Published work submitted to the Faculty of Medicine, University of the Witwatersrand, Johannesburg, for the degree Doctor of Science in Medicine. / Andrew Chakane 2018

Dissertation (PhD)--University of Stellenbosch, 2006. / Bibliography. Also available via the Internet.

No presente trabalho procura-se caracterizar biológica e ecologicamente o Aedes albopictus (Skuse) (Diptera: Culicidae) no seu mais recente ambiente geográfico, isto é, nas Américas. São descritos aspectos relacionados com a chegada e dispersão da espécie no Continente Americano. Paralelamente, são discutidos aspectos relativos ao comportamento, biologia, bionomia, ecologia, hematofagia, interação com outras espécies do gênero Aedes, bem como a importância epidemiológica do Ae. albopictus. Esses aspectos foram descritos, por serem considerados importantes pela influência que exercem na competência vetora da espécie, bem como na sua adaptação ao ambiente antrópico. Ressaltando a importância epidemiológica deste vetor para a espécie humana, citam-se dados referentes à potencialidade do Ae. albopictus participar na transmissão de arboviroses. Assim, procura-se caracterizar a importância da possível participação do Ae. albopictus na veiculação do vírus dengue, febre amarela, bem como outros arbovírus que afetam o ser humano nas Américas. Paralelamente, discute-se a possibilidade do Ae. albopictus atuar como vetor da Dirofilaria immitis (Leidy) ao homem. / In the present work, biological and ecological features of Aedes albopictus (Skuse) (Diptera: Culicidae) are discussed based on literature records. The main focus of this study is the epidemiological importance of the presence of this species in Americas. Aspects relative to colonization and dispersion of Ae. albopictus in Americas are discussed. Dispersion, ethology, biology, ecology, haematophagy, bionomics as well as epidemiological importance, and relationships among Ae. albopictus and other members of the genus Aedes are discussed. These aspects are included because of their influence in the vector competence of Ae. albopictus, and also in its adaptation to the human environment. Because of the epidemiological importance of Ae. albopictus to humans, data relative to the potentiality of this species to participate in the transmission of arboviruses are discussed. In this sense, the importance of Ae. albopictus as vector of dengue, yellow fever, and other arboviruses that cause human diseases is described. Besides, it is discussed the potentiality of Ae. albopictus to act as a vector of Dirofilaria immitis (Leidy) to humans.

No presente trabalho procura-se caracterizar biológica e ecologicamente o Aedes albopictus (Skuse) (Diptera: Culicidae) no seu mais recente ambiente geográfico, isto é, nas Américas. São descritos aspectos relacionados com a chegada e dispersão da espécie no Continente Americano. Paralelamente, são discutidos aspectos relativos ao comportamento, biologia, bionomia, ecologia, hematofagia, interação com outras espécies do gênero Aedes, bem como a importância epidemiológica do Ae. albopictus. Esses aspectos foram descritos, por serem considerados importantes pela influência que exercem na competência vetora da espécie, bem como na sua adaptação ao ambiente antrópico. Ressaltando a importância epidemiológica deste vetor para a espécie humana, citam-se dados referentes à potencialidade do Ae. albopictus participar na transmissão de arboviroses. Assim, procura-se caracterizar a importância da possível participação do Ae. albopictus na veiculação do vírus dengue, febre amarela, bem como outros arbovírus que afetam o ser humano nas Américas. Paralelamente, discute-se a possibilidade do Ae. albopictus atuar como vetor da Dirofilaria immitis (Leidy) ao homem. / In the present work, biological and ecological features of Aedes albopictus (Skuse) (Diptera: Culicidae) are discussed based on literature records. The main focus of this study is the epidemiological importance of the presence of this species in Americas. Aspects relative to colonization and dispersion of Ae. albopictus in Americas are discussed. Dispersion, ethology, biology, ecology, haematophagy, bionomics as well as epidemiological importance, and relationships among Ae. albopictus and other members of the genus Aedes are discussed. These aspects are included because of their influence in the vector competence of Ae. albopictus, and also in its adaptation to the human environment. Because of the epidemiological importance of Ae. albopictus to humans, data relative to the potentiality of this species to participate in the transmission of arboviruses are discussed. In this sense, the importance of Ae. albopictus as vector of dengue, yellow fever, and other arboviruses that cause human diseases is described. Besides, it is discussed the potentiality of Ae. albopictus to act as a vector of Dirofilaria immitis (Leidy) to humans.

