Title – Cytogenetic abnormalities In the Male Partners of couples with recurrent pregnancy loss and subfertility.

Supervisors – Dr Kalum Wetthasinghe & Ms. Imalki Kariyawasam

Study Setting – Human Genetics Unit, Faculty of Medicine, University of Colombo

Synopsis – Reproductive failure is a common phenomenon experienced worldwide. Subfertility and recurrent pregnancy loss (RPL) are two of the most common reproductive problems. Recent studies have recognised that male and female factors can equally contribute to both diseases. The prevalence of cytogenetic abnormalities in the male partners of couples with RPL and subfertility was investigated in patients referred to the Human Genetics Unit, Colombo, Sri Lanka.

The study was conducted on 20 males, 14 of them had a history of RPL and 6 suffered from subfertility. Controls were not used in the study. Cytogenetic analysis was performed on peripheral blood lymphocytes using G-banding technique. Karyotyping was performed. Chromosomal aberrations were detected in 78.8% of patients experiencing RPL and 33.3% of subfertile patients. All detected anomalies were structural anomalies. The anomalies included the polymorphic variants Yqh-, Yqh+, 9qh-, 15pstk+, 21pstk+ and 22ps+. The karyotypes 46,XY,t(1;9)(q42;q13) and 46,XY,inv(9)(p13;q13), which displayed non-polymorphic variants, were detected in the recurrent miscarriage patient group. The study also showed that G-banding was a successful procedure for cytogenetic analysis of Sri Lankan male patients.

Title – Analysis of antimicrobial activity of Ethanol extracts of Trachyspermum roxburghianum (Asamodagam) leaves and seeds

Supervisors – Prof. Chandrani Wijeyaratne (Department of Botany, University of Sri Jayawardanepura)

Study Setting – Spectrum Institute of Science and Technology, Colombo

Synopsis – The clinical efficacy of most existing antibiotics is currently threatened due to the advent of multi-drug resistant pathogens with the emergence of strains with reduced susceptibility to most drugs. Infectious diseases caused by resistant microbes are associated with prolonged hospitalisations, mortality and morbidity. As this has become a major global health concern, the discovery and de-velopment of novel antimicrobial agents of clinical significance is crucial. Plant extracts and their metabolites are now being considered as potential antimicrobial agents. T.roxburghianum is a common medicinal plant popularly recognised as ‘Asamodagam’ in Sri Lanka.

This study dealt with preliminary phytochemical screening of seeds and leaves for the presence of phenolic compounds, flavonoids, tannins and saponins. Phytochemicals are the compounds re-sponsible for anti-inflammatory, antimicrobial and other therapeutic effects in plants. Leaf ex-tracts of T.roxburghianum were found to be a rich source of phytochemicals than the seed ex-tracts. Antimicrobial activity of both extracts were tested using the Kirby Bauer disc diffusion assay against a fungal strain (C.tropicalis) and two bacterial strains (gram negative E.coli and gram posi-tive S.aureus). Amoxicillin was used as a positive control for the bacterial strains, with Fluconozole for the fungal strain and Ethanol as the negative control.

Results exhibited dose dependant antimicrobial activity from both extracts which were tested at five different concentrations. The highest activity was observed against C.tropicalis from both ex-tracts overall.

This study displayed the antimicrobial potential of T.roxburghianum which can be exploited and investigated further as a potential pharmaceutical treatment.

Title – Isolation, identification and characterization of thermophilic bacteria isolated from Nelum-wewa hot well spring, Polonaruwa

Supervisors – Dr. Minoli Perera

Study Setting – Spectrum Institute of Science and Technology, Colombo

The study was conducted to isolate, identify and characterize the thermophilic bacteria from Nelumwewa, Polonnaruwa, Sri Lanka by conventional and molecular methods (16s rRNA sequencing). In addition, these isolates were screened for the production of enzymes and their extracellular release. A total of 8 different isolates were selected based on variable colony formation and morphological features. Screening for potential enzyme production showed that all of the isolates identified were capable of producing industrially valuable enzymes. Furthermore, most of the isolates showed a capacity for producing at least two biotechnologically important enzymes. Among the 8 selected strains, Bac:4 was further characterized by 16s rRNA sequencing and showed 97% similarity with Bacillus licheniformis strain 6A.

This study therefore confirmed that the isolated strain Bac:4 is a Bacillus sp. and can be classified as a thermophilic bacteria. It has potential to serve as a source of thermo stable enzymes such as thermo stable protease, which can be utilized in industrial and pharmaceutical applications. Further investigation of microbial population in hot springs around Sri Lanka is important to characterize the biodiversity in these regions and to isolate and characterize other bacterial species of industrial, and thus economic, importance.

Title – Comparison of the effectiveness of different bacterial DNA isolation kits and the phenol-chloroform method

Supervisors – Dr. Minoli Perera

Study Setting – Spectrum Institute of Science and Technology, Colombo

Molecular diagnosis of pathogenic bacteria using clinical samples is a pivotal step in diagnosis and treatment of disease. Extraction of genomic DNA and downstream molecular applications such as PCR are utilised for identification of microorganisms such as E. coli which are responsible for gastrointestinal diseases. In this study, three bacterial DNA extraction methods were compared to determine the most efficient method.The three DNA isolation methods include the standard phenol chloroform method, DNeasy Blood and Tissue Kit (spin column based method) (Qiagen Inc.

Valencia, CA, USA) and Wizard® Genomic DNA Purification Kit (solution based method) (Promega, Madison, WI, USA). The primary aim of this study is to determine the most effective method of DNA extraction with regard to the purity of DNA obtained. Determining which method provides the highest yield was the secondary objective. The isolated DNA was then subjected to agarose gel electrophoresis and U.V spectroscopy (OD260/OD280 ratio) to determine purity and yield.

PCR was performed to determine if amplification of extracted DNA was possible. The highest mean absorbance ratio of 1.7175 was obtained by DNeasy Blood and Tissue Kit. The phenol chloroform method provided a mean absorbance ratio of 1.675 which was considered of good quality and Wizard® Genomic DNA Purification Kit had the lowest mean absorbance ratio, 1.475.Hence the results indicate that the DNeasy Blood and Tissue Kit is capable of producing DNA of high quality and the phenol chloroform method can be used as an alternative to the expensive DNA isolation kits.

