Browsing by Author "Wanigatunge, R. P."

Now showing 1 - 6 of 6

Assessment of human health risk of cylindrospermopsin by consuming Oryza sativa (rice) from selected CKDu endemic areas in Sri Lanka
(Faculty of Science, University of Kelaniya Sri Lanka, 2022) Bandara, W. M. S. N.; Wanigatunge, R. P.; Rajapaksha, A. U.; Vithanage, M. S.; MaganaArachchi, D.N.
The proliferation of toxic cyanobacterial blooms in freshwater bodies can seriously threaten human health. Many studies have been carried out on cyanotoxins in water sources, but studies on cyanotoxins in terrestrial plants are limited. Cyanotoxins in water sources can enter terrestrial plants when irrigated with cyanotoxin-contaminated water and cultivated in cyanotoxin-contaminated soil. As the main staple food in Sri Lanka, Oryza sativa (rice) is widely grown island-wide, mainly in the country's dry zone. The rice fields are irrigated with raw water from the reservoirs throughout their cultivation. Previous literature reported the presence of cyanotoxins, and potential cyanotoxin-producing cyanobacterial species in reservoirs in the dry zone in Sri Lanka. Moreover, cyanotoxins are hypothesized as one of the risk factors for chronic kidney disease of unknown aetiology (CKDu) in Sri Lanka. Cylindrospermopsin (CYN) is a cyanotoxin and negatively affects on several organs in the human body, including the kidneys. Therefore, the current study investigated CYN in O. sativa samples (n=102), including the most cultivated variants, samba (n=20), nadu (n=61), rathu kekulu (n=12) and keeri samba (n=9) collected from CKDu endemic Girandurukotte and Dehiattakandiya areas. As a control, rathu kekulu samples (n=22) were collected from CKDu non endemic Sewanagala area. CYN was extracted and analysed by high-performance liquid chromatography (HPLC) with reference to the CYN standard (SIGMA ALDRICH 32087). Estimated Daily Intake (EDI) of CYN was then calculated to determine the potential human health risk, for an adult by consumption of CYN-contaminated rice. Results of the HPLC analysis revealed that the mean ± SE values of CYN concentrations in samba, nadu, rathu kekulu, keeri samba rice from CKDu endemic areas and rathu kekulu from the control area were 6235.74 ± 1289.15, 6821.44 ± 694.80, 6538.66 ± 1797.81, 3702.51 ± 1321.02, and 3460.99 ± 359.91 μg/kg, respectively. Mean ± SE of estimated daily intake when exposure to CYN via consumption of samba, nadu, rathu kekulu, keeri samba rice in CKDu endemic areas and rathu kekulu from the control area were 30.45±6.29, 33.31±3.39, 31.93±8.77, 18.08±6.44 and 16.90±1.75 μg/kg of body weight per day, respectively. All these values exceeded the provisional Tolerable Daily Intake (TDI) of CYN value established by the World Health Organization (0.03 μg/kg of body weight per day), which can pose a health risk to consumers. The present study revealed that the consumption of O. sativa in the studied areas has a potential risk of accumulation of CYN in the human body. However, it emphasized the importance of investigating the uptake of CYN into rice, in large sample sizes in the study areas.
Cyanotoxins uptake and accumulation in crops: Phytotoxicity and implications on human health
(Toxicon, 2022) Weralupitiya, C.; Wanigatunge, R. P.; Gunawardana, D.; Vithanage, M.; Magana-Arachchi, D.
