International Postgraduate Research Conference (IPRC)
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Item Evaluation of the Pyrethroid Resistance based on Voltage-Gated Sodium Channel (VGSC) Mutations in Aedes aegypti populations of Colombo, Gampaha and Kandy Districts in Sri Lanka(International Postgraduate Research Conference 2019, Faculty of Graduate Studies, University of Kelaniya, Sri Lanka, 2019) Ranathunge, T.; Udayanga, L.; Sarasija, S.; Karunathilaka, S.; Nawarathne, S.; Rathnarajah, H.; Dulficar, F.F.; Shafi, F.N.; Dassanayake, R.S.; Gunawardene, Y.I.N.S.Many countries focus on chemical based vector control strategies to restrict the disease transmissions, where pyrethroid insecticides are widely used as the first line of defense against Ae. aegypti. However, the constant use of insecticides have proven to induce insecticide resistance in mosquitoes. The knockdown resistance (kdr) occurs due to mutations in the Voltage Sensitive Sodium Channel (VSSC) or mutations in the Voltage-Gated Sodium Channel (VGSC), coded by the VSSC gene. Only three kdr mutations namely, the V1016G, S989P, and F1534C have been confirmed as commonly occurring amino acid substitutions among mosquito populations in Southeast Asia. Therefore, to extend this observation, current study was conducted to evaluate the prevalence of V1016G and F1534C mutations among Ae. aegypti mosquito populations in three different geographical regions of Sri Lanka. Immature (both pupae and larvae) stages of Ae. aegypti mosquitoes were collected from Colombo, Gampaha and Kandy districts from March to December 2018 and samples were transported to the Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya. A total of 855 Ae. aegypti larvae were collected from all districts and polymerase chain reaction (PCR) assay for molecular genotyping of mutations was performed for collected all Ae. aegypti larvae (III instar), to identify the prevalence of kdr mutations in the three Ae. aegypti populations. The frequencies of the resistant and susceptible kdr alleles were determined by using the Hardy–Weinberg Equilibrium for each of the point mutation. The Ae. aegypti populations from Colombo, Gampha and Kandy districts showed 40.07% (123/307), 39.58% (114/288) and 19.58% (47/240) of V1016G and F1534C mutations, respectively. The wild type (RR) genotype remained predominant within all the three districts, whereas the homogenous (SS) mutation genotype occurred only in minority. Further, the F1534C was predominant in Ae. aegypti populations of all districts. Among the kdr mutation population, heterogeneous genotyping (RS) for both V1016G and F1534C was prominent, while SS genotyping for V1016G mutation was not observed in the Kandy district. The findings clearly denote that long-term insecticide applications and multiple use of pyrethroids has led to the progression of insecticide resistance among local Ae. aegypti populations. Therefore, evaluation of the prevalence levels of these kdr mutations highlights the necessity for shifting towards novel vector control strategiesItem Potential Use of a Combined Sterile Insect Technique (SIT) and Wolbachia-Based Approach for the Control of the Dengue Vector Aedes albopictus in Sri Lanka(19th Conference on Postgraduate Research, International Postgraduate Research Conference 2018, Faculty of Graduate Studies,University of Kelaniya, Sri Lanka, 2018) Wijegunawardana, N.D.A.D.; Chandrasena, T.G.A.N.; Abeyewickreme, W.; Dassanayake, R.S.; Xi, Z.; Bourtzis, K.; Gunawardene, Y. I. N.S.This study addresses the possibility of using a novel and environmental friendly strategy of combined sterile insect technique (SIT) and a Wolbachia-based approach in Aedes albopictus population control efforts in Sri Lanka. The above strategy combines irradiation and the intracellular endosymbiont bacteria Wolbachia, in insects which are vertically transmitted from mother to offspring. Releasing sterile male mosquitoes in the wild will introduce sterility for suppression of Ae. albopictus wild mosquito population. The strategy aims at the releasing of sexually incompatible Wolbachia triple infected and irradiated male mosquitoes on a large scale to the natural environment. By integrating irradiation, any accidentally released female mosquito will be made fully sterile, and hence the risk of population replacement of the target population with triple infected mosquitoes is eliminated. Therefore, the combined SIT/IIT approach