Title – Antibiotic Resistant Bacteria in the Wild Bird Faeces

Supervisors –Mrs. Sajeevee Sarathchandra

Study Setting –Spectrum Institute of Science and Technology, Colombo

The emergence and detection of antibiotic-resistant bacteria from the wild, especially commensal bacteria such as Escherichia coli (E. coli) and enterococci have become an enormous global health concern. Therefore, this study intended to isolate E. coli from the faeces of feral pigeons from the districts of Galle and Colombo and to determine their resistance to eight selected antibiotics.A total of sixty-four fresh faecal samples of wild pigeons and domesticated pigeons were collected and subjected to selective culturing, a series of biochemical test, and PCR. The identity confirmed E. coli samples were subjected to eight selected antibiotics, and E. coli from all sixty samples showed resistance to six of the antibiotics. E. coli from twenty-one and twenty-nine wild pigeon faecal samples showed resistance to Ciprofloxacin and Tetracycline respectively.

E. coli from all thirty and nineteen domesticated pigeon faecal samples showed resistance to Tetracycline and Ciprofloxacin respectively. In general, Ciprofloxacin, Gentamicin, and Tetracycline were observed to be the most effective against E. coli among the eight selected antibiotics.

Moreover, the habitat of pigeons was not observed to have an impact on the effectiveness of the antibiotic against E. coli. However, for Chloramphenicol, a variance of 30% was observed.

Title –Estimation of Terpenoid contents of different parts of Gyrinops walla (walla patta) tree from different locations of Sri Lanka.

Supervisors –Dr. Pathmasiri Ranasinghe

Supervisors –Senior Deputy Director, Herbal Technology Section, Modern Research & Development Centre, Industrial Technology Institute – Malabe.

Study Setting –Industrial Technology Institute, Malambe

Gyrinops walla is endemic and known to be the only resinous tree which produces Agarwood in Sri Lanka. The resin which is extracted from Gyrinops walla possesses a unique fragrance beneficial for the manufacture of expensive perfumes and cosmetics. High demands lead to illegal over exploitation of the plant and to drastic declination of Gyrinops walla tree population in Sri Lanka.

The present study was conducted to investigate the terpenoid content in leaves, stems and roots of Gyrinops walla extracted with hexane and dichloromethane. Extracted samples were subjected to colorimetric analysis under three assays; Total Phenolic Content, Total Flavonoid Content and Total Terpenoid Content using Folin–Ciocalteu, aluminium chloride and concentrated sulphuric acid methods. Terpenoid composition was identified using Gas chromatography mass spectrometry (GC-MS).

Results showed significant differences of TPC, TFC and TTC in two different fractions for all three samples (p<0.05) and highest contents were reported in DCM fraction for all three samples. GCMS indicated nine sesquiterpenes. The results concludes that terpenoids are present in leaves, stems and roots of the plant. With the same objective, Thin Layer Chromatography analysis was performed. Significant data values were not obtained for this analysis since the optimization of the solvent system used in TLC is time consuming and complicated.

Title – Identification and characterization of antibiotic resistant uidA and sul1 gene of Escherichia coli in Muthurajawela estuary in Sri Lanka

Supervisors –Mrs. Sajeevee Sarathchandra

Study Setting –Spectrum Institute of Science and Technology, Colombo

Distribution of Escherichia coli and K. pneumoniae in the surface microlayer and the underlying water in different locations along the Negombo estuary has been elucidated. Several physical parameters were measured onsite in order to determine the association between survival pattern of E. coli and the spatial and temporal variations. Biochemical and microbiological tests were conducted to confirm the presence of E. coli and K. pneumoniae and to isolate the microbes.

The study was done to analyze the antibiotic resistant profiles of the isolates against six different antibiotics, antibiotics: amoxicillin, cefuroxime, ciprofloxacin, clindamycin, nalidixic acid and tetracycline. The antibiotic resistant genes, uidA and sul1 gene in E. coli and K. pneumoniae was detected by polymerase chain reaction using specific primers. The purified PCR products with 16s rRNA primers were sent for sequencing to Gene Tech. Online similarity searches were identified using BLAST software. Statistical tests including paired t-test and analysis of variance (ANOVA) was performed to explain the difference in E. coli and K. pneumoniae distribution between the water layers and to compare the distribution in different locations throughout the study.

Analysis has confirmed that there is a significant difference between the surface microlayers, but the variability E. coli distribution in underlying water is high. Results obtained confirmed the availability of antibiotic resistant genes in high fractions in estuarine system which are influenced by the rainfall and the water temperature. The frequency of multi-resistant bacterial communities present in low dense areas is high, therefore the selective pressure exerted on E. coli is comparatively high. Presence of uidA gene in E. coli is detected in urban areas was 90% where the estuary is exposed to various antibiotics, isolates were highly resistant to nalidixic acid and clindamycin. Another promising finding was that sul1 gene is abundant in areas where the natural ecosystem is not disturbed, and those E. coli strains showed resistant to cefuroxime. ene sequencing was performed by Gene Tech and In conclusion, uidA gene is an important target which can be used to detect E. coli and thereby to assess the water quality or the level of faecal contaminants in estuarine ecosystems.

Title –Cytogenetic abnormalities in the female partners who face recurrent pregnancy loss and subfertility

Supervisors –Dr. Kalum Wetthasinghe & Mrs. Imalki Kariyawasam

Study Setting –Human Genetics Unit, Faculty of Medicine, University of Colombo

Recurrent pregnancy loss (RPL) and subfertility are a result of reproductive failure. A significant proportion of couples in Sri Lanka are affected, but there is no statistical evidence. The number of RPL and subfertile patients have increased over the years. RPL is known as two spontaneous pregnancy losses which affects 1 in 100 pregnancies. Maternal age and number of successive losses are the main risk factors for RPL. Subfertility is decreased fertility within one year of unwanted conception affecting 1 in 7 couples. One of the main contributors to subfertility is female age, as this affects the quantity and quality of the oocytes, thereby decreasing the chance of conceiving. One of the main causes of both, RPL and subfertility is chromosomal abnormalities. Previous studies report balanced translocations to be of the highest incidence in RPL patients. The objective of the study was to identify the chromosomal abnormalities specific to the Sri Lankan population. This will help in the future management and diagnosis of couples facing RPL or subfertility.

Fourteen females facing RPL and six subfertile females were included in this study from the years 2017 and 2018. Pre-clinical genetic counselling and the following steps were conducted in the Human Genetics Unit, Faculty of Medicine, University of Colombo, Sri Lanka. Conventional cytogenetics is the use of a staining technique known as Giemsa banding (G-banding) to study the chromosomes and identify any abnormalities. Peripheral blood samples that were obtained from the patients were subject to the standard 72-h culturing technique. Briefly, the steps of this technique involved culturing and harvesting of the cells, slide preparation and Leishmann staining and karyotyping analysis. Two chromosomal abnormalities were detected in the females studied. Polymorphic variants were also found. However, the highest incidence was that of normal karyotypes.

