with substantial similarity to GenBank sequences (coding loci) gave similar results, providing support for various subfamilies and tribes, more support for eastern genera and little support for western genera. Each locus was a mosaic of species or population markers, sometimes providing discriminatory power for allopatric populations of a species, such as C. macrocheilus, while not discriminating other species. Upper Klamath Basin species were noteworthy in their lack of autapomorphies, but had similar numbers of derived informative sites as other catostomins. Upper Klamath Basin species consistently shared ancestral or equivocal informative sites either with moxostomatins or a variable group of western species and shared derived sites with other western species, especially C. occidentalis. The data suggest that Upper Klamath Basin species have retained a largely ancestral genome at these loci. Thus, the failure of this technique to uncover significant variation in Upper Klamath Basin species may be a reflection of their plesiomorphic genome at these loci and not necessarily hybridization.
Graduation date: 2004

The taxonomy and biogeography of the southern African pheasant shell fauna are poorly known. Thirty–one nominal taxa referable to Phasianelloidea have been described or recorded in this region, but no systematic revision of these has ever been undertaken. Morphological evidence suggests that 16 taxa represent valid species, 13 are synonyms and two represent incorrect identifications. DNA sequence data from mitochondrial COI and 16S markers are used to assess the validity of the described nominal southern African Tricolia species. Phylogenetic analyses recovered seven distinct clades. Tricolia adusta, T. elongata, T. formosa, T. kochii, T. saxatilis and T. neritina were recovered as distinct species. Tricolia africana and T. capensis are genetically indistinguishable. However, morphological characters of the shell are clearly diagnosable. This could be due to incomplete sorting (ancestral polymorphism) reflecting recent speciation with rapid morphological and ecological divergence co–incident with geographical separation. Similarly, there is little genetic differentiation between T. bicarinata, T. insignis and T. kraussi. In this case the similarity is also supported by morphological data as the three species are conchologically close with intergrading shell characters, and might even be one species exhibiting ecogeographic variation in shell form. Monophyly of the southern African Tricolia species is not supported as well as the relationship between these and the European Tricolia pullus. In the last chapter a molecular phylogeny based on sequence data from mtDNA (COI and 16S), nuclear (18S and 28S) and the combined data (COI, 16S, 18S and 28S) is presented for the Phasianelloidea. Bayesian inference analyses performed on the combined data support the monophyly of Tricolia sensu stricto, Eulithidium and Phasianella. Tricolia sensu lato is not monophyletic, as its southern Australian and Indo–West Pacific species do not cluster with its southern African and Eastern Atlantic representatives. The position of Hiloa and Gabrielona within the Phasianelloidea is unresolved. Phylogenetic reconstructions using bayesian inference support monophyly of the Phasianelloidea.
Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2010.

Plantago ovata Forssk. (Plantaginaceae) is a winter annual species which, in North America, inhabits desert and Mediterranean habitats of the southwest United States, northwest Mexico and the Channel Islands of California and Mexico. In the eastern hemisphere P. ovata inhabits desert regions ranging from the Canary Islands, across northern Africa to western India. The wide disjunction between P. ovata in the western and eastern hemispheres poses an interesting question as to the origin and biogeography of the species. Previous authors have hypothesized that P. ovata was introduced to North America over the Bering land bridge, from Asia, during the Miocene, or introduced anthropogenically from Europe during the 18th century by Spanish settlers. In this study we examined sequence data from the chloroplast trnL-trnF, trnS-trnG and psbA-trnH regions, the nuclear ribosomal internal transcribed spacer (ITS) and a region 5' of the TCP region of a CYCLOIDEA gene. Using a molecular clock based on an ITS calibration within the Plantago genus, and a clock for plant chloroplast, we date a non-anthropogenic introduction event, from the Old World to North America, approximately 200,000-650,000 years ago. This is consistent with a Pleistocene origin, and does not support a Miocene origin of the disjunction. Based on a morphological survey of 552 specimens, from throughout the world range of P. ovata, we suggest the recognition of four subspecific taxa. Molecular phylogenetic analysis of chloroplast DNA and nuclear ribosomal DNA ITS sequences support this taxonomic treatment. Furthermore, phylogenetic sequences of the CYCLOIDEA gene support the morphological data. Both suggest the origin of North American P. ovata as a result of hybridization between Old World P. ovata varieties. This event provides further evidence that hybridization may serve as a predictor of invasiveness in plants.
Graduation date: 2005