offers a safe and bio-secure strategy for the population suppression of Ae. albopictus in Sri Lanka. The overall work involved assessment of the density and species abundance of wild mosquito populations, introgression, and characterization of Wolbachia triple infected Ae. albopictus line in a Sri Lankan genetic background and determination of minimum sterility dose for the males and females of the introgressed line. Results revealed that Ae. albopictus mosquito was the dominant vector species in the study area (Narangodapaluwa PHI area) with an average ovitrap index of 75.25% (range; 56.9% - 94.7%) over the study year. The frequency of Wolbachia infection among the mosquito population was ~17% (13/78). The most prominent Wolbachia super group infecting the wild mosquito species was the B group and Ae. albopictus was naturally infected with both strains A and B. Eighty-five new alleles and 8 complete allelic profiles were submitted to the Wolbachia MLST database. All sequencing data were deposited in the Genebank under the accession numbers MH447376-MH470292 and MH756095-MH777430. Successful transfer of Wolbachia triple infection to Ae. albopictus into Sri Lankan genetic background was achieved through an introgression experiment and characterization of the newly developed Ae. albopictus line expressed partial CI and revealed competitive fitness cost for fecundity, fertility, and longevity compared with wild-type while there was no cost for pupation rate, adult emergence and sex ratio. Therefore, integration of an irradiation (dose; 30Gy) is recommended to achieve complete male and female sterility prior to mosquito release (minimum sterility dose for males - 28 Gy and females - 30 Gy). This minimum sterility dose is lower than the irradiation dose applied for SIT and had not resulted in a significant effect on male competitiveness. Therefore, a combined SIT and Wolbachia based approach in Ae. albopictus dengue vector control is recommended as an additional tool in integrated vector management in Sri Lanka.Item Determination of appropriate positioning of the ovitraps for dengue mosquito surveillance(Faculty of Graduate Studies, University of Kelaniya, 2015) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Chandrasena, T.G.A.N.; Dassanayake, R.S.; Gunathilaka, P.A.D.H.N.; Abeyewickreme, W.Three months ovitrap survey was conducted to assess the suitable position in placing the ovitraps for dengue vector mosquito surveillance and this study was initiated due to loss of valuable data from our previous studies as a result of physical damage of the ground kept ovitarps. Thirty four households in the Ragama Medical Officer of Health area in Gampaha District were selected to conduct the ovitrap survey during the period of May to July, 2015 to select the most appropriate positioning of the ovitrap. The conventional black plastic ovitraps (3.2x 2.7 cm) were used in this purpose to collect aquatic stages of Aedes mosquitoes while placing plywood paddle (4 x 0.5 cm) over the upper rim of each coded ovitrap. A total of 136 ovitraps were used in the study site providing four ovitraps (2 each indoor & outdoor) for each house while one of the ovitraps of indoor and outdoor being hung and other being kept on the ground. In positioning ovitraps, the outdoor ones were kept 3m away from the house while leaving indoor ovitraps in the living room in close proximity to racks/hanging clothes or partially shaded places. Following collection of samples at each week, ovitraps were washed thoroughly, refilled with new water and a new paddle, and corresponding data were recorded and analyzed. These analyses revealed that number of larvae and the number of Aedes mosquito eggs present in the two different ovitrap positions (Ground kept vs Hung) were not significantly different; in spite of significant difference (P=0.001) between the outside and inside placements. Further, significantly higher values were observed for both number of mosquito eggs and larvae present in each ovitrap kept outside (60 and 13 respectively) than those placed inside (32 and 3 respectively). Furthermore, slightly higher values were observed for hung ovitraps (49 and 9 respectively) than ones kept on the ground (43 and 7 respectively). Finally, ovitrap placed above the ground level was selected in continuing the routine ovitrap survey, as there was considerable reduction of mechanical damage to the latter thus facilitating continuous data collection.