Medicine
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This repository contains the published and unpublished research of the Faculty of Medicine by the staff members of the faculty
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Item Optimization of Aedes albopictus rearing procedures for combined sterile insect techniques (SIT) and Wolbachia-based laboratory studies in Sri Lanka(Springer Science and Business Media Deutschland GmbH, 2020) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Abeyewickreme, W.; Chandrasena, T.G.A.N.; Dassanayake, R.S.; Manamperi, A.The establishment of a laboratory colony is an essential first step for conduction of laboratory studies on the biology and control of Aedes albopictus mosquitoes. Therefore, with the objective of generating high quality mosquitoes as research material for utilization in ongoing vector control studies and to disseminate authenticated, high-quality Ae. albopictus mosquito rearing information to the research community, maintenance of an Ae. albopictus mosquito colony was initiated at the Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Sri Lanka. A self-mating colony was established from eggs of the F1 progeny of individuals collected as free-living larvae in Narahenpita (Western Province), Sri Lanka. The mean temperature of 28 °C (± 2 °C) and relative humidity of 80% (± 5%) was constantly maintained inside the insectary. Lighting was provided by fluorescent lights, regulated with 12:12 h continuous dark and light period. Pest insects were controlled manually. Mosquitoes were maintained on bovine blood provided via an artificial membrane feeding system and a continuous supply of 10% sugar supplements. Larvae were maintained in deoxygenate water and fed with International Atomic Energy Agency (IAEA) recommended diet of tuna meal, bovine liver powder, and brewery yeast in a ratio of 37.5:27:10.5 in 1 L. Data on fecundity, fertility, larvae death, pupation, adult emergence, adult mosquito longevity were recorded. Adhering to bio-safety, all discarded materials were boiled thoroughly and incinerated if required. This report on the establishment and maintenance of a laboratory colony of Ae. albopictus will be of value for identifying the critical requirements essential under artificial conditions.Item Maternal transformation of Wolbachia isolated from infected mosquito hosts to Aedes aegypti using micro-injection based procedure: an approach towards integrated dengue vector control(Moleclar Medicine Unit, Faculty of Medicine, University of Kelaniya, Sri Lanka, 2015) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Chandrasena, T.G.A.N.; Dassanayake, R.S.; Ruanareerate, T.; Kittayapong, P.; Abeyewickreme, W.BACKGROUND: The maternal transformation of Wolbachia (Wb) to Aedes mosquitoes is an approach that has been widely used in Aedes mosquito population control strategies in the integrated dengue vector control efforts. This study details usage of such approach in the maternal transformation of Aedes aegypti using the Wb isolated from infected mosquito hosts using micro-injection based procedure for the first time in Sri Lanka. METHODS: Twenty five ovaries of each adult female Aedes albopictus and Culex quinquifaciatus mosquitoes (14-18 days old) were dissected and Wb was isolated and suspended in 20 μl of 1X Phosphate Buffered Saline (PBS). The isolated samples were kept on ice to maximize the bacteria viability. The exudate was then used in the trans-infection of Ae. aegypti mosquitoes. Infection of Wb into the mosquito Ae. aegypti was carried out by maternal/adult microinjection. To create stable trans-infected lines, the regions near the 3rd leg emerged from the thorax of Ae. aegypti were microinjected to incorporate Wb into the developing germ line and then to facilitate the transmission of Wb to offspring. The volume used for injection based on the swelling of the abdomen, which was monitored throughout the experiment while maintaining the injection pressure, time and number of injecting times. Virgin female Aedes mosquitoes survived after microinjection was fed with 1 ml of 10% sugar solution containing 10,000 units Penicillin/Streptomycin per 100 ml. Survival rate was monitored at 1, 3 and 24 hour/s following microinjection procedure. Survived female mosquitoes resulted from injected line (generation 0 [G0]) was placed in vials with three uninfected male mosquitoes to establish iso-female lines. After egg laying 3 times, G0 females were sacrificed and total DNA was extracted, and PCR was carried out (wsp primers 81F and 691R) using DNA extracted from Ae. aegypti to