Foi realizado um estudo das relações filogenéticas de Stygarctidae e Digitopoda (Tardigrada, Archaeotardigrada, Macrocephala) seguindo-se os princípios e métodos henniguianos. Foram selecionados na literatura os caracteres morfológicos utilizados para os dois grupos: 61 caracteres (43 binários e 18 multiestado) para 17 táxons terminais, de Stygarctidae, ao nível de espécie, e 40 caracteres (31 binários e 9 multiestado) para 10 táxons superiores de Digitopoda. As análises manuais produziram cladogramas totalmente resolvidos com 101 passos evolutivos e índice de consistência 0,86 para Stygarctidae, e 79 passos evolutivos e índice de consistência 0,63 para Digitopoda. Também foram feitas análises numéricas, com o auxílio do programa Hennig 86, comparando-se os resultados destas com as análises manuais. O algoritmo utilizado foi o mhennig*. O consenso de duas árvores de Stygarctidae apresentou topologia idêntica à do cladograma supracitado, com 98 passos evolutivos, índice de consistência 0,77 e índice de retenção 0,82. No caso de Digitopoda, o consenso de duas árvores apresentou uma topologia radicalmente diferente do resultado manual, com 66 passos evolutivos, índice de consistência 0,68 e índice de retenção 0,63. Também foram analisados dois caracteres multiestado, cada um com duas condições apomórficas, para três táxons terminais (nível de espécie) de Orzeliscinae. Neste caso, o cladograma obtido apresentou quatro passos evolutivos e índice de consistência 1. Stygarctidae foi mantido como táxon monofilético, apresentando a seguinte topologia entre os seus gêneros: ((Parastygarctus + Stygarctus) + ((Mesostygarctus + Pseudostygarctus) + Megastygarctides)). Digitopoda foi subdividido em dois táxons monofiléticos, apresentando a seguinte topologia: (Neostygarctus + (Neostygarctus + Batillipes + (Batillipes + (Halechiniscinae + Orzeliscinae) + ((Halechiniscinae + Orzeliscinae) + Dipodarctus + (Dipodarctus + (Floractinae + Tanarctinae))))) + Renaudarctus + (Renaudarctus + (Euclavarctinae + Styraconyxinae))). Halechiniscidae não é monofilético e Halechiniscinae é um táxon mais restrito, que inclui apenas Halechiniscus e Chrysoarctus. Orzeliscinae é redefinido de forma a incluir Paradoxipus, grupo-irmão de Opydorscus + Orzeliscus. Styraconyxinae inclui também Archechiniscus. Archechiniscinae não é válido. Neostygarctidae, Renaudarctidae, Neoarctidae e Megastygarctidinae foram eliminados do sistema de Tardigrada / Relationships among the subgroups of Stygarctidae and Digitopoda (Tardigrada, Archaeotardigrada, Macrocephala) were invetigated according to hennigian principles and methods. Morphological characters were selected from the literature for these two groups: 61 characters (43 binary and 18 multistate) for 17 species of Stygarctidae and 40 characters (31 binary and 9 multistate) for 10 major groups of Digitopoda. Manual analysis produced fully resolved cladograms with 101 evolutionary steps and consistency index = 0,86 for Stygarctidae, and 79 evolutionary steps and consistency index = 0,63 for Digitopoda. Numerical analysis were done using the software Hennig 86, for comparison with manual analysis. The algoritm mhennig* was used. The consensus tree of two Stygarctidae trees showed identical topology with the manual cladogram, with 98 evolutionary steps, consistency index = 0,77 and retention index = 0,82. The consensus tree of two Digitopoda trees showed a complete different topology from the manual cladogram, with 66 evolutionary steps, consistency index = 0,68 and retention index = 0,63. There were also analysed two multistate characters, each one with two apomorphic conditions, for three Orzeliscinae species. In this case, another fully resolved cladogram with four evolutionary steps and consistency index = 1 was obtained. Stygarctidae was maintained as a monophyletic taxon, showing the following system for its genera: ((Parastygarctus + Stygarctus) + ((Mesostygarctus + Pseudostygarctus) + Megastygarctides)). Digitopoda, branched into two monophyleitc taxa, showed the following system: (Neostygarctus + (Neostygarctus + Batillipes + (Batillipes + (Halechiniscinae + Orzeliscinae) + ((Halechiniscinae + Orzeliscinae) + Dipodarctus + (Dipodarctus + (Floractinae + Tanarctinae))))) + Renaudarctus + (Renaudarctus + (Euclavarctinae + Styraconyxinae))). Halechiniscidae is not a monophyletic taxon and Halechiniscinae is a more inclusive taxon comprising only Halechiniscus e Chrysoarctus. Orzeliscinae includes Paradoxipus, the sistergroup of Opydorscus + Orzeliscus. Styraconyxinae is monophyletic with the inclusion of Archechiniscus. Archechiniscinae is not valid. Neostygarctidae, Renaudarctidae, Neoarctidae and Megastygarctidinae are not supported in the system of the Tardigrada

