Chromogenic Agar; the gold standard for identification and analysis of pathogens

By: Kassapa Ellepola

‘Traditional standard growth media’ for microorganisms, since its introduction in the 1800’s has dominated for more than two centuries as the most widely used growth media for bacteria and fungi. The discovery of Agar, extracted from Red Algae became the best solidifying agent for microbiological media and not much has changed in these media apart from their chemical composition depending on the purpose (general purpose, enrichment, selective etc) of the medium.

Dr. Alain Rambach in 1979 pioneered the invention of ‘chromogenic culture media’ that at the time was believed would make a promising change in the medical field as well as in industry. Chromogenic Agar is a medium that assists the isolation and identification of certain pathogenic microorganisms with the aid of colony color, colony morphology and colony growth. Pathogenic microorganisms are fastidious. Therefore he equipped the medium with special nutrients that enhance their growth while inhibiting the growth of less fastidious organisms.

The normal procedure to grow fastidious organisms will be to carryout enrichment before growing in standard culture media. The proven technique is to carry out Enrichment in broths before plating out. This has been practiced for a long time. But with the use of chromogenic agar, will this be necessary!

Detection systems of conventional culture media combine various carbohydrates, amino acids, and glycosides with a suitable indicator-system. The incorporation of enzyme substrates as a detection system has led to the development of numerous primary isolation media.

Chromogenic enzyme substrates consist of chromophors linked to an enzyme-recognizing-part, such as a carbohydrate, amino acid or phosphate. Specific enzymes produced by the target microorganism cleave the chromogenic substrate liberating the chromophor, which highlight the micro-organism by coloration of the grown colony.

There is a diversity of chromogenic agar that is available for microbiological work. One such medium is the chromogenic Agar for identifying ‘methycillin resistant Staphylococcus aeureus’ (MRSA). There are chromogenic media designed for identifying Neisseria sp as well.

Infections due to Salmonella have become a principle health issue related with the food industry as well as with water quality. Rapid identification methods are needed for prevention of outbreaks. ‘Salmonella Chromogenic media’ has been designed based on Salmonella’s ability to utilize one of two chromogenic substances in the medium while others in family Enterobacteriaceae can utilize both.

‘Salmonella Chromogenic Agar Base’ combines two ‘chromogens’ for the detection of Salmonella sp. ‘Magenta-caprylate’ and ‘X-gal’. β-D-galactosidase acts on X-gal as it is a substrate for the enzyme . Lactase negative Salmonella species hydrolyse the chromogen, Mag-caprylate giving magenta colored colonies. The medium contains bile salts to inhibit the growth of Gram-positive organisms. The medium may also contain novobiocin to inhibit Proteus growth and cefsulodin to inhibit growth of Pseudomonads.

Not only for bacteria, but it also works for fungi such as Candida sp which its different strains are colored differently in a chromogenic agar plate.

Hospitals and nursing care institutes find it very important to find immediate identification of pathogens. Specialized chromogenic agar can be used for this purpose as it would easily inhibit undesirable organisms while enhancing the growth of the fastidious pathogens while giving colored colonies for pathogens. Use of chromogenic agar will aid the food industry as well.

Using chromogenic Agar is distinctively advantageous than traditional methods. Within 24 hours after inoculation, the chromogenic agar medium allows the presumptive identification of the pathogen with the aid of colony colors. But colonies grown on a traditional agar plate does not allow direct identification. Therefore other methods such biochemical tests should be employed which may involve costly tool kits and would take another day or two to give the results.

Improved accuracy compared to traditional agars, faster detection of targeted organisms, cost effective methodologies, ability to easily integrate other molecular methodologies and accurate selection of colonies without missing or mixing colonies are few more benefits.

Although classical media serves as an inexpensive option financially, its results are less reliable. The modern molecular biological approaches are very accurate but could be costly and laborious. Chromogenic Agar falls in-between both these techniques making it a middle cost and appropriately accurate option.

Initially nobody including industries has showed interest in the Chromogenic medium developed for E.coli in 1979, patented by Dr. Rambach. Therefore he stopped his work in chromogenic field for 10 years and he was more interested in genetic engineering. After 1989 he again developed ‘Rambach Agar Salmonella’ for identifying Salmonella sp and ‘CHROMagar E.coli’ for identifying E.coli.

  

More than three decades after its invention, chromogenic agar caters to only 10% of the world’s usage. Expansion of awareness among the microbiology community will lead to the maximum utility of this technology.

References:

• http://ezinearticles.com/?How-Microbiologists-Use-Chromogenic-Agar-Media&id=3549221
• http://www.oxoid.com/UK/blue/prod_detail/prod_detail.asppr=CM1007&c=UK&lang=EN&org=124&img=CM1007C&sec=
• http://www.internationalpbi.it/docs/PBI/seminari/relazioni/Seminario-CHORMagar-PBI-AlainRambach.pdf
• http://www.chromagar.com/p-30_years_of_innovation.html?PHPSESSID=9f63481a27c30dc4a35d928f18c18482#.ULc9TuT28_h
• Z.Samra, M. Heifetz, J. Talmor, E. Bain and J. Bahar J. Clin. Microbiol. 1998, 36(4):990.
• http://www.internationalpbi.it/docs/PBI/seminari/relazioni/Seminario-CHORMagar-PBI-AlainRambach.pdf
•  http://www.biosynth.com/index.asp?topic_id=137

Bio-deterioration of Cosmetics and Personal Care Products

By: Kassapa Ellepola

Bio-deterioration can be defined as an undesirable change occurring in a natural or processed material of economic importance, which is brought about by the activity of living organisms especially the microorganisms.

