NUR'AIN BINTI ZULKEFLI - 111399

LAB 4 - SOURCES OF CONTAMINATION AND INFECTION

→ Introduction.

     Contamination can be defined as the presence of a minor and unwanted constituent in material, physical body , natural environment and many more. Meanwhile, infection is the invasion of body tissue by disease-causing microorganisms , their multiplication and the reaction of body tissue to the microorganims and the toxins they produce. Infection are caused by virus, prions, bacteria and viroids, though larger organisms like macroparasites  and funi also can be also infect. Airbone disease is caused by pathogenic microbial agent and transmitted through the air. These viruses and bacteria can be aerosolized through coughing, sneezing, laughing or through close personal contact. These pathogens ride on either dust particles or small respiratory droplets and can stay suspended in air and capable of travelling distance on air currents.


Resident microorganism are usually non-pathogenic, and either commensals (are not harmful to their host ) or mutualistic (offer a benefit.The benefits bacteria can offer include preventing transient pathogenic organisms from colonizing the skin surface , either by competing for nutrients, secreting chemicals against them, or stimulating the skin's immune system.        
Most research has been upon those resident microorganism upon the 2 square meter of human skin. Many of them are bacteria of which there are around 1000 species upon skin human. Most are found in the superficial layers of the epidermis and the upper parts of hair follicles. However, resident microorganism can cause skin diseases and enter the blood system creating life-threatening diseases particularly in immunosupressed people.
Transient microorganism are different from resident microorganism in that they do not take up permanent residence in the gastrointestinal tract. Instead, they established small colonies for brief periods of time before dying off or being flushed from the intestinal system via normal digestive process, or by peristaltic bowel action. However, in taking up temporary residence , they contribute to the overall function and condition of the digestive system. For example, the lives of some of the most important resident microorganism involved in human digestion and intestinal health depend on by product produced by the visiting transient.

→ Objective

  
 To determine the microorganisms in the air and from healthy humans.

→ Results

EAR

HANDS








AIR




NORMAL BREATHING







VIOLENT COUGH 

→ Discussions

In a healthy animal, the internal tissue , e.g. blood, brain, muscle, etc., are normally free of microorganisms. However, the surface tissues, i.e., skin and mucous membranes, are constantly in contact with environmental organisms and become readily colonized by various microbial species. The mixture of organisms regularly found at any anatomical sites is referred to as the normal flora, except by researchers in the field who prefer the term "indigenous microbiota". The normal flora of human consists of a few eukaryotic fungi and protists, but bacteria are the most numerous and obvious microbial components of the normal flora. The staphylococci  and corynebacteria occur at every site listed. Staphylococcus epidermidis is highly adapted to the diverse environments of its human host. S. aureus is a potential pathogen. It is a leading cause of bacteria disease in humans. It can be transmitted from the nasal membranes of an asymptomatic carrier to a susceptible host. Many of the normal flora are either pathogens or opportunistic pathogens. The asterisks indicate members of the normal flora that may be considered major pathogens of humans. Besides, Pseudomonas aeruginosa is the quintessential opportunistic pathogen of humans that can invade virtually any tissue. It is leading cause of hospital-acquired (nosocomial) Gram-negative infections, but its sources is often exorgenous (from outside the host).Colonies of Pseudomonas aeruginosa growing on an agar plate. The most virulent Pseudomonas species produce mucoid colonies and green pigments such as this isolate.

Violent coughing is the syndrome affecting canines. Characterized as inflammation of the upper respiratory system, it can be caused by viral infections, such as canine distemper, canine adenovirus , canine parainfluenza virus , canine respiratory coronavirus or Orthomyxoviridae Influenzavirus, or bacterial infections, such as Bordetella bronchiseptica.

Hand infections are common as the naked hand is the body's first point of physical contact with the environment.Staphylococcus aureus, a gram-positive cocci, is the causative organism in up to 60% of all hand infections. A wide variety of other micro-organisms may be involved, either individually or as a polymicrobial type of infection. This includes gram-negative organisms such as  Enterobacter, Streptococcus, Pseudomonas aeruginosa, anaerobes and fungal organisms.

