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This repository contains the published and unpublished research of the Faculty of Medicine by the staff members of the faculty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 Recent chikungunya outbreak in Sri Lanka 2006-2007(Faculty of Tropical Medicine, Mahidol University, 2007) Abeyewickreme, W.; Bandara, K.B.A.T.; Dayanath, M.Y.D.; Sumanadasa, D.; Hapuarachchi, H.A.C.; Gunawardena, N.K.; Hapugoda, M.D.; Wijesiriwardena, B.; de Silva, S.; Perera, T.BACKGROUND: Chikungunya(CHIK) is a viral disease transmitted by Aedes mosquitoes. Cases with symptoms of CHIK had been reported from several parts of Sri Lanka in 2006-2007. Laboratory testing of samples is a prime requirement for confirmation of transmission. OBJECTIVES: To confirm CHIK infection in suspected patients by rapid Reverse Transcription Polymerase Chain Reaction Assay(RT-PCR), find out manifestations specific for CHIK infection and study the transmission of CHIK virus by vector mosquitoes. METHODOLOGY: Serura. samples and information on clinical manifestations were collected from 189 chikungunya-suspected patients from different geographical areas in Sri Lanka from September 2006 to September 2007. Samples were tested for Chikungunya RNA by RT-PCR. Amplified products were visualized by agarose gel electrophoresis. Adult mosquitoes were also collected from chikungunya case-reported stations. They were tested for Chikungunya RNA through RT-PCR-followed by agarose gel electrophoresis assay. RESULTS: Of the CHIK-suspected patients reported from all parts of the island 86/189 (45.5%) were positive for CHIK virus. Of the PCR positive 06, all had fever with either arthralgia or arthritis or both. Headache (95.3%) and backache (84.6%) were also common among above patients. Eight percent (4/50) of both species of Aedes mosquitoes were RT-PCR positive. DISCUSSION: RT- PCR is important in early diagnosis of the infection and differentiation from dengue fever. The most common clinical symptoms observed were fever with either arthralgia, arthritis or both. Both Aedes aegypti and Aedes. albopictus are important in transmitting the disease.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 A Case report of dengue and chikungunya co-infection in Sri Lanka(The Parasitology and Tropical Medicine Association of Thailand, 2008) Abeyewickreme, W.; Hapuarachchi, H.A.C.; Bandara, K.B.A.T.; Hapugoda, M.D.; Williams, S.Dengue fever and chikungunya are arboviral diseases transmitted by Aedes mosquitoes. Though dengue has been an important communicable disease in Sri Lanka for many years, chikungunya has not been reported in Sri Lanka since late 1960s. However, in November 2006, an outbreak suggestive of chikungunya erupted in the country. We report here the first laboratory confirmed case of dengue and chikungunya co-infection in Sri Lanka. The objective is to confirm the co-infection of dengue and chikungunya in a clinical case reported in November 2006. Clinical history of high fever, severe headache, nausea, loss of appetite, severe arthralgia and mild oedema of knees, small joints of hands and feet for 3 days suggested the possibility of dengue and chikungunya in a 70 year old male. There was no skin rash or bleeding manifestations. Laboratory investigations performed included total white blood corpuscle count/differential count (WBC/DC), platelet count (PLT), serum, haemoglobin (Hb%) and packed cell volume levels (PCV). Reverse Transcription- Polyrnerase Chain Reaction (RT-PCR) technology was used to confirm the presence of either dengue or chikungunya. Viral RNA was extracted from serum samples collected during the first five days of infection using QiAmp Viral RNA Kits and amplified products were visualized by 2% agarose gel electrophoresis and ethidium bromide staining. WBC/DC analysis showed a leucopaenia (WBC count 3.04 x 103 per μl) with relative lymphocytosis (51.0%). The total PLT was 115 x 103 per μl. Hb% was 14.3 g/dl with a PCV of 43.8%. The presence of both infections was confirmed by RT-PCR which amplified 225 bp and 354 bp products for dengue and chikungunya respectively. This was the first laboratory confirmed case of dengue and chikungunya co-infection, which was also