Although polymorphic variants were known to have no clinical significance, an association between polymorphic variants and RPL and infertility is now becoming clear. An association between chromosomal abnormalities and reproductive failure is evident. This study also suggests that polymorphic variants play a role in RPL and subfertility. This research area should be investigated further with the inclusion of a control group. This will make the results valid in the context of scientific evidence.

Title –Establishment of callus cultures for Gyrinops walla (Walla Patta) using leaf materials as explants.

Supervisors –Dr. Pathmasiri Ranasinghe, Senior Deputy Director, Herbal Technology Section, Modern Research and Development Centre, Industrial Technology Institute, Malabe.

Study Setting –Industrial Technology Institute, Malabe

Gyrinops walla is an endemic plant to Sri Lanka. The genus Gyrinops consist of 08 different species of trees that belongs to the family of trees Thymelaeaceae. The rapid increment in international demand for Agarwood over the past decade led to the over exploitation of the plant and smuggling of Gyrinops walla led to the rapid decline the species.

The present research study was conducted in the context of establishment of callus culture under optimized and most effective media composition and thereby induction of mass propagation of Gyrinops walla considering its current status.

Leaves of Gyrinops walla were used as explant materials. Leaf surface sterilization protocol was optimized with different concentrations of sterilizing chemical under varied exposure times.

Results of establishment cultures indicated a survival rate of 82% of explants was under the treatment procedure which involved 70% ethanol and 50% Clorox (5%) at 45 seconds and 15 minutes exposure times. Experimental procedures were carried out with the involvement of three plant growth hormones, BA, 2,4-D and Picloram under varied concentrations.

The cultured explants were incubated as light and dark conditions. The results indicated, calli was induced in explant cultures supplemented with higher concentrations of plant growth hormones. Calli was induced under both light and dark conditions.

Title –Antimicrobial Activity of Basidiomycetes Grown in Sri Lanka

Supervisors –Prof. Chandrani Wijeyaratne (Department of Botany, University of Sri Jayawardanepura)

Study Setting –Spectrum Institute of Science and Technology, Colombo

Synopsis –Basidiomycetes, a high group of fungi commonly called ‘mushrooms’, have the potential to be a source of novel antimicrobial compounds due to their natural metabolites. This research tested the antibacterial and antifungal activity of three edible mushrooms grown in Sri Lanka – Pleurotus ostreatus, P. abalonus and Calocybe indica- in varying masses of 6 mg, 8 mg and 10 mg, against three microbial pathogens, specifically Escherichia coli, Staphylococcus aureus and Candida albicans. The metabolites present within the extracts were identified via Gas-Chromatography Mass Spectrometry (GC/MS).

Ethanolic extraction followed by the Kirby- Bauer disc diffusion assay revealed that P. ostreatus inhibited the growth of both S. aureus and E. coli, while P. abalonus only inhibited S. aureus. C. indica did not inhibit the growth of any tested organism and Candida albicans was not inhibited by any extract. Statistical analysis by One-way ANOVA showed that there was no significant difference in inhibition by varying the masses of P. ostreatus against E. coli and P. abalonus against S. aureus. However, 10 mg was found to be the most suitable for the inhibition of P. ostreatus against S. aureus.

GC/MS revealed the presence of Octaethylene glycol monododecyl ether, Hexaethylene glycol monododecyl ether and Hexaoxacyclooctadecane – which are implicated in antiviral, antibacterial and antioxidant activity – in all three extracts and a further two, Isonitrosoacetophenone and Octaethyleneglycol, in P. ostreatus alone, which may have contributed to its increased antimicrobial activity. The hypothesis that certain Basidiomycetes grown in Sri Lanka show antibacterial activity was accepted and that they show antifungal activity was rejected based on this study.

Title – Preliminary Screening on forest leaf litter leachate as a novel source of small molecules for drug leads

Supervisors –Dr. Pathmasiri Ranasinghe, Senior Deputy Director, Herbal Technology Section, Modern Research and Development Centre, Industrial Technology Institute, Malabe.

Study Setting –Industrial Technology Institute, Malabe

Synopsis –In this current world the need for medicines is continuously increasing. Scientists and researchers are in a hunt to find lead compounds to make such medicines, in a way that is fast and efficient. It is obvious that extracting out such compounds is way less cheap than making them synthetically from scratch. The natural forest is also called the pharmacy due to its abundant source of resources as they produce and have a huge possibility to make a wide spectrum of organic molecules as defence against diseases. In addition to that about seventy percent of the drugs being prescribed today comes from such natural forest. Researches had been direct extracting individual plants to look for drug leads. This method can be time consuming and waste of resource. Statistics show that only one percent of the forest had been thoroughly examined for seeking medicines.

This study gives in a new approach to find drug leads faster and more efficiently in a way maximum of the resources available could be used to seek for a lead compound. By using the decomposed leaf leachate further increases the chance of compounds being found due to interaction of microbes. it was found that this research does seem promising because a rich amount of poly phenols (15.41±0.49 mg/g of GAE of dry extract) and greater amount of antioxidant activity was seemed to be seen (34.412 ±4.88 mg/g of trolox of dry extract). The alpha amylase inhibition was seemed to be low. And was concluded that it can’t be used for amylase inhibition. Polyphenols and the high antioxidant capacities can contribute to being the cure for various diseases. Exhibited a promising antioxidant potential that can be explored in the development of novel drugs for oxidative stress-related diseases.

Therefore the forest leaf leachate did show some potential for drug making. Further studies includes needing to define the precise mechanism of action, bioactive compounds. Also on long run this could be used to figure out and isolate which specific type of tree or bio molecule actually contributed to react against a specific condition.

Title –Study on the in-vitro glycation process of isolated Human Hemoglobin as an alternative to BSA for novel bio-assay development.

Supervisors –Dr. Pathmasiri Ranasinghe, Senior Deputy Director, Herbal Technology Section, Modern Research and Development Centre, Industrial Technology Institute, Malabe.