Bacteria cause most ear infection. The most common type are Streptococcus pneumoniae (also called pneumococcus ), Haemophilus influenza, and Moraxella catarrhalis. Viral infection can also lead to ear infections. The respiratory syncytial virus (RSV) and flu (influenza) virus are the most frequent types found.

→Conclusion

Every human is colonized by billions of microorganisms. These microorganism, some are vital to our wellbeing, constitute our resident or normal microflora.

Many common infection can spread by airbone transmission, at least in some cases including anthrax,chickenpox,influenza,measles,pertussis, smallpox and tuberculosis.Airbone microbes, allergens and chemicals cause respiratory disease-inflammation in the nose, throat, upper airway to the lung. Many infection are acquired by inhalation of pathogens that may remain in the respiratory system but also invade the rest of the body through lymphatic and blood circulations.

→ References.

http://en.wikipedia.org/wiki/Infection

http://en.wikipedia.org/wiki/Contamination

http://en.wikipedia.org/wiki/Airborne_disease

http://en.wikipedia.org/wiki/Skin_flora

http://www.organicprobioticsandantioxidants.com/probiotics/transient-micro-organisms/

http://textbookofbacteriology.net/normalflora.html

http://en.wikipedia.org/wiki/Kennel_cough

http://www.sma.org.sg/smj/4704/4704pe1.pdf

LAB4 NURUL SAFFA BINTI ZAINAL ABIDIN 

SOURCES OF CONTAMINATION AND INFECTION

INTRODUCTION :

It is difficult to find a spot on the earth’s surface where bacteria do not around. They are most abundant in the air of crowded cities, but also are found in the air of mid-ocean. The air of high mountain tops is comparatively free from bacteria as also the air of the Artic regions. Even the sea water near the poles contains few microbes. Bacteria grow with great rapidity in the warm moist climate oh the tropics, but fortunately their development in hot countries is greatly hindered by disinfecting effect of the actinic rays of the tropical sun.

Bacteria of the skin :Bacteria which are capable of infecting the body cells and causing suppuration are constantly found upon the skin in countless numbers. These round streptococci infest especially the oil glands of the skin and the hair follicles. The so-called "skin worms" or comedones consist chiefly of colonies of bacteria.

The etire alimentary tract harbours multitudes of bacteria. The saliva swarms with bacteria which are ready to develop enormously if favourable conditions are supplied and may attack the teeth and gums. So long as the blood is maintained in a stage of high resistance the saliva will protect it by inhibiting or hindering the growth of bacteria. regardless of their origins,most of the transient are non pathogenic and are quickly killed by various defencess against them.

OBJECTIVES :

To determined the microorganism in the air and from healthy humans .

RESULTS :

ear

air

hands

normal breathing

violent cough

DISCUSSIONS :

Indeed bacteria, and micro-organisms of all sorts, can be found all over the planet.  Bacteria have been found to live in deserts, where most of the year they exist as dormant spores.  Algae have been isolated from snow banks in Antarctica.  Some bacteria can live in the acid stomachs of mammals, while others live in the alkali stomachs of insects.  Microbes even have been isolated from the boiling hot environment around deep ocean volcanoes.

Tiny forms of life have adapted to a whole spectrum of environments.  But it is important to keep in mind that not every microorganism can live in every environment.  There are different species of plants and animals.  The bacteria that live in the rocky tundra aren't the same as those that live in hot springs. Any place that offers the basic nutrients required for life will almost always have microbes. "Almost always" because changes in local environments may make that place sterile.

In the air samples, the researchers uncovered at least 1800 diffrent types of microbes, including those such as the diarrhea-causing Acrobacter and Ulcer-inducing Heliobacter general that can be dangerous to human health. Previous effort to deteremine microbe counts in the atmosphere had relied on culturing the iar to see what grew.