the first confirmed report of chikungunya since 1969 in Sri Lanka. As clinical and biochemical manifestations of this patient suggested the probability of a mixed infection of dengue and chikungunya, the confirmation was achieved by a RT-PCR assay. This report highlights the importance of using molecular assays to confirm mixed viral infections during their early stages, especially infections such as dengue which can result in fatal complications.Item Confirmation of 2006 chikungunya outbreak in Sri Lanka using RT-PCR(Malaysian Society of Parasitology and Tropical Medicine, 2007) Abeyewickreme, W.; Bandara, K.B.A.T.; Perera, H.; Dayanath, M.Y.D.; Hapuarachchi, C.Chikungunya, a mosquito-borne viral infection caused by a single-stranded RNA virus of the family Togaviridae, is considered as a rare, non-fatal disease. During February to October 2006, an epidemic of over 1.3 million suspected cases was reported in India and neighbouring countries causing a significant economic loss due to crippling manifestations of this infection. With the outbreak of many viral fevers including dengue and dengue haemorrhagic fever, in October–November 2006, patients with manifestations suggestive of chikungunya such as high fever, headache, arthralgia and arthiritis (particularly, in ankle, knee and small joints of hands) were reported in many parts of Sri Lanka. As no chikungunya cases had been officially reported in the island since 1969, laboratory investigations for the presence of chikungunya virus was a prime requirement for confirmation of the outbreak. A total of 60 venous blood samples collected from suspected patients from different geographical regions of Sri Lanka were analysed using a reverse transcriptase-polymerase chain reaction (RT-PCR) technique to confirm the presence of chikungunya virus. Viral RNA was extracted from samples collected within 1-4 days of fever by using a Qiagen RNA extraction kit. RT-PCR was performed using chikungunya specific oligonucleotides. Both positive and negative controls were included in each set of reactions. The amplified products (354 bp) were visualized by running in a 1.5% agarose gel followed by ethidium bromide staining. Of the 60 samples, 33 (55%) were positive for chikungunya. They were distributed among almost all the geographical regions, highlighting the presence of a wide-spread epidemic in the country.Item Re-emergence of chikungunya in Sri Lanka: First confirmation of the 2006 outbreak by molecular diagnosis(Sri Lanka Association for the Advancement of Science, 2007) Perera, E.D.T.; Wijesiriwardena, B.; Hapugoda, M.D.; Bandara, K.B.A.T.; Dayanath, M.Y.D.; Wellawaththage, L.C.; Gunawardena, N.K.; Hapuarachchi, C.; Abeyewickreme, W.Chikungunya virus infection is clinically similar to many other acute febrile illnesses, such as dengue infection, malaria, west nile fever and leptospirosis. Rapid confirmation of the outbreak by laboratory diagnosis is important to ensure public health safety by appropriate patient management and controlling the disease. Molecular diagnosis by Reverse Transcriptase- Polymerase Chain Reaction (RT-PCR) assists rapid diagnosis. The objective of the present study was to determine the clinical manifestations of chikungunya confirmed patients in Sri Lanka. Venous blood samples and clinical information were collected from 66 chikungunya suspected patients having fever of less than 4 days from different geographical areas in Sri Lanka during the period September 2006 to February 2007. Serum samples were tested for chikungunya RNA by RT-PCR. Amplified products were visualized by agarose gel electrophoresis. Among 66 suspected patients, 51% (34/66) were positive for chikungunya by RT-PCR assay and 55.9% (19/34) were females. All age groups were affected similarly with the mean age of 41 years (range = 4 months to 80 years). Of the PCR positive 34, all had fever with either arthralgia or arthritis or both. Most of them had only pain in the joints without swelling (arthralgia only); 67.6% (23/34) in knee, 55.9% (19/34) in ankle, 50% (17/34) in wrist, 44.1% (15/34) in elbow and 52.9% (18/34) in small joints. Arthritis of ankle joint 35.2% (12/34) was more frequent compared with arthritis of the knee joint17.6% (6/34). PCR