A família Rallidae é representada por aves cosmopolitas popularmente conhecidas como saracuras, sanãs, carquejas, galinhas-d`água e frangos-d`água. Compreende cerca de 135 espécies distribuídas em 33 gêneros, dos quais 13 são monotípicos. As relações filogenéticas baseadas em caracteres morfológicos e dados moleculares indicam diferentes afinidades entre os membros da família, principalmente na posição dos gêneros Rallus, Porphyrio, Gallinula e Fulica. Neste estudo, focado em espécies Neotropicais da família, uma nova análise filogenética baseada em caracteres osteológicos é proposta. Uma amostra de 100 esqueletos de 13 gêneros e 31 espécies foi analisada. No total 50 caracteres foram codificados, dos quais 17 são cranianos e 33 pós-cranianos para a construção de uma matriz e subseqüente análise filogenética de acordo com o princípio da parcimônia. Foram calculados árvores de consenso estrito e consenso de maioria. A primeira gerou 151 árvores igualmente parcimoniosas com 99 passos. A análise com método de pesagem sucessiva dos caracteres obteve melhores resoluções entre as espécies amostradas. A topologia do cladograma permite a validade de determinados gêneros como entidades monofiléticas, como Rallus, Porphyrio, Aramides, Gallinula e Fulica. O posicionamento de Porphyrio como um ramo basal dentro da subfamília Rallinae foi suportado e suas relações interespecíficas demonstram que as espécies do Novo mundo são mais proximamente relacionadas do que P. porphyrio, permitindo a inclusão taxonômica de Porphyrula. A relação próxima entre as espécies do gênero Gallinula e Fulica foi corroborada, no entanto, G. melanops é um ramo basal do clado que inclui as espécies de Fulica, indicando que uma mudança taxonômica é necessária. A relação entre as espécies de Rallus e Pardirallus é distante e não suporta a inclusão das espécies de Pardirallus em Rallus. As maiores discordâncias da filogenia proposta em comparação com estudos moleculares referem-se à posição interna dos membros de Porphyrio e suas relações com outros gêneros / The family Rallidae is represented by cosmopolitan birds commonly known as wood-rails, crakes, coots, gallinules and swamp hens. It comprises around 135 species, distributed in 33 genera, of which 13 are monotypics. The phylogenetic relationships based on morphological characters and molecular parameters indicate different affinities among family species, mainly the position of the genera Rallus, Porphyrio, Gallinula and Fulica. In this study, focused on the Neotropical species of the family, a new phylogenetic analysis based on osteological characters is proposed. A sample of 100 skeletons of 13 genera and 31 species was analyzed. A total of 50 characters was codified, of which 17 were cranial and 33 post-cranial to provide a matrix construction and a subsequent phylogenetic analysis according to the principle of parsimony. A strict consensus and a majority rule consensus tree were calculated. The former generated 151 equally parsimonious trees with 99 steps. The successive weighting approach analyses of characters obtained better resolutions around the sampled species. The cladogram topology allows the acceptance of some genera as valid monophyletic groups, such as Rallus, Porphyrio, Aramides, Gallinula and Fulica. The position of Porphyrio as a basal branch within the subfamily Rallinae was supported and the interspecific relationships show that New World species were more closely related than P. porphyrio, allowing the taxonomic inclusion of Porphyrula. The close relationship between the species of Gallinula and Fulica was corroborated, but G. melanops is a basal branch of a clade that includes Fulica species, indicating that a taxonomic change is needed. The relationship of Rallus and Pardirallus is distant and so does not support the inclusion of Pardirallus species in Rallus. The major discordances of the proposed phylogeny in comparison with molecular studies concern the internal position of the Porphyrio members and their relationships with other genera.