Study Setting –Industrial Technology Institute, Malabe

Synopsis –The research was based on the hypothesis that BSA (Bovine Serum Albumin), which is currently used as a target protein in anti-glycation assays, had structural differences to Human blood protein. It was assumed that this could cause a disparity between results of an in-vitro assay and an in-vivo human trial of an anti-glycative drug. The study was focused on determining whether Human hemoglobin would be a better alternative to BSA as a target protein in anti-glycation assays.

Blood samples of 10ml were collected from healthy volunteers for this purpose. The volunteers were in the age group of 22-24 years of age and there was an even balance of male and female. Hemoglobin was extracted by breaking the red blood cells by homogenization and then precipitating the RBC lysate thus obtained using saturated ammonium sulphate solution. Afterwards the precipitate was repeatedly washed with a phosphate buffer of pH 6.4. This pH was the isoelectric point of the human hemoglobin protein. After several washes with the isoelectric buffer, the remaining precipitate again dissolved in phosphate buffer of pH 7.4 and freeze dried.

The freeze-dried isolate was dissolved in PBS and its fluorescence was measured at 280nm / 335nm excitation/ emission wavelength. The readings obtained indicated the presence of hemoglobin but did not indicate its purity, in fact there was evidence that albumin was present in the isolate. The isolate was then incubated with glucose at varying concentrations and temperatures of 37˚C and 60˚C. BSA was also incubated in parallel with the isolate and fluorescence readings were obtained at the end of these incubations. The obtained readings indicated very low levels of glycation in the hemoglobin extract in comparison to the BSA incubation. This meant that that this extract could not be used as an alternative to BSA in a bioassay because a large glycation index is required to properly identify the proportion of glycation inhibited in an anti-glycation assay.

Title –Comparison of Three Genomic DNA Extraction Methods To Obtain High Quality DNA From Coconut And Rubber Leaf

Supervisors –Mrs. Imalki Karyawasam

Study Setting –Spectrum Institute of Science and Technology

Synopsis –DNA isolation from any living organism should be pure, intact and of high quality, as it is very important for any studies relevant to molecular biology. DNA integrity and purity are very important aspect in downstream applications such as random amplified polymorphic DNA (RAPD), simple sequence repeats (SSR), sequence related amplified polymorphism (SRAP) and amplified length polymorphism (AFLP). Two crude methods and one kit method was used and compared in this study.

The DNA selected for this study was from Rubber and Coconut leaves both of which have high secondary metabolites. These secondary metabolites obstruct the isolation and the purity of the DNA extraction by degrading the DNA sample The DNA extraction was optimized in various ways and run on 1% agarose gel. To measure DNA purity, both gel electrophoresis and spectrophotometer was used. Using the results of these techniques, the quality of the DNA extraction between protocols was compared and analysed. In addition, PCR was run to observe if the DNA extracted was pure and up to standard to be used in PCR experiments. Pairwise ANOVA with t tests was used to statistically analyse the data obtained. There was a significant difference in the absorbance results obtained between the two crude methods (Doyle & Doyle 1987; and Porebski et al, 1997) and the PROMEGA kit. When the pairwise comparison was done for concentration of DNA, all three methods showed significant differences. The PCR bands obtained for all three methods were not satisfactory therefore further optimization needs to be carried out in order to perfect the DNA extraction process for PCR.

In conclusion, the crude method for DNA extraction is recommended as opposed to the kits if DNA quality is of priority. If the quantity of DNA is more relevant, then the PROMEGA kit is recommended.

Title –Biotransformation of ‘Ma Dan’ (Syzygium cumini) by semi solid-state fermentation using Saccharomyces cerevisiae.

Supervisors –Dr. Pathmasiri Ranasinghe, Senior Deputy Director, Herbal Technology Section, Modern Research and Development Centre, Industrial Technology Institute, Malabe.

Study Setting –Industrial Technology Institute, Malabe

Synopsis –Biotransformation is a process where the structural composition in a chemical is modified using enzymes or organisms. ‘Ma Dan’ (Syzygium cumini) fruit is utilized for making fermented beverages in recent years. It is a seasonal fruit. S. cumini has a number of phytochemical constituents. This research study evaluates the bioactivity and antioxidant properties between fermented and non-fermented S. cumini fruit extract in fruit pericarp only (Group A) and the mixture of fruit pericarp with seeds (Group B). Total soluble solids, pH, Acidity and alcohol content was determined separately in the samples. Total phenolic compounds were determined by Total Phenolic Content (TPC) assay.

Antioxidant activity was determined by Ferric Reducing Antioxidant Power (FRAP) and Diphenyl-2-picryl-hydrazyl (DPPH) assay. The phytochemical constituents and antioxidant properties are enhanced in fermented samples than the non-fermented samples in both groups.As a future prospective, this research study can be optimized by performing assays more than one time and determine the Colony Forming Unit (CFU) in the starter culture before adding to the samples. ‘Ma Dan’ fruits must be washed with salt (NaCl) or food preservative rather than using tap water only.

Title –Screening for gene polymorphism rs77990309 in leptin (LEP) gene in patients with Polycystic Ovarian Syndrome in Sri Lanka.

Supervisors –Prof. W.S.S. Wijesundera (Department of Biochemistry & Molecular Biology, Faculty of Medicine, University of Colombo)

Study Setting –Human Genetic Unit, Faculty of Medcine, University of Colombo

Synopsis –Polycystic Ovarian Syndrome (PCOS) is a heterogeneous endocrine disorder characterized by anovulation and hyperandrogenism affecting fertility in women. The aetiology of PCOS is still not clearly understood as it is a multifactorial disease with genetic and environmental influence. Leptin, an adipocyte secreted hormone that signals to the hypothalamus about the triglyceride stores in the body to regulate appetite has recently been discovered to also modulate the reproductive axis. The link between leptin hormone and PCOS is seen by obesity, leptin resistance and endocrine dysfunction which are characteristic of both PCOS and leptin irregularity.

The aim of the study was to assess the prevalence of G>A nucleotide polymorphism on 5’ UTR of leptin gene (rs7799039) in patients with classic PCOS in Sri Lanka. PCR – RFLP was done on the DNA extracted from whole blood of 38 patients with PCOS to assess the genotype of the polymorphism in LEP gene.

In the current study, the presence of mutant form (A/A or G/A) of the polymorphism was more prevalent than the wild type in the patients with PCOS. A significance difference was seen between the serum FSH levels of patients with the wild type (G/G) and mutant (A/A or G/A) genotypes. The mutant genotypes of rs7799039 was associated with higher serum FSH level relative to the wild type genotype. The polymorphism was not associated with BMI or age of the patients.