In the case of Cosmetics and Personal Care Products, bio-deterioration can cause them to be objectionable for use and also could be therapeutically inactive. These products contain a number of chemicals which support microbial growth. Mainly chemicals such as sugars as sweetening agents, polymers such as starch and cellulose used as thickening agents and for use in suspensions, and oils and fats used in the preparation of creams and emulsions can be considered. Also the growth of microorganisms on many of these products causes the formation of foul smelling odors, and the separation of emulsions into their component parts.

Bio-deterioration of Cosmetics and Personal Care Products can be caused by bacteria, yeasts and fungi which are extremely metabolically diverse as they are with the capacity of genetically modifying themselves to adapt to a broad range of environmental conditions.

Microorganisms can be introduced in to the product via the raw materials, the processing plant, packaging materials and from operatives or by elsewhere in the environment. But the growth and multiplication of these organisms can be supported by the formulation of the product, ambient temperatures as well as by the humidity factor.

The application of cosmetics can occasionally cause irritation and this may be a result of high levels of microbial contamination. The contaminants may produce foreign proteins which can evoke allergic reactions. Particularly the ‘eye’ is susceptible to infection from contaminated cosmetics and other personal care products.

Cellulose polymers which are commonly found in cosmetics can act as potential targets for microbial attack and sometimes support extensive growth under suitable conditions. As a result, cellulosic substances can be converted from a stiff gel into a running liquid, thereby rendering the cosmetic unfit for use. Three different types of enzymes (exo-beta-1, 4-glucanase, endo-beta-1,4-glucanase and beta-glucosidase) are involved in the degradation of crystalline cellulose. Enzyme production is basically done by three fungi belonging to the genera Aspergillus, Fusarium and Penicillium.

Microorganisms also possess a significant ability to inactivate biologically active molecules. Antibiotics in cosmetics may be destroyed by microbial action, for example penicillin will be destroyed by any β-lactamase producing organism. Microbial enzymes which inactivate chloramphenicol is also known. Many preservatives, disinfectants and surface active agents are metabolized by bacteria, although usually at concentrations below the normal ‘in use’ concentration. There have, however, been a number of reports of the degradation of simple preservatives such as benzoic acid at ‘in use’ concentrations.

Visible growth of microorganisms can be observed in liquid formulations such as sediments, turbidity or pellicles. These are good indicators of bio-deterioration. Color changes due to alterations in the components of the product may result from the pH, redox or other changes caused by the metabolic activities of an organism. The color changes may also be due to pigment production, as an example the Pseudomonas genus has the ability to metabolize a broad range of products and produce soluble pigments ranging in color from blue to green to brown. These organisms can render conditions that favor the growth of anaerobic organisms. Some oxidative yeasts can cause a rise in pH by utilizing organic acids, encouraging the bacterial growth.

Gas production is also a visible indicator of bio-deterioration of the products especially when glycerol, a common ingredient in many cosmetics is fermented by some common waterborne organisms.

Apart from those changes, the physicochemical condition of emulsions can be changed. The oil phase can be hydrolyzed and also an increase in the viscosity and aggregation of particles may be observed.

Olfactory effects can also change due to bio-deterioration. Off odors can be formed due to the production of gases such as Ammonia, amines and also hydrogen sulphide. These odors can be disastrous to cosmetics and toiletry preparations which depend on specific perfumes. The smell of mould is one of the common olfactory warnings. Actinomycetes growing on products cause an earthy odor due to ‘geosmin’. Spoilage by yeast gives a typical alcoholic odor.

The texture may also feel different where creams can get a lumpy or ‘gritty’ texture and there might be changes in the viscosity which can be felt when applied to the skin.

Oil/ water emulsions are very much susceptible to spoilage as the water allows the microorganisms to spread. The spread occurs throughout the continuous phase of water and also at the boundaries. The concentration of microorganisms depends on the relative solubility of the oil in water.

The degradation of creams depend on the presence of water. As water is one of the major products of metabolism, the degradation process becomes autocatalytic. Many preservatives, disinfectants and surface active agents are metabolized by bacteria.

Recently there is a tendency to use natural ingredients such as animal proteins and vitamins. These ingredients are highly nutritious to microorganisms. Glycerol which is abundant in creams can be metabolized by microorganisms such as Klebsiella producing gases in poorly preserved products. Mould growth can occur in products such as antifungal, calamine, baby and hair creams and other cosmetic formulations including moisture and cleansing creams.