Normal breathing may contain bacteria named streptococcus pneumonia because S. Pneumoniais found normally in the upper respiratory throat and nasal passage. S.pneumonia can cause sinusitis,septic arthritis osteomyelitis,peritonitis and endocarditis. S.pneumonia is transmitted directly from a person through close contact via respiratory droplets. The organism frequently colonizes the nasopharynx of healthy people.

If we having coughing the bacteria that exist is Haemophilus influenza and Bordetella Pertusis bacteria . In infants and young children, H.influenza causes bacteremia,pneumonia and acute bacterial meningitis. On occasion it causes cellulitis,osteomyelitis,epiglottitis and infectious arthritis.for Bordetella Pertussis is spreads by air borne droplets which are transmitted  by sneezing or coughing. Complication of pertusis incude pneumonia and ear infection.

The bacteria normally found on your skin are what we call resident bacteria. they exist on the skin of normal healthy people and are ussually not harmful. they're always thereand can't be removed completely. other bacteria are transfered to your skin in one way or another. let's call these transient bacteria.The main pathogemic bacteria which are more likely to be found on the hands are Staphlococcus, Corynebacteria ,Streptococcus, E.coli, Myobacteria and Haemophilis,all in different concentrations. these bacteria are also found in other parts of the body like nose,eyes,mouth,gut and vagina,though the rate of incidence might vary.The first two types of bacteria are pathogens that are rensponsible for causing a large part of bacterial infectious in humans. these are found in almost all parts of the body, transmitted mainly from nasal membranes to other hosts. Staphylococcus cause boils, urinary tract infections and in serious casses food poisoning. Corynebacteria is commonly known to cause acne,and also associated with diptheria and more commonly in people using prosthetic diet.different Streptococcus strains caused varied infections affecting throat, lungs in case pneunomia, are rensponsible for causing dental carries, and can seriously damage the heart and kidney also.

CONCLUSIONS : 

Bacteria have exploding populations and can reproduce every 20 minutes. There are so many bacteria that you have on your body and at the air. Bacteria might be good or bad, depending on what they do to you or for you, and they are here to stay. Some bacteria spend their lives in the small folds of the skin on hair or under fingernails. Others cause body odor. Still called pathogens,cause disease.  

Microorganisms can enter the air when a human or animal sneezes, or by the wind picking up the light particles and blowing them to humans. When a human sneezes microorganism leave the lung at around 200 mile per hour. Some og the microorganism that are growing in the mucus in the respiratory tract enter the air with the moisture particles that are sneezed out of the lungs. These microorganism can be breathed into the lung of another person and that person coud get sick.

REFRENCES :

http://www.scientificamerican.com/article.cfm?id=microbe-census-reveals-ai

http://www.pediatriconcall.com/forpatients/commonchild/infectious_diseases/whooping_cough.asp

