International Postgraduate Research Conference (IPRC)

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Author: search.filters.author.Jayaratne, D.L.

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    A Phage Indicator Model for In-Vitro Evaluation of Antiviral Drugs Used in Ayurvedic and Traditional Medicine
    (19th Conference on Postgraduate Research, International Postgraduate Research Conference 2018, Faculty of Graduate Studies,University of Kelaniya, Sri Lanka, 2018) Dayarathna, M.T.A.; Jayaratne, D.L.
    Finding new antiviral drugs for the emerging new viruses have become an outstanding scope of study. The analysis of anti-phage activity evaluation model based on plaque reduction assay has been performed by numerous researchers in evaluating antiviral properties of Western and Ayurvedic medicines. This has been performed by using phages with their corresponding hosts. This study was carried out to validate the antiviral sensitivity of a phage-indicator model to evaluate antiviral drugs used in Ayurvedic and traditional medical treatments in Sri Lanka The host bacterium Escherichia coli was isolated with its corresponding coliphage from sewage effluents. Prior to the antiviral assay, the antibacterial effect of standard antiviral drugs, Acyclovir, Lamuvidine, Ribavarin and Oseltamivir on E. coli was determined. According to the results up to 80mg/ml for Acyclovir, Lamuvidine and Ribavarin did not show any antibacterial effect while Oseltamivir showed only up to 20mg/ml. The antiviral assay was performed in two approaches where the corresponding test drug was applied to a young culture of host bacterium and incubated overnight prior to the inoculation of the bacterium by phage suspensions. As the second approach, the test drug was applied to a phage suspension and incubated overnight prior to the inoculation of phage into the bacterium. According to the reduction of plaques on agar plates after performing the two approaches the mode of antiviral activity was determined. There was a significant reduction of plaque formation observed when the host bacterium was treated with the western antiviral drugs, suggesting that their antiviral effects occur during the viral replication stages in the host cell and this complies with the available literature which describes the mode of action of these Western antiviral drugs. The model was also used to evaluate traditional herbal extracts to determine the presence of antiviral properties. Accordingly, the second approach revealed that the aqueous black tea extract showed an antiviral effect during the adsorption stages while crude papaya leaf extract in the replication stages of the virus life cycle. However, the Ayurvedic product ‘Sudarshana Churnaya’ showed no effects in either the two stages of the virus life cycle. With these results it is concluded that this model is feasible, and efficient against laborious cell culture techniques in screening antiviral drugs
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    Use of molecular features for identification of isolated fungal pathogens of big onion damping off disease and Trichoderma spp. isolated from soil
    (Faculty of Graduate Studies, University of Kelaniya, 2015) Gunaratna, L.N.R.; Deshappriya, N.; Jayaratne, D.L.
    Big onion (Allium cepa L.) is one of the economically important spices grown in Sri Lanka. Damping off disease caused by Fusarium sp. during nursery stage of growth poses a major factor that affect the yield significantly. Application of fungicides decrease incidence of damping off disease considerably, but this is neither economical nor environmental friendly. Thus, disease management practices have to be directed towards biological control strategies. Trichoderma spp. have been extensively studied as biological control agents for controlling numerous soil-borne fungal pathogens. In the present study, isolation and identification of fungal pathogens associated with damping off disease of onion and Trichoderma spp. present in soil of the same onion fields was carried out with a view to using the Trichoderma spp. in the management of damping off pathogens. Pathogens associated with damping off were isolated from diseased and healthy seedlings (7-30 days old) collected from the fields in the Matale and Anuradhapura districts. Seedlings were surface sterilized and plated in Potato Dextrose Agar (PDA) supplemented with tetracycline. Soil samples collected from the same fields were used for the isolation of Trichoderma spp. using the Warcup method. Based on morphological characteristics and using identification keys, the fungal pathogens isolated from seedlings were identified as Fusarium, Curvularia, Alternaria and Sclerotium spp. and 14 fungal species isolated from soil samples were identified as Trichoderma spp. Although fungi can be identified using morphological features, the use of molecular biological methods tend to be more accurate. Therefore, the identity of isolated fungal species was confirmed by molecular biological methods. Genomic DNA of Fusarium spp., Alternaria spp., Trichoderma spp. were extracted. Molecular characterization of these DNA was carried out using Polymerase Chain Reaction (PCR) where the Internal Transcribed Spacer (ITS) region of rDNA gene was amplified using ITS-1 and ITS-4 primer pairs. The products were subjected to agarose gel electrophoresis. The procedures were repeated 3 times. Results showed 550 bp size bands characteristic of Fusarium spp. and 570 bp products specific to Alternaria spp. confirming the previous identity using culture based methods. Fungal species isolated from soil showed products of 600 bp which corresponds to Trichoderma sp. Molecular characterization of the potential biocontrol agents i.e. Trichoderma spp. and A.cepa L. pathogens using PCR amplification of ITS region confirmed the preliminary identities carried out using culture based methods.