Browsing by Author "De Silva, D.P.N."

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    Antibacterial Efficacy of Mangrove Rhizophora mucronata Extracts on Aquatic Bacterial Pathogens
    (Uva Wellassa University of Sri Lanka, 2013) Bokalawella, S.H.K.; De Silva, D.P.N.; Jayamanne, S.C.; Gunasekara, A. J. M.; Piyadasa, H.T.N.I.
    Medicinal plants contain components of therapeutic value and are used as remedies for human/animal diseases. They are environmentally friendly due to less toxicity and have minimal health hazards (Opara and Wokocha, 2008). Mangrove plant extracts and its major chemical classes displayed various levels of antibacterial, antifungal, antifouling, antiviral and free radical scavenging effects due to their bioactive compounds (Bandaranayake, 2002). Bacterial infections cause severe economic losses to the aquaculture industry which increased the use of antimicrobial agents significantly as a treatment (Alderman and Michel, 1992). Improper usage and presence of drug residues lead to the development of resistant strains of pathogenic bacteria. Therefore, novel antibacterial compounds of natural products which degrade easily need to be synthesized to overcome above consequences. Aim of this study was to find out the antibacterial efficacy of Mangrove Rhizophora mucronata extracts on aquatic bacterial pathogens, to be used as a remedy to treat fish diseases. Methodology Fresh matured leaves of six mangrove plant species (Avicennia marina, Lumnitzera racemosa, Bruguiera sexangula, Acanthus ilicifolius, Excoecaria agallocha and Rhizophora mucronata) were collected from mangrove reserve at Kadolkele, Negombo and were brought to the laboratory for making extractions. Initially, preliminary trials were conducted for identification of most suitable species with antibacterial properties against four aquatic pathogens Listeria monocytogenes) in vitro by well diffusion method. Upon identification of Rhizophora mucronata having the highest inhibitory zone for tested bacteria, further analysis were performed using various parts such as roots, barks, flowers and fruits of R. mucronata. The solvent was extracted by mixing 8 g of each part of mangrove in powder form, with 125 ml of 95% methanol and evaporating methanol using rotary flash evaporator (model 519-00000-00- 3). Antibacterial efficacy was assessed against all four species of fish pathogens, using was used as the positive control against Rhizophora mucronata extractions of 350 mgml each. After 24 hour incubation at 25 o C, the diameters of the inhibitory zones were obtained and statistically analyzed by using One way-ANOVA incorporated in Minitab version 16. Results and Discussion All six mangrove species showed antibacterial activity against Listeria monocytogenes, Pseudomonas fluorescens, and Pseudomonas aeruginosa significantly (p<0.05) but insignificant for Shigella flexneri (p>0.05). Highest inhibition zones were observed in Rhizophora mucronata leaf extract against all bacteria examined for the study (Table 1). Rhizophora mucronata extracts (root, bark, fruit and flower) also exhibited positive inhibition zones for all the tested bacterial species cultured in vitro. Root and bark extracts possessed higher antibacterial potential when compared to the fruit and flower extracts. Rhizophora leaf extracts showed highest inhibition for Listeria monocytogenes and Pseudomonas fluorescens while root and bark extracts were effective against Shigella flexneri and Pseudomonas aeruginosa respectively (Figure 1). The overall mean comparative evaluation on antibacterial efficacy of Rhizophora mucronata extracts displayed the highest in leaves followed by the root, bark and fruit extracts. The minimum antibacterial activity was observed in the flower extracts.
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    Determination of the Effect of Papaya (Carica papaya) Leaf Powder as a Low Cost, Immunogenic, Herbal Feed Additive on Zebrafish (Danio rerio
    (Uva Wellassa University of Sri Lanka, 2021) Prameesha, H.A.; De Silva, D.P.N.; Guruge, W.A.H.P.
