Identification and phylogenetic analysis of Aedes species (Diptera: Culicidae) and arboviruses associated with them across tropical and temperate regions of South Africa (2015-2018)

Abstract

Emerging and re-emerging diseases have increased worldwide in incidence in the past decades. Of these emerging diseases 60.3% are caused by zoonotic pathogens of which 22.8% are arboviruses or arthropod borne viruses. Arboviruses are transmitted by hematophagous insects, especially moquitoes. Multiple factors such as human population growth, climate change and adaptations of certain Aedes mosquito vector species to urban environments and anthropophilic have been attributed to causing this rise in arboviral infections. In Southern Africa, zoonotic arboviruses belonging to the families Flaviviridae (genus Flavivirus), Togaviridae (genus Alphavirus), and those in the order Bunyavirales, family Phenuiviridae: (genus Phlebovirus) and Peribunyaviridae (genus Orthobunyaviruses), have proven, in the past, to be of both medical and veterinary importance. Recent detection of neurological cases in South Africa, most likely, due to flaviviruses, alphaviruses and orthobunyaviruses in the Simbu serogroup, has rekindled interest in these zoonotic diseases. This interest is also warranted because of lack of recent information on arboviral prevalence in mosquito species, distributions, abundance, and ecology, especially of Aedes species, the likely primary vectors of these arboviruses in Southern Africa. To update this lack of information, this study t reports on zoonotic arboviruses circulating in selected areas in the north-eastern provinces of South Africa in mosquitoes with a focus on Aedes. Many Aedes species are morphologically quite difficult to identify especially when they are old, and scales rubbed off in the process of trapping. To aid in the identification of Aedes in this study we provide molecular barcodes for Aedes species occurring in in South Africa and define their phylogenetic relationship with other mainly Afrotropical Aedes mosquitoes based on the cytochrome oxidase I gene sequences. The first Chapter provides a comprehensive review of the literature and describes the importance of arboviruses worldwide and in South Africa, highlighting the role of Aedes mosquitoes as vectors. In Chapter 2, what is known about the broad patterns of Aedes mosquito species diversity, abundance, and distribution in different habitats across selected sites in five different provinces in South Africa is described. The sites selected were chosen because of evidence of neurological cases in humans and animals in recent years likely due to arboviral infections. In total, 61,737 adult mosquitoes were collected from January 2014 to May 2018, using three kinds of carbon dioxide baited trap types About 16% (11,440) were Aedes species, of which, 14 species were recognised or suspected vectors of mosquito-borne diseases because of positive infections, including Aedes mcintoshi which was the most abundant Aedes species captured. The effect of the climatic conditions on the mosquito population dynamics were also investigated. Aedes species were present in the sites following the peak of the rainfall and were mostly captured in temperatures between 18°C and 27°C. Chapter 3 focuses on determining the blood meal source present in engorged Aedes mosquitoes sampled to give an assessment of blood feeding tendencies that would serve useful to determine their vector status. Aedes species were identified feeding on a broad range of livestock, and wildlife, only two specimens were identified as feeding on avian species.Chapter 4 focuses on interpretations of cytochrome oxidase subunit 1 (COI) gene sequences to identify Aedes species in South Africa and to analyse the relationship among the species. A total of 52 COI sequences were aligned representing 21 Aedes species. In several cases these were the first African aedine species uploaded in NCBI GenBank. Neomelaniconion species clustered together, except for Ae. aurovenatus. Finally, the data also suggested that Ae. cumminsii present in South Africa belongs to the subspecies ssp. mediopunctatus. In Chapter 5 results of arboviral infections in Culicidae mosquitoes captured from the selected sites, particularly Aedes species is provided. Arboviral infection or prevalence screening was performed using multiple genus specific polymerase chain reactions (PCR). Alphavirus and Orthobunyavirus were detected in different Culicidae genera, including Aedes, Culex, Anopheles and Mansonia. There were no isolations of pathogenic flaviviruses in mosquitoes. The only alphaviruses detected in mosquitoes were Middelburg, Sindbis and Ndumu viruses during the period of the study. Shuni virus was the only member of Orthobunyavirus genus, detected. Even though, the main aim was to identify pathogenic viruses, several insect-specific viruses belonging to Alphavirus and Flavivirus genera were also detected and these are described in Chapter 6. The numerous arboviruses detected in Culicine mosquitoes, including Aedes species, demonstrate that some species are likely maintaining natural cycling of these arboviruses. Noteworthy, is that mosquito species positive for arboviruses are often the most abundant in the selected sampling locations and that these species blood feed mostly on the larger vertebrates present in the area. Outbreaks possibly occur when the prevalence of certainmosquito species are high due to favourable climatic conditions. Highest arbovirus detections occurred in peri-urban, rural, and conservation areas, indicating that livestock and wildlife likely play an important role in the amplification of these arboviruses. This study highlights the importance of a continues mosquito-based surveillance for arboviruses in South Africa, and the role that Aedes species might be playing in the circulation of these arboviruses. Surveillance for the species that tested positive for pathogenic arboviruses during the arbovirus season may act as an early warning system and can also help to avoid spill over in animals and humans in the area surveyed.