Philosophiae Doctor - PhD / The Ruschieae is the most diverse and speciose tribe within the large subfamily Ruschioideae (Aizoaceae), with approximately 71 genera and a distribution centred in the arid parts of the Greater Cape Floristic Region (GCFR) of South Africa. Recent phylogenetic analyses provided the first insights into generic relationships within the tribe, with a number of novel generic relationships discovered. The tribal phylogeny recovered 12 large clades, of which the Conophytum-clade was one the most morphologically diverse based on leaf and capsule characters. The Conophytum-clade is an early-diverging lineage of the Ruschieae and includes the following 10 genera: Cheiridopsis N.E.Br., Conophytum N.E.Br., Enarganthe N.E.Br., Ihlenfeldtia H.E.K.Hartmann, Jensenobotrya A.G.J.Herre, Namaquanthus L.Bolus, Octopoma N.E.Br., Odontophorus N.E.Br., Ruschianthus L.Bolus and Schlechteranthus Schwantes. The present study presents an expanded phylogenetic analysis of the Conophytum-clade, with the sampling of the majority of species in the genera and a representative sampling (56% of species) of the speciose genus Conophytum. Phylogenetic data for up to nine plastid gene regions (atpB–rbcL, matK, psbJ–petA, rpl16, rps16, trnD– trnT, trnL–F, trnQᶷᶷᶢ–rps16, trnS–trnG) were produced for each of the sampled species. The produced plastid data was analyses using maximum parsimony, maximum likelihood and Bayesian inference. The combined plastid phylogenetic analyses were used in combination with morphological, anatomical and palynological data to assess generic and subgeneric circumscriptions within the clade. Upon assessment of generic circumscriptions in the Conophytum-clade, the number of recognised genera in the clade decreased from ten to seven. Arenifera A.G.J.Herre, which had not been sampled in any phylogeny of the Ruschieae, and Octopoma were recovered as polyphyletic, with species placed in the Conophytum-clade, while the type species was placed in the xeromorphic clade of the tribal phylogeny. The species of Arenifera and Octopoma placed in the Conophytum-clade were subsequently included in Schlechteranthus upon assessment of generic circumscriptions between the taxa. Two morphological groupings were recognised within Schlechteranthus, one including the species of Schlechteranthus and the other including species previously recognised as Arenifera and Octopoma. These two morphological groupings were treated as subgenera, with the erection of the new subgenus Microphyllus R.F.Powell. A detailed taxonomic revision of subgenus Microphyllus is presented with a key to species, descriptions of the species (including a new species: S. parvus R.F.Powell & Klak), known geographical distributions and illustrations of the species. In addition to the changes mentioned above, the expanded sampling and phylogenetic analyses of the Conophytum-clade recovered Ihlenfeldtia and Odontophorus embedded in Cheiridopsis. The species of Ihlenfeldtia were recovered with species of heiridopsis subgenus Aequifoliae H.E.K.Hartmann, while the species of Odontophorus were recovered as polyphyletic within the Cheiridopsis subgenus Odontophoroides H.E.K.Hartmann clade. Cheiridopsis was subsequently expanded to include the species of Ihlenfeldtia and Odontophorus, with these species accommodated in the subgenera of Cheiridopsis. The phylogenetic placement and relationship of these species was supported by the shared capsule morphology. The expanded sampling of the clade did not resolve the phylogenetic relationship of the monotypic genera Enarganthe, Jensenobotrya, Namaquanthus and Ruschianthus, with these genera unresolved in the Conophytum-clade. These genera however, exhibit a unique combination of morphological characters, such as a glabrous leaf epidermis and variation in pollen exine and colpi structure, in contrast to the other genera of the clade. The assessment of the generic circumscription of these genera, based on the molecular, morphological, anatomical and palynological data suggested that the generic statuses of these monotypic genera should be maintained. The expanded phylogenetic sampling of the morphologically diverse and speciose genus Conophytum recovered the genus as monophyletic. This monophyly was supported by the unique floral type in Conophytum, with the fused petaloid staminodes forming a tube. None of the sectional classifications were recovered as monophyletic but the phylogenetic analyses did recover a few clades which more or less corresponded to the current sectional classification of the genus. A number of clades were also recovered which included species from a range of different sections. Diverse leaf and floral traits were shown to have evolved numerous times across the genus. This was particularly interesting with regards to the selected floral traits, as the phylogeny indicated a number of switches in floral morphologies across the genus. The floral diversity was