Title – Antimicrobial activity of Trachyspermum roxburghianum.

Supervisors –Prof. Chandrani Wijeyaratne (Department of Botany, University of Sri Jayawardanepura)

Study Setting –Spectrum Institute of Science and Technology, Colombo

Synopsis –In this research, ethanol extracts of leaves, seeds and roots of a commonly used plant in herbal therapies, named Trachyspermum roxburghianum was used to investigate the anti-microbial activity against a gram positive bacteria, gram negative bacteria and two species of Candida using Kirby Bauer Disc Diffusion Assay (KBDDA) on Mueller Hinton Agar. Phytochemical screening of the plant extracts were also performed using standard methods. KBDDA revealed that all three extracts of the plant at a concentrations of 100mg/mL to inhibit Staphylococcus.aureus. However at 250μg/mL only the roots extract inhibited S.aureus. And GC/MS detected the compound Isonitrosoacetophenone to be present in roots sample. This compound is known to have anti-oxidant and anti-fungal properties. Phytochemical screening investigation revealed seeds extract to be the most potent as it contains all five phytochemical constituents tested (saponins, flavonoids, tannins, phenolic compunds and quinones) while roots extract was the least (Only quionones was present). Statistical tests, ANOVA was performed to explain the difference between the zones of inhibition of S.aureus for 100mg/mL of T. roxburghianum in KBDDA. Analysis confirmed there is difference between the means and greatest zones of inhibition were found in the seeds extract for S.aureus.

Conducting further investigations done on anti-microbial activity against various other microbes could be of help to enhance the effectiveness of using this plant as a therapeutic ingredient.

Title –Characterization of Phytoplankton communities in mangrove ecosystems associated with basin estuaries in the western coast of Sri Lanka

Supervisors –Mrs. Azra Sartaj & Prof. Pathmalal Manage (University of Sri Jayawardanepura)

Study Setting –Faculty of Science and Department of Zoology, University of Sri Jayawardenapura

Synopsis –The current study characterized the phytoplankton communities in the Chillaw basin estuary, Negombo basin estuary and Madu Ganga basin estuary and assessed the linkage between water quality parameters, the Shannon-Weiner diversity index of the respective locations and phytoplankton populations. A total of 64 species belonging to 12 taxonomic groups were identified in the assessed locations.

The most dominant taxonomic groups in the Chillaw estuary were Cyanophyceae (89%), Chlorophyceae (7%), Bacilliariophyceae (2%) Leptocylindraceae (1%), and Trebouxiophyceae (1%). The most dominant taxonomic groups found in the Negombo estuary were Cyanophyceae (93.7%), Trebouxiophyceae (24.4%), chlorophycea (2.21%) and Bcilliariophycea (0.5%). The most dominant taxonomic group in the Madu Ganga estuary were Cyanophyceae (98.2%), Dinophyceae (0.47%), Chlorophycea (0.35%) and Bacilliariophycea (0.17%). The study identified key species that were indicator organisms of pollution and the pollution indicators displayed a qualitative link with the pollutants. Protoperidium sp were identified as indicator of resuspension and phosphorous pollution, Oscillatoria sp. were identified as indicators of ammonia pollution and Synococcus sp. were identified as indicators of organic pollution.

The assessed Chillaw and Negombo estuaries were moderately polluted, while the Madu Ganga estuary was highly polluted. In could be concluded that mangrove estuarine ecosystems have a diverse range of phytoplankton species and that they can be used as effective indicator organisms.

Title –The isolation, characterization and analysis of the bacterial diversity in selected Sri Lankan Mangrove Ecosystems.

Supervisors –Ms Fathina Fara

Study Setting –Spectrum Institute of Science and Technology, Colombo

Synopsis –Mangrove soil has distinctly variable conditions of salinity, light and temperature and provides a unique ecological niche to a diverse multitude of microorganisms. A bacterial characterization study was carried out to isolate and identify the bacteria associated with mangrove soil in two mangrove locations in the Eastern and Western coasts of Sri Lanka. The two locations, Puttalam and Batticaloa were selected based on their exposure to human activity and vegetation. Soil and water samples were collected from two sites in each location.

Physical parameters such as temperature, pH and salinity were measured. The soil was also chemically analysed for nitrates, total phosphorous, organic carbon and potassium, through an external lab. The soil samples were then serially diluted, plated and sub cultured to obtain pure colonies. Morphologically dominant colonies were selected for further testing. The bacterial isolates were tested using Macroscopical, Microscopical and Biochemical tests. The samples from Puttalam were predominantly gram positive bacteria, while samples from Batticaloa consisted of mostly Gram Negative Bacteria.

The detailed identification tables in the ‘Bergey’s Manual of Systemic Bacteriology’ were used to identify the isolates to a genus or if possible, species level. Bacterial genera isolated from the mangrove soil samples in Puttalum included Pseudomonas, Bacillus, Listeria, Acinetobacter, Corynebacterium, Micrococcus, Arthrobacter, Aneurinibacillus, Paenibacillus and Burkholderia. In comparison, the genera identified from the Batticaloa samples included Proteus, Pseudomonas, Micrococcus, Acinetobacter, Klebsiella, Paenibacillus, Listeria, Citrobacter, Acinetobacter and Burkholderia. The enzymatic and biochemical characterization of the isolated bacteria indicated a diverse range of bacteria within the Sri Lankan mangroves. The isolates were tested for amylase, catalase, citrase, urease and protease production. 85% of the isolates from sample 2 in Batticaloa showed the production of Urease, while only a single isolate produced urease in sample 2 from Puttalam.

In addition, while none of the isolates produced citrase in Sample 1 from Puttalam, around 77% of the isolates did from sample 2 in Batticaloa. Species level identification was possible for some isolates, and species such as Micrococcus luteus, Listeria innocua, Burkholderia pickettii, Acinetobacer baumanii and Pseudomonas stutzeri were identified. Overall, it is evident that the mangrove ecosystem in Sri Lanka is home to a unique and diverse range of bacteria with useful characteristics.