    Fish diseases including bacterial diseases cause significant loss in aquaculture. Antibiotic treatments are expensive and lead to antimicrobial resistance. The objective of this study was to identify the potential of a low cost herb which enhance fish immunity using Carica papaya leaf powder as a feed additive on zebrafish infected with bacteria and determined the effect on immunity. Ninety healthy fish were divided into three groups with three replicates (30 fish per group) exposed to Aeromonas hydrophila in water for a week followed by feeding trial. Three groups were fed with commercial (control), antibiotic and papaya leaf incorporated fish feed with 2% of body weight per day. Differential white blood cell (WBC) count was taken to identify the effects on disease response. Antibiotic susceptibility test was performed to identify, antibiotic resistance of bacteria and antibacterial activity of aqueous papaya leaf extract. Growth parameters and growth performances of experimental fish were calculated. WBC counts were measured before exposure to bacteria, one week after infectious challenge, one week after feed trial. One way ANOVA followed Tukey Pairwise comparison test used for data analysis. Fish exposed to A. hydrophila showed haemorrhages on the base of pectoral fin, fin rot and body discoloration. As per the results, monocyte count was decreased and lymphocyte count was increased in all three tanks after infection but one week after receiving the antibiotic and papaya feed, lymphocyte counts were decreased significantly compared to the control, while monocyte count and neutrophil counts were increased compared to the control. Accordingly, initial increase of lymphocyte count was observed due to immune response against bacteria as they involved in antigen-antibody reactions. Lowered monocyte count was observed due to recruit them for wound healing. Lowered lymphocyte count was observed due to the reduction of pathogens in response to antibiotic and papaya feed. Neutrophils play a role in phagocytizing bacteria, increased in papaya fed fish indicated a positive immune response. The results indicated that papaya leaf incorporated feed increased the immune response of zebrafish against A. hydrophila, which can be used as a low cost, herbal feed additive in ornamental fish industry. Keywords: Zebrafish; Carica papaya; Aeromonas hydrophila; Immunity; Herbal antibiotic
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    The Effect of Coriandrum Sativum as Feed Additives on Growth Performance, Immunity and Disease Resistance of Southern Platy Fish (Xiphophorus Maculatus)
    (Uva Wellassa University of Sri Lanka, 2021) Bowalage, B.W.G.S. L.; Coswatte, A.C.W.W.M.C.L.K.; De Silva, D.P.N.
    Ornamental fish production and trade is a profitable alternative in the aquaculture industry. Nutrition is one of the key factors to obtain a higher productivity while maintain immunity of ornamental fishes. These fishes live in an environment surrounded by stress factors. Increasing immunity via feed additives may help them to thrive such stressors. Therefore, this study was carried out to assess the effect of coriander as feed additives and immune enhancers, on growth performance, immunity and disease resistance of southern platy fish (Xiphophorus maculatus). Sixty healthy platy fish, of 2 months old with an average weight of 0.17 ± 0.01 g were used in the experiment. Fish were fed with coriander incorporated commercially available feed for one month. The experiment was triplicated and controls were also maintained without incorporating coriander to commercially available feed. The weight gain, length and specific growth rate (SGR) were measured. The infection trial was carried out using Aeromonas hydrophila. Immune response and disease resistance of fish were determined by analyzing white blood cell (WBC) count. Results revealed that the, mean length and SGR obtained from coriander fed fish were significantly higher than the control (p<0.05). The fish fed with Coriander incorporated feed obtained 20% of neutrophil count while fish fed without coriander incorporated feed obtained 10% of neutrophil count. In conclusion, there was a positive effect on disease resistance and immunity by the coriander incorporated feed on southern platy fishes and significant length and weight gain indicated the feasibility of using coriander as feed additives and immune enhancers in aquaculture. Keywords: Coriandrum sativum; Disease Resistance; Innate Immunity; Ornamental fish; Specific Growth Rate; White Blood Cell
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    Investigation of Disease Incidence and Management Practices to Build a Database to Fill the Knowledge Gap of Ornamental Fish Farmers in Sri Lanka
    (Uva Wellassa University of Sri Lanka, 2021) Dinushika, M.P.G.; Dias, P.C.B.; Sampath, J.K.H.; De Silva, D.P.N.