Two of the most challenging goals of evolutionary biology are to reconstruct the evolutionary relationships among all extant species and to understand the process by which new species form. Accomplishing these goals will require accurate computational methods for reconstructing phylogenetic trees, general analytic models of speciation, and powerful statistical tools for studying the process of speciation in natural systems. In the first chapter, I study the effects of improper model assumption on estimates of phylogeny. Using DNA sequence data simulated under a variety of models of sequence evolution, I demonstrate that use of oversimplified models can result in erroneous phylogeny estimates. This result suggests that if the models currently utilized are oversimplified then current estimates of phylogeny may be inaccurate and more complex models need to be developed and employed. In the second and third chapters, I study one process thought to be important in completing the final stages of speciation: reinforcement. Using simulations of a hybrid zone, I show that the process of reinforcement can result in patterns other than reproductive character displacement. I also show that speciation by reinforcement is more likely when the genes involved in reproductive isolation are sex-linked. In the fourth chapter, I develop a statistical method of quantifying the degree of isolation between species undergoing divergence. Using genotype data obtained from natural hybrid zones, this novel method can be used to estimate the fitness of hybrids during different stages of their life cycle. This approach offers a new approach to empirical biologists studying extrinsic postzygotic isolation in natural systems.

Scarabaeid pests in South Africa and especially KwaZulu-Natal are characterised by a very long larval life cycle and short pupal and adult periods. However, it has nearly always been the adults of the species that have been identified, with very little attention paid to the larval identification of the species. This is unfortunate as it is nearly always the larval stage that is found to be associated with crop damage. Accurate identification of the species of these larvae is important for the management of scarabaeid pest species, as it unlocks the necessary information on the biology and ecology of many species, which allows the adaptation of control methods for different species. Inadequate keys for the taxonomy of larvae of these groups, as well as the lack of morphological taxonomists working on these groups have been identified as constraints. When a species is difficult to identify using traditional taxonomic methods, DNA diagnostic tools can be useful. Chapter 2 investigated the feasibility of identifying scarabaeid larvae using mitochondrial DNA data. Variation in the base pair sequence of the mitochondrial cytochrome c oxidase sub unit I (cox 1) gene was used. DNA sequences of cox 1 from scarabaeid larvae collected from sugarcane fields were compared with sequences from scarabaeid adults of known species in order to identify the species attacking sugarcane. Neighbour-joining and maximum parsimony analyses of 658 bp cox 1 sequences identified groups of larvae that linked to adult specimens. The major groupings delimited specimens belonging to the subfamilies Dynastinae, Melolonthinae and Rutelinae. Within-group sequence divergence ranged from 0 - 3.4 % and divergence between sister groups ranged from 2.6 - 25.1 %. The recorded divergence range within and between tribes was 0 - 21.3 % and 17.3 - 28.5% respectively. Similarly, the divergence range observed within and between genera was 0 - 19.2 % and 17.1 - 25.4% respectively. The maximum sequence divergence observed within subfamilies was 23.7 % and divergence between subfamilies ranged from 16.8 - 26.7 %. Examination of pairwise sequence divergence levels as well as node support allowed 68% of the unidentified larval specimens to be associated with identified adult specimens. Phylogenetic analysis matched identified adult mtDNA with unidentified larval mtDNA. This allowed the identification of those larvae through morphological characteristics unique to certain species. To create a field key to the subfamilies of Dynastinae, Melolonthinae and Rutelinae the most useful character distinguishing larvae of different species was the raster but additional morphological characteristics were included. These relationships between larval and adult scarabaeid specimens from sugarcane were examined using various phylogenetic tools. The data set included a total of 19 morphological characters as well as 166 partial cox 1 gene sequences. Maximum parsimony analyses were performed on morphological, molecular and combined data. The same morphological and molecular data sets were run both separately and as a combined analysis with MrBayes. In both types of analyses the morphological data performed poorly and crude groupings resulted, dividing taxa to tribe level only. Molecular data showed greater resolution than the morphological data and taxa were separated into groups equivalent to species and morphospecies designated in Chapter 2. A partition homogeneity test indicated that both data types could be combined. It is recommended that both morphological and molecular data be utilised in identification of scarabaeid sugarcane pests and that a character-based approach be implemented. Further molecular data from other genes should be included to test the accuracy of these results. The keys produced during this study will allow workers to focus on a single species biology, and subsequently allow an analysis of between species interactions, and within species control. These advances are a start to the improvement of knowledge of the species composition of scarabaeid larvae in sugarcane fields, thus making management and biological control of these pests a greater possibility. Further recommendations for future work are discussed in Chapter 5.
Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2008.