assessed in complex species communities of Conophytum across the GCFR, where up to 11 species of Conophytum are found occurring sympatrically, and floral traits were shown to be different across the species within the communities. Pollination competition and adaptation were suggested as possible drivers of floral diversity in the genus, with differences in phenology, anthesis and floral morphology within the species complex communities. The unique floral type of Conophytum has enabled the species to develop a diverse range of specialised flowers, with a variety of structures, scents and colours, resulting in the diverse floral morphologies found across the genus. The complex Conophytum species communities included both closely as well as distantly related species, suggesting the soft papery capsules of Conophytum are wind dispersed. This adaptation to long distance seed dispersal resulted in a significantly higher phylogenetic diversity in Conophytum when compared to its sister genus, Cheiridopsis. A population genetics study of Conophytum also suggested that the capsules may be wind dispersed, with an indication of genetic connectivity between the geographically isolated populations of C. marginatum Lavis across the Bushmanland Inselberg Region. Although the capsules are dispersed by wind, the seeds are released from the hygrochastic capsules by runoff during rainfall events. The relationship between seed dispersal and runoff is evident from the genetic structure of populations of C. maughanii N.E.Br. and C. ratum S.A.Hammer that occur on the tops and the surrounding bases of the inselbergs, as the drainage pattern was found to directly influence population structure in these species. In addition, the AFLP analyses provided insight into the conservation of the flagship species C. ratum. The summit populations of this species were shown to sustain the populations at the base of the Gamsberg. This finding is especially important, as the distribution of the species is restricted to the Gamsberg inselberg, where mining has already commenced as of this year. / National Research Foundation (NRF)

Members of the Mylagaulidae have been known for over a century to bear nasal horns; the only rodents, extinct or extant, ever to have done so. This striking feature is known from five of the over 30 species of mylagaulid rodents discovered across North America and Eurasia, all relatively large animals that were likely less fossorial than their relatives. We describe herein a sixth new species of horned mylagaulid. This new taxon from Sioux County, Nebraska, offers the opportunity to reassess the phylogenetic relationships of Mylagaulidae and test several evolutionary hypotheses. Our analyses demonstrate that horns evolved only once in Mylagaulidae, in the common ancestor of Ceratogaulus, first as short horns exapted from the thickened nasals of fossorial ancestors, and later as taller horns. The horns evolved following a positive allometric scaling with body mass that suggests a response to predation pressure in these nearly blind animals. The evolution of tall horns also corresponds to a jump in body mass. The largest mylagaulids are not horn-bearing species, however. Additional analyses of the complex pattern of body mass evolution we reveal will be necessary to explain the evolution of the largest head-lift digging rodents in Earth history. https://zoobank.org/urn:lsid:zoobank.org:pub:81FE999A-F79E-4BD4-9A81-2C7D3D5D81CD.

The phylogenetic relationships of Pseudophryne and the closely related monotypic Metacrinia are resolved with the use of morphological characters and comparison with representatives species of Crinia, Uperoleia, and Taudactylus as defined out-groups. Characters describing musculature are not sufficient to resolve the relationships, but do provide support when used in combination with osteological and external characteristics. When all data are considered, parsimony and maximum likelihood analyses yield the same hypothesis of relationships within Pseudophryne + Metacrinia. Thus, a monophyletic lineage of Pseudophryne + Metacrinia is supported by four synapomorphies: (1) the absence of toe fringes, (2) wide frontoparietal fontanelle, (3) m. abductor indicis longus arising from the humerus and radioulna, and (4) neopalatine in contact with the maxilla. The columella is shown as a relatively plastic characteristic, present ancestrally but lost in the common ancestor to Pseudophrye+Metacrinia and then reappearing within that taxon as well as one outgroup species (Crinia riparia). Analyses of size data reveal shape trends among the outgroup taxa that differ from the in-group of Pseudophryne and Metacrinia. Interpretations of shape differences are congruent with the placement of Uperoleia as more closely related to Pseudophrye. This study suggests support for re-synonymy of Metacrinia nichollsi with Pseudophryne, but formal change of status depends on access to more complete data.

