Conference Papers

Permanent URI for this collectionhttp://repository.kln.ac.lk/handle/123456789/6561

This collection contains abstracts of conference papers, presented at local and international conferences by the staff of the Faculty of Medicine

Browse

Search Results

Now showing 1 - 10 of 25

Molecular evidence of hantavirus infection among clinically suspected patients with hae-morrhagic fever with renal syndrome (HFRS)
(Sri Lanka College of Microbiologists, 2013) Muthugala, M.A.R.V.; Manamperi, A.A.P.S.; Gunasena, S.; Hapugoda, M.D.; Butch, G.
INTRODUCTION: Hantavirus disease is an emerging zoonotic viral infection with high fatality. Transmission is by inhalation of aerosols generated from virus contaminated rodent excreta. There are two major clinical forms, haemorraghic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Clinical features of HFRS, often mimic leptospirosis. Large number of cases of leptospirosis like illness has been reported in Sri Lanka annually. Although there were serological evidence of different types of hantavirus infection in Sri Lanka, diagnosis of hantavirus is not routinely performed. Due to the genetic and antigenic diversity, an assay that could detect a wide range of hantaviruses need to be established. OBJECTIVES: To establish, evaluate and validate a genus specific hantavirus RT-PCR assay. To diagnose hantavirus infection among clinically suspected HFRS patients in three selected hospitals.To describe clinical manifestations of hantavirus infections in the study population. METHODOLOGY: Genus specific conventional RT-PCR assay was established using panhanta primers and evaluated, optimized and validated using synthetic genes of 12 known hantavirus species as reference samples. Assay was able to detect a wide range of hantaviruses at minimum detection limit of 70 copies/ reaction. Molecular diagnosis of hantavirus infection was carried out in three hospitals in Colombo and Gampaha districts. Study was conducted from 01st of January 2011 to 31st of April 2011 and 61 adult patients were recruited to this study. Hantavirus RT-PCR was performed on all collected samples after extraction of RNA by TRIzol® method. RESULTS: Of 61 tested samples, 05 were positive for hantavirus genome. These results were confirmed at reference laboratory as well and species identification result is pending. Of 58 tested samples, 06 samples were positive for hantavirus IgM by in-house ELISA. All PCR positive samples were positive for hanta virus IgM. All patients with hantavirus infection had clinical and biochemical features of liver involvement in addition to fever, thrombocytopenia and renal involvement. CONCLUSION: Established RT-PCR assay was able to detect a wide range of hantaviruses and by using it molecular evidence of hantavirus infection was demonstrated in humans in Sri Lanka. Further studies are required to describe the disease epidemiology and to identify natural hosts in the country.
Clinical and virological features of dengue in 2010
(Sri Lanka College of Microbiologists, 2011) Hapugoda, M.D.; Manamperi, H.; Gunasena, S.; Athapaththu, A.M.M.H.; Premawansa, G.; Wellawaththage, C.; Jayarathna, T.D.S.S.; Abeyewickreme, W.
INTRODUCTION: Dengue is an important viral infection in Sri Lanka. All 4 serotypes co-circulate in Sri Lanka. OBJECTIVE: To study the clinical and virological features of dengue in 2010. DESIGN, SETTING AND METHODS: A hospital-based study was carried out at North Colombo Teaching Hospital, Ragama in 2010. Patients clinically suspected of having dengue, with fever less than 5 days were recruited. Acute and convalescent blood samples were collected within 7 days after obtaining informed written consent. Demographic, clinical information and laboratory results were obtained. Acute serum samples were tested using molecular (RT-PCR and Semi-Nested PCR) and serological (ELlSAs and HAI) assays. Convalescent samples were tested by serological assays. RESULTS: Of 209 patients enrolled, 93 % (195/209) were laboratory confirmed as recent positive cases of dengue viral infection; of these, 5% (9/195) were classified as dengue fever; 85%(1G5/195) dengue haemorrhagic fever (DHF) and 0.5% (1/195) dengue shock syndrome. Mean platelet value and packed cell volume (PCV) in laboratory confirmed dengue patients were 56,107/mm3 (range 10,000-306,000) and 42%(range 34-61 %) respectively. Patients infected with DHF showed both primary (n=45) and secondary (n=102) infections. Interestingly, secondary