Potomac Horse Fever (PHF) is a disease of great concern to many horse owners in the Potomac River area of Maryland and Virginia. It is caused by a rickettsia, Ehrlichia risticii. The involvement of an arthropod vector has been suspected because of the seasonal epidemiology of the disease. This research was an attempt to identify and evaluate potential arthropod vectors. A seasonal activity study of biting arthropods attacking horses in endemic areas of Maryland and Virginia identified five potential vectors: (1) Simulium jenningsi (Diptera: Simuliidae), (2) Stomoxys calcitrans (Diptera: Muscidae), (3) Culicoides obsoletus (Diptera: Ceratopogonidae), (4) C. variipennis, and (5) Dermacentor variabilis (Acari: Ixodidae). These five arthropod species were given status as potential vectors because they were collected feeding on horses just prior to and throughout the PHF season. Simulium jenningsi and D. variabilis have the closest seasonal association with the occurrence of PHF as presented in this study. D. variabilis was determined to have the greatest potential due to its reported association with other rickettsial diseases. A series of laboratory and field studies were designed to examine the potential role of D. variabilis in the transmission of E. risticii. We first attempted to transmit E. risticii by feeding adult D. variabilis collected from an endemic farm on susceptible horses. Other laboratory studies included mouse to horse and mouse to mouse transmission attempts using ticks fed on mice inoculated with E. risticii. A serological survey of 105 trapped field rodents (host of immature D. variabilis) on endemic farms in Maryland showed all specimens collected to be negative for PHF antibodies. These studies and others gave no indication of D. variabilis's involvement in the transmission of the disease in nature. The other species mentioned above were not examined. / Ph. D.

L’empreinte écologique de l’homme sur les milieux naturels forestiers d’Afrique centrale n’a cessé de s’accroître au cours des dernières décennies, et rares sont les écosystèmes qui ne sont pas exploités. Ainsi, même des milieux a priori hostiles comme les grottes constituent des ressources répondant aux besoins primaires des populations environnantes (chasse, pêche, lieu de culte), mais permettant également le développement d’activités économiques (exploitation minière et écotourisme). Cette anthropisation est susceptible d’augmenter l’exposition des populations humaines à une multitude d'agents infectieux circulant au sein de la faune cavernicole. La présence d'animaux sauvages ou domestiques dans l'environnement immédiat des grottes contribue également à augmenter un tel risque. Parmi ces agents infectieux, certains peuvent être transmis par contact direct avec le réservoir animal, d’autres peuvent nécessiter l’intervention d’insectes hématophages. Le rôle de ces insectes dans l'épidémiologie de nombreux pathogènes est bien connu en Afrique en milieux épigés, mais demeure largement sous-investigué dans les environnements cavernicoles. Dans ce travail de thèse, nous nous sommes proposés de réponde à cette problématique. Dans six grottes du Gabon nous avons entrepris une étude de la diversité des diptères hématophages, en réalisant un inventaire taxonomique et l'étude de la dynamique temporelle des assemblages d'espèces. Dans un second temps, nous avons entrepris un criblage moléculaire d'agents infectieux parasitaires, bactériens et viraux en ayant recours à des techniques de détection par PCR nichée et séquençage à haut débit.Les travaux menés nous ont permis de réaliser un inventaire taxonomique de la faune diptérienne hématophage colonisant les grottes explorées. Notre étude a permis de découvrir une part importante de sa diversité de diptères hématophages cavernicoles, représentant plus de 60% des espèces de diptères (tout milieux confondus) actuellement au Gabon. Nos données ont montré que la composition des communautés de diptères était différente d'une grotte à une autre, et que ces dernières présentaient des variations au cours du temps en lien avec les variations microclimatiques des grottes. Le criblage d'agents infectieux chez les diptères collectés a permis 1) d'explorer la diversité parasitaire, virale et bactérienne qu'ils hébergent et 2) d'évaluer leur implication dans la transmission / The man's ecological mark on natural forest environments of central Africa has been increasing in recent decades, and only few natural areas remain non exploited yet. Thus, even supposedly hostile environments such as caves meet the primary needs of surrounding populations (hunting, fishing, shamanic practices), or economical needs (mining and ecotourism). That anthropization is likely to increase the exposure of human populations to multiple infectious agents carried by cave fauna. The presence of wild or domestic animals in the surroundings of caves also increase that risk. Among these infectious agents, some can be transmitted by direct contact with animal reservoirs, whereas others may require the involvement of blood-sucking insects. The role of these insects in the epidemiology of many pathogens is well known in Africa, particularly in epigeic environments, but remains largely investigated in caves. In this thesis work, we proposed to address that issue. In six caves of Gabon we firstly explored the diversity of blood-sucking Diptera, and we studied temporal dynamics of species assemblages. Secondly, we undertook a molecular screening of haemosporidia, bacteria and viruses using nested PCR and high-throughput sequencing.The work carried out has enable to make a taxonomic inventory of the Diptera fauna of the investigated caves. Our study revealed a significant diversity of the blood-sucking Diptera fauna, accounting for more than 60 percent of blood-sucking Diptera species currently known in Gabon, in all types of areas. Our data showed that the composition of Diptera communities was different from one cave to another, and that Diptera assemblages varied over time according to micro-climate fluctuations within caves. The screening of infectious agents in Diptera collected helped 1) to explore the parasitic, viral and bacterial diversity they host, and 2) to assess their involvement in transmission.