http://www.oldandsold.com/articles11/bacteria-4.shtml

http://www.iknow2.net/what-kinds-of-bacteria-are-on-your-hands/

LaB 4: NuRaIn BiNtI aRzMi~111400

~~~Lab 4: Sources Of Contamination And Infection~~~

~Introduction~

              Airborne microorganisms are usually carried on dust particles, although some (fungal spores, for example) may be carried directly by air current. It is important for microbiologists to be aware of the potential for contamination by airborne microorganisms. Carefully observation of simple precautions dramatically reduces the risk of contamination of the cultures.

            Every human is colonized by billions of microorganisms. These microorganisms, some of hitch are vital to be our wellbeing, constitute our resident or normal microflora. Resident microorganisms are nourished by the chemicals and moisture excreted by human body. In moist areas such as the armpits, there are may be one million bacteria per centimeter cube, on the drier skin of forearm, there may be 10000 bacteria per centimeter cube.

           Resident microorganisms are neither nonpathogenic or are prevented from infecting the body by an array of mechanical and chemical defenses. Some resident microbes are however opportunistic pathogens which may cause infection if the body’s defenses are breached for example if the skin is broken.

          Transient microorganisms are picked up from our environment, for examples from faecal contact or from soil and usually fail to become permanent skin residents. One of the most important reasons for failure to gain permanence is that the established residents are better able to compete for nutrients. Since transient generally originate in other environment, they are poorly adapted to conditions on the skin and usually disappear within 24 hours of arrival.

         Microorganisms in the upper respiratory tract are either normal residents or transients. As with the skin, the normal microflora largely consists of nonpathogens opportunistic pathogens.

       

 Large numbers of transients enter the upper respiratory tract as we breathe or eat. They may also come from our own hands or from improper sanitation during food preparation. Regardless of their origin, most transients are nonpathogenic and are quickly killed by various defenses arrayed against them.

~Objective~

To determine the microorganisms in the air and from healthy human

~Result~

µ     Air

µ     Hands

µ     Ears

µ     Normal breathing

µ     Violent coughing

~Discussions~

         Microbes are found everywhere from the tallest mountains to the deepest part of an ocean on earth, but they are mostly far too small to be seen by the naked eye. This activity allows students to discover that microbes are found in a range of different habitats, to explore the variety of microorganisms around us and to compare the range of microorganisms that are found in different places. There are about 2,000 species of bacteria identified and even more where that they come from including some pathogens, waffing in the air over.

 In ours earth, the air also have the bacteria. The bacterium is Bacillus cereus that is a gram-positive, spore-forming bacterium. It causes two types of food poisoning that are the diarrheal type by enterotoxin production in the small intestine and the emetic type by toxin which is formed in food. It spores are both highly resistant to a large number of stresses and very hydrophobic, which causes them to adhere easily to food processing equipment. In our ear, the bacterium is Staphylococcus aureus. It is gram-positive, coagulate-negative cocci that a part of our normal flora. Consequently, it is a true opportunistic pathogen, as it requires a major breach in the host’s innate defenses. The infections are associated with intravascular devices (prosthetic heart valves, shunts, etc.) but also commonly occur in prosthetic joints, catheters and large wounds.

There is also a bacterium that is totally not cleaned when we wash ours hands. The bacterium is Staphylococcus aureus. S. aureus is a facultative anaerobic Gram-positive coccal bacterium. It is frequently found as parts of the normal skin flora on the skin and nasal passage. S.aureus is the most common species of staphylococcus to cause Staph infections and also can cause a range of illness, from minor skin infections such as pimples, impetigo, boils(furuncles), cellulitis folliculitis, carbuncles, scalded skin syndrome and abscesses to life-threatening disease such as pneumonia, meningitis, osteomyelitis, endocarditis, toxic shock syndrome(TSS), bacteremia and sepsis.