When ointments and oils are considered, normally they will not support the growth of microorganisms as they are anhydrous material. But the containers they are filled into may contain air spaces which could allow the growth of moulds which utilize atmospheric moisture. Maize and olive oil can be spoiled by bacteria and fungi.

Shampoos are contaminated by a range of Gram-negative waterborne bacteria. They can cause slimy sediments, pellicles, discoloration and odor. The loss of lathering properties can also take place. Shampoos also have a potential to be spoiled by bacteria such as the Pseudomonads.

Powder products can be spoiled by visible growth of moulds in damp conditions and it is very important that topical preparations do not contain Clostridia spores. Precautions should be made on powders and other solid cosmetics that are applied in the region of the eyes.

Solid cosmetics such as lipsticks can be subjected to mould ‘blooms’ where moulds can grow on the lipstick while it is inside the lipstick case. This may be a result of it been moistened by breath when used or when moistened from perhaps saliva.

Soap gives off moisture and if wrapped with an impermeable plastic over paper or card, may become discolored due to mould growth on damp paper.

Therefore bio-deterioration of cosmetics and personal care products must be prevented as it is significantly important with relation to health and economy. The disease causing microorganisms must be removed from the product. The presence of many thousands of organisms however indicates that spoilage will occur. Regular monitoring during development and manufacturing, establishing the type and minimum number of organisms which are achievable for each specific product should be well defined.

Reference:

• Dart, R. K., Microbiology for the analytical chemist, Royal Society of Chemistry (Great Britain), Information Services, pg: 16, 17 (14.01.2010)
• Harrison, L.A. 1987, Microbial degradation of cellulose polymers used in cosmetics and toiletries, Int J Cosmet Sci. Apr;9(2):73-84.
• Smart, R., Spooner, D.F., 1972, Microbiological spoilage in pharmaceutical and cosmetics, J. Soc. Cosmet. Chem. 23:721-737

Remembering Saddhatissa Wadigamangawa

By Kassapa Ellepola

This piece of writing is for the remembrance of ‘Saddhatissa Wadigamangawa’ whose 11^th death anniversary falls on the 14^th of December 2011.

‘Saddhatissa Wadigamangawa’ as known by many, was an eminent politician from Sri Lanka. He was a member of the National State Assembly in the latter part of the 1970’s, a Trade Union leader and an activist, teacher and also an author. He was born in 1928 and passed away in the year 2000, completing 72 valuable years of service to the country and its people.

He was born on the 4^th of February, 1928 in Kuliyapitiya, Twenty years before Sri Lanka achieved independence. He received his primary and secondary education from St. Paul’s College Kandy and from St. Anne’s College, Kurunegala. The political career of Saddhatissa Wadigamangawa was well influenced by his primary and secondary education as well as from his family background; Wadigamangawa’s being a renowned Family in the North Western Province. After the completion of his studies, he joined the profession of teaching and was appointed as an English Teacher at the Saranath Vidyalaya, Kuliyapitiya in 1952. He also taught at the Methodist College in Bandarawela.

He was married to Padma Ilankoon from Pilessa, Kurunegala and was father to six children.

He was influenced by the attempt of the world’s Communist movement to achieve global democracy and decided to join the Sri Lanka Communist Party with the aspiration of serving the people and his country. He was involved in the establishment of a branch of the Sri Lanka Communist Party in Kuliyapitiya in 1952. As a result of his leadership abilities, he was soon appointed as the Divisional Secretary of the Communist Party in the Kurunegala District and later on as a Central Committee member, who led many promising struggles to win people’s rights.

He was also a prominent figure in the Trade Union sector. He has held the positions of Kuliyapitiya Branch Chairman of the ‘Sri Lanka National Teacher’s Union’ (SLNTU), as the Divisional Chairman of the SLNTU of the Kurunegala District and he was also a Central Committee member of the SLNTU. His service as the District representative of the Kurunegala ‘Teacher Transfer Panel’ and as the Kurunegala district representative of the Education Service Cooperative Union is always reminisced by the teachers and educationists in the Kurunegala District. He committed himself in solving problems of teachers and addressed issues in the teaching profession throughout his career.

His services are also recognized by a lot of dance and music artists in the Kurunegala District and also by Sathkoralaya Literary Union where he has been a committee member. He is been recognized by many as an individual to have had shown his abilities in a various amount of fields. He was also fighting for the rights of the farmer community where he was the first ever elected General Secretary of the ‘Lanka Jathika Govi Peramuna’ led by Dr. S.A. Wickramasinghe.

As he was recognized as a prominent leader of the Communist Party who led the party and its members against the United National Party (UNP), he was expelled from his teaching career in 1965. He was reappointed as a teacher in 1970 after the ‘United Front’ acquired power.

He also did an eminent service as a community leader to the people in the Kurunegala and Puttalam Districts, to refurbish Economic and Cultural Development in urban and rural areas.