    The freshwater ornamental fish industry is one of the economically important, home entertainment industry in the world. During the last few decades, the ornamental fish industry in Sri Lanka has become one of the highest potential exporters in the global market. However, fish diseases are one of the major constraints to the failure of the ornamental industry. To avoid economic losses and to prevent disease outbreaks proper identification and diagnosis of fish diseases are crucial. Hence, this study was aimed to evaluate the present status of farmer’s knowledge of fish disease; management measures to avoid disease outbreaks and prevalence of fish diseases with seasonal and regional variation in the freshwater ornamental fish industry. Sixty farmers cultivating various ornamental fish were selected using stratified and simple random sampling techniques from the National Aquaculture Development Authority (NAQDA) website. Data were collected using a telephone-interviewed method through a pretested questionnaire survey. Collected data were analyzed using the descriptive statistical methods in SPSS. . Major disease incidence caused by parasites (33.57%), followed by bacterial (23.57%), fungal (16.43%), viral (5%), and non- infectious diseases (21.43%). The most frequently observed disease was white spot disease (11.4%), followed by Columnaris (10.8%), Dactylogyrus sp. (9.5%), and fin rot (8.9%). The prevalence of fish diseases varied with season. The average prevalence of fish diseases was highest in the southwest monsoon period (49%). Common treatment methods that farmers used to treat the diseases were isolation (20.9%), water exchange (19.9%), adding salt (15.7%), methylene blue (13.6%), copper sulphate (11.5%), and antibiotics (7.1%). However, most of the farmers were able to identify diseases by using behavioral changes (40.5%) and symptoms (37.3%) although they were incapable of recognizing the disease-causing agents. Preventive and prophylactic measures were practiced by 91% of the farmers, the majority of them tend to find solutions through online sources, and some of them get advice from either colleagues or veterinarians. Furthermore, insufficient materials or lacks of awareness regarding fish diseases prevent farmers from gaining new reference knowledge. Thus, the results obtained from this study can be used to fill the knowledge gap by developing an awareness tool that enhances their knowledge on fish health management. Keywords: Ornamental fish; Fish Disease; Seasonal variation; Awareness tool
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    A Study on Toxicity Effect of Mancozeb Fungicide Residual on Zebrafish (Danio rerio) Embryo
    (Uva Wellassa University of Sri Lanka, 2018) Hiroshini, W.C.; Liyanage, N.P.P.; Jayamanne, S.C.; De Silva, D.P.N.
    Mancozeb fungicide which belongs to ethelene bisdithiocarbamates (EBDCs) chemical group causes potential hazard to human and environment. Considerable amount of this pesticide leaches through soil, air and accumulate in aquatic environment. The fungicide breaks down into ethylenethiourea (ETU) within two days and residuals remain in the aquatic environment. In this study, the effect of Mancozeb residuals on zebrafish embryo was performed according to OECD guideline and investigating the lethal end points, median lethal concentrations (LC50) and morphological deformities at different concentrations. Range finding test was carried out initially, using different concentrations to determine the test concentration for toxicity. The embryo toxicity test for Mancozeb residual was performed over four days period according using 16-cell stage fertilized zebrafish embryos. Ten test concentrations (5, 2.5, 1.25, 0.64, 0.32, 0.16, 0.08, 0.04, 0.02 and 0.01) mg L -1 Mancozeb residual was tested on 60 embryos pre-treatment. Lethality rate and morphological abnormalities were recorded at 24, 48, 72 and 96 hours post fertilization (hpf). LC50 for 24, 48, 72 and 96 hours of exposure to Mancozeb residual was determined as 1.4, 1.15, 1.15, 1.15 mg L-1 respectively. A significant difference was observed between Mancozeb residual and control group (P < 0.05). Coagulation of the embryo, non-detachment from the tail bud, lack of heartbeat and lack of somite formation were observed as lethal end points. Yolk sac and pericardial edema, degeneration of tail region, malformation of head and heart, retarded growth, twisted tail, shrinkage of chorion and spinal curvature were observed as morphological deformities. This study demonstrated that Mancozeb residual at 1.4 mg L-1 was lethal at 24 hpf level and 1.15 mg L-1 was lethal at 48, 72 and 96 hpf. The results indicated that Mancozeb residuals affect zebrafish (Danio rerio) embryo indicating its potential to cause environmental toxicity.
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    Toxicity effects of trace metals on Zebra fish (Danio rerio) embryo
    (Uva Wellassa University of Sri Lanka, 2015) Fasmina, M.F.A.; Jayamanne, S.C.; Liyanage, N.P.P.; De Silva, D.P.N.
