Faculty of Science

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Browsing Faculty of Science by Subject "Aedes aegypti"

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    Improving mosquito (Aedes aegypti) sex-sorting methods for sterile insect technique using RNAi gene knockdowns
    (2023-04) Rempel, Kadri
    Doublesex (dsx) is an alternatively spliced mosquito (Aedes aegypti) gene that controls sex development by producing male (DSXM) or female (DSXF) transcription factors that regulate gene expression in a sex-specific manner. As both DSXF and DSXM bind the same DNA sequence, their ability to regulate differential gene expression is predicted to be modulated by other proteins that interact with DSX. A previously conducted protein-protein interaction study identified several proteins that bind to DSXF and their role in altering female development was explored in this study by knocking down their corresponding transcripts using RNA interference during the larval stages of development. Knockdowns of the genes nop-14, wdr-48, and rnmt was achieved by feeding mosquito larvae HT115 strain E. coli transformed with a pl4440 plasmid containing a dsRNA template specific to each of three target genes. The knockdowns were carried out at their normal rearing temperature, 28oC, and at 22oC to slow mosquito development. Knockdown of one of the genes, wdr-48, at either temperature, significantly increased the time to pupation of females compared to males. This delayed development of females could prove useful in the sex sorting of males and females during the pupal stage by providing a method to select only males for a sterile male insect (SIT) population control program for Ae aegypti. Of note, the 22oC growth conditions provided a larger difference in the pupation times between the sexes than the 28oC conditions did, and thus may provide even better sex-sorting for mosquito SIT.
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    Sex-sorting Aedes aegypti for sterile insect technique (SIT) by knocking down four different gene targets, expressed in the guts of female larvae
    (2023-04) Felix, Sharon
    Mosquitoes are vectors for diseases that affect approximately 0.7 billion people globally. Most of these diseases have a high incidence in tropical often developing countries, where they prove either fatal or severely debilitating. Despite huge investments of time and money to find solutions, we have been unable to control this disease vector. Additionally, some of the techniques previously and currently in use to prevent these diseases, have serious drawbacks such as detrimental effects on non-target species, humans and the environment. We need eco-friendly alternatives to these harmful chemicals, that are just as efficient while also being species-specific to avoid harmful effects on other beneficial insect species. Sterile insect technique (SIT) is a creative technology that does just that. It involves the mass release of sterile males of a species to outcompete wild males and mate with wild females, eventually resulting in the decline of a targeted insect population. However, before these sterile males are released, they must be separated from females. An efficient way to sex-sort males from females can use RNA interference (RNAi). RNAi is a natural defence mechanism that exists in eukaryotic cells, that can be used to knock down female-biased or female-specific genes. Targeted females will develop slower than their male counterparts due to this knockdown and this will enable their separation from a pool of male individuals. For my honours thesis, I targeted four such female-biased genes - AAEL014604, AAEL014797, AAEL009313 and AAEL005884 and measured development (in days), survival and pupal size for each of these treatments for both male and female mosquitoes. None of my treatments resulted in significant results but one of these (AAEL014797) showed great promise for an improved SIT.
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    The importance of discs large homolog 5 and microRNA-34 expression to the development of the mosquito Aedes aegypti
    (2023-04) Carroll, Lara; Wilkins, Olivia (Biological Sciences); Whyard, Steve
    Understanding how the expression of particular genes and regulation of genes affects development in different organisms is key to advancing our knowledge in developmental biology. The epithelial-mesenchymal transition (EMT) process is commonly observed during the growth and development of various animal tissues. One gene involved in EMT regulation and directly related to development in humans is discs large homolog 5 (dlg5). If the role of this gene is conserved across species, it opens many medical application possibilities to help treat a multitude of medical issues, such as cancer. microRNAs are short, single-stranded non-coding RNAs that regulate gene expression by binding to the 3’UTR of complementary target mRNAs. microRNA-34 (miR-34) is a microRNA of interest with relation to dlg5 as it is predicted to bind with dlg transcripts. miR-34 has also shown clinical potential in the past with regulating cell proliferation. This study looks at the dlg5 gene and miR-34 in the mosquito Aedes aegypti to see if the expression of this gene and microRNA is evolutionarily conserved to that of Drosophila melanogaster. A developmental expression qRT-PCR analysis, analyzing the relative dlg and miR-34 transcript levels in different temporal stages of A. aegypti development, was conducted to compare the relative transcript levels of A. aegypti throughout development. Then, a dlgdsRNA RNAi-mediated knockdown by bacterial feeding was performed to see if knocking down dlg has an effect on development. The results of the qRT-PCR analysis and RNAi experiment were not significant enough to come to a justified conclusion. Therefore, further testing is required to produce meaningful conclusions regarding the developmental expression of dlg and miR-34.