Title –Association of leptin receptor (LEPR) gene polymorphisms and serum leptin levels in women with Polycystic Ovary Syndrome in Sri Lanka

Supervisors –Prof. W.S.S. Wijesundera (Department of Biochemistry & Molecular Biology, Faculty of Medicine, University of Colombo)

Study Setting –Human Genetic Unit, Faculty of Medicine, University of Colombo

Synopsis –Polycystic Ovarian Syndrome (PCOS) is a common condition affecting reproductive-aged women caused by hormonal abnormality leading to menstrual irregularity and infertility. Obesity, defined by increase in body-fat due to combined effects of genes, environment and their interactions is a common finding in PCOS. Leptin, an adipocyte-derived hormone is involved in relaying metabolic signals to the hypothalamus to regulate food intake and energy expenditure.Leptin modulates the Hypothalamus-Pituitary–Gonadal (HPG) axis. Dysregulation in HPG signaling due to hyperleptinemia leads to altered androgen levels in the ovary leading to PCOS. Soluble leptin receptor, encoded by LEPR gene is the major leptin-binding protein which is primal for transportation, sequestration and regulation of hypothalamus signaling.

Genomic DNA of a selected study population of 36 females with classic PCOS in Sri Lanka was extracted from whole blood. The frequency of Single Nucleotide Polymorphism (SNP) rs 1137100 of LEPR gene in the study population was determined by genotyping using Tetra-primer Amplification Refractory Mutation System (ARMS) PCR and the serum leptin levels were determined through ELISA. The association between the SNP and serum leptin levels in patients with PCOS was determined through statistical analysis. The mutant alleles were more prevalent in the population and no significant association was found between the polymorphisms and other parameters assessed.

Title –Isolation and characterisation of fungal species in selected mangrove environments in Sri Lanka.

Supervisors –Ms. Fathina Fara

Study Setting –Spectrum Institute of Science and Technology, Colombo

Synopsis –The project involved collecting soil samples from mangrove environments located in Puttalam in the North-Western province of Sri Lanka and Batticaloa in the Eastern province of Sri Lanka. Soil samples were analysed to determine the soil type, pH of sample, temperature and salinity. Chemical tests were performed on soil samples to identify the mineral composition. The effect of soil composition on the distribution of were observed as the collected samples were exposed to various anthropogenic activities as they were obtained from different locations.

Following sample collection, the samples were immediately stored in a cool dry environment under suitable conditions. Serial dilution and spread plate methods were used to isolate fungal species from the soil samples. Duplicates were maintained and the cultures were incubated at 31°C which is the optimum temperature for growth. Pure cultures were isolated and maintained to identify and characterize to a species level through macroscopic, microscopic and biochemical analysis. Mangroves in the Eastern province had a higher diversity of fungi in soil, mostly due to their pristine nature compared to those in the North-Western Province.

Title –Preliminary study on isolation and culturing of bacteria and fungi from leaf litter samples in Peak Wilderness Sanctuary (Sri Pada)

Supervisors –Mrs. Azra Sartaj & Dr. Pathmasiri Ranasinghe (Senior Deputy Director, Herbal Technology Section, Modern Research and Development Centre, Industrial Technology Institute, Malabe)

Study Setting –Industrial Technology Institute, Malambe

Synopsis –The main objective of this research is to find the antimicrobial and antiglycation properties of microbes found in decaying leaf litter samples collected from Peak Wilderness sanctuary (Sri Pada). Therefore, the results from this research will help to improve the knowledge base providing effective solutions to deal with the above challenges.

The experimental procedure for this research follows; gaining approval from Department of Wildlife prior to collection of samples. Samples were collected from 2 different sites. The composite samples were washed with sterilized distilled water. The washout was enriched with sterilized sorbitol and glycerol and then concentrated using a freeze drier. The concentrated washout was used to culture the plates. As bacteria and fungi were cultured and investigated, selective media such as Nutrient agar and Sabouraud Glucose agar media with chloramphenicol were used respectively. Selected microbial colonies were mass cultured into respective broth cultures. The freeze dried products of the broth cultures were used to perform the colorimetric analysis. All the extracts were screened for radical scavenging activity, anti-amylase activity and anti-glycation activity.

The results were statistically analysed and discussed. The only extracts which showed greater than 60% activity will be selected and used to study further about the composition in the future. The microbes responsible for this, can be characterized using LC-MS/MS or GC-MS/MS analysis. Therefore, this research aims to give a more accurate account on using these properties derived from the responsible microbes for therapeutics.

Following research has been carried out

Iron Status, Vitamin A Status, Total Energy Intake And Associated Factors Among Female Field Hockey Players

Compromised micronutrient or energy intake can affect aerobic endurance of athletes. The objectives were to assess body mass index, aerobi c endurance, iron and vitamin A status and nutritional intake of national level female field hockey players and to identify associations between analyzed parameters. A purposive sample of national level professional, adult female field hockey players (n=31) were recruited. A pretested questionnaire for general socioeconomic data and sport specific data was administered and a 24-hour dietary recall was done. Height and weight were assessed using standard techniques and BMI was calculated. Hemoglobin concentrations (Cyanmethemoglobin method), serum ferritin levels (ELISA)and serum vitamin A concentrations (HPLC) and aerobic endurance ('beep test') were assessed.

Following research is being carried out –I am investigating the role of a scaffold protein (AKAP12) in blood vessel development. Scaffolding proteins are important because they are critical regulators of key signaling pathways by providing a framework that organizes signaling proteins into specific complexes in space and overtime, thereby favoring one cellular outcome over the other. Abnormal blood vessel development is the underlying factor in many diseases including ischemia and stroke (vascular insufficiency) and tumor growth and retinopathies (vascular overgrowth). We believe that AKAP12 has a critical role in regulating new blood vessel development.My work involves culturing vascular endothelial cells, performing assays to study endothelial cell migration and blood vessel sprouting, performing immunofluorescent labeling assays to examine cellular structure using fluorescence and confocal microscopy, and using genetic engineering tools such as CRISPR Cas9 to rigorously investigate the role of AKAP12 in our model systems.

Title of Internship –Antimicrobial Activity of Different Nanostructured Coatings on Glass Surface

Supervisors –Dr. Abhilasha Misra

Study Setting –Department of Biotechnology of the Graphic Era deemed to be University, Dehradun, Uttarakhand, India

Synopsis –Glass plastic and metals are materials that are widely used in a range of applications and industries. Unfortunately, it is contaminated by microbes during daily use of tools made of these materials such as biosensors, surgical equipment, protective apparels of hospitals, food packaging and storage, industrial and marine equipment. These microbes on the surfaces form a structure called biofilm. Formation of these biofilms lead to hospital associated infections.

Even though the risk percentage of the infections due to implantations are very low the results are of great concern. As a result, superhydrophobic coatings have been developed. Techniques such as dip coating and spin coating are used to coat the material with superhydrophobic layer. Research were done to explain the antimicrobial activity of superhydrophobic coatings. It was proved that due to the air layer trapped between the surface and the superhydrophobic coating bacterial growth is inhibited.