Fungi are vital microbes present throughout the biosphere. Many species are essential decomposers in the ecosystem, breaking down organic materials and nourishing other lives. Moreover, some have directly influenced human civilization by providing beneficial products, such as edible mushrooms, brewer's yeast, baker's yeast, and antibiotics. However, it's important to note that this group of organisms can also have a "dark side". Each year, fungal pathogens cause approximately 150 million severe infections and 1.7 million deaths. The high rate of infection is compounded by the limited availability of antifungal drugs and the increasing prevalence of antifungal resistance. In response to the global burden of fungal diseases, the World Health Organization published a list of priority fungal pathogens in 2022 and highlighted strategies such as surveillance, sustainable research investments, and public health interventions to combat the increasing fungal threats. My PhD research has focused on surveillance and genomic analyses of several human fungal pathogens, particularly Candida auris. Candida auris is an emerging multidrug-resistant yeast that causes systemic infections with high mortality rates. While initially recognized as a nosocomial pathogen, our genomic analyses of strains isolated from clinical environments, tropical wetlands, fruit surfaces, and dog ears revealed potential transmission routes between diverse environments and patients, including a potential driver for the prevalence of antifungal resistance. Furthermore, our research indicated limited genetic exchange within and between lineages of Candida auris. Through genome-wide association analyses of global Candida auris strains, several known and novel genomic variants were identified associated with susceptibility to azoles, echinocandins, and amphotericin B. Overall, our studies underscore the importance of continuous surveillance to understand potential routes of Candida auris transmission and the urgent need for innovative approaches to treat multidrug-resistant Candida auris infections. / Thesis / Doctor of Philosophy (PhD)

No description available.

Thesis (MSc)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: The Afroedura nivaria complex is one of the six recognized species complexes within a southern African endemic genus, Afroedura. The A. nivaria complex is a morphologically conservative group of medium-sized geckos endemic to South Africa though they are unevenly distributed in the Eastern Cape, Free State and KwaZulu-Natal provinces. The complex comprises the following five species: A. nivaria (Boulenger 1894), A. amatolica (Hewitt 1925), A. karroica (Hewitt 1925), A. tembulica (Hewitt 1926) and A. halli (Hewitt 1935). These nocturnal and rupicolous geckos shelter in narrow rock crevices on outcrops. It is currently unknown whether a) the described species are valid and b) if additional lineages are present on isolated outcrops. I investigated the hypothesis that endemics with a narrow distribution, that is, A. amatolica and A. tembulica are valid species but that isolated populations in the widespread species (A. nivaria, A. karroica and A. halli) demonstrate genetic variation at the species level. Fragments of two mitochondrial genes (16S rRNA and ND4) and a single nuclear marker (KIAA) were sequenced and analysed using Bayesian inference, maximum parsimony and maximum likelihood. All analyses strongly supported the genetic distinctiveness of the described species. The A. nivaria complex is not monophyletic, A. karroica appeared to be outside the species complex and A. pondolia (thought to be outside the A. nivaria complex) consistently nested within A. nivaria complex. Additional clades recovered in the phylogeny within A. halli and A. nivaria had large genetic divergences and no spatial overlap. Narrowly distributed A. amatolica showed to have two highly diverged clades. Clades recovered in the phylogeny highlight geographical structuring. These findings suggest the existence of up to four additional cryptic lineages within the complex. I used morphometric data (ecologically relevant morphological traits) to investigate whether the genetic lineages would present morphological conservatism. Multivariate analyses of 19 variables showed variation within the A. nivaria species complex was accounted for mostly by differences in locomotor apparatus (limbs