Thesis (PhD (Botany and Zology))--University of Stellenbosch, 2006. / The well-known, and often phytopathogenic, ophiostomatoid fungi are represented in South Africa by the two phylogenetically distantly related genera Ophiostoma (Ophiostomatales) and Gondwanamyces (Microascales). They are commonly associated with the fruiting structures (infructescences) of serotinous members of the African endemic plant genus Protea. The species O. splendens, O. africanum, O. protearum, G. proteae and G. capensis have been collected from various Protea spp. in South Africa where, like other ophiostomatoid fungi, they are thought to be transported by arthropod vectors. The present study set out to identify the vector organisms of Protea-associated members of mainly Ophiostoma species, using both molecular and direct isolation methods. A polymerase chain reaction (PCR) and taxon specific primers for the two Protea-associated ophiostomatoid genera were developed. Implementation of these newly developed methods revealed the presence of Ophiostoma and Gondwanamyces DNA on three insect species. They included a beetle (Genuchus hottentottus), a bug (Oxycarenus maculates) and a psocopteran species. It was, however, curious that the frequency of these insects that tested positive for ophiostomatoid DNA was very low, despite the fact that ophiostomatoid fungi are known to colonise more than 50% of Protea infructescences. Subsequent direct isolation methods revealed the presence of reproductive propagules of Ophiostoma spp. on four Protea-associated mite species (Oodinychus sp., two Tarsonemus spp. and Proctolaelaps vandenbergi). These mites are numerous within Protea infructescences and Ophiostoma spp. were isolated from a high frequency of these individuals. The Oodinychus sp. mite was found to vector most of the Protea-associated Ophiostoma species. It was thus postulated that the mites (in particular the Oodinychus sp.) act as primary vectors of the Protea-associated Ophiostoma species. The association between Oodinychus mites collected from P. repens and O. splendens proved to be mutualistic. Mites feeding on this fungus showed significantly higher population growth than mites feeding on any of the other fungal species tested. The short- and long-distance dispersal methods of these mites were also investigated. Firstly the ability of mites to move from drying infructescences to moist and sheltered areas such as provided by intact infructescences on the same plant was investigated experimentally. Significantly more mites were found to actively disperse from drying infructescences to artificially manufactured infructescences containing moistened filter paper shreds than to artificially manufactured infructescences containing dry filter paper shreds. The frequent fires associated with the habitat of these mites would, however, require movement over larger areas than what would be possible through self-dispersal. Dispersal of mites via air currents was thus investigated using sticky traps, but no Ophiostoma-vectoring mites were captured in this way. Self-dispersal aided by air currents could thus be ruled out, and our investigations shifted to vectored dispersal. Numerous insects emerging from Ophiostoma-containing P. repens and P. neriifolia infructescences were collected using specially designed emergence cages. Scanning electron microscopy and stereo-microscopy revealed that all three Ophiostoma-vectoring mite genera were phoretic on the beetle G. hottentottus. Tarsonemus spp. and P. vandenbergi were also phoretic on the beetles Trichostetha fascicularis and T. capensis associated with P. repens and P. neriifolia flowers. Mites collected from the surface of these beetles were found to vector reproductive propagules of various Ophiostoma spp. This thus seems to be the only method of long-distance dispersal of these mites and subsequently also the Protea-associated Ophiostoma species. Molecular phylogenetic reconstruction based on large subunit, ITS and beta-tubulin DNA sequence data suggests a polyphyletic origin for the Protea-associated members of Ophiostoma, which proposes multiple invasions of this unusual niche by these fungi. These studies also revealed the presence of four new species of Ophiostoma associated with Protea spp. The new species O. palmiculminatum, O. phasma, O. gemellus and Sporothrix variecibatus were thus described. Ophiostoma palmiculminatum is associated with P. repens infructescences and the Oodinychus mites collected from them. Ophiostoma phasma was collected from various Protea and mite species. Ophiostoma gemellus and Sporothrix variecibatus were initially only isolated from mites, but have subsequently also been isolated from Protea spp. The present study clarifies many aspects pertaining to the phylogeny and ecology of the interesting members of Ophiostoma associated with Protea hosts. As such this study will form the platform for further studies on the co-evolution of these insect / mite / fungi / plant associations.