He left the ‘Communist Party’ and joined the ‘Sri Lanka Freedom Party’ (SLFP) led by Sirimavo Bandaranaike and contested the General Election in 1977. Saddhatissa Wadigamangawa was nominated as the representative of the Anamaduwa electorate from the SLFP. He got the opportunity to enter the Sri Lankan National State Assembly in 1977. He was a member of the National State Assembly from July 1977 to February 1980. When all democratic and left wing political parties were washed off in the 1977 election by the United National Party (UNP) where they had a 5/6 power over the opposition, Wadigamangawa still managed to be selected to the National State Assembly. He was one of only eight to be elected to parliament from the SLFP in the election which is considered a landmark of Sri Lankan political history. This election result showed that the people in Anamaduwa were in great favor of Saddhatissa Wadigamangawa.

However, he lost in the 1980 By-election; it was only from a narrow margin of 1787 votes. The ruling UNP appointed Ashoka Wadigamangawa, a close relative of Saddhatissa Wadigamangawa as the UNP candidate. But political analysts say that the very well established UNP government who had a stronghold of 5/6 in the National State Assembly from the previous 1977 election, put all its collective effort including money, manpower, ministers including President J.R. Jayawardena himself was positioned in Anamaduwa for campaigning to win the electorate from Saddhatissa Wadigamangawa, which they succeeded but only from a narrow margin.

Saddhatissa Wadigamangawa is considered a very close friend of Late Minister Anura Bandaranaike and also J.R.P. Sooriyapperuma. He is talked about with great respect among people in different levels of the Sri Lankan Society and was regarded as a charismatic person by many politicians from different parties.

He has represented the country in many delegations that toured to other countries. He led the Sri Lankan Communist Party delegation to the Soviet Union in 1967 and also led a group of the SLFP members who went to Cuba for the World Youth Conference. He was also a member of the team which visited Venezuela in 1980 for Inter Parliament Society Conferences.

He joined the Democratic United National Front (DUNF) in the early 1990’s. The party was headed by promising politicians such as late Lalith Athulathmudali and late Gamini Dissanayake. He contested the 1993 North Western Provincial Council election from the DUNF where he won and then was appointed to the post of Chairman of the North Western Provincial Council which he held till 1999 where again he managed to perform lot of services to the people in the North Western Province.

The latter part of his life was spent leisurely with the books he loved. Saddhatissa Wadigamangawa was a great admirer and follower of music, drama and dance, cinema as he himself acted in few movies and many other cultures. He was also extremely knowledgeable about them. His close friends say that when he got himself involved in any art, he gave it a new dimension and a great sense of value. In the final few months of his life he translated ‘Zorba the Greek’ a great novel by the Greek author Nikos Kazantzakis. He was also into composing poems and song lyrics.

He succumbed due to a tumor in December 14, 2000 and his funeral was held in Anamaduwa, amidst a huge gathering of admirers who has received his service and has benefitted from him.

A very close friend of Saddhatissa Wadigamangawa, W.A. Abeysinghe a renowned author and a poet in the country, in a letter of remembrance of Saddhatissa Wadigamangawa on his funeral day, quoted a few lines from the drama ‘Julius Caesar’ by the great poet William Shakespeare which he believed that would be best to describe Wadigamangawa. The dialogue of Mark Anthony says,

“His life was gentle

And elements so mixed in him

That nature might stand up

And say to all the world

This was a Man”

May he attain the blissful ‘Nibbana’.

The Highland Beauty

By Kassapa Ellepola

Who would think that a suddenly picked bunch of friends and a suddenly planned trip would give you one of your greatest times of life? Well, it became real when we took the morning train from Colombo Fort. There were five boys and four girls who clambered in the train beaming at the joy they were going to have.

The train left the Fort heading upcountry gathering its speed. The train busied its way through the greenery surrounding the upcountry. The

atmosphere got cooler and brightened our feelings. The Colombo-Badulla Railway had its old splendor, which we now have come across quite a number of times. Its beauty has lifted it seems. The mountains covered with mist indicated that upcountry is still young and handsome as it has always been.

This time we traveled further. By late evening, we reached Diyathalawa. We got off from the station where two more of our friends joined us, and followed a road down to the small Diyathalawa town which was not much more than a few government buildings and few shops. It was the ideal place for a holiday, silent and inviting.

We resided the night in a small holiday bungalow, well furnished and with a good cook. We had plenty of fresh vegetables in the locality which added taste to our meals. The nights’ stillness was challenged by the sound of continuous gunfire exploding from the guns of Army trainees at the nearby Diyathalawa Army Camp. We could see the night being lighted by the ignition emanating from the rifles. It is a tiresome period which takes days and nights of training, producing hardy soldiers.

The next morning we started sightseeing. We got our backpacks packed full with water, food and left the bungalow. We first took a bus heading to Addisham Bungalow with a transit in Haputale. The ‘Bungalow’ was situated in a reserve named ‘Thangamalai’. We had to go past a tea estate first to reach the reserve. The walk across the reserve through the shade of the trees, the cold breeze and the call of birds gave us such contentment.

We reached the ‘Bungalow’. It was a fine piece of architecture. A majestic Bungalow made of solid mantel creating splendor to the vicinity. The heavy wooden doors made of solid oak, and the beautifully carved window sills brought majesty to the building. The finely furnished house was elaborated with antiques and all the other decorations in the house were an additive to its glamour.