detect the presence of Wb DNA and its density in the latter mosquito using dilution series of extracted DNA. To select for a stable infection, only offspring from females that came positive for Wb by PCR screening was used as parental stocks. For the samples positive by wsp PCR, strain identification was carried out by performing PCR with 4 different primer sets; wPip, wAlbA, wAlbB and wsp specific for Wb. Females that tested negative for Wb were discarded along with their progeny. The resulting triple infected line will be monitored periodically by PCR to confirm infection status. RESULTS: Wolbachia-triple infected line of Ae. aegypti mosquito was created by using isolated wAlbA+wAlbB+wPip Wolbachia strains. Infection status of newly developed line was monitored by PCR up to G2 and will be continued to G8. Experimental work will be started from G5 to evaluate the use of newly develop Ae. aegypti mosquito line for Wb-based approach under laboratory condition. CONCLUSION: Direct maternal micro-injection technique was used to create Wb infected mosquito line of Ae aegypti, which could be applied in Wb-based dengue vector control activities in Sri Lanka once the line is established.Item Ovi-trap based surveillance for Aedes in dengue endemic Gampaha District, Sri Lanka during rainy season(Faculty of Tropical Medicine, Mahidol University, 2008) Hapangama, H.A.D.C.; Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Benjamin, S.; Abeyewickreme, W.Dengue is endemic in Gampaha District, Sri Lanka. Aedes aegypti and Aedes albopictus have been incriminated as vectors in urban and suburban environments. An ovitrap-based weekly surveillance was carried out to determine the distribution and abundance of Dengue vectors, in six dengue endemic areas; Galudupita, Gongitota, Wanawasala, Kerangapokuna, Mabole and Welikadamulla in the Gampaha District during May and June 2008 in both indoors and outdoors. Galudupita and Gongitota gave the highest ovitrap indices, both indoors and outdoors as 97% and 73% respectively. Of the total positive ovitraps(1320), positivity for Ae. albopictus, Ae. aegypti and both were 63.5%, 32.9% and 3.6% respectively. Highest mean number of Ae. aegypti and Ae. albopictus larvae per ovitrap in both indoor and outdoor ovitraps were observed Galudupita and Gongitota respectively. However, statistical analysis showed that ovitrap index of Ae. aegypti in six study was not significantly different from each other (P > 0.05). Outdoor mean number of Ae. aegypti larvae per ovitrap in Galudupita, was significantly higher than other sites (F=3.587, P=0.012). Indoor Ae. albopictus ovitrap index of Gongitota was significantly higher than of Welikadamula (F=2.59, P=0.046) while outdoor Ae. albopictus ovitrap index of Gongitota and Wanawasala was significantly higher than of Welikadamula F=3.19 P=.0.02). Results irnplicate that Ae. albopictus prefers more vegetation as observed in Gongitota and Wanawasala. In highly populated and urbanized Galudupita, area with less vegetation, Ae. albopictus population was less than Ae. aegypti even outdoors. Study implicates that Ae. albopictus is the predominant indoor and outdoor breeder during rainy season.Item Entomological assesment of Wuchereria bancrofti transmission following mass treatment in Gampaha District, Sri Lanka.(Faculty of Tropical Medicine, Mahidol University, 2007) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Abeyewickreme, W.Global program to eliminate Lymphatic filariasis (Lf) relies mainly on Mass Drug Administration (MDA). Success of filariasis control programs depends on careful monitoring of infection levels in human populations as well as vectors following the introduction of the drug intervention. This study was conducted to assess the current level of Lf transmission following mass distribution of Diethylcarbamazine-Albendazole in the Gampaha district, Sri Lanka. Field study was conducted in 45 sites in all Medical Officer of Health (MOH) areas of the Gampaha district identified by the Anti Filariasis Campaign (AFC) as high risk for bancroftian filariasis. Investigation revealed 42.22% (19/45) of the sites were infested with mosquitoes positive for Wuchereria bancrofti. Only the sites in urban and semi urban areas were positively infested while rural areas were free of infection. An infection rate of 5.26% was observed among the mosquitoes caught from households and the larval density was 8.7 per positive mosquito. According to a study conducted by the AFC in a sample population (14 sites), the prevalence of Lf was 0.038%. Data recorded by the AFC in 1994 indicated that the infective rate, positivity of mosquitoes and microfilaria density were 90%, 3.05% and 23 respectively. The present study confirms that the level of transmission of W. bancrofti has not decreased in the Gampaha district, despite the MDA Programme been implemented since 2002. Therefore, a proper screening-programme combined with anti filarial treatment and. vector control programme is urgently required to minimize filarial morbidity and interrupt filarial transmission within the country.Item Patterns of monthly Culex mosquito density variation in Gampaha district, Sri Lanka.