infection was not significantly correlated with DHF (x2-0.3:p=0.6). DEN-1 was responsible for the majority of cases, with a minority due to other three serotypes; all serotypes contributed to severe disease. CONCLUSION: DEN-1 was responsible for the majority of cases in 2010 but it circulated at a low level during previous epidemics. Majority of patients had severe clinical symptoms. In this epidemic, the clinical presentation of dengue differed according to the geographic region and viral serotype. ACKNOWLEDGMENTS: Financial assistance and technical co-operation by International Center for Genetic Engineering and Biotechnology (ICGEB CRP SRL 08/02), National Science Foundation (NSF/RG/2009/BT/01) and International Atomic Energy Authority (lAEA/SRL/5/042) is acknowledged.
Comparison of recombinant protein and cell lysate antigens for detection of anti-chikungunya (CHIK) IgM antibody
(Sri Lanka College of Microbiologists, 2011) Athapaththu, A.M.M.H.; Khanna, N.; Inouve, S.; Gunasena, S.; Abeyewickreme, W.; Hapugoda, M.D.
INTRODUCTION: Chikungunya (CHIK) virus specific antigen which has high specificity and low cross reactivity with other related diseases is required for laboratory confirmation. OBJECTIVE: To compare two antigens for detection of anti-CHIK antibody. DESIGN, SETTING AND METHODS: In this study, two antigens {viral cell lysate and recombinant protein) were evaluated for detection of anti-1 CH IK antibody by using IgM ELISA. A novel recombinant | protein antigen was designed based on envelope domain, a critical antigenic region of the major structural protein, I This protein was expressed in Escherichia coli and resultant protein was affinity purified and ~10mg with >95% of purity per liter of culture was obtained. Cell lysate antigen was prepared using a crude culture fluid. Two antigens were evaluated separately using a panel of well characterized serum samples obtained from the Dept. of Virology (WHO Reference Centre for Viral Reference and Research), Institute of Tropical Medicine,] Nagasaki University. RESULTS: Atotal of 64 serum samples confirmed as positives andl 22 confirmed as negatives were used to evaluate the antigens. Specificity and sensitivity of the recombinant protein antigen was 48% and 90% respectively. Specificity and sensitivity of the viral lysate antigen was] 17% and 100% respectively. Conclusion Viral lysate antigens can cause biohazard risk, high production cost and cross reactivity with other organisms of the same genus/family. Recombinant protein antigen which shows high specificity and sensitivity used in this study is important to overcome problems associated with viral lysate antigen. Testing of a large number of samples is needed to reconfirm this finding. ACKNOWLEDGMENT: Financial assistance and technical co-operation by International Center for Genetic Engineering and Biotechnology (ICGEB CRP SRL 08/02), National Science Foundation (NSF/RG/2009/BT/01) and International Atomic Energy Authority (IAEA/SRL/5/042) is acknowledged.
Detection of dengue virus in Aedes albopictus mosquitoes by Reverse Transcription Polymerase-Chain Reaction-Liquid Hybridization (RT-PCR-LH) based assay.
(Sri Lanka College of Microbiologists, 2003) Hapugoda, M.D.; Gunasekera, M.B.; de Silva, N.R.; Gunasena, S.; Prithimala, L.D.; Dayanath, M.Y.D.; Abeyewickreme, W.
Dengue is an important public health problem. In this study an RT-PCR-LH assay was developed for the detection of dengue virus in Ae.albopictus, a vector of dengue. Laboratory bred Ae.albopictus (adults inoculated with dengue prototypes were tested by RT-PCR-LH assay. RT-PCR products of NS3 gene of 4 dengue prototypes were hybridized in liquid phase with 32P) labelled cocktail of dengue serotype-specific ologonucliotides. Semi-Nested-PCR agarose gel electrophoresis (Semi-Nested-PCR-AGE) assay with dengue type specific oligonucliotides was carried out for typing of RT-PCR products. Wild-caught Ae.albopictus (larvae (n=89 pools) and adults (n=69 pools) collected from dengue case reported stations during the period of 1999-2002 were also tested by RT-PCR-LH and typed by Nested-PCR-AGE assay). A DNA band (470bp) specific for dengue virus was observed in all pools of Ae.albopictus (inoculated with dengue prototypes in RT-PCR-LH assay. When RT-PCR products of dengue prototypes inoculated mosquitoes were typed by Semi-Nested-PCR-AGE assay, bands of 169,362, 265, 426 bp sizes corresponding to DEN1, DEN2, DEN3 and DEN4 respectively were