The research was mainly focused on evaluating the antimicrobial activity of different coating on glass against E.coli. Methyltrimethoxysilane and Tetraethyl orthosilicate sols were prepared. The glass slides were coated using these sols by dip coat method. The coated glass piece was let to incubate for 5 days at 37C with periodic observations. Gram staining was used to distinguish the growth of E. coli on the surface of the glass. At a 24 hour interval for 5 days the gram staining was done to compare the growth of the inoculated e. coli. The stained glasses were observed under the light microscope under 100 x lens.

In conclusion we could say that the MTMS coating gave a better result compared to the TEOS coating. E. coli growth was inhibited on the MTMS surface. This could be due to the sir trapped between the coating and the surface which prevent adhesion of the bacteria. It is possible that the air layer got removed after 96 hour, as bacterial growth was seen on the 5th day of incubation. TEOS coating showed bacterial growth after 24 hours. Hence it was concluded that TEOS had no antimicrobial activity.

Title of Internship –Inter-relationship between Diabetes Mellitus and Vitamin D Deficiency in patients with Osteoarthritis: A Retrospective Cross-Sectional Study in a South Indian Tertiary Healthcare Facility

Supervisors –Dr. Sonal Sekhar

Study Setting –Department of Pharmacy Practice Kasturba Hospital, Manipal, Karnataka

Synopsis –AIESEC is a global exchange organization that facilitates cross-cultural internships for youth. Through AIESEC’s Global Talent program and a joint partnership between McMaster and Manipal University, I was able to pursue an 8-week internship at Kasturba Hospital, specifically in the department of pharmacy practice.

The aim of my research project, titled “Inter-relationship between Diabetes Mellitus and Vitamin D Deficiency in patients with Osteoarthritis: A Retrospective Cross-Sectional Study in a South Indian Tertiary Healthcare Facility” was to test for an association between vitamin D deficiency (VDD) and Diabetes Mellitus in patients with Osteoarthritis.

The study was conducted using a retrospective cross-sectional analysis of the prevalence of diabetes, VDD and Osteoarthritis in patients, with patient data obtained from the Kasturba hospital database. This was ultimately aimed at contributing further evidence to a growing body of contradictory research pertaining to the relationship between vitamin D deficiency and diabetes. Given the high prevalence of both conditions in India, this endeavor was anticipated to be of particular public health significance.

Overall, this opportunity to conduct my own clinical research project was of significant academic value. Gaining exposure to all phases of the research process provided me with new and deepened insights into the work and careers of scholars in both medicine and public health.

Title of Internship –Study of the vegetation and soil analysis under soil forest ecosystem in Doon valley, Uttarkand, India.

Supervisors –Dr. Archana Bachheti

Study Setting –Department of Biotechnology of the Graphic Era deemed to be University, Dehradun, Uttarakhand, India

Synopsis –Uttarkand, located at the foothills of Himalayas is filled with lush green vegetation and comprises of a diverse range of fauna and flora. The area is covered by moist deciduous forest present at an elevation of 1,500-3,000m. As a consequence of deforestation a large range of the Himalayan forest is restricted to protected areas and sanctuaries. The vegetation of the area has provided an interesting field for investigation throughout the years. The excessive exploitation of the forests in the area, natural disasters and development processes has accelerated the deterioration of the biodiversity of the forest. Globally human activity has been considered as the main reason for the alteration of land cover.

In the present study the chemical and physical properties of soil and the phytodiversity was analyzed from an Asarori forest range, situated near clement town, Dehradun. The forest site analyzed in the present study shows significant signs of disturbance, which could be attributed to its close proximity to human settlement. Although the dominant species of the study was Sal the IVI of the species indicates that the study area is under severely disturbed. Further the high IVI of invasive species such as Lantana camera and Murraya koenigii indicates that the site is disturbed by invasive species, threatening the pure state of Sal stands. The physical and chemical parameters of soil indicates that the study site is composed of fertile soil that can support vegetation which is in coherence with the data obtained via the phytosociological study.

Title of Internship –Pre-Formulation and Formulation studies on anti-hypertensive drugs

Supervisors –Dr. Anup Naha, associate professor of Manipal University

Study Setting –Manipal College of Pharmaceutical Science, Manipal University, Manipal, India.(The program was arranged by AIESEC of Colombo North, Sri Lanka with AIESEC at Manipal University )

Synopsis –Practical techniques required for formulation and pre-formulation and properties of excipients added to oral tablets were studied, before starting work, from the practical manuals at Manipal University premises and additional literature review.

Calculation and determination of bulk density, tapped density, Carr’s index, Hausner ratio and Angle of repose for pre-formulation of tablets. Evaluation studies on formulated tablets such as, weight variation, test for hardness, friability test, tablet Disintegration and tablet Dissolution were performed.

The above-mentioned studies were performed for varying concentrations of excipients on 300mg drug containing direct punched oral tablets and powder. The anti-hypertensive drug used was Lercanidipine and the excipients contained were starch/lactose and talc. All the above pre-formulation and formulation studies were performed on powder and tablets containing starch as the excipient with talc at 5%, 10% and 25% concentrations and lactose as the excipient with talc at 5%, 10% and 25% concentrations.

The two excipients showed varying properties when compared to each other and also at different concentrations, while starch seemed to be the more suitable and economical excipient to be used in oral direct punched tablets of anti-hypertensive drugs.

An Edinburgh Napier degree with a spectrum of opportunities

Ashinsa from Sri Lanka was lucky enough to visit us from one of our transnational education partners, the Spectrum Institute of Science and Technology. She shares her Edinburgh Napier experience and her thoughts on Edinburgh as she hopes to return in the near future.

Gaining an Edinburgh Napier degree from the comfort of my own home I’ve loved biology ever since I was a child and I knew biology was what I wanted to pursue my future career in. Growing up in Sri Lanka, I wanted an undergraduate programme that allowed me to stay in Colombo for the entirety of its duration. Thus, I found Spectrum Institute of Science and Technology (SIST), which is the partner of Edinburgh Napier University in Sri Lanka. The microbiology and biotech programme they offered seemed very interesting and covered a variety of topics.

Future career goals I would definitely like to continue my higher studies. However, before I do, I hope to work in Sri Lanka for a few years, preferably in environmental microbiology or a similar field. I feel as though ‘real life’ experience is necessary moving forward. In 2-3 years (ideally), I hope to do a PhD, possibly outwith Sri Lanka. I feel my course will be immensely helpful in gaining experience in laboratory techniques and correct research methodology for future use.