(Faculty of Tropical Medicine, Mahidol University, 2009) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Manamperi, A.; Abeyewickreme, W.BACKGROUND: The ecology, development, behavior, and survival of mosquitoes and the transmission dynamics of the diseases they transmit are strongly influenced by climatic factors. OBJECTIVE: The objective of this study was to identify the population density variation of Culex mosquitoes within a period of one year in Gampaha district, Sri Lanka together with potential climatic factors that influenced the Culex population density variation. METHODOLOGY: Culex mosquitoes were routinely collected on monthly basis from 9 sites in Gampaha district. Climate data was obtained from the Department of Meteorology. RESULTS: An exponential growth of Culex population densities was observed in all sites starting in December to February during the study period. The maximum density occurred in January and decreased from March until July. It again increased during August and thereafter decreased until December. Among the study sites the maximum Culex density (mosquitoes/man-hour) was observed in Hekiththa ranging between 89 to 22 and the minimum was from Kurukulawa ranging from 6 to 1. Climatic data suggest that temperature is a limiting factor for the Culex population growth while it was strongly influenced by the rain fall pattern. DISCUSSION: Similar Culex population density variation pattern was observed in all sites but exhibited enormous variation between sites, probably due to different local conditions. Also it was suggested that estimation of W, bancrofti transmission levels in Culex mosquitoes should be practiced in field settings where high mosquito density was observed. Since mosquito density appears difficult to be analyzed by individual dissection use of pool-screen PCR-ELISA would be a better method.Item Polymerase Chain Reaction and mosquito dissection as tools to monitor filarial Infection levels following mass treatment in Gampaha District, Sri Lanka(Elsevier, 2008) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Manamperi, A.; Bandara, K.B.A.T.; Liyanage, T.; Abeyewickreme, W.BACKGROUND: Mass Drug Administration (MDA)-based Global Lymphatic filariasis (Lf) eradication programmes are aimed at stopping transmission of Wuchereria bancrofti by its mosquito vector. The study was designed to compare one year post treatment (mass distribution of Diethylcarbamazine-Albendazole) infection rates of Wuchereria bancrofti in Culex quenquifaciatus, the main vector of Lf in Sri Lanka using Conventional dissection techniques and a Polymerase Chain Reaction (PCR) assay based on parasite specific Ssp1 repeat which amplifies a fragment of 188 bp. METHODS: Field study was conducted in 45 sites in all Medical Officer of Health (MOH) areas in the Gampaha district, Sri Lanka; identified by the Anti Filariasis Campaign (AFC) as having high-risk for bancroftian filariasis transmission. Indoor-resting mosquitoes were collected by aspiration from 20 houses per each site. Part of the mosquitos were used for dissection and the remainder was used for PCR to detect the filarial parasites in mosquito. RESULTS: Mosquito dissection data revealed 42.22% (19/45) of the sites were infested with mosquitoes positive for Wuchereria bancrofti, indicating 8 transmission active MOH areas (53.33%; 8/15). An infection rate of 5.26% was observed among the mosquitoes caught from households and the larval density was 8.7 per positive mosquito. PCR investigation revealed that 46.67% (21/45) of the sites were positive for W. bancrofti DNA, indicating 11 transmission active areas (73.33%; 11/15). The PCR was found to be more sensitive compared to microscopy in detecting the filarial parasite in field collected mosquito samples with respect to the MOH areas. CONCLUSION: The PCR technique employed offers scope for detection of the filarial parasites with higher sensitivity and specificity; is efficient and rapid. This technique applied for the first time in Sri Lanka, can be adopted