Grams staining and yoghurt making

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IB Biology Option F SL

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Grams staining and yoghurt making

  1. 1. Investigating prokaryotes<br />IB Biology<br />
  2. 2. Label this cell!<br />
  3. 3. Cell Wall<br />Determine shape of bacteria<br />Strength prevents osmotic rupture<br />20-40% of bacteria<br />Unique to bacteria<br />Some antibiotics effect directly<br />Penicillin<br />
  4. 4. Cell Wall <br />Peptidoglycan polymer <br /> -amino acids + sugar<br />Amino acids cross link sugars<br />
  5. 5.
  6. 6. Gram Staining<br />Technique named after Hans Christian Gram (Danish scientist)<br />Developed technique in 1884<br />Used to differentiate between bacteria that show same clinical symptoms<br />
  7. 7. The characteristic compound found in all true bacterial cell walls is peptidoglycan. <br />The amountof peptidoglycycan is among one of the differences between the Gram Positive and Gram Negative cell walls. <br />Gram-positive cell wallsGram-negative cell<br />Thick peptidoglycan<br />90% peptidoglycan<br />Teichoic acids<br />1 layer<br />Not many polysaccharides<br /><ul><li>Thin peptidoglycan
  8. 8. 5-10% peptidoglycan
  9. 9. No teichoic acids
  10. 10. 3 layers
  11. 11. Outer membrane has lipids, polysaccharides</li></li></ul><li>Figure 4.13b, c<br />
  12. 12. The process includes the use of:<br /> a primary stain (crystal violet) <br /> a mordant (helper) iodine solution, <br /> a decolorizer (95% ethanol), <br /> a counterstain (safranin). <br />
  13. 13. Patient No. 986214<br />Admitted at 01.03 this morning after complaining of severe stomach cramps and nausea<br />Case history revealed that the patient bought a tamale from a street vendor on his way home from a late shift at the Jumex factory yesterday night<br />After feeling okay for 24 hours after consumption of the tamale the cramping began <br />
  14. 14. Your task<br />The doctor wants to rule out bacterial food poisoning from Salmonella<br />Bacterial cultures from the patient´s blood and stomach contents have been prepared<br />Your job as the hospital microbiologist is to test the sample for presence of Salmonella<br />
  15. 15.
  16. 16. Salmonella toxins<br />Enterotoxin:<br />releasedbymicroorganism in thelowerintestine<br />alterspermeability of intestinal wall<br />waterleaksinto intestinal tractcausingdiarrhoea<br />Endotoxin:<br />Salmonella enters intestinal cellsthroughphagocytosis<br />endotoxinsdamagethecellsfromtheinside<br />cause fever and inflammation in largeamounts<br />
  17. 17. Diagnosis<br />Is the patient severely infected with Salmonella??<br />Or has he got something else??<br />You decide!!<br />Lactobacillus acidophilus (G +ve)<br />A good bacteria present in everybody’s digestive system<br />Salmonella (G -ve)<br />A bacteria that causes severe food poisoning<br />
  18. 18. GRAM STAIN PROCEDURE<br />1. Gather all equipment and supplies <br />Gram stain<br />Microscope slides<br />Heat source<br />Sterile swab<br />Culture or specimen<br />Microscope<br />
  19. 19. GRAM STAIN PROCEDURE<br />2. Sterilise your bench, wash your hands and light your bunsen burner<br />
  20. 20. GRAM STAIN PROCEDURE<br />3. Choose an isolated colony off of the agar plate and obtain bacteria with a sterile swab.<br />
  21. 21. GRAM STAIN PROCEDURE<br />4. Place the swab on the microscope slide and spread the colonies in a circular motion.<br />
  22. 22. GRAM STAIN PROCEDURE<br />5. Heat fix the microorganisms to the slide by passing the slide through the bunsen flame a couple of times<br />
  23. 23. GRAM STAIN PROCEDURE<br />6. Place slide on staining tray or hold with forceps above the sink.<br />
  24. 24. GRAM STAIN PROCEDURE<br />7. Flood the surface of the slide with Crystal Violet stain and let sit for one minute.<br />
  25. 25. GRAM STAIN PROCEDURE<br />8. Rinse the slide with distilled water.<br />
  26. 26. GRAM STAIN PROCEDURE<br />9. Flood the slide with Gram’s Iodine and time for one minute.<br />
  27. 27. GRAM STAIN PROCEDURE<br />10. Rinse the slide with distilled water.<br />
  28. 28. GRAM STAIN PROCEDURE<br />11. Flood the slide with Gram’s decolorizer and time for 30 seconds.<br />
  29. 29. GRAM STAIN PROCEDURE<br />12. Rinse the slide with distilled water.<br />
  30. 30. GRAM STAIN PROCEDURE<br />13. Flood the slide with the counterstain, Safranin, and let sit for one minute.<br />
  31. 31. GRAM STAIN PROCEDURE<br />14. Rinse the slide with distilled water.<br />
  32. 32. GRAM STAIN PROCEDURE<br />15. Blot the slide and read with the oil immersion lens of the microscope. Look for Gram-negative and Gram-positive bacteria.<br />
  33. 33. Diagnosis<br />Is Patient No. 986214 severely infected with Salmonella<br />Or has he just got something else!<br />You decide!!<br />Lactobacillus acidophilus (G +ve)<br />A good bacteria present in everybody’s digestive system<br />Salmonella (G -ve)<br />A bacteria that causes severe food poisoning<br />
  34. 34. Making Yoghurt<br />
  35. 35. Fermentation of milk<br />Lactobacillus<br />Lactose lactic acid<br />The milk becomes more and more acidic, lowering the pH and making it taste sour<br />The souring of milk is a fermentation process<br />which happens when the bacteria in the milk use the milk sugar (lactose).<br />It is their food source during anaerobic respiration.<br />
  36. 36. Product- yoghurt<br />Yoghurt-making uses 2 species of bacteria to ferment milk (often low-fat). They are added as a starter culture and produce lactic acid.<br />Lactobacillusacidophilus acidifies the milk<br />Streptococcus thermophilus turns the milk creamy<br />When the pH drops to 4.4 the proteins in milk coagulate (thicken) producing yoghurt.<br />
  37. 37. Yoghurt production<br />mixing<br />Milk products<br />Heat treatment at 90oC<br />1.<br />2.<br />3.<br />Removes oxygen<br />Kills harmful bacteria<br />Changes milk proteins<br />Cooled to 40oC<br />Yoghurt bacteria added<br />Best temperature for growing yoghurt bacteria<br />Ferment milk sugar (lactose) into lactic acid.<br />1.<br />2.<br />Give yoghurt its acid taste<br />Credit: museum<br />of brands<br />Fermentation for 6 hours<br />Storage tank at 4.5oC<br />Slows down further bacterial fermentation<br />
  38. 38. Make your own!<br />You will need:<br />Skimmed UHT milk (this milk is sterile!)<br />Live yoghurt (starter culture)<br />Dried milk powder(makes it creamy!<br />
  39. 39. Method<br />Take a clean pot and label with your name and the date<br />Fill 2/3 of the pot with UHT<br />Dissolve 1 teaspoon of milk powder into the milk<br />Add 1 teaspoon of yoghurt to the milk<br />Test the pH and make a note of it<br />Put the lid on and place in the water bath at 35 degrees C<br />

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