Browsing by Author "Bandara, K.B.A.T."

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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.
Chikungunya outbreak in 2008 in Ratnapura district, Sri Lanka - clinical and socio-economic analysis
(Sri Lanka Association for the Advancement of Science, 2008) Sumanadasa, S.D.M.; Hapuarachchi, C.; Bandara, K.B.A.T.; Wellawaththage, L.C.; Abeyewickreme, W.
Since 2006, Sri Lanka has experienced several outbreaks of chikungunya fever (CHIK) affecting several thousands of people. Today, CHIK has become one of the most important vector-borne diseases in the country. The objective of this study was to analyse the clinical manifestations and socio-economic status among CHIK patients reported from Pallebedda and Godakawela areas in Ratnapura district during the outbreak in February and March 2008. After obtaining the informed written consent, venous blood samples were collected from 80 suspected patients. A medical officer carried out clinical examination of each patient. Clinical information along with socio economic data of the patients was recorded in an interviewer-administered questionnaire. Serum samples were tested for CHIK by a Reverse-Transcription Polymerase Chain Reaction (RT-PCR) assay. Of eighty patients tested, 51% (n=42) were positive for CHIK. All positive patients had fever for less than 5 days duration. Majority of them (95%, n=40) had severe arthralgia with arthritis of small joints of hands and feet (81%, n=34). Moreover, a generalized, Itchy maculopapular rash was present in 78% (n=33) of them. The appearance of skin rash only after 4-5 days of fever was characteristic in the majority of patients. The mean age of positive patients was 38 years and consisted of 48% (n=20) of males. Many (43%, n=18) of them were farmers having a mean monthly family income of Rs. 4867.00. Analysis of educational status revealed that 60% (n=26) of family members had educated up to G.C.E. O/L whereas only 26% (n=12) had completed G.C.E. A/Ls. Twenty eight (67%) positive patients had at least one or more CHIK infected family members in addition. Moreover, 95% (n=40) of them were surrounded by infected neighbours indicating active, intense transmission in the area. According to the results, the most predominant clinical features of CHIK were fever either with severe arthralgia or arthritis of small joints of hands and feet. Skin rash, though characteristic, appeared to develop 4-5 days after the infection. CHIK has mainly affected the most productive labour force in these areas with majority belonging to the middle class farming community with a low monthly income. Hence, the sources of income of the affected families were severely hampered by the CHIK outbreak. Therefore, non-fatal, CHIK may have a negative impact on the socio-economic status of the affected communities. "The staff of the Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Dr Richard Perera and the staff of Godakawela Hospital and Dr. Susanth Kariyawasam and the staff of Pallebadda Hospital are acknowledged".
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
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.
Evidence for emerging sulfadoxine-pyrimethamine resistance of Plasmodium falciparum isolated in the Northern Province of Sri Lanka
(Sri Lanka Association for the Advancement of Science, 2005) Hapuarachchi, H.A.C.; Dayanath, M.Y.D.; Abeysundara, S.; Bandara, K.B.A.T.; Abeyewickreme, W.; de Silva, N.R.
Genetic evidence of emerging sulfadoxine-pyrimethamine resistance of Plassmodium falciparum isolates in an operational area in the Northern Province of Sri Lanka
(Sri Lanka Association for the Advancement of Science, 2004) Hapuarachchi, H.A.C.; Dayanath, M.Y.D.; Abeysundara, S.; Bandara, K.B.A.T.; Abeyewickreme, W.; de Silva, N.R.
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 Available
Large-scale entomological assessment of Wuchereria bancrofti transmission by dissection and PCR-ELISA in Gampaha district, Sri Lanka
(Sri Lanka Association for the Advancement of Science, 2008) Wijegunawardana, N.D.A.D.; Gunawardene, Y.I.N.S.; Manamperi, A.; Hapuarachchi, H.A.C.; Bandara, K.B.A.T.; Abeyewickreme, W.