The invasive nature of cyanotoxin-producing cyanobacteria and the adverse effects concerning their toxic impacts have gained heightened scientific attention of late. The persistence of cyanotoxins in irrigation water leads to bioaccumulation in plants, the development of phytotoxic effects, and the threat of groundwater contamination. The accumulation of cyanotoxins in plants is caused by several factors leading to severe toxic effects, including reduced plant growth and seed germination, enhanced oxidative stress, lowered rate of mineral uptake, decreased photosynthetic efficiency, and loss of chlorophyll content. The uptake and accumulation of cyanotoxins in plants can be concentration-dependent, as reported in a myriad of studies. Even though several studies have reported phytotoxic effects of cyanotoxin contamination, field-related studies reporting phytotoxic effects are particularly inadequate. Paradoxically, at realistic conditions, some plants are reported to be tolerant of cyanotoxins. Furthermore, the breadth of adverse impacts of cyanotoxins on human health is significant. Cyanotoxins cause major health effects including cancer, oxidative stress, organelle dysfunction, DNA damage, and enzyme inhibition. This review intends to present compelling arguments on microcystins (MCs), cylindrospermopsins (CYN), β-N-methylamino-L-alanine (BMAA), and anatoxin-a (ANTX-a), their uptake and accumulation in crop plants, phytotoxic effects on plants, and potential health implications to humans. The accumulation of cyanotoxins implants cultivated as food crops, resulting in phytotoxic effects and adverse impacts on human health are serious issues that require scientific inputs to be addressed.
Detection of microcystins (cyanotoxin) in selected drinking water wells in the Gampaha district, Sri Lanka
(Faculty of Science, University of Kelaniya Sri Lanka, 2023) Sinhapitiya, S. P. A.; Herath, H. M.; Bandara, W. M. S. N.; Rajapaksha, A. U.; Vithanage, M. S.; Magana-Arachchi, D. N.; Wanigatunge, R. P.
The prevalence of toxic cyanobacteria and various cyanotoxins, such as microcystins (MCs) in drinking water sources have gained considerable attention worldwide due to their potential health risk. Approximately 80% of the Sri Lankan rural population and 34% of the urban population rely on well water for their daily needs. Therefore, people could be exposed to cyanotoxins by consuming well water. This study investigated the presence of potentially toxic cyanobacteria and microcystin variants in the household well water samples collected from the 13 divisional secretariats in the Gampaha district, the second-most populated district in Sri Lanka. Twenty-six well water samples were collected, and physicochemical parameters such as temperature, salinity, pH, conductivity, and Total Dissolved Solids (TDS) were measured in triplicates, in situ using a multi-parameter. All samples met the Sri Lankan Standards (SLS) for drinking water in terms of pH, salinity, temperature and TDS. One well water sample collected from Wattala did not meet the SLS standards in terms of conductivity (829.00±2.05 μS/cm), indicating the well’s unsuitability for water consumption. Morphological identification of cultures originating from well water samples in cyano-specific BG11 medium indicated the presence of nine cyanobacterial genera, including Gloeobacter, Myxosarcina, Dermocarpa, Xenococcus, Synechococcus, Pseudanabaena, Chroococcus, Lyngbya and Geitlerinema. Among them, Synechococcus and Pseudanabaena are reported to be microcystin producers. The presence of extracellular MCs in water samples was detected by High-Performance Liquid Chromatography (HPLC). Extracellular MCs were extracted from water samples using 70% methanol. MC variants and their concentrations in each well water sample were determined using MC-RR-YR-LR standard mixture (Cat no. 33578; SIGMA ALDRICH) by comparing peak retention times and the area of the peaks. HPLC analysis revealed the presence of the microcystin variant MC-LR only in two water samples collected from wells located in the Divulapitiya and Katana areas with MC-LR concentrations of 195.1 μg/L and 278.3 μg/L, respectively. These values exceeded the standard guideline value of 1 μg/L for MC-LR set by the World Health Organization for drinking water. This indicates the potential health risk for consumers of those wells because there is a potential for hepatotoxic MCs to cause acute and chronic illnesses in humans.
Morphological characterization and molecular identification of three fungal pathogens isolated from Solanum melongena L. in Sri Lanka.
(Journal of Agricultural Sciences – Sri Lanka, 2023) Koshila, H. V.; Wanigatunge, R. P.; Dias, R. K.; Edirisinghe, P.