observed. The DNA band specific for dengue virus (470bp) was also observed in 6 pools of wild-caught adults in RT-PCR-LH assay. They were found to be infected with DEN3 (265bp DEN3 specific DNA band was detected) by Semi-Nested-PCR-AGE assay. None of the wild-caught larvae showed dengue specific DNA band (470bp) in RT-PCR-LH assay). RT-PCR-LH with Semi-Nested-PCR-AGE assays are useful for the detection and typing of dengue virus in Ae.albopictus. Ae.albopictus (in Sri Lanka is competent in transmitting DEN3 and possibly other serotypes. Detection of dengue virus for the first time in Ae.albopictus in Sri Lanka confirms earlier observations that it may play an important role in transmitting dengue). Acknowledgements: Financial assistance by the International Atomic Energy Agency (Technical Co¬operation grant no SLR/ 06 / 024) and University of Kelaniya (Research grant no RP/03/04/06/01/00) is gratefully acknowledged.
Enzyme-linked Immunosorbent Assay (ELISA) using recombinant protein antigens for detection of anti-chikungunya antibodies
(Faculty of Tropical Medicine, Mahidol University, 2010) Athapaththu, A.M.M.H.; Khanna, N.; Abeyewickreme, W.; Gunasena, S.; Hapugoda, M.D.
OBJECTIVES: Chikungunya is a mosquito borne viral infection that has caused great medical and public health problems in South East Asia during last few years. Currently available laboratory diagnostic kits depend on Enzyme-Linked Immunosorbent Assay (ELISA) based on whole viral antigens caused biohazard risk, high production cost and cross reactivity with other organisms of the same genus/family. These problems can be avoided by using recombinant protein antigens in ELISAs. METHODOLOGY: Two novel recombinant protein antigens based on Envelope (E) domain, a critical antigenic region of the major structural protein of chikungunya virus were expressed separately in a bacterial expression system (Escherichia coli). Two proteins were purified under denatured conditions. They were evaluated as potential diagnostic intermediates for detection of and-chikungunya antibodies in Immunoglobulin M (IgM) and Immunoglobulin G (IgG) ELISAs separately using a panel of serum samples confirmed by the gold standard assay, Heamagglutination Inhibition (HAI) assayRESULTS: These 2 protein antigens: El and E2 showed more than 60% positivity in IgG ELISAs and IgM ELISAs. A field validation using a large number of serum samples should be done for further confirmation of these results. It can be concluded that these 2 novel recombinant protein antigens can be used as a diagnostic intermediate to detect anti-chikungunya antibodies. ACKNOWLEDGEMENTS: Financial assistance from the International Centre for Genetic Engineering and Biotechnology (1CGEB CRP/ SRI08-02) is gratefully acknowledged
Shifting of circulating serotypes in dengue outbreaks during 2009/2010 in Sri lanka
(Faculty of Tropical Medicine, Mahidol University, 2010) Manamperi, N.H.; Athapaththu, A.M.M.H.; Premawansa, G.; Wellawaththage, C.; Jayarathna, T. D. S. S.; Abeyewickreme, W.; Hapugoda, M.D.
OBJECTIVES: Sri Lanka has experienced explosive outbreaks of dengue infection in 2009 and 2010. It has been identified that DEN- 3 and DEN- 2 were the predominant serotypes with DEN-1 and DEN- 4 circulating at a lower level in previous dengue outbreaks during 2003-2006, Objective of this study was to identify the circulating serotype/s during 2009 - 2010 outbreaks. METHODOLOGY: A prospective study was carried out at North Colombo Teaching Hospital, Sri Lanka during December 2009-August 2010. Clinically suspected dengue patients, with fever less than 5 days were recruited. An interviewer administered questionnaire was filled for each patient, by a Medical Officer. Venous blood samples confirmed for the presence of dengue virus by RT-PCR were typed by Semi-Nested PCR. RESULTS: Out of the 209 patients recruited in the study 80 (38%) were positive for dengue virus by RT-PCR. Of the positives, 43 (54%) were typed and circulation of all 4 serotypes was observed- Of the 43 positives, presence of DEN-1, DEN-2, DEN-3 and DEN-4 serotypes was 34 (79%), 3 (7%), 2 (5%) and 3 (7%) respectively DEN-1 was the predominant serotype in the recent epidemics which was circulating at a low level in previous epidemics. In DEN-1 infected patients, the mean platelet value was 58,588/ rnm3 and the mean PCV value was 41.4%. Associated symptoms such as headache, retro-orbital pain, neck pain and limb pain were present in 94% (32/34), 59% (20/34), 24% (8/34J and 91% (31/34) patients respectively. Bleeding manifestation developed in 47 % (16/34) patients. The mortality rate ranged from 0.7%- 1.0% during the recent outbreaks. Acknowledgement: Financial and technical assistance from the International Centre for Genetic Engineering and Biotechnology (ICGEB CRP/ SRI08-02) and the International Atomic Energy Agency (IAEA SRI 5/042) is gratefully acknowledged.