The cliff beside gave a panoramic view over the surrounding area. Occasionally, the mist covered the area hiding the gigantic mansion drenching the well kept loan and the rosy garden in mist. Roses in an assortment of colors were neatly grown in beds, indicating the sensation of extreme gardening. The place revealed the true glamour of the upcountry.

We left the place and took a bus to Kalupahana, which was situated from an hour’s ride from the Haputale town. Our intension was to see the ‘Bambarakanda Falls’, which is written down as the tallest waterfall in the country and we considered it as a great privilege to witness this important place.

It took a tiresome walk through a hilly road to reach the falls. The road was well made and passed a quiet village with only few houses. We could get a glimpse of the waterfall above the mist covered green tree tops. It had such elegance that it looked like a bride whose white veil has been put over the cliff. It added advancing beauty to the surrounding area. The mild atmosphere created a lively ecosystem which made the area so pleasant.

To reach the bottom of the waterfall we had to carve our way through a small thicket and over some small streams. We tried our best to avoid leeches that tried to climb up our legs, but was not so successful. We speeded up through the trees, deliberate to see the waterfall closer. The deafening sound of the mass of water pounding on the pond was getting near.

Around a corner with a blink of the eye we saw it at last! We looked up at the waterfall with awe! It deceived us making us think that the water was raining down from the sky as the blue sky decorated its background. The whole surrounding was covered with mist generating from the waterfall. We were drenched with the mist. It was a sight to witness for a lifetime….

We were hurried to get changed in to our swim suits and we had a great time in the water. The water hit the pond below with massive force. We splashed ourselves in the water and had a great time until we felt tired and hungry. We climbed out and sat under the shade of some conifers where we found a great place which gave a splendid view over the waterfall. The lunch also tasted great with the background sound of the waterfall and the sight of it.

We decided to climb further more in to the conifer forest in search of another infamous waterfall named ‘Lanka Falls’. We kept further through the dark forest treading on the aging dry roots and sticks which covered the forest bed. The place had the most mystique look with it and we thought that the giant trees were looking down at us, the only souls who had infiltrated the sleeping ancient forest.

We kept through a foot path which carved its way through grass which had grown shoulder height. We finally hiked our way to the waterfall which was situated in a dale. A sound of water echoed around the place filling our ears. We saw it through the trees. After few minutes, we finally reached it!

It was small, not more than fifteen meters high but looked immensely beautiful. The pure white water leaped down to the pool below with elegance and style. The basin below took exactly the shape of the geographical area of Sri Lanka and we could guess now why the falls has received its name. The place was picturesque and we wasted our time with our cameras.

We believed that our virtue brought us to witness such a gorgeous place. We had pride when we turned back and left the place that we were few of the lucky people who has been here. We said farewell to the two beautiful waterfalls and the charming surrounding, gathering plenty of sweet memories to cherish for a lifetime.………….

The Sand, waves and Paradise

By Kassapa Ellepola

The Chill in the dawn was overwhelmed by the thrill inside the group of boys who had gathered in the Fort Railway Station, who were bound to take a train ‘Down South’.

The train gathered its speed across the awakening Capital of Sri Lanka and was taking the railway track heading Down South, to Galle. The railway ran along the South-Western coastal line. It was always a nice feeling to hear the graceful sound of the ocean, to get the first glimpse of the blue color and the ivory colored beaches through the tropical trees. The waves climbed up the beautiful beach regularly and again were dragged back by the ocean. Waves created white foam on the beaches which disappeared with the succession of another hurrying wave.

The train travelled across eye-catching sceneries of the South-Western coast passing beautiful beaches such as Mount Lavinia, Beruwala, Polhengoda and Kaluthara.

The train was not crowded and it was a very comfortable journey. It gave a pleasant sensation to stare out of the window towards the sea. The wind kept blowing towards us carving a smile in the faces.

After a four hour ride from the Capital, we reached our destination, ‘Galle’. Galle served as a strategically important naval base and also as a harbor from the period of Dutch and British.

The highlight of the city was the Galle fort, constructed on a headland beside the Galle bay; it is a majestic building complex with ancient buildings constructed by the Dutch. The fort was fortified by a high strong wall with guard towers at the corners. The Fort only had one entrance from the mainland. A majestic clock tower loomed from the inside overlooking the area.

The wall around the fort allows a walk on top of it which gave a great view over the coastal area. The grey color of the clouds added beauty to the ocean and the waves came hitting the jagged rocks surrounding the fort. Ships were seen as tiny dots in the horizon. Some parts of the wall opened into steep cliffs where you can see the heavy waves battering the strong wall.