and feet) and head dimensions. These traits are mostly related to microhabitat usage and/or dietary specialization in lizards. There were no significant differences for body dimensions between species within the complex, indicative of morphological conservatism. It appears genetic divergence has been achieved among the different clades within A. nivaria complex, but with much similarity in phenotype being retained because of fragmented but similar habitats occupied. / AFRIKAANSE OPSOMMING: Die Afroedura nivaria kompleks is een van ses herkende spesies komplekse binne die endemiese suidelike Afrika genus, Afroedura. Die A. nivaria kompleks is ‘n morfologiese konserwatiewe groep bestaande uit medium grootte geitjies endemies tot Suid Afrika, alhoewel hulle oneweredig verspreid is in die Oos Kaap, Vrystaat en Kwazulu-Natal provinsies. Die kompleks bestaan uit die volgende vyf spesies: A. nivaria (Boulenger 1894), A. amatolica (Hewitt 1925), A. karroica (Hewitt 1925), A. tembulica (Hewitt 1926) and A. halli (Hewitt 1935). Hierdie geitjies kom snags voor en skuil tussen nou skeure op klip koppies. Dit is tans onbekend of a) die beskryfde spesies geldig is en b) of die addisionele afstammelinge voorkom op geisoleerde koppies. Met die studie het ek die hipotese ondersoek dat endemiese spesies met ‘n noue verspreiding (A. amatolica en A. tembulica) geldige spesies is, maar dat spesies met ‘n wye verspreiding (A. nivaria, A. karroica and A. halli) genetiese variasie op spesie vlak wys. Fragmente van twee mitochondriale gene (16S rRNA and ND4) en ‘n enkele nuklêre merker (KIAA) se basispaaropeenvolgingsdata was verkry en geanaliseer deur Bayesian inferensie, maksimum parsimonie en maksimum waarskynlikheid. Alle analise het die genetiese kenmerkendheid van die beskryfde spesies sterk ondersteun. Die A. nivaria kompleks is monofileties, A. karroica het geblyk om buite die spesies kompleks voor te kom en A. pondolia (voorheen beskryf as buite die A. nivaria kompleks) het voortdurend binne die A. nivaria kompleks voorgekom. Addisionele klades afkomstig vanaf die filogenië van A. halli en A. nivaria het vir beide spesies groot genetiese divergensie met geen ruimtelike oorvleuling gewys. Afroedura amatolica, met sy noue verspreiding, het twee hoogs divergente klades getoon. Die klades onthul deur die filogenie beklemtoon ‘n geografiese struktuur. Hierdie bevindings blyk die bestaan van tot vier ekstra kriptiese afstammelinge binne die kompleks. Ek het morfometriese data (ekologiese relevante morfologiese eienskappe) gebruik om vas te stel of die genetiese afstammelinge morphologies konserwatief sal wees. Meerveranderlike analises op 19 veranderlikes het variasie binne die A. nivaria spesies kompleks getoon. Hierdie veranderinge was meestal gevind in die beweeglikheidsapparatuur (ledemate en voete) en kop dimensies. Die verskeie eienskappe hou meestal verband met die mikrohabitatte wat gebruik word en/of dieët spesialisering in akkedisse. Daar was geen noemenswaardige verskille in liggaamsdimensies tussen spesies in die kompleks nie, beduidend op ‘n konserwatiewe morfologie. Dit wil blyk of genetiese divergensie tussen die verskeie klades van die A. nivaria kompleks bewerkstellig is met ooreenstemming in die fenotipes as gevolg van gefragmenteerde maar soortgelyke habitat verbruik.

Thesis (PhD)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Grapevine fanleaf virus (GFLV) is one of the oldest, most widespread and devastating viruses infecting grapevine, and occurs globally where Vitis vinifera is grown. In South Africa (SA) GFLV is predominant in the Breede River Valley, one of the highest wine producing regions in SA. To date, only three GFLV isolates have been completely sequenced internationally, and limited sequence information is available for SA GFLV isolates. In this study, the first full-length GFLV genome sequence from a South African isolate, GFLV-SAPCS3, was determined. Full-length sequences were used for phylogenetic analysis and revealed that the SA isolates are separate from other sequenced GFLV isolates. Full-length sequences were also used to investigate putative intra- and interspecies recombination events involving GFLV-SAPCS3 RNA1 and RNA2 between GFLV and Arabis mosaic virus (ArMV) isolates. Using two different recombination analysis software packages, the most notable of the putative recombination events involving GFLV-SAPCS3 