Tetravalent dengue specific domain III based chimeric recombinant protein as dengue diagnostic intermediates for the detection of both anti-dengue immunoglobulin M(IGM) and imunoglobulin G(IGG) antibodies in human serum samples.
(International Water Management Institute, 2006) Hapugoda, M.D.; Abeyewickreme, W.; Gunasena, S.; Khanna, N.
BACKGROUND: Dengue infection is an important mosquito borne viral infection caused by four serotypes of dengue virus with explosive outbreaks occurring in many tropical areas. Laboratory diagnosis of the disease mainly depends on Enzyme-Linked Immunosorbent Assay (ELISA) based on whole viral antigens which cause biohazard risk, high production cost and cross reactivity with other flaviviruses. OBJECTIVES: To produce a recombinant protein antigen to overcome problems associated with whole dengue viral antigen/lysate or recombinant whole envelope protein. STUDY DESIGN: We have designed and expressed a single recombinant tetravalent protein antigen which contains Domain III of envelope protein from all four serotypes of dengue virus, linked with each other through penta glycine linkers. This synthetic gene was expressed in Escherichia coli and protein was purified using a single affinity chromatographic step. We developed Immunoglobulin M (IgM) and Immunoglobulin G (IgG) ELISAs using this novel protein as the capture antigen. The antigen was validated as a diagnostic reagent on serum samples. RESULTS: 30 mg of recombinant protein per litre of culture could be purified. Both ELISAs developed using this novel recombinant protein showed an excellent agreement with a commercially available IgM ELISA (MRL diagnostic) and haernagglutination inhibition assay respectively. Conclusions: Findings of this study suggests that this single dengue specific tetravalent recombinant protein antigen can be used as a diagnostic intermediate for detection of dengue infection.
Dengue vector surveillance in a dengue hot-spot in Sri Lanka
(Faculty of Tropical Medicine, Mahidol University, 2007) Sumanadasa, S.D.M.; Hapugoda, M.D.; Perera, D.; Bandara, S.; Mansoor, M.A.; Peris, I.; Abeyewickreme, W.
BACKGROUND: In South Asia, dengue has been declared as one of the most, fast-spreading vector-borne diseases. Therefore, mosquito surveillance is important for early detection of outbreaks along with implementation of prompt control activities. OBJECTIVES: To identify entomological risk factors with regard to transmission of dengue in a dengue hot-spot. Seventy five human dwellings in Vehara in the Kurunegala District of the Western Province were selected based on high disease incidence during 2000-2004, high Aedes as well as human population density and increased building activities. Entomological surveillance was done during May-August, 2007. RESULTS: The house Index ranged from 2.67% to 5.33% for Aedes aegypti while it for Aedes albopictus was 1.33% to 6.60%. The container index ranged from 23.67% to 29.33% for Ae. aegypti and from 1.33% to 18% for Ae. aibopictus. Man biting rates of 0.43-5.78 bites/man/hour were estimated for Ae, aegypti, while it ranged between 0.49 and 1.33 for Ae. aibopictus. The most common breeding place for Aedes species was plastic baskets (16%, n=12). DISCUSSIONS: Vector surveillance showed that the predominant vector species present in the study area was Ae. ageypti. Aedes mosquito larval densities and adult biting rates were sufficient to promote outbreaks of dengue in this study area. Community must be educated regarding effective measures to protect them from dengue. Their cooperation should be elicited in the early detection and elimination of vector species by source reduction, environmental management and personal protection measures.
Correlation between clinical and laboratory diagnosis of dengue in Sri Lanka.
(Faculty of Tropical Medicine, Mahidol University, 2007) Hapugoda, M.D.; Khan, B.; de Silva, N.R.; Gunasekera, J.; Abeyewickreme, W.