In another side of the Fort a lighthouse loomed. You can get down to a small beach from there, where the beach is protected from a stretch of rocks midway in the sea. It allowed slowing down the waves, making the basin an ideal spot for bathing. The temptation to bathe was too much that we could not resist. It was two hours passed noon when we got into our swimsuits and jumped into the water. It was a great spot for bathing and we spent time playing and challenging the wave’s whole evening. As it became dark, the lights flashed from the lighthouse helping the boats and ships at sea. It was a remarkable experience to bathe in the night, looking at the sky filled with stars and looking at the flash of light emitting from the lighthouse souring across the sky. We saw the multicolored lights of the ships and boats at bay. After hours of bathing we left the wonderful beach and went to a friends’ place to spend the night.

The next morning we went to the Unawatuna beach which is famous for its beauty. A twenty minute bus ride from the Town and a five minute walk brought us to the beach. The beach was protected by a small bay in the shape of a half circle which was covered with soft sand. The beach was so beautiful but the waves were rough making it hard for us to bathe. Unawatuna is a tourist attraction famous for diving, surfing and snorkeling.

Our next destination was Roomassala, a rocky cliff which overlooked the other side of the Galle harbor. Few minutes back again in a bus towards Galle and after a thirty minute walk on a hilly area, we reached our destination. Roomassala gave a magnificent view over the coastal area. It was a fascinating place where the geography of the area was quite different from the surroundings from Galle. The area was much cooler and also the trees were different which we thought had much greenery.

Roomassala was famous for the ‘Jungle Beach” where you have to climb down to the beach from top of the cliff through a small jungle. It was a darling little beach deserted and unspoiled by humans, not known by many travelers. The beach had rough sand compared to Unawatuna. We had a stroll around the beach and we waited resting as it was an ideal location.

Then we again climbed up through the jungle and went to the ‘Japanese Peace Pagoda’, a marble white monument newly constructed on a cliff in Roomassala. The scenery from the top of it was magnificent where you can see miles into the blue ocean.

We left the place after making a quick decision to visit the Polhena beach which accounted be a great place for bathing. Polhena was very close to Matara and we had almost an hour’s ride on Bus. We passed the southern coastal line of the country as the bus raced along the smooth road passing the towering coconut trees alongside the coast. We came to a stop at Polhena.

After a fifteen minute walk we were in ‘paradise’. It was a large unspoiled beach, which had sand as white as marble and with blue water. We were so hurried to get in the water. We spend hours and hours in the water having the fun of a lifetime.

The sight of stilt fisherman few miles across the sea indicated the unique note and heartbeat of the southerners’ lifestyle. It was such nice scenery to be overlooked.

A chat with a local fisherman revealed that there was a dear little island few miles from the mainland. We decided to give it a visit and the fisherman took us in his boat. It was a jolly boat ride where we peaked from the sides of the boat to see the shallow bottom of the sea covered with corals.

We finally brought ourselves near a dear little island which had a mystic look with it. It had a monument in the middle which loomed over the stretch of jungle covering it. We were waiting for someone to raise the question. No sooner one of our friends asked “What is that?” from the boatman.

“It’s an ancient Pagoda made in the times of the kings, but we still don’t think that proper historical research has been carried out about this place.” He said.

We jumped down from the boat. Soon as our feet touched the ground and as our eyes hit the surrounding landscape, we knew that we were in heaven. It was a tiny island which was extremely beautiful. Sea shells were plenty on the beaches, birds soured the sky above the island. The beach around the island was rocky and we jumped from rock to rock and went around the island. There was also a strange looking monument made of stone which reminded us of the statues of Easter Island. The middle of the island was covered with a thicket which hided the track to the pagoda. Through the thicket we searched our way towards the pagoda. It was almost in ruins. We could imagine the grandeur and the ancient history of the place. We thought the top of it would be an ideal spot to see the surrounding of the Island. We climbed to the top and got a marvelous view of the island. It was ultimately the paradise, isolated from the rest of the world.

Although we wanted to stay back, alas we couldn’t. The sun slowly hid behind the horizon. The sky took its unique beautiful bluish color and its color was reflected in the water. We turned back thanking mother nature for such a lovely creation. We clambered in the boat. The sun took its final dive into the horizon sinking the world in darkness as the boat took us to the mainland. It was time to leave the paradise………..

Worshiping the Sun; the Beginning

By Kassapa Ellepola

It is called friendship, the bond made between special people. Filled with respect and creamed with understanding, true friendship leads to new experiences, adventures and amazements.

It was December, the month it all began. The cold breeze invites you to test courage and stamina. It should be climbing up the highest mountain or swimming down to the bottom of the sea. We opted to climb high, to Adams’ peak.

We had a group of eight, six boys and two girls. A readily made simple plan including destination, mode of travel and food was more than enough. Travelling light was our policy.

Train travelling in the Colombo-Badulla route is promising for its scenic beauty which managed to trigger us as well, to take the ‘train’ to Hatton.

We caught the train to ‘Badulla’ from ‘Fort’, one morning in December. We were quick to jump in and catch two rows of seats facing each other, comfortable enough to have a chat. We sat down with excitement. The train puffed, gave a loud awakening whistle indicating its departure and pulled forward.