Purpose: Solanum melongena L. (brinjal/ eggplant) is a popular vegetable in Sri Lanka. Like many solanaceous crops, brinjal fruits and plants are susceptible to various diseases. The objective of this study was to isolate, confirm pathogenicity and characterize fungal pathogens causing leaf blight, yellowing and necrosis in S. melongena plants in selected localities in Gampaha district, Sri Lanka. Research Method: Fungi associated with symptomatic S. melongena plants collected from three localities in Gampaha district, Sri Lanka, were isolated onto potato dextrose agar and their pathogenicity was tested on healthy S. melongena plants. Fungal isolates that were confirmed to be pathogenic were subjected to morphological and molecular characterization. Findings: Fifteen fungal isolates were obtained from infected leaves of S. melongena plants, and four fungal isolates were confirmed to be pathogenic on S. melongena. Isolates H32a and H32b causing similar disease symptoms were identified as Lasiodiplodia theobromae with 99.81% sequence similarity by the analysis of the internal transcribed spacer region (ITS1-5.8S-ITS2). The remaining two pathogenic isolates were identified as Pseudopestalotiopsis theae and Diaporthe eugeniae, with 100% and 99.82% sequence similarities, respectively. Originality/ Value: To our knowledge, this is the first report of leaf necrosis by L. theobromae, leaf yellowing by P. theae and leaf blight by D. eugeniae associated with S. melongena plants in Sri Lanka
Phytochemical and antifungal screening of Monoon longifolium leaf acetone extract
(Faculty of Science, University of Kelaniya Sri Lanka, 2024) Amarasiri, V. H. N. H.; Wanigatunge, R. P.; Edirisinghe P., Edirisinghe P.
Prolonged and indiscriminate use of fungicides in agriculture can have adverse effects on human health and the environment while also causing pathogen population shifts rendering their efficacy. Hence, the development of eco-friendly alternatives has become a necessity. Monoon longifolium is a widely distributed plant in Sri Lanka, known for its pharmaceutical and antimicrobial properties. Thus, this study aimed to investigate the phytochemical composition and antifungal activity of M. longifolium leaf acetone extract against three fungal pathogens of Solanum melongena (brinjal). Phytochemicals in cleaned, dried leaves of M. longifolium were extracted into acetone, and their total phenolic (TPC) and flavonoid (TFC) content, and antioxidant activity by DPPH radical scavenging were determined. Preparative thin layer chromatography (TLC) was performed with ethyl acetate: methanol: water (5:1:5) solvent system, followed by individual biochemical testing for each band. GC-MS analysis were performed to assess the phytochemical diversity in the M. longifolium crude extract, while the antifungal potential of these compounds was determined with WAY2DRUG PASS online server. Antifungal activity of the crude extract at 2000 and 3000 ppm concentrations was evaluated against three pathogenic fungi of S. melongena i.e. Diaporthe eugeniae, Pseudopestalotiopsis theae, and Lasiodiplodia theobromae by poisoned plate method by calculating the growth inhibition percentage. Captan, sterile distilled water and DMSO were used as positive, negative and experimental controls respectively. All antifungal screening tests were triplicated and analyzed by Kruskal-Wallis nonparametric and Dunn-Bonferroni post-hoc tests (p = 0.05). TPC and TFC of the crude extract were 23.16 ± 1.67 mg GAE /g and 137.6 ± 7 mg QE/g, respectively. DPPH radical scavenging activity of the extract had an IC50 of 0.135 mg/mL, while IC50 for the ascorbic acid standard was 0.251 mg/mL. TLC resulted in seven bands (Rf values 0.11, 0.25, 0.40. 0.49. 0.59, 0.69 and 0.89) and biochemical tests indicated the presence of seven phytochemical groups, i.e. alkaloids, flavonoids, tannins, phenols, saponins, terpenoids and polyphenols. Phenols and saponins were present only in the bands with Rf value of 0.89 and 0.11 respectively. The other five bands contained multiple phytochemical groups. GC-MS analysis identified 19 prominent volatile compounds with peak areas higher than 1%, of which β-caryophyllene oxide indicated the highest antifungal potential (PA = 0.647) according to PASS analysis. The highest antifungal activity at 3000 ppm was against P. theae (82.66 ± 1.5%) though not significantly different from L. theobromae (78.88 ± 0%, p > 0.05). The lowest activity was against D. eugeniae (63.77 ± 0.5%) at 2000 ppm while no difference was observed between 2000 and 3000 ppm concentrations against any fungal pathogen. The outcome of this study signifies the potential to use M. longifolium leaf extracts in control of fungal phytopathogens, warranted through field experiments.
