ICAPS-2021
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Item Molecular characterization of Sri Lankan tea [Camellia sinensis (L.) O. Kuntze] genotypes with diverse origins using SSR markers for future breeding programmes(Faculty of Science, University of Kelaniya, Sri Lanka., 2021) Kottawa-Arachchi, J.D.; Ranathunga, M. A. B.; Sharma, R. K.; Amarakoon, A. M. T; Gunasekare, M. T. K.; Chaudhary, H. K.; Attanayake, R. N.The tea plant [Camellia sinensis (L.) O. Kuntze] is one of the most popular non-alcoholic beverage crops worldwide. Tea germplasm is the most valuable and fundamental resource for breeding high yielding, climate resilient, and quality tea cultivars. Therefore, efforts were made to characterize the existing genetic diversity of Sri Lankan tea using simple sequence repeat (SSR) markers. A total of 91 tea accessions including exotic, estate selections, improved quality and diverse cultivars were selected for the diversity estimations. Eleven informative highly polymorphic SSR markers selected based on inferences of previous studies (EST, miRNA and traits associated SSR markers) were used in this study. Genetic relationships were analyzed by a neighbor-joining (NJ) tree constructed using DARwin software. Genetic structure analysis was performed by the Bayesian clustering model using STRUCTURE analysis and the optimum delta K was generated. Genetic diversity analysis grouped all the accessions into three major clusters. Further, structure analysis complemented the cluster analysis and revealed three genetic populations (China types with Korean origins, Assam types with ASM4/10 ancestral relationship and mixture of different origins) in the Sri Lankan tea germplasm. Newly recommended TRI5001, TRI5002, TRI5003 and TRI5004 cultivars have shown affinities with five Assam introductions and grouped together in the same cluster. Interestingly, five high yielding TRI developed cultivars (TRI2023, TRI2025, TRI3013, TRI3055 and TRI3073) having an ancestral relationship with the parent ASM4/10 were also grouped together. Likewise, seven accessions of Azerbaijan origin were remained together. The estate selections were scattered in the three clusters. The inferences of the current study provide molecular evidence for the genetic diversity and population structuring of the tea germpalsm and will help the selection of the potential parental group for the future cultivar development programme of Sri Lanka.Item Bioactive properties and metabolite profile of an endolichenic fungus, Hypoxylon lividipigmentum(Faculty of Science, University of Kelaniya, Sri Lanka., 2021) Weerasinghe, W. R. H.; Shevkar, C. D.; Silva, R. S. De; Attanayake, R. N.; Weerakoon, G.; Kate, A. S.; Kalia, K.; Paranagama, P. A.Endolichenic fungi (ELF) serve as a novel source of secondary metabolites. Hypoxylon lividipigmentum is an ELF isolated from the lichen Opegrapha medusulina, collected from mangrove plant Xylocarpus granatum from Negombo lagoon, Sri Lanka. The fungus was identified to the species level using morphological and DNA barcoding techniques. Ethyl acetate extract of the fungus was subjected to in vitro assays to determine antioxidant, anti-inflammatory, tyrosinase inhibitory and antibacterial potency. Liquid Chromatography-Mass Spectrometry (LCMS) dereplication was conducted on the crude extract in order to detect the secondary metabolites present. The extract reported a IC50 value of 18.34±1.37 µg/ml on par with the positive control BHT, in DPPH radical scavenging assay. It also exhibited moderate anti-inflammatory activity with an IC50 value of 81.08±1.05 μg/ml. Tyrosinase inhibitory activity was fairly comparable with an IC50 value of 121.20±2.55 μg/ml. Agar well diffusion assay was conducted to determine antibacterial activity against aerobic bacterial species Escherichia coli, Bacillus subtilis, Staphylococcus aureus and the anaerobic bacterial species Streptococcus mutans. Suppression of growth was shown only against B. subtilis. Five major mass peaks were observed during the study of LCMS profile of the extract. After a thorough dereplication process, two masses could be presumed to be from novel scaffolds. Since none of the mass peaks could be dereplicated within the species or genus level, it could be speculated that the chemical profile of Hypoxylon lividipigmentum was previously poorly explored in literature thus making it an interesting organism to study further for novel metabolites.Item Biodeterioration of low density polyethylene by Montagnula scabiosae(Faculty of Science, University of Kelaniya, Sri Lanka, 2021) Perera, T. W. N. K.; Attanayake, R. N.; Paranagama, P. A.Endolichenic Fungi (ELF) are a group of fungi adapted to live within lichen thalli. Present study investigates the ability of Montagnula scabiosae, an ELF, for biodeterioration of low density polyethylene (LDPE) by determining the activity of three extracellular fungal enzymes, reported to have potential roles in depolymerizing polyethylene, using enzymatic assays. Liquid medium screening assays, using four potato dextrose agar (PDA) plugs (0.5 cm diameter) of ELF cultures, inoculated into Erlenmeyer flasks containing sterile mineral salt medium broth (MSMB). Eight sterile LDPE strips (10 cm x 3 cm) were added into each flask and incubated at room temperature (28 ± 2°C) for 21 days. MSMB flasks with plain PDA plugs and sterile LDPE strips were used as the controls. Solid medium biodeterioration studies also were conducted using three PDA plugs of the ELF cultures inoculated into mineral salt medium agar (MSMA) plates. Eight sterile LDPE strips were aseptically placed on the surface of each MSMA plate and incubated at room temperature for 45 days. MSMA plates with plain PDA plugs and sterile LDPE strips were used as the controls. Both liquid and solid media assays were carried out in four replicates. LDPE biodeterioration was analyzed based on reductions in weights and tensile properties, changes in peaks of Attenuated Total Reflection-Fourier Transform Infrared (ATR-FTIR) spectra, changes in carbonyl index (CI) and Scanning Electron Microscopic (SEM) images. A distinguishable LDPE deterioration ability was shown by Montagnula scabiosae, isolated from the host lichen Pyrenocarp sp. Results showed that isolate reduced weights of strips by 13.22% in liquid medium and by 12.06% in solid medium. Tensile strength of liquid medium treated strips, reduced by 6.64% and solid medium treated strips, by 3.00%. The control LDPE strips on ATR-FTIR analysis showed four prominent peaks at 2916 cm-1, 2848 cm-1, 1463 cm-1 and 720 cm-1 and a few minor peaks. Changes visible in the ATR-FTIR spectra of treated strips, manifested new peaks, fluctuations in percentage transmittance and deletions of existing peaks. CI value of the LDPE strips in liquid medium were found to be increased by 2.70% and in solid medium increment was 2.31%. SEM images of treated strips showed erosion zones on LDPE matrix. Qualitative enzymatic assays using Agar plate-based screening methods, indicated the presence of laccase and peroxidases, in this ELF isolate. In the quantitative enzymatic assays, M. scabiosae showed laccase, lignin peroxidase and manganese peroxidase activities as 1.07×10-7 kat L-1, 1.22×10-8 kat L-1 and 2.10×10-8 kat L-1 respectively. Remarkable LDPE deterioration ability shown by this ELF isolate indicates that it has a high potential in degrading LDPE waste.