Entomological surveys are important tools for monitoring progress of lymphatic filariasis (Lf) eradication programs. In this study, dissection of Culex quinquefasciatus was compared with a Polymerase Chain Reaction - Enzyme Linked Immunosorbent Assay (PCR-ELISA) for pooled mosquitoes to assess filarial infection levels in the major vector of Wuchereria bancrofti in Gampaha district, following mass-treatment programme with diethylcarbamazine (DEC) and albendazole. Mosquitoes were collected in 30 sentinel and 15 non-sentinel sites in 15 Medical Officer of Health (MOH) areas of Gampaha district known for the presence of W. bancrofti transmission. Captured mosquitoes were dissected to determine the W. bancrofti larvae (L1, L2, L3). PCR was carried out using Deoxyribonucleic acid (DNA) extracted from mosquito pools (15 body parts/pool) utilizing primers specific for the Wb-SspI repeat. PCR products were analyzed by hybridization ELISA using fluorescein-labeled wild type specific probes. The prevalence of infected/infective mosquitoes in PCR pools (3pools/site) was estimated using the PoolScreenTM algorithm and a novel probability-based method. The prevalence of infected mosquitoes with L1-L2 larvae of W. bancrofti ranged from 0%-8.54% by dissection and point estimates of infection prevalence as assayed by PCR-ELISA, ranged from 0% - 25.4%. Mosquitoes collected from all MOH areas (80%, N = 12), except for Minuwangoda, Dompe and Ragama, were positive for W. bancrofti larvae, with a prevalence rate ranging from 0.78% to 16.97% in both methods. Of 30 sentinel sites, 43.3% (N = 13) were positive for W. bancrofti transmission whereas it was evident in 40% (N = 6) of non-sentinel sites. The proportion of positive pools detected by the PCR-ELISA assay was higher than that obtained by the dissection indicating that PCR-ELISA assay is more sensitive than the dissection method in detecting infected/infective mosquitoes. Also results of this study showed that autochthonous transmission of W. bancrofti continues in the Gampaha district despite completion of the 5 year mass drug administration (MDA) programme. Therefore, we emphasize the use of more sensitive tools such as PCR-ELISA to monitor the impact of the MDA programme on disease transmission. This study also emphasizes that control measures should be further continued until the microfilareamic population is reduced to a level which could interrupt transmission in the area. Financial assistance received from WHO/SEARO/TDR (grant no. SN 1152) and University of Kelaniya (Grant no. RP/03/04/06/01/2006) is acknowledged
Molecular Diagnosis for confirmation of Infectious Diseases in Sri Lanka in 2009
(Sri Lanka Association for the Advancement of Science, 2009) Hapugoda, M.D.; Bandara, K.B.A.T.; Dayanath, M.Y.D.; Wellawaththage, C.; Hapuarachchi, H.A.C.; Abeyewickreme, W.
Confirmation of infectious disease outbreaks in Sri Lanka is an important national requirement. Many clinicians as well as general practitioners find it difficult to confirm diagnosis of some infectious diseases only on clinical grounds. Molecular assays can rapidly confirm diagnosis at the early phase of diseases when aetiological agents are present and before antibody titers are at detectable levels. PCR-based assays are more sensitive and more specific than all conventional methods. Overall objective of this study was early, rapid and definitive laboratory confirmation of the aetiology of chikungunya, dengue, Japanese Encephalitis (JE), leishmaniasis, leptospirosis, malaria and West Nile Virus (WNV) through molecular assays. A rapid mobile investigation team equipped with the case definition, questionnaires, sample collection methods and diagnostic methods for each disease was established. This group visited outbreak areas and collected clinical and laboratory information and clinical samples from suspected patients at the early stage of symptoms: 1-5 days. Clinical samples were laboratory tested by disease specific molecular assays (PCR/RT-PCR). Clinical parameters of each disease were analyzed. Only chikungunya, dengue and leptospirosis outbreaks out of the above mentioned diseases were reported during the preceding six months in 2009. The team collected blood samples from clinically suspected cases of chikungunya (n=430), dengue (n=116) and leptospirosis (n=23) from different parts of the island. Molecular assays confirmed infections only in 81% (350/430) for chikungunya, 7% (8/116) for dengue and 9% (2/23) for leptospirosis in selected suspected patients. Reports of the confirmation of the disease outbreak by molecular assay were sent to the relevant health authority within two days to highlight the magnitude of the infection. These results showed importance of aetiological confirmation of infectious diseases by molecular assays. In conclusion, molecular diagnosis using a single clinical sample is important for rapid, definitive and early confirmation of aetiology of a particular infectious disease outbreak when serological methods are of little value at the early stage of infection. This is important for cost effective and efficient control of the outbreaks through proper clinical management.
Molecular Phylogeography and Clinical manifestations of Russell’s viper (Daboia russelii) in Sri Lanka
(Faculty of Graduate Studies, University of Kelaniya, 2015) Dissanayake, D.A.; Dissanayake, D.S.B.; Bandara, K.B.A.T.; Kularatne, S.A.M.; Karunathilake, D.G.K.J.K.; Rajapakse, R.P.V.J.
