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Isolation and identification of individual microbes and growth and monitoring of microbes.

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Introduction

This assignment is divided into two sections, the first deals with the isolation and identification of individual microbes. The second part looks at the growth and monitoring of microbes. Section 1 Identification of microbes: A case study Task 1 Practical: Aseptic technique Task 2 Write a step-by-step guide on how to perform an aseptic technique, include a reason and explanation of the precautions taken. Make reference to the health and safety legislation covering micro-organisms. All of these steps are to ensure that cross contamination does not occur and cause the results to be misleading. � Restrict micro-organisms present in specimens or cultures to the vessels in which they are contained. � Prevent environmental micro-organisms (normally present on hand, hair, clothing, laboratory benches or in the air) from entering specimens or cultures and interfering with the results of the studies 1. Make sure bench is clutter free. So that belongings do not get contaminated / or the sample doesn't get contaminated. 2. Hair should be tied back. To avoid contamination or accidents, such as hair catching fire from a Bunsen burner. 3. Lab coats should be worn. To avoid cross contamination 4. Bags and coats should be stowed away under the bench or in a cloakroom. To avoid accidents / cross contamination 5. Wipe bench down with alcohol. To sterilise working area before and after practical 6. Collect all equipment and set it up. So that you are ready to begin and can work in an organised manner with out the risk of contaminating a clean area. 7. Keep the lids of the petri dishes closed. So that they do not get contaminated with micro-organisms that are in the atmosphere 8. Wipe any slides that will be used with alcohol. And place them in on the bench that has been cleaned with the alcohol. So that the are sterile 9. Heat the inoculation loop in a Bunsen flame until it glows red and is sterilised. ...read more.

Middle

Such as acid-fast staining for mycobacteria and nocardia, and immunofluorescent stain for Legionella. The Gram stain procedure was originally developed by the Danish physician Hans Christian Gram to differentiate pneumococci from Klebsiella pneumonia. The procedure entails adding a solution of iodine to cells that have previously been stained with crystal violet or gentian violet. This procedure produces "purple coloured iodine-dye complexes" in the cytoplasm of the bacteria. The cells that were previously stained with crystal violet and iodine are next treated with a decolonising agent; 95% ethanol or a mixture of acetone and alcohol. The difference between Gram-positive and Gram-negative bacteria is in the permeability of the cell wall to these "purple coloured iodine-dye complexes" when treated with the decolourising solvent. Gram-positive bacteria keep the purple iodine-dye complexes after treatment with the decolourising agent; Gram-negative bacteria do not retain complexes when decolourised. To visualize decolourised Gram-negative bacteria, a red counter-stain such as safranin is used after decolourisation. The right preparation of the smear sample is essential. Make a thin film of the sample on a clean glass slide, using a sterile loop. Air dry and then heat fix the slide by passing it several times through a flame (the slide should not become too hot to touch). To be visible on a slide, organisms that stain by the Gram method must be present in concentrations of a minimum of 104 to 105 organisms/ml of unconcentrated staining fluid. At lower concentrations, the Gram stain of a clinical specimen seldom reveals organisms even if the culture is positive. Smears that are not properly fixed tend to be washed away during staining and washing resulting in the absence of stained bacteria. Source: www.meddean.luc.edu Section 2 Micro-organisms are extremely useful in many areas of our lives, they are used to produce enzymes, to make medicines and vaccines and in bread and beer production. To be used in manufacturing industries the microbes have to be grown on a large scale and in industrial fermenters or bioreactors. ...read more.

Conclusion

Organisms cultured from body surfaces or any environmental source must be examined in unopened containers, or killed before examination as described above. Sterilization and disposal. All cultures must be heated to kill micro-organisms before disposal. This is best done using a pressure cooker or autoclave, in conjunction with autoclavable bags. The caps of all screw-topped bottles must be loosened before cultures and media are sterilized. It is very important that instructions for use of the autoclave are followed in order to achieve and maintain sufficiently high temperatures for a long enough time. Pressure cookers are unlikely to be equipped with appropriate instructions for sterilization and those for some autoclaves, designed for use with surgical instruments, state that the equipment is unsuitable for sterilizing liquid media. Such autoclaves can be used for microbiological preparations but advice on their correct operation should be sought. Teachers and technicians should be trained to follow safe working practices. Seals and safety valves should be checked before each use. Heating autoclaves or pressure cookers with Bunsen burners is not recommended. Rapid cooling and the release of steam to lower the internal pressure quickly to atmospheric pressure is dangerous because it may shatter glassware and/or cause liquid media to boil over. Equipment should be allowed to cool unaided before opening. Further information may be sought from the advisory bodies such as MISAC. Sterilization cannot be achieved by the use of chemical disinfectants. If, in exceptional circumstances, chemical disinfection of cultures is contemplated prior to disposal, use a freshly-made solution of a disinfectant that is not degraded when in contact with organic matter (see 'Spills' above). Cultures and equipment must be opened under the surface of the solution and left for at least 12 hours. Again it is essential to follow disinfectant instructions carefully. Chlorate(I) solution is inactivated by large amounts of organic matter, although if a culture might contain viruses this is often the preferred disinfectant. ?? ?? ?? ?? Assignment 2 Melanie Harris MICROBIOLOGY BTEC National Diploma Health Studies 27/04/2007 Page 1 of 3 Unit 13 ...read more.

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