ABSTRACT
Background & objectives: Dengue fever (DF) is associated with significant morbidity and mortality in the tropical and sub-tropical regions of the world. Since there are no effective antiviral drugs for treatment, clinicians often rely on the accurate diagnosis of dengue fever to begin supportive therapy at early stages of the illness. The objective of this study was to develop an in-house dengue virus serotype 2 (DENV-2) non-structural protein- 5 (NS5) based indirect ELISA. Methods: DENV-2 was raised in Vero cells and the viral proteins were separated and subsequently the NS5 protein was eluted. Serum samples from primary and secondary dengue fever patients; and acute and convalescent samples from Japanese encephalitis (JE) and West Nile virus (WNV) cases were used to validate the ELISA. Results: The assay was found to be 100 per cent specific in detecting DENV-2 specific antibodies from patient’s serum. However, in terms of sensitivity, the assay could detect IgM antibodies only from 90 per cent of the primary dengue samples. The IgM/IgG ratio of the primary and secondary samples was 7.24 and 0.64, respectively. Interpretation & conclusions: The results indicate that the DENV-2 NS5 ELISA is dengue group specific and can be used to differentiate dengue infection from other circulating Flavivirus infections. This NS5 ELISA can also be used to distinguish between primary and secondary dengue fever on the basis of IgM/IgG ratios. Further studies with larger sample sizes and different DENV serotypes are required to validate the ELISA.
ABSTRACT
Objective:To obtain luteolin and apigenin rich fraction from the ethanolic extract ofCynodon dactylon (L.) (C. dactylon) Pers and evaluate the fraction’s cytotoxicity and anti-Chikungunya potential using Vero cells.Methods:The ethanolic extract ofC. dactylon was subjected to silica gel column chromatography to obtain anti-chikungunya virus (CHIKV) fraction. Reverse phase-HPLC and GC-MS studies were carried out to identify the major phytochemicals in the fraction using phytochemical standards. Cytotoxicity and the potential of the fraction against CHIKV were evaluatedin vitrousing Vero cells. Reduction in viral replication was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) after treating the viral infected Vero cells with the fraction.Results:Reverse Phase-HPLC and GC-MS studies confirmed the presence of flavonoids, luteolin and apigenin as major phytochemicals in the anti-CHIKV ethanolic fraction ofC. dactylon. The fraction was found to exhibit potent viral inhibitory activity (about 98%) at the concentration of 50 μg/mL as observed by reduction in cytopathic effect, and the cytotoxic concentration of the fraction was found to be 250 μg/mL. RT-PCR analyses indicated that the reduction in viral mRNA synthesis in fraction treated infected cells was much higher than the viral infected control cells.Conclusions:Luteolin and apigenin rich ethanolic fraction fromC. dactylon can be utilized as a potential therapeutic agent against CHIKV infection as the fraction does not show cytotoxicity while inhibiting the virus.
ABSTRACT
In this study, we report the evaluation of In-house fl avi virus immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA), which can be used as a screening test to determine the infecting fl avivirus serotype over the current serological methods. A panel of 88 sera (inclusive of well characterized dengue, Japanese Encephalitis (JE) and West Nile virus (WNV) positive and negative samples tested and confi rmed by commercial kit) was used for evaluation of the kit. The sensitivity and specifi city of the In-house capture assay versus the commercial kit for the sero-diagnosis of dengue was 100% and 87% respectively, for JE IgM, it was found to be 90% and 100% respectively, and for West Nile it was 87.5% and 90.9%. Based on the study, we concluded that this fl avivirus-serotyping ELISA provides rapid results and may be used as an accurate alternate to other serological tests for the specifi c diagnosis of fl avivirus infections.
ABSTRACT
Background: Dengue infection is emerging as a serious public health problem in Tamil Nadu. An enhanced surveillance system can generate information on the epidemiology of the disease, which is essential for planning and development of relevant control/preventive measures against Dengue. Materials and Methods: A prospective descriptive study was undertaken between January 2011 to December 2011, by testing suspected Dengue patients attending Thanjavur Medical College and Trichy Hospital (TMCH, a major Government referral hospital in Thanjavur District, Tamil Nadu, India) to define the magnitude of Dengue burden, the natural history of this disease in terms of clinical presentation and outcome of the infections in hospitalized Dengue patients. The sera collected from suspected patients were analyzed for Dengue specific IgM and IgG antibodies by IgM antibody capture enzyme linked immunosorbent assay (ELISA) using NIV kit and IgGPanBio Duo Rapid Immunochromatographic Card Test (Brisbane, Australia). The clinical case definition by World Health Organization was adopted to categorize the Dengue cases. Results: The total number of samples screened during the period was 200, out of which 79 (39.5%) were positive for IgM and IgG antibodies and 65 (32%) for IgM antibodies only. By clinical evaluation, Dengue fever was diagnosed in 43 patients, 18 had hemorrhagic manifestations and four patients had progressed to DSS. Though (DSS + DHF) was present in 22 patients, all of them recovered well. Conclusion: In developing countries like India, building of laboratory with advanced capacity for diagnosis and combat-mode ready preparedness for the management of Dengue cases in emergency situation may reduce Dengue-related mortality.