An integrative approach to understanding effects of valproate in C. elegans

Obesity is one of the biggest public health problems today, leading to type II diabetes, coronary heart disease, hypertension, sterility and cancer (Hashmi et al., 2013). $C.elegans$ has been used extensively in obesity research to dissect the underlying principles. The overarching goal of my thesis has been to identify molecular signatures linking phenotypic and molecular responses of the nematode $C.elegans$ to a number of experimental conditions that induce alteration in lipid metabolism. More specifically, I developed experimental procedures in $C.elegans$ to study the effects of exercise, diet and a $C.elegans$ antiageing drug valproate. Using a modified lipid assay, I found for the first time that valproate induce lipid accumulation in $C.elegans$ in a dose dependent manner. In this study obesity is induced in $C.elegans$ by treating with either cholesterol or sodium valproate. Using microarray technology, expression profiles associated with the induction of obesity are studied. In addition, $C.elegans$ were treated with monosodium glutamate which is found to decrease lipid mass. Exercise is a recommendation for obese people and hence a $C.elegans$ model system is developed to study effect of exercise. Effects of valproate on $C.elegans$ with respect to lipid quantity are studied in detail, with emphasis on insulin signalling pathway. Expression changes associated with insulin mutants are compared with the valproate exposure expression profile to understand how much of the valproate exposure could be explained by insulin signalling. I also found for the first time that valproate cause a global down regulation of nuclear components. A hypothesis on lipid accumulation on valproate exposure is presented. TGF-β pathway is also found to be involved in fat metabolism. Finally I report an interesting phenotype for the first time in which the TGF-β pathway receptor mutant $sma$-$6$ shows paralysis in response to valproate

Evaluation of salt content and effectiveness of excessive salt reduction methods in selected commercially available dried fish types in Sri Lanka

High salt intake elevates the risk of non-communicable diseases such as high blood pressure, cardiovascular diseases and stroke worldwide. Sri Lanka has recorded in 2010 as the country with highest average fish & fish products consumption in South Asia. In the current study, salt in ten types of commonly available dried fish namely; sprats, prawns, smoothbelly sardinella, queen fish, cat fish, sail fish, shark, skipjack tuna, Bombay duck and trenched sardinella was analyzed and determined salt reduction methods with minimal protein loss. Four salt reduction methods were tested; Method 1: washed with water at Room Temperature (RT) for five times; Method 2: washed with water for five times at RT and kept in hot water for 5min; Method 3: washed with water for five times at RT and boiled for 5min; Method 4: washed with water for five times at RT and kept in coconut water for 5min. Using Volhard method, sodium chloride was analyzed while protein was determined using Kjeldahl method. All four methods showed significant reduction of salt level in tested dried fish (p < 0.05). Among the tested salt reduction methods, Method 3 showed the highest salt reduction for all types of dried fish except smoothbelly sardinella and cat fish.The highest salt mean value of 28.8% was recorded in queen fish and the lowest mean value of 12.8% was recorded in smoothbelly sardinella. The highest protein loss was recorded in Method 3. To reduce considerable amount of salt, the easiest and fairly effective method is method 1 except for prawns and Shark. Although higher salt reduction showed in method 2 and 3, they are not recommended due to high protein loss, high energy expenditure and reduction of freshness of dried fish. Method 4 can be applied for all dried fish types because it is economical and reduces considerable amount of salt, removes less amount of protein comparatively. The results revealed that all tested dried fish except smoothbelly sardinella contain significantly high amount of salt (p < 0.05) than the standard value specified (12%) by the Sri Lanka Standards Institution (SLSI).Keywords: Dried fish, protein loss, salt-intake, salt reduction, non-communicable disease

Antimicrobial Activity of Cultivable Endophytic and Rhizosphere Fungi Associated with “Mile-a-Minute,” Mikania cordata (Asteraceae)

Endophytic and rhizosphere fungi are understood to be aiding the host plant to overcome a range of biotic and abiotic stresses (nutrition depletion, droughts, etc.) hence, they remain to be reservoirs of plethora of natural products with immense use. Consequently, this investigation of endophytic and rhizosphere fungi isolated from Mikania cordata (a perennial vine that is well established in Sri Lanka) for their antimicrobial properties was performed with the aim of future derivation of potential beneficial pharmaceutical products. Leaves, twigs, and roots of M. cordata were utilized to isolate a total of 9 endophytic fungi out of which the highest amount (44%) accounted was from the twigs. A sample of the immediate layer of soil adhering to the root of M. cordata was utilized to isolate 15 rhizosphere fungi. Fusarium equiseti and Phoma medicaginis were endophytes that were identified based on colony and molecular characteristics. The broad spectrum of antimicrobial activity depicted by F. equiseti (MK517551) was found to be significantly greater (p≤0.05, inhibitory against Bacillus cereus ATCC 11778, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 25853) than P. medicaginis (MK517550) (inhibitory against Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 25853) as assessed using the Kirby-Bauer disk diffusion method. Trichoderma virens and Trichoderma asperellum were rhizospere fungi that exhibited remarkable antimicrobial properties against the test pathogens chosen for the study. T. asperellum indicated significantly greater bioactivity against all four bacterial pathogens and Candida albicans ATCC 10231 under study. The ranges of minimum inhibitory concentrations (MICs) of the fungi depicting antimicrobial properties were determined. The results obtained suggest that F. equiseti, P. medicaginis, T. asperellum, and T. virens of M. cordata harness bioprospective values as natural drug candidates. This is the first report on isolation and evaluation of the antimicrobial properties of endophytic and rhizosphere fungi of Mikania cordata