as a diagnostic tool for the detection of filarial parasites in the vector population in surveillance to enable effective control of filariasis in the country. Acknowledgements: Authors acknowledge the WHO/SEARO/TDR (grant no. SN 1152) and University of Kelaniya (Research grant no. RP/03/04/06/01/2006) and to Ms. H.M.Renuka and Mr. H.P.Anura U. Pathirana, Mr. M.I.M.Peris and Mr. Y.L.Rassapana for their support during field study activities. © 2008 Elsevier Inc.Item Night blood survey of a selected high-risk population for lymphatic filariasis(Sri Lanka Association for the Advancement of Science, 2007) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Abeyewickreme, W.; Gunawardena, N.K.; Hapuarachchi, H.A.C.; Abeysundara, S.Human infection with Wuchereria bancrofti causes a disabling parasitic disease known as lymphatic filariasis, which is a major public health and socio-economic problem in many parts of the world. Little is known about the prevalence of filariasis among high-risk populations for filariasis. Objective of this study was to determine such prevalence of lymphatic filariasis among Mahara prison inmates whom the Anti Filaria Campaign (AFC) has identified as a high-risk group. All inmates of Mahara Prison were screened for Microfilariae (Mf) except those in special cells, by night blood film microscopy to determine the prevalence of infection from February to May 2007. All inmates were males of greater than 15 years. Of the 423 inmates screened, 15 were positive for Mf, giving a Mf positive rate of 3.55% in the study population and a mean Mf density of 5 Mf/60 æl blood, ranging between 4 to 9.2 Mf /60 æl of blood with a standard deviation of 2.49. The highest number of infected inmates was residents of Colombo and Gampaha districts where transmission is currently taking place. This is one of the few studies undertaken to date to determine the prevalence of bancroftian filariasis among inmates of a prison, a neglected population in Sri Lanka. This study indicates that the Mf rate of bancroftian filariasis in this study population is far greater than the 0.18% currently reported in the country. Therefore, an intensive programme is recommended to contain the spread of infection within this study population. For this, a proper screening programme combined with antifilarial treatment and vector control programme is urgently required. Acknowledgements: Authors wish to acknowledge the financial assistance received from WHO/SEARO/TDR (grant no. SN 1152) and University of Kelaniya (Research grant no. RP/03/04/06/01/2006). Authors wish to thank Dr. Ravi Mudaliage, Senior Medical Officer, Prison's Hospital, Mahara, Ragama for his support and encouragement during field study activities. Authors also wish to thank Mr. M. Y. D. Dayanath, Ms. N.M. Ashoka Malanie, Mr. M.I.M.Peris, Mr. Y.L.Rassapana and other staff members of the Molecular Medicine Unit and Department of Parasitology, Faculty of Medicne, University of Kelaniya, Ragama for their assistanceItem Geographic information system (GIS) mapping of lymphatic filariasis endemic areas of Gampaha District, Sri Lanka based on epidemiological and entomological screening(SEAMEO Regional Tropical Medicine and Public Health Project, 2012) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Manamperi, A.; Senarathne, H.; Abeyewickreme, W.The objective of this study was to develop a site directed geographic information system (GIS) map of lymphatic filariasis (LF) in Gampaha District, Sri Lanka as a guide for targeted control activities. Epidemiological and entomological screening of LF was carried out in nine pre-identified endemic areas in Gampaha District, using night blood screening and pool-screening PCR-ELISA. In total, 1,073 subjects (286 children, 787 adults) from 9 sites were examined. Positive cases were detected at 2 sites, with prevalence rates of 0.5% (Hekiththa) and 3.4% (Peliyagoda); the prevalence of microfilaria (mf) among adult Culex quinquefasciatus mosquitoes surveyed was 30%. The overall prevalence of mosquitoes with L1-L2 larvae of W. bancrofti ranged from 0% to 8.31% using dissection and point estimates of infection prevalence, and ranged from 0 to 32.4% using PCR-ELISA. The largest number of human cases was found at altitudes of 2.5-3.5 min highly populated areas, where transmission appears to have taken place. Questionnaires indicated that limited community awareness of LF may be a reason for the fairly static infection prevalent among the local population. The GIS mapping of LF cases shows a considerable prevalence of LF and marked variability by geographic site in Gampaha.