        In normal breathing, the bacterium is Streptococcus pneumonia. That is a normal inhabitant of the human upper respiratory tract. The bacterium can cause pneumonia, usually the lobar type, paranal sinusitis and otitis media or meningitis which is usually secondary to one of the former infections. It also causes osteomyelitis, leptic arthritis, endocarditis, peritonitis, cellulitis, and brain obsesses. Currently the leading causes of invasive bacterium disease in children and elderly. While in violent coughing, the bacterium is called Haemophilus influenza. This bacterium is small (1 µm X 0.3 µm), pleomorphic, gram-negative coccobacillus. It is a nonmotile, non–spore-forming, fastidious, facultative anaerobe. Some strains of H influenzae possess a polysaccharide capsule. The other encapsulated strains H influenzae occasionally cause invasive disease similar to that of Hib. H influenzae type A (Hia) has been known to cause invasive disease (eg, meningitis) clinically indistinguishable from that caused by Hib. The nonencapsulated, or NTHi, strains cause mucosal infections, including otitismedia, conjunctivitis, sinusitis, bronchitis, and pneumonia. Less commonly, these strains cause invasive disease in children but account for half of the invasive infections in adults.

~Conclusions~

When we are walking into a room we can begin to stir up the microbial soup that fills it. Without our thinking, one person can send about 37 million bacteria into the air every hour, according to new research that measured microbes suspended in a room when it was occupied and unoccupied. We are exposed to the microorganisms. These microorganisms can be list as bacteria, viruses, mould and fungi and are present around us all of the time. These bacterium are also part of life's delicate balance and normally do not cause us harm. Airborne microbes cause a lot of illnesses and diseases in humans that we do not know.  

Microorganisms in the air can make themselves in human or animal body when a human or animal sneezes, or by the wind picking up the light particles and blowing them where humans are. When a human sneezes microorganisms leave the lungs at around 200 miles per hour. Some of the microorganisms that are growing in the mucus in the respiratory tract enter the air with the moisture particles that are sneezed out of the lungs. These microorganisms can be breathed into the lungs of another person and that person could get sick

~References~

1.  http://www.lbl.gov/Science-Articles/Archive/ESD-air-bacteria.html

2.  http://www.buzzle.com/articles/helpful-and-harmful-types-of-bacteria.html

3.  http://aem.asm.org/content/73/5/1481.full

4.  http://en.wikipedia.org/wiki/Staphylococcus_aureus

5.  http://web.uconn.edu/mcbstaff/graf/Student%20presentations/S%20epidermidis/sepidermidis.html

6.  http://textbookofbacteriology.net/S.pneumoniae.html

7.  http://emedicine.medscape.com/article/218271-overview

LAB 4: SOURCES OF CONTAMINATION AND INFECTION

LAB 4: YASMIN SYAFIKAH BT

         RAZALI  (111435)

INTRODUCTION

                Airborne microorganisms are usually carried on dust particles, although some (fungal spores, for example ) may be carried directly by air current. It is important for microbiologists to be aware of the potential for contamination by airborne microorganisms. Carefully observation of simple precautions dramatically reduces the risk of contamination of the cultures.

                 Every human is colonized by billions of microorganisms. These 

microorganisms, some of hitch are vital to be our wellbeing, constitute our resident or normal microflora. Resident microorganisms are nourished by the chemicals and moisture excreted by human body. In moist areas such as the armpits, there are may be one million bacteria per centimeter cube, on the drier skin of forearm, there may be 10000 bacteria per centimeter cube.

               Resident microorganisms are neither nonpathogenic or are prevented from infecting the body by an array of mechanical and chemical defenses. Some resident microbes are however opportunistic pathogens which may cause infection if the body’s defenses are breached for example if the skin is broken.

               Transient microorganisms are picked up from our environment, for example from faecal contact or from soil and usually fail to become permanent skin residents. One of the most important reasons for failure to gain permanence is that the established residents are better able to compete for nutrients. Since transient generally originate in other environment, they are poorly adapted to conditions on the skin and usually disappear within 24 hours of arrival.

                    Microorganisms in the upper respiratory tract are either normal residents or transients. As with the skin, the normal microflora largely consists of nonpathogens opportunistic pathogens.Large numbers of transients enter the upper respiratory tract as we breathe or eat. They may also come from our own hands or from improper sanitation during food preparation. Regardless of their origin, most transients are nonpathogenic and are quickly killed by various defenses arrayed against them.

OBJECTIVE

To determine the microorganisms in the air and from healthy human

RESULT

AIR

HANDS

EAR

NORMAL BREATHING

VIOLENT COUGHING

DISCUSSION

                  