positive patients manifested more symptoms compared with PCR negative patients; 85.3% (29/34) headache, 79.4% (27/34) backache, 58.8% (20/34) nausea and 61.8% (21/34) vomiting. Compared with dengue, most of the chikungunya patients did not have dermatological manifestations. This is the first confirmation of the 2006 chikungunya outbreak in Sri Lanka. Some of the patients who had symptoms suggestive of chikungunya, tested by PCR were negative. These patients were probably suffering from other illnesses such as dengue. Acknowledgements: The International Atomic Energy Agency (TC SRL 6/028) for technical cooperationItem Potential use of blood-fed mosquitoes as evidence in forensic casework(Sri Lanka Association for the Advancement of Science, 2007) Marasinghe, M.P.L.R.; Sirisena, D.M.; Bandara, K.B.A.T.; Hapuarachchi, H.A.C.; Abeyewickreme, W.Medico - criminal entomology has been an important source of evidence in forensic investigations for many years. However, it has not been widely used to provide direct evidence for personal identification in forensic casework so far. In this study, we made an attempt to determine the usefulness of human DNA extracted from blood-fed mosquitoes as evidence in forensic casework. Approximately 1500 adult female mosquitoes of same age from four different species, i.e. Culex quinquefasciatus, Armigerus sabalbatus, Aedes aegypti and Anopheles tessellatus were fed with the same volume of human blood and maintained under the same environmental conditions. Three batches (N=1, 5 and 10) of randomly selected blood-fed mosquitoes were crushed and blotted onto filter paper strips separately at two hour intervals subsequent to blood meals up to 48 hours to determine the longitudinal variation in the extraction of polymerase chain reaction (PCR) amplifiable human DNA from mosquitoes. Human DNA was extracted from filter papers using a chelex-100 extraction protocol and amplified by PCR technique using human primers. Amplified products were run in 1.5% agarose gels. PCR amplifiable human DNA could be extracted from mosquitoes of Cx. quinquefasciatus and An. tessellatus up to 48 hours subsequent to blood meals. However, this duration was up to 46 and 42 hours in Ae. aegypti and Ar. Sabalbatus, respectively. The amount of amplifiable DNA was inversely proportionate to the post-feeding duration, but increased with number of mosquitoes at each time interval. Amplifiable human DNA could be extracted even from a single mosquito of four different species even after 48 hours from a blood meal with slight variation among species. However, as the amount of DNA decreases with time interval, more mosquitoes will have to be used for extraction in late sample collections. Similarly, it may be required to use a highly sensitive PCR protocol when analyzing such samples. Blood-fed mosquitoes collected even after 2 days from a crime scene may be used as a source of direct evidence for personal identification in forensic casework.Item Re emergence of Chikungunya virus in South-east Asia: virological evidence from Sri Lanka and Singapore(Cambridge University Press, 2010) Hapuarachchi, H.C.; Bandara, K.B.A.T.; Sumanadasa, S.D.; Hapugoda, M.D.; Lai, Y.L.; Lee, K.S.; Tan, L.K.; Lin, R.T.; Ng, L.F.; Bucht, G.; Abeyewickreme, W.; Ng, L.Chikungunya fever swept across many South and South-east Asian countries, following extensive outbreaks in the Indian Ocean Islands in 2005. However, molecular epidemiological data to explain the recent spread and evolution of Chikungunya virus (CHIKV) in the Asian region are still limited. This study describes the genetic Characteristics and evolutionary relationships of CHIKV strains that emerged in Sri Lanka and Singapore during 2006-2008. The viruses isolated in Singapore also included those imported from the Maldives (n=1), India (n=2) and Malaysia (n=31). All analysed strains belonged to the East, Central and South African (ECSA) lineage and were evolutionarily more related to Indian than to Indian Ocean Islands strains. Unique genetic characteristics revealed five genetically distinct subpopulations of CHIKV in Sri Lanka and Singapore, which were likely to have emerged through multiple, independent introductions. The evolutionary network based on E1 