Fossils are needed to calibrate the molecular clock used for dating phylogenies. This has led to a re-examination of the identification and phylogenetic placement of fossils used for calibration. Until recently, fossil pollen has been neglected for calibration due to a belief that pollen lacks sufficiently useful characters. Consequently, taxonomy of extant pollen has also been neglected, and this applies particularly to the pollen of Myrtaceae, which was last fully revised in the 1950s. This study surveys pollen morphology across Myrtaceae, assessing the phylogenetic signal in pollen characters and assessing suitability of fossil pollen for calibration. Fossils identified as suitable were used to calibrate a new dated molecular phylogeny, which was then applied to investigating the evolution and biogeography of Myrtaceae. A study of pollen from over 200 living taxa using Scanning Electron Microscopy (SEM) and 500 taxa using Light Microscopy (LM) found nine distinct pollen types within Myrtaceae that potentially showed a relationship to molecular tribal classification of the family. Optimising pollen characters onto a molecular phylogram, constructed from two chloroplast (matK and ndhF) and one nuclear (ITS) loci from 111 taxa, indicated the potential use of colpal morphology in diagnosing Myrtaceae pollen groups. However, exine pattern, apocolpial island presence and pollen width are homoplasious and relatively uninformative. A review of all formally described Myrtaceidites fossil species identified nine distinct morphotypes, including six that could be used to calibrate molecular dating. One new morphospecies, Myrtaceidites leptospermoides, was described for fossil pollen with syncolpate colpi and a granulate exine pattern. The fit of 26 pollen fossils onto the new molecular phylogeny was measured using parsimony optimisation of characters from extant Myrtaceae pollen. Eight Myrtaceidites fossils were identified as appropriate for calibration based on their placements on the tree. These fossils were used to calibrate a Bayesian phylogenetic analysis and this led to older estimates than have been previously found for the crown ages of tribes such as Eucalypteae and Myrteae, showing the potential of pollen for calibration. A Bayesian phylogenetic analysis using a partitioned relaxed clock and 12 fossil calibrations was used to test biogeographic hypotheses by optimizing the modern geographic location of extant taxa using parsimony reconstruction. Of the 22 tested disjunct sister-groups, up to five could possibly be explained by vicariance, four likely resulted from overland dispersal via new land connections, and 13 were too young for vicariance by continental drift and therefore inferred to be the result of long distance dispersal and establishment (LDDE) events. Holocene fossil pollen from Bega Swamp, New South Wales, Australia, were compared with pollen of twenty-five extant Myrtaceae species from the surrounding Bega Swamp area using visual judgement and a Lucid key constructed for the purpose. It was found that Eucalyptus pauciflora, together with other Eucalyptus species, has occurred in the Bega Swamp area for over 12,500 years. However, interpreting past vegetation composition using Eucalyptus is problematic because extant taxonomic (e.g. subgenus Symphyomyrtus) or ecological (e.g. alpine or wet forest) groups do not form pollen with distinct pollen types and therefore fossils cannot be confidently assigned to any extant Eucalyptus group.