                    Microbes are found everywhere, but they are mostly far too small to be seen by the naked eye. This activity allows students to discover that microbes are found in a range of different habitats, to explore the variety of microorganisms around us and to compare the range of microorganisms that are found in different places. The air we breathe is teeming with more than 1,800 kinds of bacteria, including harmless relatives of microbes. The bacteria that can be exist in the culture that is exposed to air is the Bacillus cereus. Bacillus  cereus causes two types of food poisoning in humans including diarrhoeal syndrome and emetic syndrome. Food poisoning results from its production of enterotoxins in the gastrointestinal tract. Bacillus species are aerobic, sporulating, rod-shaped bacteria which are ubiquitous in nature.

                    The wash water from our hand may contain Staphylococcus aureus. S. aureus can cause a range of illnesses, from minor skin infections, such as pimples,, scalded skin syndrome, and abscesses, to life-threatening diseases such as pneumonia. S. aureus reproduces asexually by binary fission. S. aureus can infect other tissues when barriers have been breached example skin or mucosal lining. This leads to furuncles and carbuncles. In infants, S. aureus infection can cause a severe disease such as staphylococcal scalded skin syndrome. In our ears maybe contain bacteria such as Staphylococcus epidermidis. S. epidermidis is not usually pathogenic, patients with compromised immune systems are often at risk for developing an infection. These infections can be both nosocomial or community acquired, but they pose a greater threat to hospital patients. This phenomenon may be the result of continuous use of antibiotics and disinfectants within hospitals, leading to evolutionary pressure toward more virulent strains of the organism. S. epidermidis causes biofilms to grow on plastic devices placed within the body. Infection can also occur in dialysis patients or anyone with an implanted plastic device that may have been contaminated. Another disease it causes is endocarditis. This occurs most often in patients with defective heart valves. In some other cases, sepsis can occur in hospital patients.

                       Normal breathing may contain bacteria named Streptococcus pneumonia because S. pneumonia is found normally in the upper respiratory tract, including the throat and nasal passages. S pneumoniae infection is also an important cause of sinusitis, septic arthritis, osteomyelitis, peritonitis, and endocarditis and an infrequent cause of other less-common diseases. S. pneumoniae is transmitted directly from person to person through close contact via respiratory droplets. The organism frequently colonizes the nasopharynx of healthy people, particularly young children, without causing illness. Transmission is thought to be common, but clinical illness occurs infrequently among casual contacts. If we having violent coughing the bacteria that exist is Haemophilus influenza. H. influenzae seems to occur in humans only. In infants and young children, H. influenzae type b (Hib) causes bacteremia, pneumonia, and acute bacterial meningitis. On occasion, it causes cellulitis, osteomyelitis, epiglottitis, and infectious arthritis.

CONCLUSION

                       Merely walking into a room can begin to stir up the microbial soup that fills it.  One person can send about 37 million bacteria into the air every hour, according to new research that measured microbes suspended in a room when it was occupied and unoccupied. We are constantly exposed to microorganisms. These microorganisms are classed as bacteria, viruses, mould and fungi and are present around us all of the time, are part of life's delicate balance and normally do not cause us harm. Airborne microbes cause a lot of illnesses and diseases in humans.  

                     Microorganisms can enter the air when a human or animal sneezes, or by the wind picking up the light particles and blowing them where humans are. When a human sneezes microorganisms leave the lungs at around 200 miles per hour. Some of the microorganisms that are growing in the mucus in the respiratory tract enter the air with the moisture particles that are sneezed out of the lungs. These microorganisms can be breathed into the lungs of another person and that person could get sick.

REFERENCES

      1.           http://www.livescience.com/19431-microbes-bacteria-indoor-air.html

      2.             http://www.nuffieldfoundation.org/practical-biology/microbes-all-around-us

      3.             http://microbewiki.kenyon.edu/index.php/Bacillus_cereus

LAB 3-NUR'AIN BINTI ZULKEFLI-111399

LAB 3: PREPARATION AND STERILIZATION OF CULTURE MEDIA

INTRODUCTION

          

Culture media are available as powders. They require only the addition of water. Nutrients medium is a general purpose preparation for culturing microorganisms which are not nutritionally fastidious.  The broth contains:

3.0 g/L “Lab-lemco” powder (a beef extract)

2.0 g/L yeast extract

5.0 g/L peptone (a nitrogen source)

5.0 g/L sodium chloride

15.0 g/L agar powder

The agar has the same composition, except that it contains 15g/L agar. The final pH of both media is 7.4