The opportunity to travel to Edinburgh I was given the opportunity to visit Edinburgh through the Small Project and Mobility Grant, awarded by Edinburgh Napier. The project aims to provide students with a glimpse of student life in Edinburgh and allowed me to compare the similarities and differences between the programme in Colombo and Edinburgh. It was the first time I travelled entirely on my own, which was an interesting experience. Edinburgh Napier at a glimpse "Students at the University should consider themselves very fortunate." Edinburgh Napier University is wonderful! The faculty and staff were very welcoming and friendly, often going out of their way to make my visit special. While I was based at Sighthill, I had the pleasure of seeing all 3 campuses. Merchiston, with its lovely tower and Craiglockhart, with its sprawling lawn and War Poets display. Both were very memorable. The facilities Sighthill had to offer, from coffee shops to student lounges, were quite a novel experience. The University definitely exceeded my expectations. I was blown away by the facilities the campuses have to offer. Everything is provided for Edinburgh Napier students, from Wi-Fi, computers, study rooms, cafes and lounges. Everything one would need to lead a life as a student is available. Students at the University should consider themselves very fortunate.

Project Title I –

Development of a standard plant tissue culture laboratory.

Specific Objectives –

• To establish a modern plant tissue culture laboratory (PTCL) under international standards.
• Commercialization of plants (orchids, indoor plants, aquatic plants, bonsai varieties etc.).
• To develop new plant varieties (hybrids).
• Development of Hydroponic Systems and Temporary Immersion Plant Bioreactor Systems.
• To provide the best tissue culture certificate course on basic tissue culture techniques, innovation of new culture methodologies and commercialization of products (on local and export market).

Project Title II –

Development of a bacterial treatment model to facilitate biodegradation and detoxification of textile dyes present in waste water derived from apparel industries in Sri Lanka.

Research Problem -v

To develop a bacterial treatment model for the treatment of specific, identified toxic chemicals and decolorization and detoxification of textile dyes (azo dyes) released from the apparel industries of Sri Lanka.

Specific Objectives –

• Characterization of dyes used in apparel industries in Sri Lanka.
• Characterization of types of apparel wastes and sludge waste.
• Determine the selected dyes that is treated chemically which makes the highest contribution to the sludge waste and characterize the chemical.
• Isolation of microbial strains from different sources. (textile industrial waste effluents, garbage dump sites etc.)
• Biochemical characterization of the selected strains
• Molecular identification of the selected strains/ Sequence analysis of the selected isolates.
• Test the isolates with the selected laboratory preparations based on dye and sludge characterizations.
• Find the most effective strain or the combination of strains on the selected chemical preparations.
• Optimizing the activity of the selected strains for the degradation of the identified compounds.
• Preparation of the industrial inoculum and developing a pilot physical model for treatment plants.
• Optimization of the pilot physical model upon industrial requirements.
• Analysis of treated waste effluents.
• Determination of sludge characteristics upon treatment.
• By product analysis – Biogas, toxicity.
• Preparation of final product.

Title of the internship: Effect of Natural Compounds on Inhibiting Bacterial Biofilms

Supervisors: Prof. T. S. Murali, Ms. Anjali Warrier (Research Scholar)

Study Setting: Department of Life Science, Manipal Academy of HIgher Education, Karnataka, India.

Synopsis: Bacterial biofilms act as a major impediment in the treatment of chronic wound infections. Biofilms are difficult to eradicate due to their enhanced antibiotic resistance properties and virulence potential compared to their planktonic counterparts. Recently, several natural compounds have been screened for their effectiveness in reducing biofilm levels when treated in synergy with antimicrobials.

This research was mainly focused on understanding the various techniques involved in biofilm inhibition, expression analysis of specific genes involved in quorum sensing, testing the biofilm inhibitory concentration of different natural compounds, screening natural compounds for their effect on biofilms by both common microbial testing and molecular techniques.

Minimum Inhibitory Concentration ((MIC) tests were done for four selected bacterial strains with twelve different antibiotics to select the most sensitive and effective antibiotic to the specific bacteria. Followed by biofilm assay and micro broth dilution. Micro broth dilution is a common method used to determine lowest concentration of antibiotics that is effective. In this method, a serially diluted broth with antibiotics is used. 96 well plate was used where the bacteria were inoculated into the wells in the presence of antibiotics of different concentrations.

Molecular techniques including primers to target the genes forming biofilms such as Reaction mix for LasI, LasR, PelA, PelB were designed for P. aeruginosa (PA21 and PA333). Temperature gradient PCR was performed to determine the annealing temperature of the primers. Finally, quantification of gene expression was carried out using qT-PCR. Real-time PCR can quantify gene expression and confirm differential expression of genes. The expression of biofilm forming genes of Pseudomonas aeruginosa were analyzed using RT-PCR.

Title of Internship – Produce and purify the exopolysaccharides from three bacterial strains by adopting submerged fermentation process

Supervisors –Subbalaxmi S (Assistant Professor – Senior Scale)

Study Setting – Department of Biotechnology, Manipal Institute of Technology, Karnataka, India.

Synopsis – Bacterial extracellular polymeric substances (EPS), which are basically bacterial metabolites, have currently become a subject of great concern of modern day microbiologists and biotechnologists. Among these metabolites, bacterial exopolysaccharides or EPS, in particular, have gained a significant importance. Polysaccharides are the most diverse families of biopolymers. Bacterial exopolysaccharides are contextually limited to all forms of polysaccharides synthesized and secreted into cellular external environment which may remain loosely attached to the surface (capsule) or completely detached. Bacteria produce these biopolymers with varied chemical properties via utilization of simple to complex substrates.

Initially in the study, bacterial growths and kinetics were observed three strains of bacteria (S. pas, Lactobacillus helveticus, and Lactobacillus bifermentans). The cell concentrations were collected by measuring the dry biomass of the bacteria. In order to obtain dry mass, the samples were centrifuged and the precipitate was dried in hot air oven and the cell concentrations were obtained. To analyse exopolysaccharides contents in substrate solutions, DNS and EPS assays were conducted, sugar intake and EPS productions were observed. Readings were obtained using spectrophotometer or calorimeter.

In the latter part of the study, proteins were isolated using ammonium sulphate precipitation. Then, antimicrobial testes were carried out to test the antimicrobial activity of the exopolysaccharides of obtained samples.