Symposium on Dengue - 2015
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Papers presented at the Symposium on Current Research Activities on Dengue, organized by the Molecular Medicine Unit, on 13th of November 2015
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Item Effect of γ-radiation on fertility and survival of Aedes albopictus (skuse) males in the laboratory for Sterile Insect Technique(Moleclar Medicine Unit, Faculty of Medicine, University of Kelaniya, Sri Lanka, 2015) Harishchandra, J.; Abeyewickreme, W.; Hapugoda, M.; Premaratne, R.G.; Gilles, J.R.S.BACKGROUND: It is believed that Aedes albopictus, one of the dengue vectors in Sri Lanka can be controlled using Sterile Insect Technique (SIT) when integrated with other conventional control methods. The objective of this study was to determine the effective dose of gamma radiation for producing sterile males of Ae. albopictus mosquitoes. METHODS: A batch of male pupae (n=32) aged 24-48 hours in F1 was irradiated using a gamma-ray irradiator (Gamma 220, Atomic Energy of Canada Ltd., Co60) with 25 Gy in duplicates. Following the same procedure, different doses (30 Gy, 40 Gy, 50 Gy, 60 Gy and 70 Gy) were given to each pupal batch in duplicates. Then they were transferred to laboratory cages (30 cm X 30 cm X 30 cm) for emergence and supplied with 10% sucrose solution. Adult emergence rate and male longevity were recorded. Virgin females from the same cohort (F1) were introduced into each cage for mating and fed blood starting 5 days after emergence. Females were then isolated in to individual tubes and hatching rate of individual egg batches was determined after two weeks of egg maturation in hatching solution containing 0.25 g BNB, 0.05g BY in 700 ml distilled water. Spermachecae of female mosquitoes were dissected and insemination rates were calculated after egg laying. RESULTS: Male mosquito pupae in F1 showed low mortality (0-3.12%) immediately after exposing to radiation (0-48 hours). After 21 days of the irradiation, probability of survival of male mosquitoes in F1 were 0.578, 0.494, 0.453, 0.313, 0.328, 0.381 and 0.219 at 0Gy, 25 Gy, 30 Gy, 40 Gy, 50 Gy, 60 Gy and 70 Gy respectively (Kaplan Meier survival analysis). Log Rank test indicated significant differences of survival of control males with males irradiated at 40Gy, 50 Gy, 60 Gy and 70 Gy. The survival of males irradiated at 25 Gy and 30 Gy did not differ from each other and from the control. The survival of males irradiated at 40 Gy, 50 Gy, 60 Gy were not significantly different among each other. The mean hatching rate (Mean+SE) of the F2 progeny of Ae. albopictus males (F1 progeny) were 10.89+2.76%, 7.36+1.75%, 3.09+0.71%, 0.79+0.27%, 0.66+0.23% and 0% when irradiated at 25 Gy, 30 Gy, 40 Gy, 50 Gy, 60 Gy and 70 Gy respectively. The control group showed a hatching rate of 64.26 + 7.12%. It was found that insemination rate of the irradiated males among different doses and controls were above 90% in F1 based on spermatheca dissection of blood-fed females (F1). CONCLUSION: 50 Gy is recommended as the most suitable γ radiation dose to produce 99% sterility in Ae. albopictus males which has 0.328 survival probability after 21 days of irradiation. This dose can be used to produce sterile males of Ae. albopictus for population suppression.Item Laboratory studies on larvivorous potential of five fish species against Aedes ageypti (Linnaeus) and Ae. albopictus (Skuse)(Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Sri Lanka, 2015) Kusumawathie, P.H.D.; Jayasooriya, G.A.J.S.K.; Gunathilaka, P.A.D.H.N.; Hapugoda, M.; Abeyewickreme, W.BACKGROUND: The dengue vectors, namely, Aedes aegypti (Linnaeus) and Ae. albopictus (Skuse) are container breeders, thus, container management is a key element for elimination of vector breeding sites. Use of predator fish is a potential tool for dengue vector control in some container habitats. However, in order to select suitable fish species, larvivorous potential of different fish species needs to be determined. Therefore, the objective of the study was to determine the larvivorous potential of some fresh water fish species against the dengue vectors. METHODOLOGY: Five fish species, namely: Poecilia reticulata, Aplocheilus dayi, Oreochromis niloticus, Rasbora daniconius and Puntius bimaculatus were collected from their natural breeding places during the field surveillance conducted from January-October, 