indicated that the GFLV-SAPCS3 RNA2 5’ UTR might have evolved from an interspecies recombination event between GFLVF13- type and ArMV Ta-type isolates. The presence of satellite RNAs (satRNA) associated with South African GFLV isolates was also investigated. In a collaborative study (see Chapter 4 for details), more than a 100 GFLV- infected grapevine plants were screened for satRNAs. SatRNAs were present in only two plants, containing isolates GFLV-SACH44 and GFLV-SACH47. The full-length nucleotide sequences of the GFLV-SACH44 genomic RNAs 1 and 2, and the associated satRNA were determined. No significant sequence variation could be detected between the GFLV isolates that had the presence of a satRNA and those that had not. The GFLV-SACH44 RNA2 5’ UTR also had the same conserved sequence that was found in GFLVSAPCS3, which suggests that GFLV-SACH44, like GFLV-SAPCS3, may have arisen from a common ancestor, which may have originated from an interspecies recombination event. The GFLV-SACH44 satRNA was found to be more closely related to the ArMV large satRNA than to the satRNA associated with GFLV-F13. A full-length cDNA clone of GFLV-SACH44 satRNA was constructed and its replication and systemic spread in herbaceous hosts, when mechanically co-inoculated with two GFLV isolates as helper viruses, was demonstrated. Replication of the GFLV-SACH44 satRNA cDNA clone was however abolished when co-inoculated with an ArMV helper virus, even though it is phylogenetically more closely related to ArMV satRNAs. The full-length satRNA clones were modified to be used as vectors for expression and/or silencing of foreign genes, by inserting the green fluorescence protein (GFP) full-length or partial sequences downstream of the open reading frame of the satRNA. These constructs were cloned into a binary vector to allow for agro-infiltration into plants. Full-length cDNA clones of GFLV-SAPCS3 RNA1 and RNA2 were constructed to be used in conjunction with modified GFLV-SACH44 satRNA full-length clones. The full length GFLV-SAPCS3 RNA1 and RNA2 clones were however not infectious in Nicotiana benthamiana after agro-infiltration and therefore the evaluation of the modified satRNA expression and silencing constructs had to be aborted. Attempts to understand this failure revealed that, among other point mutations, four frameshifts had occurred in the RNA1 full-length clone, rendering the transcripts untranslatable, and hence noninfectious. Strategies to correct the mutations are discussed. Once these mutations have been corrected this study can continue in evaluating the use of the satRNA component for expression and silencing analysis. / AFRIKAANSE OPSOMMING: Grapevine fanleaf virus (GFLV) is een van die oudste, mees wydverspreide en mees verwoestende virusse wat wingerd affekteer en word wêreldwyd waar Vitis vinifera verbou word, gevind. In Suid Afrika (SA) kom GFLV veral in die Breederivier vallei, een van die mees produktiewe wyn-produserende areas in SA, voor. Tot dusver is daar net drie GFLV isolate waarvan die volledige nukleïensuurvolgorde internasionaal bepaal is. Die nukleïensuurvolgorde informasie vir SA GFLV isolate is redelik beperk. In hierdie studie was die eerste volledige nukleïensuurvolgorde van ‘n SA GFLV isolaat, GFLVSAPCS3, bepaal. Die volledige nukleïensuurvolgordes was vir filogenetiese analise gebruik, en vermeende intra- en interspesie rekombinasie gebeurtenisse, wat GFLVSAPCS3 RNA1 en RNA2 betrek, tussen GFLV en Arabis mosaic virus (ArMV) isolate was ondersoek. Twee verskillende rekombinasie-analise sagteware programme was gebruik en die noemenswaardigste van die vermeende rekombinasie gebeurtenisse, met betrekking tot GFLV-SAPCS3, het aangedui dat die GFLV-SAPCS3 RNA2 5’ ontransleerde streek (UTR) waarskynlik van ‘n interspesie rekombinasie gebeurtenis tussen ‘n GFLV-F13-tipe en ‘n ArMV-Ta-tipe isolaat ontwikkel het. Die teenwoordigheid van satelliet RNAs (satRNAs), wat met SA GFLV isolate geassosieer is, was ook ondersoek. Meer as ‘n 100 GFLV ge-infekteerde wingerd plante was in ‘n samewerkingsprojek (sien Hoofstuk 4 vir besonderhede) getoets vir die teenwoordigheid van satRNAs. SatRNAs was net in twee plante teenwoordig, in isolate GFLV-SACH44 en GFLV-SACH47. Die vollengte nukleïensuurvolgordes van GFLVSACH44 RNA1, RNA2 en geassosieerde satRNA was bepaal. Geen beduidende volgorde variasie tussen die GFLV isolate wat satRNAs bevat het, en