Hemocup blood feeder: An affordable and simplified blood-feeding device for maintenance of Aedes aegypti mosquito colonies in Sri Lanka.

Dengue is a mosquito-borne viral disease mainly transmitted by Aedes aegypti and disease control is primarily reliant on mosquito vector control strategies. In the failure of conventional vector control strategies, new strategies are being developed which specifically require the maintenance of mosquito colonies in the laboratories. Blood-feeding is an essential part of the routine colony maintenance of Ae. aegypti. Therefore, the current study was focused on developing a simplified artificial membrane-feeding device, "Hemocup" feeder out of affordable material. viz., plastic cups, styrofoam insulation system, parafilm-M, and preheated water to facilitate the Ae. aegypti artificial blood feeding. The performance of the device was compared to that of a commercially available blood-feeding device, "Hemotek", by assessing the blood-feeding rate, fecundity, and egg hatchability. Similar blood feeding rates were observed for Hemocup and Hemotek methods (91.8 ± 1.6 and 94.3 ± 1.6 respectively>0.05) as well as comparable fecundity between the two methods (20.8 ± 0.7 and 22.0 ± 1.5 respectively; p > 0.05). Furthermore, there was no statistically significant difference in egg hatchability between the two methods (91.9 ± 1.4 and 93.8 ± 1.4, respectively; p > 0.05). The results indicate that this simple Hemocup blood-feeding system can be used for routine colonization of laboratory strains of Ae. aegypti and for mass-rearing purposes.

Multiple dengue virus serotypes resistant transgenic Aedes aegypti fitness evaluated under laboratory conditions.

Dengue viruses (DENV) are the wildest transmitted arbovirus members of the family Flaviviridae, genus Flavivirus. Dengue viruses are composed of four serotypes, DENV1, 2, 3, and 4, and these viruses can cause dengue fever and dengue haemorrhagic fever or dengue shock syndrome, when infecting humans. RNA interference (RNAi) is a self-defence mechanism, which can be used to prevent invasions of RNA viruses to the host. Genetically engineering a host with an RNAi molecule that targets a single virus serotype may develop escape mutants, and can cause unusual dominance over other serotypes. Therefore, the simultaneous targeting of multiple serotypes is necessary to block DENV transmission. Here, we report the development of transgenic Aedes aegypti based on a bioinformatically designed multiple miRshRNA (microRNA-based shRNA) DNA sequence under the control of a blood-meal induced promoter, Carboxypeptidase A, to induce RNAi for DENV in Aedes aegypti, and demonstrate the expression of a synthetic multiple shRNA polycistronic cluster having RNA interference sequences to target DENV genomes. The transgenic mosquitoes have lower rates of infection, dissemination, and transmission for DENV2 and DENV4 compared to wild mosquitoes, with a significant reduction of dengue copy number and antigen levels in the midgut. These levels of DENV were low enough to make transgenic mosquitoes stop the DENV transmission from infected host to a susceptible host and refractory to DENV2 and DENV4 infection. Such multiple resistance in Ae. aegypti has not been documented previously. Laboratory fitness measurement of transgenic Ae. aegypti showed results comparable to other reported transgenic mosquitoes.