The train pushed forward through awakening Colombo, passing towering buildings and busy people. When the train gained its speed, the buildings gradually became shorter and instead were replaced with smaller houses. We left all our despair behind with the concrete in Colombo and were amazed to find ourselves enjoying our hearts out, shouting, chatting and laughing. Maybe it was the greenery which did the trick! It was always great to travel with good hearted friends.

The train slowly climbed the hills in the upcountry. The hills offered us comfort with soft breeze. The Colombo-Badulla rail route shows its unique glamour. The train traveled through a number of pitch dark tunnels, some very long. The surrounding was filled with neatly maintained ‘Tea’ estates, which added renowned beauty to the upcountry. The sight and sound of waterfalls made a pleasant sound which climbs down from high mountain cliffs beside the railroad. Cameras kept clicking all the time to retain at least few beautiful memories of these extraordinary sceneries.

Hatton was a dear little railway station where the train stopped. We clambered down with our luggage. It was obvious that all needed a small rest after a long train ride. After a small rest you can climb into a bus bound to ‘Nallathanniya’ which takes you through a winding route overlooking the Norwood reservoir. The bus ride kept our adrenalin pumped all the way up to ‘Nallathanniya’ as our hopefully experienced driver took turns in the winding roads at sixty miles per hour. (But alas after two days back at home we watched news to see the same bus tumbled down a cliff)

‘Nallathanniya’ was a busy village with a number of small shops and restaurants. We gathered into one for lunch and psychically made a toast with the fish curry for a safe journey. There is always time to waste in some ornamental shops, which we did and had some ‘oil cake’ tasting, promising the venders to buy some from them, which was a lie made up only to fill our hungry tummies.

We washed ourselves up from a small stream to refresh ourselves before climbing.

Traditionally, a leader is appointed to guide the group and as one of us had been here a several times we appointed him as our ‘Crew leader’. We were bound to respect him and his decisions throughout the journey.

Climbing can be started around two in the evening. First we came across a ‘Kovil’ where we followed some rituals and each member was given a colorful band to be worn around the wrist which indicated that we were assured with the protection of ‘God Saman’, the landlord of the holy land of Adams’ Peak.

We climbed up step by step uphill. We did not feel tired as there was plenty of chattering and laughing. We came across the ‘Makara thorana’ (Dragons’ gate), a stone made gateway which indicated that you are entering a graceful and spiritual land belonging to God Saman and that you should behold. It was a grand monument and few minutes were spared to read the placard beneath it.

Again it was the climb. Small periods of rest were needed and we stopped to take a zip of ‘Belimal’ (An Ayurvedic beverage) halfway from a shop and again started climbing.

The air around you keeps becoming cold when you climb high. We heard the soft sound of a Stream which was of the famous ‘Seethe Gangula’, the ritualistic river where generations of pilgrims had had bathed. We got into our swimming costumes and had a great time in the surprisingly cold water. It was so much fun to bathe in such cold water and we felt very comforted and strengthened to climb the next steps.

Resting shelters are made to rest and you can stay in at them to have a tea and a tidbit.

You cannot avoid the dark when climbing this magnificent mountain. But the moonlight along with the lights here and there kept us in the right path. We witnessed many kinds of landmarks, manmade and natural along our journey. The natural sceneries at night were the best which made heaven on earth.

We did the required rituals along the route accordingly to make the journey a complete one. After a very steep climb in the cold night, we finally made it to the top. Feeling proud and overjoyed, we were enjoying our price. The cold which overwhelmed us did not manage to freeze our hearts boiling with happiness.

We were in the top of the world in a cold night filled with feelings of love and friendship. Isolated from the rest of the world, we were dreaming our own thoughts. The endless night of experience and happiness will remain in our hearts forever.

The Sunset in Adams’ Peak is special. It is blessed. It makes wishes come true for believers. The sun peaked out from the end of the world to a new day filling the world with its might and adding strength to the world and its people.

We climb down with strengthened hearts, new bonds and with blessings, for to-be continued journeys..........

‘Metagenomics’, the key to exploring Unculturable Microorganisms

By Kassapa Ellepola

Since the invention of the simple microscope and the first observation of microorganisms in a pond water sample by Antonie Van Leeuwenhoek, after more than three centuries of microbial studies, it would be surprising to know that only around 1% of microorganisms has still been isolated and identified. This fact mainly exists due to the presence of ‘Unculturable Microorganisms’ whose culturing techniques has still not been properly understood. Genomic analyses of these organisms are possible only after the isolation of pure cultures. Lack of knowledge in culturing techniques has been a setback for microbiologists throughout the 20^th century, until the discovery in the science of ‘Metagenomics’.

Simply ‘Metagenomics’ (Environmental genomics, Ecogenomics or Community genomics) can be referred to as the study of genetic material recovered directly from the environmental samples, instead of from cultures.

The modern aspects of metagenomics first appeared in publication in 1998 and the term was first used by scientists such as Jo Handelsman, Jon Clardy and Robert M. Goodman.

Genetic Material isolated from environmental samples are investigated, classified and is manipulated in metagenomics. This process can be simply summarized into four main steps (see Figure 1) which consists of

1. The isolation of genetic material from the environment
2. manipulation of the genetic material isolated,
3. library construction, and
4. finally the analysis of genetic material in the metagenomic library.