die GFLV isolate sonder satRNA was waargeneem nie. Die GFLV-SACH44 RNA2 5’ UTR het ook die gekonserveerde volgorde, wat in GFLV-SAPCS3 teenwoordig was, gehad en dit dui daarop dat GFLV-SACH44, soos GFLV-SAPCS3, van dieselfde stamvader, wat tydens ‘n vorige rekombinasie gebeurtenis ontstaan het, mag ontwikkel het. Die GFLVSACH44 satRNA was meer naverwant aan die ArMV satRNAs as aan die satRNA, wat met GFLV-F13. ‘n Vollengte cDNA kloon van die GFLV-SACH44 satRNA was ontwikkel en die replisering en sistemiese verspreiding in sagte plante, nadat dit met twee GFLV isolate as helper virusse saam ge-inokuleer was, was gedemonstreer. Replisering van die GFLV-SACH44 satRNA cDNA kloon was egter ontwrig toe dit saam met ‘n ArMV helper virus saam ge-inokuleer was, al is dit filogeneties meer verwant aan ArMV satRNAs. Die vol-lengte satRNA klone was gemodifiseer om as vektore vir uitdrukking en/of uitdowing van transgene te dien, deur om vol-lengte of gedeeltelike groen fluoressensie proteïen (GFP) nukleïensuurvolgordes aan die einde van die satRNA leesraam te koppel. Hierdie konstrukte was in ‘n binêre vektor gekloon om agroinfiltrasie in plante toe te laat. Vol-lengte cDNA klone van GFLV-SAPCS3 RNA1 en RNA2 was ontwikkel om in samewerking met die gemodifiseerde GFLV-SACH44 satRNA konstrukte gebruik te word. Die vol-lengte GFLV-SAPCS3 RNA1 en RNA2 klone het egter nie in Nicotiana benthamiana gerepliseer na agro-infiltrasie nie, daarom was die evaluasie van die gemodifiseerde satRNA konstrukte gestaak. Pogings om die mislukking te verstaan, het daarop gewys dat, behalwe punt mutasies, vier leesraam versteurings in die RNA1 vollengte kloon voorgekom het, wat ontransleerbare transkripte, en dus nie-repliserende konstrukte tot gevolg gehad het. Strategieë om die mutasies te korrigeer is bespreek. Sodra die mutasies gekorrigeer is, kan die studie voortgaan om te evalueer of die satRNA komponent vir uitdrukking en uitdowing analise gebruik kan word.

The entire stretch of the Ohio River is under fish consumption advisories due to contamination with polychlorinated biphenyls (PCBs). In this study, natural attenuation and biostimulation of PCBs and microbial communities responsible for PCB transformations were investigated in Ohio River sediments. Natural attenuation of PCBs was negligible in sediments, which was likely attributed to low temperature conditions during most of the year, as well as low amounts of available nitrogen, phosphorus, and organic carbon. Moreover, surface sediments were relatively oxidized, as indicated by the prevalence of aerobic bacteria such as beta- Proteobacteria, alpha-Proteobacteria, Sphingobacteria, and Nitrospira in 16S rRNA sediment clone libraries. On the other hand, several reductive dechlorinators were detected in sediments, including Dehalococcoides, Desulfitobacterium spp. which suggested that reductive dechlorination might be possible in sediments under certain biogeochemical conditions. Considerable amounts of PCBs were transformed by reductive dechlorination (80% in 177 days by pattern N) when sediments were maintained under anaerobic conditions, amended with nutrients and organic carbon, and incubated at 25 ºC in lab microcosms. Analysis of 16S rRNA clone libraries from these treatments revealed that Bacteroidetes, Chloroflexi and Firmicutes were enriched and Proteobacteria were depleted compared to clone libraries from treatment without organic amendments. Reductive dechlorination was decreased in sediments incubated at 10 and 40 ºC, and was not affected by FeSO4 amendments compared to unamended sediments incubated at 25 ºC. Transformations of PCB-153 were investigated in sediments under anaerobic, aerobic and sequential anaerobic and aerobic conditions. Transformations were only observed in treatments with an anaerobic phase, which occurred by reductive dechlorination by pattern N. Neither PCB-153 nor dechlorination products PCB-99 or PCB-47 were transformed under aerobic conditions. Analysis of 16S rRNA clone libraries revealed that Bacteoridetes, Chloroflexi, and Firmicutes were enriched under anaerobic conditions and Proteobacteria were enriched under aerobic conditions. Results from this study revealed that natural attenuation and biostimulation were not effective at removing PCBs from Ohio River sediments. Hence, other remediation methods will need to be employed to decrease PCB levels in this ecosystem.