2015. Each fish species was placed in separate fish stock cement tanks at the Regional Office of the Anti- Malaria Campaign in Kandy until they were used for the study. In order to determine the larvivorous potential of the fish, each species was placed in separate enamel trays containing de-chlorinated water and allowed to acclimatize for 30 minutes. A batch of 25 Ae. aegypti and Ae. albopictus larvae was introduced to each tray superlatively and the number of larvae remaining was counted after 24 hours of observation period. Once all the batch of larvae consumes by the fish species, another batch of 25 larvae from respective species was introduced during the observation period. The whole experiment was repeated for 6 times. RESULTS: The mean values of Ae. aegypti larvae consumed by one fish of O. niloticus, P. reticulata, A. dayi, R. daniconius and P. bimaculatus were 97.6, 66.9, 49.3, 62.5 and 47.5 respectively while, one fish of O. niloticus, P. reticulata, A. dayi, R. daniconius and P. bimaculatus consumed 98.4, 62.3, 51.2, 62.5 and 45.4 of mean Ae. albopictus larvae respectively. CONCLUSIONS: All five fish species showed larvivorous activity. However, O. niloticus showed the highest larvivorous potential followed by P. reticulata. Therefore, these two fish species may be considered as a supplementary method to control immature stages of dengue vectors.Item Molecular diagnosis and transmission of dengue in Sri Lanka(Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Sri Lanka, 2015) Hapugoda, M.; Abeyewickreme, W.; de Silva, N.R.; Gunasena, S.; Meegoda, D.; Manamperi, N.BACKGROUND: Dengue virus is an important vector-borne viral infection in Sri Lanka. Laboratory confirmation of suspected dengue cases is important for over/under estimation of cases. Early rapid diagnosis of dengue viral infection helps monitoring the disease, hospital admission when necessary and reduces case fatality. Detection of dengue viruses in mosquitoes is useful for studies on transmission of dengue virus. Study on risk factors for dengue is useful to understand spatial and temporal dynamics of transmission the disease. METHODOLOGY: A novel molecular-based assay; Reverse Transcription-Polymerase Chain Reaction-based Liquid Hybridization (RT-PCR-LH) was developed and validated for detection of dengue virus in clinical and mosquito specimens. Severity of dengue with circulating serotype was also analyzed. Wild-caught mosquito samples were collected from 236 dengue case-reported stations during outbreaks and a hot-spot during a period of 31 months. Epidemiological, environmental and entomological and other possible risk factors affecting transmission of dengue were analyzed. RESULTS: As an early (<5 days of fever) laboratory diagnostic method for dengue virus, the novel assay had 100% and 46% sensitivity for detection of confirmed and suspected dengue patients respectively. The assay developed in this study was found to be more sensitive than the other diagnostic techniques for early definitive laboratory diagnosis of dengue infection. Patients with definitive dengue correlated only with few signs and symptoms, indicating that laboratory confirmation is critical to avoid over estimation. A high sensitivity of 2 fluorescent focus unit of dengue virus/reaction was achieved and the assay was highly specific for dengue virus. The assay could detect dengue virus in 7% of field-caught Aedes albopictus specimens. A high density of Ae. albopictus was also associated with the dengue case-reported stations/hot-spots. Both vector species were susceptible to the 4 dengue serotypes under the laboratory conditions and DEN-3 under the field conditions. Geographical Information System (GIS) based risk mapping and database including epidemiological, climatic condition and entomological surveillance information were developed for the hot-spot in Kurunegala during the period of 2000-2003, which can be used as an early warning system. In depth study on socio-economic and other related factors affecting transmission of dengue was studied in the District of Gampaha. CONCLUSION: Ae. albopictus acts as an important vector of transmission of dengue in some urban and semi-urban areas. GIS-based risk maps developed are important to predict impending epidemics so that limited resources could be utilized in a cost effective manner to control the disease. Some socio-economic factors directly affecting transmission of dengue.