Production of certain extracellular enzymes by some bacteria and amplification of cellulase gene from Bacillus species
(4th International Research Symposium on Pure and Applied Sciences, Faculty of Science, University of Kelaniya, Sri Lanka, 2019) Jayasinghe, J. A. S. M.; Medhavi, P. I. H. R.; Magana-Arachchi, D. N.; Wanigatunge, R. P.; Herath, H. M.
Bacteria have received attention, due to their ability to produce extracellular enzymes beneficial in various industries. In the present study, extracellular enzyme production by two thermophilic bacteria (Meiothermus ruber, Tepidimonas ignava) and eight other bacterial isolates (Bacillus thuringiensis, Bacillus amyloliquefaciens, Bacillus pumilus, Bacillus aryabhattai, Pseudomonas stutzeri, Pseudomonas aeruginosa, Sphingomonas sp., Burkholderia lata) was investigated. Extracellular amylase, protease, pectinase and cellulase production was studied in vitro in media containing starch, skimmed milk, citric pectin and carboxymethylcellulose respectively, at 28 °C, 35 °C, 45 °C and 55 °C. Hydrolyzing Capacity Index (HCI) at day seven was calculated to identify the isolates, which hydrolyzed a substrate with minimal colony formation. Such isolates would have a higher potential in industrial applications. HCI values were analyzed using one-way ANOVA and Tukey’s multiple comparison tests. All isolates, except thermophilic M. ruber, produced at least one extracellular enzyme within 1-3 days. T. ignava, B. thuringiensis and P. aeruginosa produced amylases. All isolates except B. aryabhattai and M. ruber produced proteases. B. thuringiensis, Sphingomonas sp., B. amyloliquefaciens and P. stutzeri produced cellulases. Pectinases were produced only by B. lata. Thermophilic T. ignava produced amylases and proteases at 28 oC and 35 oC but did not produce any enzyme at 55 °C, the temperature of the Maha Oya hot springs from which it was isolated. B. amyloliquefaciens, P. stutzeri, P. aeruginosa, B. pumilus, Sphingomonas sp. and B. lata produced proteases, which were stable at higher temperatures; 45 °C and 55 °C. It was the only enzyme to be produced at those temperatures. According to the HCI values, B. thuringiensis and P. stutzeri were the most efficient degraders of starch and cellulose, respectively. P. stutzeri, Sphingomonas sp. and B. lata were the best protein degraders. A gene coding for glycoside hydrolase (a cellulase) was amplified from bacteria by PCR using primers designed for Bacillus licheniformis ATCC 14580. Although expected amplicon size was ~1683 bp, amplicons of apporiximately 500 bp, 600 bp and 1000 bp were generated from cellulase producing B. thuringiensis. According to the information available in NCBI, B. thuringiensis has glycoside hydrolase gene of 738 bp suggesting that those amplicons could also be some glycoside hydrolase genes of different lengths. This should be confirmed by DNA sequencing. PCR product was generated by the same primers for B. aryabhattai as well, although it did not produce cellulases in vitro. It could be due to non-expression of the particular gene at the experimental conditions used in this study. These Bacillus species are perceived as sources of purified cellulases and the particular genes would be useful also in transformation of other organisms for industrial purposes