Culture media must be stored at the specific temperature, under specific conditions. All prepared culture media should be stored away from light and exposure to direct sunlight should be avoided at all the times.

An is an instrument used to sterilize equipment and supplies by subjecting them by high pressure saturated steam at 121 degree celcius for araond 15-20 minutes depending on the size of the load and the contents.

OBJECTIVE

To prepare   sterile  nutrient agar for culturing microorganisms.

DISCUSSIONS

A widely-used method for heat sterilization is the autoclave. Plant tissue culture media are generally sterilized by autoclaving. The time required for sterilization depends upon the volume of medium in the vessels. Additional sterilizing time is usually required for liquids and instruments packed in layers of cloths, as they may take longer to reach the required temperature. Following sterilization, liquids in a pressurized autoclave must be cooled slowly to avoid boiling over when the pressure is released. To ensure the autoclaving process was able to cause sterilization, most autoclaves have meters and charts that record or display pertinent information such as temperature and pressure as a function of time. Nutrient media (agar) that contain thermolabile components can be prepared in several steps. That is, a solution of the heat-stable components is sterilized in the usual way by autoclaving, then cooled to 35-50 degree celcius under sterile conditions: in a seperate operation, solutions of thermolabile components are filter-sterilized.

REFERENCES

http://www.cabri.org/guidelines/micro-organisms/M203Ap1.html

http://en.wikipedia.org/wiki/Autoclave

http://www.sigmaaldrich.com/life-science/molecular-biology/plant-biotechnology/tissue-culture-protocols/media-sterilization.html

http://en.wikipedia.org/wiki/Sterilization_(microbiology)

http://www.phytotechlab.com/pdf/SterilizingNutrientMedia.pdf

LaB 3 : NuRaIn BiNtI aRzMi~111400

LAB 3: PREPARATION AND STERILIZATION OF CULTURE MEDIA


Introduction


Culture media are available commercially as powders; they only require the addition of water. Nutrient medium is general purpose preparation for culturing microorganism not nutritionally fastidious. The broth contains:
3.0 g/L “Lab-lemco” powder (a beef extract)
2.0 g/L yeast extract
5.0 g/L peptone (a nitrogen source)
5.0 g/L sodium chloride
15.0 g/L agar powder
The agar has the same composition, except that it contains 15 g/L agar. The final pH of both media is 7.4.
Process that use moist heat and pressure so that all parts of the material to be sterilized reach 121’C for 15 minutes is called autoclaving. This process is in essence, a large pressure cooker; a chamber which may be sealed off against surrounding air. For sterilization, the materials are placed in the chamber, the door is sealed, and pressurized steam is forced into the chamber. The incoming steam displaces cooler air through an exhaust valve; this valve closes when the cell cooler air has been vented.
Then steam is continually forced into the chamber until the pressure reaches 103 kPa above atmospheric pressure, at sea level, this pushes the temperature in the chamber to 121’C. The high pressure prevents solutions from boiling over at this temperature. Large volume required more time.


Objective


To prepare sterile nutrient agar for culturing microorganisms


Discussions


      Plant tissue culture media are generally sterilized by autoclaving at 121 °C and 1.05 kg / cm2(15-20 psi). The time required for sterilization depends upon the volume of medium in the vessel. It is advisable to dispense medium in small aliquots whenever possible as many media components are broken down on prolonged exposure to heat. There is evidence that medium exposed to temperatures in excess of 121 °C may not properly gel or may result in poor cell growth.
      Several medium components are considered thermolabile and should not be autoclaved. Stock solutions of the heat labile components are prepared and filter sterilized through a 0.22 µm filter into a sterile container. The filtered solution is aseptically added to the culture medium, which has been autoclaved and allowed to cool to approximately 35-45 °C. The medium is then dispensed under sterile conditions. Experimentation with your system is recommended.
      Liquid media which are sterilized in their final containers should be cooled down to room temperature as rapidly as possible. Screw caps should then be tightened.
      Containers of agar media which have been sterilized should be placed in a 50°C water bath and the medium dispensed as soon as it reaches this temperature, or within a maximum of 3 hours in the bath. The medium should be mixed thoroughly, without bubble formation and aseptically dispensed into sterile containers. Do not expose dishes of agar media to sunlight; it causes excessive condensation on the lids and may cause the formation of inhibitory substances by photo-oxidation.


References


• http://www.sigmaaldrich.com/life-science/molecular-biology/plant-biotechnology/tissue-culture-protocols/media-sterilization.html
• http://www.cabri.org/guidelines/micro-organisms/M203Ap1.html