In the first step, a representative sample is collected from the environment. The sample contains many different types of microorganisms. Chemical methods such as alkaline conditions or physical methods such as sonication can be used to brake open the cells to free the DNA. The DNA should be separated from the rest of the sample. Using physical and chemical properties of DNA, isolation can be done using density centrifugation, affinity binding, and solubility/precipitation techniques.

The need for manipulation of the genomic DNA, isolated from the organisms is due to the large size of the DNA which makes it extremely hard to handle, especially when introducing into vectors and model organisms. This is achieved through cutting the DNA into smaller, linear fragments using enzymes called ‘Restriction Endonucleases’. (enzymes that cut DNA at a particular sequence of base pairs). Then these small fragments can be ligated into vectors which are small units of DNA used to insert the DNA fragments into the cells. The DNA is transcribed to produce the mRNA and then is translated to produce the proteins. The vectors are designed with a selectable marker which creates abnormal growth ability (such as resistance to a particular antibiotic) in model organisms. This helps the selection of organisms containing vectors and the ones which do not.

The third step is the transformation of the metagenomic DNA containing vector, into cells of a model organism, typically Escherichia coli. This allows the DNA from organisms that would not grow under laboratory conditions to be grown, expressed, and studied. The insertion of foreign DNA into cells and stable expression of proteins is called ‘Transformation’ and it can be achieved through chemical, electrical, or biological methods.

Differently sized metagenomic DNA fragments may exist which has the same restriction enzyme cutting sequence. Although, the vectors are designed in such a way that only one kind of DNA fragments are incorporated from the sample to each individual cell. Different Restriction Endonucleases can be selected to analyze different locations of the metagenome. The transformed cells are then grown on selective media. Cells carrying vectors can be differentiated with a growing colony consisting cloned cells that originate from one single cell. These samples of cell colonies which contain the metagenomic DNA samples on vectors are called a ‘metagenomic library’. These colonies can be used and preserved for future study of the fragments of DNA isolated from the environmental sample.

The DNA from the metagenomic library is analyzed in the fourth and final step of the procedure. The DNA is the material which expresses the characteristics of organisms. DNA determines the physical and chemical properties of organisms. Hence these properties can be employed for many potential methods of analysis. Phenotypic characteristics, such as an unusual color or shape in the model organism are an example of a character in metagenomic analysis. Although chemical properties are not readily observable, performing chemical assay on products created by the model organism leads the way to identification of chemical properties expressed from metagenomic DNA. A change in chemical properties may be due to the model organism incurring enzymatic function which it previously lacked.

The influence of metagenomics has opened doors for research in a variety of fields including agriculture, medical, environmental, earth science and energy.

Microbial communities which inhabit our bodies from in and outside, affect the human health. Investigating the human microbiome and its metagenome may lead to the discoveries in tools and guidelines to human and animal nutrition, drug discovery and preventative medicine. The metagenome may reveal characters such as digesting indigestible components of the diet and also helping in synthesizing essential vitamins and amino acids. Some of the microflora in the gut has the ability to detoxify potentially harmful chemicals contained in food. Some microbes have the ability to defend the body against pathogens. Metagenomics has the ability to derive a stunning array of biologically active chemicals and has helped researchers to learn more about common diseases like obesity.

Metagenomics has a part to play in Earth Science and global change as well. Bacteria have directly influenced the photosynthesis, chemical balance of the earths’ atmosphere and have played a crucial role in the habitability of the planet. Metagenomics has helped scientists to gain an insight of the genetic bases behind achieving complex chemical transformations of a microbial community in the atmosphere and the ocean. It also broadens the knowledge about the biogeochemical cycles and the change of energy and matter.

In the field of Agriculture, metagenomics offers an opportunity to explore how microbial communities together interact with crops and what their role is, in harnessing soil microbial community to produce healthier and more robust crops. Scientists believe that microbial communities have a crucial role to play in suppressing diseases caused by other microbes and therefore metagenomics will reveal the opportunity to study on microbial community characters on crop enhancement.

Communities of microorganisms take part more effectively than single organisms in bioremediation of natural and Xenobiotic compounds. This can be applied to remediation of environmental contaminations such as ground water contamination from gasoline and oil spills in the sea. Metagenomic analysis will help identify the particular community members and their functions which help achieve the full chemical transformations.

A Microbial community convert common agricultural wastes as corn fiber, corn stalks, wheat straw, and other biomass into potential bioenergy sources such as ethanol, hydrogen, methane, butanol, and even electric current. Scientists are trying to manipulate the amount of nutrient availability and environmental conditions to capture and store the useful by-products of such microbes. Metagenomics will open more opportunities to know about how these microbial communities function which would allow scientists to control and channel the energy sources they produce.

Scientific community remains watchful about this new science, which has opened doors to a tremendous amount of scientific exploration, yet to be discovered.

Reference:

· http://dels-old.nas.edu/metagenomics/

· http://www.scq.ubc.ca/metagenomics-the-science-of-biological-diversity/