Made available in DSpace on 2015-03-26T13:30:51Z (GMT). No. of bitstreams: 1 texto completo.pdf: 408527 bytes, checksum: 3f5ff39d24998982838ce196768d85c4 (MD5) Previous issue date: 2011-02-18 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Predators capture, kíll and consume theír prey. Thís ís the only ap- proach consídered ín rnost studíes Of predator-prey ínteractíons. However, sorne recent studíes have been concerned Wíth the índírect effects Of pre- datíorL When threatened, prey can change theír behavíor, thus reducing theír chances Of death. Frorn thís perspectíve, experíments performed wíth rnosquítoes specíes ín aquatíc envíronments shows that, ín several ínstances, preys are able to ídentífy the presence Of predators. Thís perceptíon leads to behavíoral alteratíons such as fernale ovíposítíon repellency and reduced foragíng 1arvae. However, the type Of sígnal perceíved by the prey, the írn- portance Of índuced responses to the survíval Of índívíduals, and the reasons for these evolve ín sorne species and not others, remain under díscussíon. The objectíve Was to deterrníne Whether chemícal cues of predator larva Toxorhyn- chites theobaldi (Díptera: Culícídae) are capable Of ínñuencíng the behavíor Of fernales and ímmatures ín three prey species Of the sarne farníly, although wíth Very dístínct behavíor (Culea: molli8, Limatus durhamii and Aedes al- bopictu5). We found that, the ínfusíon Where T. theobaldi larvae Was shel- tered, probably releasíng chernícal sígnals, had a lower proportíon of larvae (5.4 %) of C. molli5, cornpared Wíth control (95.6 %), índícatíng ovíposítíon repellency. Meanwhíle, the other two specíes ovíposíted índíscrímínately on ínfusíon treated and controL We also observed that the presence of traces of the sarne predator, C. mollis larvae spend rnost of theír tíme ín rest (82.7 compared Wíth no sígns of T. theobaldi (49.3 %). However. L. durhamii and A. albopictus larvae do not show behavíoral changes ín the presence of the sarne sígnals. Yet, C. mollis larvae ís preyed upon ín a smaller proportíon than the others, suggestíng that these changes ínñuence ín species Vulnera- bílíty. Frorn these results, ít ís argued that the líkelíhood of the development of defensive responses índuced by predatíon rísk ís related to several factors such as hístoríc of contact Wíth the predator, the ovíposítíon strategy of the females and the ñxed and ñexíble behavíor standard and posítíoníng of 1ar- Vae. Also ís díscussed the ímportance of natíve predators to curb the ínvasíon of exotíc species and díseases spread by these Vectors. / Predadores capturam, abatem e consomem suas presas. Esta é a única abordagem considerada na maior parte dos estudos da ínteração predador- presa. Entretanto, alguns trabalhos recentes têrn se preocupado com os efeitos indiretos da predação. Quando ameaçadas, presas podem alterar seu comportamento, reduzindo suas chances de morte. Sob essa perspectiva, experimentos realizados com espécies de mosquitos em ambientes aquáticos demonstram que, em diversas situações, presas são capazes de identificar a presença de predadores. Tal percepção induz alterações comportamentais como a repelência de ovíposição das fêmeas e redução no forrageamento das 1arvas. Entretanto, o tipo de sinal percebído pelas presas, a importância das respostas induzidas para a sobrevivência dos indivíduos, e as razões destas evoluírem ern algumas espécies, e não em outras, permanecem em discussão. O objectivo do trabalho foi verificar se pistas químicas da presença larva predadora Toxorhynchites theobaldi (Díptera: Culícídae) influenciam o comportamento tanto de fêmeas ovípositantes quanto de imaturos de três espécies de presas da mesma famílía, entretanto com comportamentos bastante distintos, sendo elas Culex molli5, Limatus durhamii e Aedes albopictus (Díptera: Culícídae). Verificamos que, na infusão onde larvas de T. theobaldi permaneceram abrigadas previamente, liberando possíveis sinais químicos, foi encontrada uma menor proporção de larvas (5,4%) de C. molli5, quando comparado com o controle (95,6%) , indicando repelência na ovíposição Enquanto isso, as outras duas espécies ovíposítaram indiscriminadamente na infusão tratada e na controle. Observamos tarnbérn que, na presença de indícios desse mesmo predador, imaturos de C. mollis passaram maior parte do tempo ern repouso (82,7%), quando compadrado com a ausência de sinais de T. theobaldi (49,3%). Entretanto, ímaturos de L. durhamii e A. albopic- tus não apresentam alterações comportamentais na presença dos mesmos sinais. Ainda, C. mollis é predado em uma menor proporção que as demais, sugerindo que essas alterações influenciam na Vulnerabilidade das espécies. A partir desses resultados, é discutido que a probabilidade da evolução de respostas defensivas induzidas pelo risco de predação está relacionada a diversos fatores como o histórico de contato com o predador, a estratégia de ovíposição das fêmeas e o padrão fixo e flexicível de comportamento e posícionarnento das 1arvas. É abordada também a importância dos predadores nativos em conter a invasão de espécies exóticas e a disseminação de doenças por elas Veiculadas.