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Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges
Dissertation (MSc (Genetics))--University of Pretoria, 2014.
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| Format: | Thesis |
| Language: | English |
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2026
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| _version_ | 1867613552037068800 |
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| access_status_str | Open Access |
| author2 | Fick, Wilma |
| author_browse | Fick, Wilma |
| author_facet | Fick, Wilma |
| collection | Thesis |
| description | Dissertation (MSc (Genetics))--University of Pretoria, 2014. |
| format | Thesis |
| id | oai:repository.up.ac.za:2263/110070 |
| institution | University of Pretoria (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:37:57.427Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UPSpace — University of Pretoria Institutional Repository |
| publishDate | 2026 |
| publishDateRange | 2026 |
| publishDateSort | 2026 |
| record_format | dspace |
| source_str | UPSpace — University of Pretoria Institutional Repository |
| spelling | oai:repository.up.ac.za:2263/110070 Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges Fick, Wilma u28022263@tuks.co.za Maritz-Olivier, Christine Ekoka, Elodie L.E. Culicoides RNA Interference Akirin African Horse Sickness Vaccine Dissertation (MSc (Genetics))--University of Pretoria, 2014. Culicoides midges are haematophagous vectors mainly known for the transmission of three veterinary and economically important animal diseases, namely African horsesickness, bluetongue and epizootic haemorrhagic disease. Currently, Culicoides control is based on environmental management and insecticides, with the first approach being labour-extensive and the second one resulting in environmental pollution and development of resistant Culicoides species. An attractive vector control strategy which is cost-efficient and less polluting involves the immunisation of vertebrate hosts with recombinant vaccines targeting the midge specific antigens. To date, midge control through immunological-based approaches has never been investigated, although several vaccines have already shown great efficacies against ticks, mosquitoes or sand flies. One arthropod antigen target that has been intensively investigated is the conserved transcription factor akirin/subolesin (AKR/SUB). Preliminary data from vaccine trials revealed that AKR/SUB recombinant vaccines directed against ticks, mosquitoes or sand flies negatively affect the vector oviposition, fertility, haematophagy, survival and/or pathogen transmission. Based on these reports, we hypothesised that akirin was an antigen candidate worth investigating for the development of a Culicoides control vaccine, if the midges possess an akirin ortholog with functional similarities to the characterised akirin orthologs. Therefore, the objectives of this study were to investigate if an akirin ortholog is present in two Culicoides species, to characterise the gene and encoded protein on sequence level, and to use this ortholog as model to develop a platform for in vitro RNAi studies in the C. sonorensis KC cell line. An akirin ortholog was successfully identified in both C. imicola and C. sonorensis at transcriptome and genome levels. Notably, no introns were detected by comparing the sequences of the akirin genes to their encoding mRNAs. In silico sequence analyses revealed that both Culicoides akirin orthologs were hydrophilic soluble proteins, suggesting localisation in the nucleus. The protein contained the typical post-translational modifications observed for transcription factors. Furthermore, it was found that the midges’ akirin sequences were phylogenetically closer to that of mosquitoes compared to ticks. Interestingly, conserved linear B-cell epitopes were shared between ticks, mosquitoes, sandflies and Culicoides akirin/subolesin sequences. An investigation of the function of the midge akirin in the C. sonorensis KC cell line revealed that neither cell viability, proliferation nor morphology were affected upon gene knockdown, as analysed by an MTS viability assay and light microscopy. In follow-up experiments, the parameters for RNAimediated silencing of Culicoides genes were optimised in the KC cell line, via akirinspecific gene knockdown. The findings and methodology established here could be exploited in future for the functional characterisation of Culicoides genes, thereby allowing a better understanding of the molecular mechanisms underlying midge biology. Additional studies on the phenotypic effect of akirin knockdown in live midges may add Culicoides spp. to the list of vectors that are currently being investigated for control via an akirin-based multi-arthropod vaccine. Genetics MSc (Genetics) 2026-05-15T17:26:13Z 2026-05-15T17:26:13Z 14/11/27 2014 Dissertation http://hdl.handle.net/2263/110070 en application/pdf |
| spellingShingle | Culicoides RNA Interference Akirin African Horse Sickness Vaccine Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title | Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title_full | Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title_fullStr | Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title_full_unstemmed | Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title_short | Identification, sequence analyses and in vitro gene knockdown of the akirin ortholog of Culicoides midges |
| title_sort | identification sequence analyses and in vitro gene knockdown of the akirin ortholog of culicoides midges |
| topic | Culicoides RNA Interference Akirin African Horse Sickness Vaccine |
| url | http://hdl.handle.net/2263/110070 |