gene sequences indicated the acquisition of an alanine to valine 226 substitution (E1-A226V) by virus strains of the Indian sublineage as a key evolutionary event that contributed to the transmission and spatial distribution of CHIKV in the region. The E1-A226V substitution was found in 95.7 % (133/139) of analysed isolates in 2008, highlighting the widespread establishment of mutated CHIKV strains in Sri Lanka, Singapore and Malaysia. As the E1-A226V substitution is known to enhance the transmissibility of CHIKV by Aedes albopictus mosquitoes, this observation has important implications for the design of vector control strategies to fight the virus in regions at risk of chikungunya fever.Item Laboratory confirmation of dengue and chikungunya co-infection(Sri Lanka Medical Association, 2008) Hapuarachchi, H.A.C.; Bandara, K.B.A.T.; Hapugoda, M.D.; Williams, S.; Abeyewickreme, W.No Abstract AvailableItem Point mutations in the dihydrofolate reductase and dihydropteroate synthase genes of Plasmodium falciparum and resistance to sulfadoxine-pyrimethamine in Sri Lanka(American Society of Tropical Medicine and Hygiene, 2006) Hapuarachchi, H.A.C.; Dayanath, M.Y.D.; Bandara, K.B.A.T.; Abeysundara, S.; Abeyewickreme, W.; de Silva, N.R.; Hunt, S.Y.; Sibley, C.H.Sulfadoxine-pyrimethamine (SP) is the second-line treatment for Plasmodium falciparum malaria in Sri Lanka. Resistance to SP is caused by point mutations in the dihydrofolate reductase (Pf-dhfr) and dihydropteroate synthase (Pf-dhps) genes of P. falciparum. We determined the genotype of Pf-dhfr and Pf-dhps and the clinical response to SP in 30 field isolates of P. falciparum from Sri Lanka. All patients treated with SP had an adequate clinical response. Eighty-five percent (23 of 27) of pure field isolates carried parasites with double mutant alleles of Pf-dhfr (C59R + S108N) and showed about 200-fold higher levels of resistance to pyrimethamine than the wild type in a yeast system. None of the isolates had either known or novel mutations at other positions in the dhfr domain. In contrast, 67% (20 of 30) of the isolates carried parasites that were wild type for Pf-dhps. In Sri Lanka, detection of the triple mutant allele of Pf-dhfr will require tracking mutations at codon 51Item Chloroquine resistant falciparum malaria among security forces personnel in the Northern Province of Sri Lanka(Sri Lanka Medical Association, 2004) Hapuarachchi, H.A.C.; Dayanath, M.Y.D.; Abeysundara, S.; Bandara, K.B.A.T.; Abeyewickreme, W.; de Silva, N.R.OBJECTIVE: To determine the occurrence and species distribution of malaria and the extent of chloroquine resistance among security forcespersonnel in a selected region of the Northern Province of Sri Lanka. DESIGN: A descriptive study. SETTING: Mannar District in the Northern Province. METHODS: Nine hundred and seventy five security personnel were screened for malaria by microscopy. Those who were positive were treated withchloroquine and were subjected to 28 day in vivo assay to determine chloroquine resistance. In vitro microtest assay was performed to determine the response of Plasmodium falciparum isolates to chloroquine in vitro. RESULTS: Of the 975 personnel screened, 181 (18.6%) were positive for malaria. P. falciparum was the predominant species (n = 125; 69.1%). The rest were due to P. vivax (n = 42; 23.2%) and mixed infections (n = 14; 7.7%). This was an inversion of the usual species distribution pattern in the country. In vivo assay revealed 38 (53.5%) P. falciparum infections as chloroquine resistant. Fifteen of 23 (65.2%) P. falciparum isolates showed evidence of resistance in vitro. None of the P. vivax infections showed evidence of chloroquine resistance. There was no significant difference in the severity of clinical disease between chloroquine resistant and sensitive infections at first presentation. Recrudescent P. falciparum infections had significantly lower mean parasite densities as well as lower clinical scores at recrudescence than at first presentation. CONCLUSION: Results demonstrate the high prevalence of malaria and chloroquine resistance in the study area and explains several contributory factors for this. There is an urgent need to review antimalarial drug policies in Sri Lanka