The Russell‘s viper, Daboia russelii has a relict distribution over large areas of Indian Sub Continent, Taiwan and the Lesser Sunda islands. Russell‘s viper is distributed with varying densities and occurrence throughout many natural and modified habitats in Sri Lanka. Bite of this species is associated with a high incidence of morbidity and mortality in Sri Lanka. There is scanty of information available about Russell‘s viper in Sri Lanka. Thus, preliminary study was carried out using clinical data obtained from the twenty eight patients admitted due to Russell‘s viper bite in General Hospitals in the Gampaha District. Further, thirty five DNA samples were collected from the tail tip of specimens of the Russell‘s viper from Gampaha, Kandy, Rathnapura, Galle, Anuradhapura and Kurunagala districts for molecular phylogeny. Then fragment of mtDNA sequencing fragments of cytochrome b genes were amplified. PCR and products were subjected. DNA sequencing for molecular analysis using raxmlGUI software. Results revealed that the clinical features following Russell‘s viper envenomation are follows; local swelling 86%, local necrosis 7%, coagulopathy 75%, neurotoxicity 32%, nephrotoxicity 14% and cardiac effects 3%. Mortality of Russell‘s viper bites are caused by complications like renal failure, neurotoxicity, respiratory failure, disseminated intravascularcoagulation, heart failure and other cardiotoxic effects. Alignment of Sequences of Cytochrome-b gene by maximum likelihood analysis revealed that two strongly supported monophyletic clades corresponding to Sri Lanka and the Indian subcontinent. However, Sri Lankan Russell‘s viper was nested within the clade of Indian Russell‘s viper group. There was genetic divergence (2%) between the Indian clades and the Sri Lankan clade. However, these haplotypes do not show any geographically significant pattern. Thus, it could be suggested that Indian and Sri Lankan Russell‘s viper are genetically closely related and origin may be India or Sri Lanka. Further studies are being carried out on morphometric, molecular taxonomic and venomous differences in different geographic location within the Sri Lanka and Asian region.
A Novel reverse transcriptase-polymerase chain reaction based-liquid hybridisation(RT-PCR-LH) assay for early diagnosis of dengue infection
(Sri Lanka Medical Association, 2003) Gunasekera, M.B.; Hapugoda, M.D.; Gunasena, S.; Subasinghe, S.A.S.C.; Bandara, K.B.A.T.; Khan, K.B.; Abeyewickreme, W.
BACKGROUND: Early definitive laboratory diagnosis of dengue is difficult with the tests in routine use at present. OBJECTIVE: To develop a reverse transcriptase-polymerase chain reaction based liquid hybridisation (RT-PCR-LH) technique for the rapid and early diagnosis of dengue. RESEARCH DESIGN: RT-PCR products of the NS3 gene of dengue virus prototypes and of a few positive sera for dengue virus by culture, were allowed to hybridise in liquid phase with a mixture of dengue specific radio-labelled oligonucleotides. The products were separated by PAGE and visualised by autoradiography. 78 suspected dengue sera were also tested by RT-PCR-LH method, and by IgM-ELISA and HAI tests, for comparison. RESULTS: Two DNA bands (approximately equal to 470 bp and approximately equal to 455 bp) specific to dengue virus, were observed. RT-PCR-LH assay takes only 24 h. Of the 78 suspected dengue acute sera tested, 45/78 were positive by RT-PCR-LH, 31/78 were positive by IgM-ELISA, and 14/78 had a HAI titre > or = 2560. Duration of fever was known in 72 cases, and infection was detected by RT-PCR-LH in 11/22 of cases with < 5 d fever and by IgM-ELISA in 1/22. In cases with 5 to 15 d fever RT-PCR-LH and IgM-ELISA/HAI titre > or = 2560 detected infection in 30/50 and 27/50 respectively. The 10 sera which were negative by RT-PCR-LH, but were positive by either IgM-ELISA or HAI titre > or = 2560 were all > 5 d fever cases. RT-PCR-LH together with IgM-ELISA were capable of detecting dengue infection in 56/78 of the suspected cases. CONCLUSION: RT-PCR-LH assay developed in this study appears to have an advantage over other diagnostic techniques for the early detection of dengue.
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 51
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.
Population data for CSF1PO, TPOX, THO1, D16S539, D7S820, D13S317, FESFPS, vWA and F13B short tandem repeat (STR) polymorphisms in Sri Lanka
(Sri Lanka Association for the Advancement of Science, 2004) Manamperi, A.; Gunawardene, Y.I.N.S.; Bandara, K.B.A.T.; Dayanath, M.Y.D.; Hapuarachchi, H.A.C.; Abeyewickreme, W.
Potential use of allele distribution at codon 51 of Plasmodium falciparum dihydrofolate reductase (pfDhfr) gene as evidence for early clinical failures to sulfadoxine-pyrimethaniine in an operational area in the Northern Province of Sri Lanka
(University of Kelaniya, Sri Lanka, 2004) Hapuarachchi, H.A.C.; Dayanath, M.Y.D.; Abeysundara, S.; Bandara, K.B.A.T.; Abeyewickreme, W.; de Silva, N.R.
Abstract available
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.
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.
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 cooperation
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.
Role of Aedes albopictus in transmitting dengue virus in some endemic areas in Kurunegala District
(University of Kelaniya, Sri Lanka, 2003) Hapugoda, M.D.; de Silva, N.R.; Abesundara, S.; Bandara, K.B.A.T.; Dayanath, M.Y.D.; Abeyewickreme, W.
Abstract Available