BACKGROUND: In Sri Lanka, diagnosis of dengue mainly depends on clinical signs and symptoms. Very few suspected patients from the state and private sector health institutions are tested by laboratory diagnostic assays compared to the number of dengue cases recorded all over the island. OBJECTIVES: To correlate clinical parameters with laboratory diagnosis in confirmation of dengue. RESEARCH DESIGN: Patients, clinically suspected of having dengue (n=201) were selected based on WHO criteria. Serum samples were tested using major 3 types of laboratory diagnostic assays; molecular, virus isolation and serology. Differences in clinical and laboratory data were analyzed on the basis of the final diagnosis assigned as dengue or non-dengue. Chi-square test was used for comparison of data. RESULTS: The proportion of laboratory diagnosed dengue patients were 80% (162/201). Mean platelet value and PCV in laboratory confirmed dengue patients were 92 247/mm3 (range 20 000-318 000) and 45% (range 31-59%) respectively. On comparison of the presence of clinical features that are used by the WHO for diagnosis of dengue, headache (129/162 vs 18/39, x2=23, p=0.00), limb pain (107/162 vs 18/39, x2=4.56, p=0.03) and external bleeding (67/162 vs 00/39, X2=27, p=0.00) showed significant association, with dengue infection. The infection was confirmed as definitive dengue in 75% (121 /162) and probable dengue in 25% (41/162). DISCUSSION: Surveillance based on clinical diagnosis may result in over estimation of the disease as clinical diagnosis is not specific enough. Laboratory confirmation of dengue suspected patients is important to measure the real incidence of the disease is needed in country like Sri Lanka.
Dengue as-a public health problem in Sri Lanka
(La Fondation pour l’Université de Lyon, 2009) Hapangama, H.A.D.C.; Gunawardene, Y.I.N.S.; Hapugoda, M.D.; Premaratna, R.; Manamperi, A.; Gunasena, S.; Abeyewickreme, W.
Dengue infection is an important global public health problem and an increasing number of persons from the South Asian region have been directly or indirectly affected by the disease. In Sri Lanka, dengue has become a major threat to public health in many urban and sub-urban' areas during past three decades. Rapid unplanned urbanization and increasing human population has increase the rate of infection and the frequency. The study area, Gampaha District is the second most populous district in the country having a population density of 1 539 persons per km2 and was the district reporting the second highest incidence of dengue in 2008. Therefore, current research efforts are focused on dengue transmission, examining the presence of sub-clinical infections, role of vector mosquitoes and Knowledge, Attitude and Practices (KAP) of the community on dengue infection in an effort to contain the disease. In the present study, dengue antibodies were detected in samples collected from clinically suspected patients and as well as in samples collected from volunteers. Volunteer sera collected around the confirmed cases had a 23.6% sero-positive rate for dengue IgM antibodies. The rate of asymptomatic recent infections was calculated to be 16.9%. In present study we have serologically confirmed the presence of subclinical infections and according to the published data this is the first confirmation of asymptomatic dengue infections in Sri Lanka. According to the entomological investigations carried out, the common breeding places for Aedes vectors were found to be discarded small containers. Even though Ae. Aegypti has been considered as the principal vector transmitting dengue fever, current studies highlighted the predominant ro!e of Ae. albopictus in the disease transmission. A previous study in Sri Lanka also suggested that prevalence and .presence of high-density of Ae. albopictus should be considered as a risk factor for endemic/epidemic dengue. In view of the above, the spread of dengue by Ae. albopictus should be a matter of great concern. Findings of KAP survey revealed that the community possessed substantially higher knowledge on the spread of dengue, vectors, vector breeding and also seriousness of the infection. However it was observed that good knowledge does not necessarily lead to good practices. Since the attitudes of the respondents were found to be good and most of them were supportive of control measures; next effort of the present study is to see how a novel community mobilized solid waste management system will be effective in dengue vector control.

Conference Papers

Browse

Filters

Settings

Sort By

Results per page

Search Results