In Vitro Determination of Antimicrobial and Hypoglycemic Activities of Mikania cordata (Asteraceae) Leaf Extracts

Infectious diseases and diabetes mellitus are counted responsible for a substantial amount of mortality among the human population. The current study was performed to detect the antimicrobial activities and hypoglycemic potential of Mikania cordata (Asteraceae) leaves extracted into aqueous media and several organic solvents (ethyl acetate and methanol). The ethyl acetate extract of Mikania cordata (MEA) leaves was observed to possess significantly (p≤0.05) greater antimicrobial capabilities (susceptible against Bacillus cereus ATCC 11778 and Staphylococcus aureus ATCC 25923) when compared with that of the methanol (MME) and aqueous extracts (MDW) which were assessed based on Kirby-Bauer disk diffusion assay. The minimum inhibitory concentration of MEA (against B. cereus, S. aureus, and Escherichia coli ATCC 25922) and MME (against B. cereus, S. aureus, E. coli, and Candida albicans ATCC 10231) lies in a similar range of 1.13 > MIC>0.56 mg/ml. In the present study, a single compound (from MEA) of Rf value 0.64 was isolated by thin-layer chromatography (TLC) that was responsible for the zone of inhibition against B. cereus (20.3 ± 0.3 mm). The results of this study also depicted the antihyperglycemic properties of M. cordata leaves which followed the same trend as the commercial drug Metformin in a glucose concentration-independent manner when tested in a glucose uptake assay by yeast cells. Therefore, it is evident that Mikania cordata is a reservoir of useful bioactive compounds which with further research will be paving the path for drug commercialization. This is the first record of TLC-based isolation of antimicrobial compounds of M. cordata and analysis of the hypoglycemic properties of M. cordata leaves

Quality Assessment of Selected Dairy Products in Sri Lankan Market

The purpose of the study was to determine the quality of selected Sri Lankan-marketed dairy products. Four brands of full cream milk powder (FCMP) (imported A and B; local C and D) and three brands of pasteurized milk (PM) attributed to the alphabetical identifies E, F, and G were tested, with raw cow’s milk (CM) as control. Fat, protein, ash, carbohydrate, moisture, and water percentage, total solids (TS), titratable acidity (TA), pH, specific gravity (SG), arsenic content, and total coliform count (TCC) were assessed. The average fat and ash content per serving of milk (SOM) of FCMP was significantly lower than the PM and CM. Highest (p>0.05) protein content (7.58 g ± 1.05) was recorded for CM. Carbohydrate and pH were not significantly different in three types of milk products. FCMP had a significantly lower (p0.05) compared to the CM (1.030 ± 0.00). All the abovementioned parameters between imported and locally produced FCMP brands were not significantly different from each other. In imported FCMP, mean moisture % was significantly higher (p>0.05) than local brands; however, in each FCMP, brand mean moisture % was statistically non-significant. Total solids in PM was significantly lower (p<0.05) than the CM. Every tested sample was free of arsenic. However, all PM brands and B of FCMP were contaminated with coliform. Total coliform count in B and E agreed with the Sri Lankan standard level. Nutritional value in SOM of PM and FCMP was less than CM, while the lowest value was recorded in FCMP. It can be concluded that all brands of powdered milk possess the recommend suggested standards in terms of both physicochemical and microbiological qualities. Though the physicochemical characteristics in PM brands agree with the standard levels, microbial hygiene is poor where coliform contamination was very high in Brand E

Microbiological study in a gneissic cave from Sri Lanka, with special focus on potential antimicrobial activities

The emergence of antibiotic resistance is a global health crisis, thus the search for novel antimicrobial compounds has become a continuous necessity. Underexplored and extreme environments, such as cave ecosystems, have been identified as a promising potential source for the discovery of novel microorganisms with novel antimicrobial compounds (AMC). This study presents the first cave microbiological investigation in Sri Lanka, with a special preference for bioprospecting of novel AMC. The cave sediment characterization demonstrated the presence of close to strong acidic conditions (pH 3.1 – 3.3) and thus indicates the possibility of isolating acidophilic microorganisms. Eight cave wall/ceiling fungal strains were isolated from Sthreepura Cave - Kuruwita and identified using both morphological and ribosomal Internal Transcribed Spacer (ITS) region sequence analysis. Interestingly, four fungal isolates (Penicillium panissanguineum, P. cremeogriseum, Aspergillus bertholletius and Trichoderma yunnanense) were found to be the first records in Sri Lanka. Of these eight isolates, three showed antimicrobial activity (AMAs) against at least one of the five tested human pathogens in preliminary screening, while A. fumigatus (SKW 404) strain showed the highest AMA against Staphylococcus aureus (ATCC 11778) assessed by agar culture plug method on Muller Hinton Agar (MHA). Crude Ethyl Acetate (EtOAc) fraction of both mycelial and Potato Dextrose Broth (PDB) extracts of A. fumigatus demonstrated similar bioactive metabolic profiles with four corresponding chemical fractions [Rf = 0.47, 0.56, 0.65, 0.82; EtOAc: Hexane (4:1, v/v)] in TLC: agar overlay bioassay. The present study indicates that there is potential for discovering novel Sri Lankan deep cave microorganisms and bioprospecting of their novel bioactive compounds. Hence, further island-wide in-depth cave microbiological investigations are required for a better understanding of the Sri Lankan cave microbiology