    Water quality deterioration is a major problem in the most of the countries, due to organic and inorganic contaminants. Among all the toxicant trace metals are threaten to the aquatic ecosystem and organisms. Metals are natural component of aquatic ecosystem. Trace metals such as Copper (Cu), Zinc (Zn) and Chromium (Cr) are important for the metabolic and other biological activities of lives whereas Mercury (Hg), Lead (Pb), Cadmium(Cd) are biologically non-essential metal that can be toxic to biota even at very low concentration. High concentration of some essential trace metals can be toxic when it exceeds the limits(Ebrahimi and Taherianfard, 2011).High accumulation of trace metal in both biotic and abiotic components causes serious health consequences. Thus, assessment of their toxicity has become an important component of water pollution monitoring. Now in most of the toxicity studies Zebra fish (Danio rerio) embryo used as an alternative model for the fish acute toxicity to determine the toxicity of pollutants. Therefore this study was focused on determination of the acute toxicity of Cu, Zn, Cd, As, Pb and Hg that produce lethal effect on zebra fish embryos during four day period. Materials and Methodology Zebra fish breeding was carried out and eggs were collected using a small pipette. The stock solution of 1000 ppm of selected Cu, Zn, Pb, Cd, As and Hg were prepared a day before the test by dissolving Copper sulfate (CuSO4.5H2O), Zinc sulfate (ZnSO4. 7H2O), Lead nitrate (Pb(NO3)2, Arsenic pentoxide (As2O5), Cadmium chloride (CdCl2. ½ H2O) and Mercury chloride (HgCl2) in deionized water. The working treatment solution was prepared daily by serial dilution of the stock solution. Initially a range finding test was carried out in the nominal concentrations of (1000, 100, 10, 1 and ) for 96 hours. Ten eggs per concentrations were used. Deionized water was used as a negative control. Each treatment had three replicates. Acute Cu, Zn, Pb, As, Cd and Hg toxicity experiments were performed for a 4-day period using Danio rerio embryo. Ten test concentrations of Cu, Cd (0.025, 0.05, 0.1, 0.2, 0.4, 0.8, 1.6, 3.2, 6.4 and 10.0 mgL⁻ ), Zn, Pb, As (2.0, 4.0, 8.0, 16.0, 32.0,64.0, 125.0,250.0, 500.0 and 1000.0 mgL⁻ ) and Hg (0.002, 0.004, 0.008, 0.016, 0.032, 0.064, 0.125, 0.250, 0.500 and 1.0 mgL⁻ ) were prepared by diluting the stock solution in deionized water. Deionized water was used as negative control and as internal plate control. As a positive control concentration of 15% ethanol(Hassan et al., 2008) was used with each embryo batch for testing. Fertilized zebra fish embryos were immersed in the test solutions before cleavage, by the 16 cell- stage. At least twice the number of eggs needed per treatment group (40 eggs) was randomly selected and transferred into the respective concentrations and controls within the 90 minutes of post fertilization. 24-well plates were filled with 2 mL per well freshly prepared test solutions. In each plate twenty wells were used for test concentrations and four were used as internal plate control. Another plate was used as positive control. The eggs in standard 24- well plate were covered by shelf adhesive foil and maintain at a temperature to 26 ± 1 °C. The mortality of the fish eggs were recorded at 24, 48, 72 and 96 hours of exposure using a stereo microscope with the indicator of lethality; coagulation of fertilized egg, lack of somite formation, lack of detachment of the tail and lack of heart beat as described in OECD/ OCDE 236, 2013; guideline for the test of chemical, Fish Embryo Acute Toxicity (FET) tests. Dead eggs were removed immediately. Three replicates were performed individually. In this study the acute toxic effect of each metal on the Danio rerio embryo was determined by Probit Analysis LC50 determination method. Data analysis was performed by using statistical software SPSS15.
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    Usability of Fish Haematological Parameters as a Biological Indicator for Freshwater Quality in Badulla District
    (Uva Wellassa University of Sri Lanka, 2016) Jayasundara, J.M.N.M.; Liyanage, N.P.P.; Wijesundera, R.R.M.K.K.; Senevirathne, J.D.M.; De Silva, D.P.N.
    Water, the fundamental of life, requires continuous monitoring because water quality deteriorates over time. Fish haematological parameters respond more quickly for environmental changes with more precise results than chemical analysis which conduct periodically. As it is profitable to authenticate eco-friendly method for assessing water quality, this study was conducted to investigate the usability of fish haematological parameters as a bioindicator for assessing freshwater quality. Comparison of physico¬chemical parameters of water quality was done in two inland static water bodies in Badulla district: Horabora and Mapakada reservoirs. Results of present study revealed that Transparency, Total Dissolved Solids, Biochemical Oxygen Demand (BUD), pH, Total hardness and Trace metal (As, Pb and Cu) levels were significantly different between Horabora and Mapakada reservoirs (p<0.05). No statistical difference was found for total alkalinity, Dissolved Oxygen level, and Surface water temperature (p>0.05). Both water bodies were identified as polluted in terms of Universal Water Quality Index, although Horabora reservoir was much more polluted than Mapakada reservoir by resulting higher BOD, Total Dissolved Solids, Total hardness and trace metal levels (Pb and As) with less clear water. Blood from Tilapia (Oreochromis mossambicus), Labeo rohita and Calla catla was used for fish haematology analysis. Red blood cell count was significantly different (p<0.05) between two different water bodies for each fish species. Eosinophil count showed a significant difference among two water bodies (p<0.05). The study revealed that a great possibility exists for using fish haematological parameters as a bioindicator for freshwater quality. Moreover, a multi-year continuous study with elaborated investigation require for clarify trends which may exist between fish haematological parameters with respect to water quality changes. Keywords: Water quality study, Haematology, Tilapia, Eosinophil, Trace metal