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Bacterial identification plating streaking how to inoculate how to observe

Bacterial identification plating streaking how to inoculate how to observe. Using sterile techniques. Media used for bacteria growth  welcoming for many bacteria We only want specific ones to grow ** Sterile technique s**

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Bacterial identification plating streaking how to inoculate how to observe

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  1. Bacterial identificationplatingstreaking how to inoculatehow to observe

  2. Using sterile techniques • Media used for bacteria growth  welcoming for many bacteria • We only want specific ones to grow ** Sterile technique s** • Sterile remain sterile as long as doesn’t touch anything that isn’t sterile • Also avoid prolonged exposure to air

  3. Sterile techniques: • Wash your hands • Keep your bench clean • Wear gloves • Flame loop, neck of tube • Keep cap facing down • Work quickly albeit efficiently • Limit talking when opening cultures http://www.parentsguidecordblood.com/vita34cleanroom50.jpg

  4. Autoclaving before after • Apparatus used to sterilize liquid and instrument • Heating up to 121oC at 15 psi for 15 minutes • Kill most microbe • Autoclave tape  chemical reaction  black stripes if autoclaving ok http://tea.armadaproject.org/Images/stoyles/stoyles_before_autoclaveJPG.JPG.jpg http://www.sterilizers.com/Images/Tecno-gaz-autoclave.jpg http://www.fibermark.com/images/prod_imgs/ds_m_01_Autoclave2.jpg

  5. Inoculation of bacteria in to Culture media Plate Broth Slant Deep http://student.ccbcmd.edu/courses/bio141/lecguide/unit2/control/images/broth.jpg http://www.mushmush.nl/images/methods/working_with_agar/slant.jpg http://82.43.123.182/globalplantclinic/images/Bacteria_plate.jpg http://student.ccbcmd.edu/courses/bio141/labmanua/lab7/images/negmotility.JPG

  6. Broth High concentration of bacteria Slant Space saving solid culture Plate Individual colonies Can be used to count bacteria Deep Look at motility & oxygen requirement Uses

  7. Pure vs mixed culture • Pure: originate from 1 bacteria strain • All colonies look the same • Mixed: originate from many bacteria strains • Colonies have different size/shape http://smccd.net/accounts/case/biol240/streakplate.html

  8. Bacteria colonies http://textbookofbacteriology.net/growth.html

  9. Composition of media • NA = Nutrient Agar • peptone, beef extract, salt, agar 2% • Many other medias available. These 2 will be used very often in this lab • Note: Peptone: enzymatic digest protein

  10. Few notes on agar • Not degraded by most bacteria • Is liquified at 100oC and remain liquid until about 40oC • If added to growth medium  medium becomes solid • Semi solid media: 0.5% agar • Broth: no agar • Solid media: 2% agar

  11. How to prepare a Petri plate • Take liquid agar (in the water bath) • Pour aseptically into the base of the Petri plate (top is larger than the base) • Wait until solidify (15 minutes)  invert • ***Plates are kept inverted so condensation does not drip onto the agar

  12. Pour plate method Pouring a plate http://www.biotopics.co.uk/microbes/pourp2.gif

  13. Objective 3:a How to inoculate a plate • Plate: provide large surface for isolation and observation of colonies • Using a sterile loop or a sterile swab streak your sample on the petri plate • Important let your sterilized loop cool before you pick up your sample

  14. Observation of your plate • You will see individual colonies (hopefully!) • Describe using the following criteria: • Colony shape • Elevation • Color • Texture

  15. http://faculty.mc3.edu/jearl/ML/ml-9.htm http://www.bact.wisc.edu/themicrobialworld/Prop.acnes_colonies.jpg

  16. Colonies morphology • Shape: round, irregular, punctiform (tiny dot) • Elevation: convex, umbonate, flat, raised • Color: translucent, shiny, dull, white • Texture: moist, mucoid, dry (or rough)

  17. Colonial morphology Margin- edge http://www.rlc.dcccd.edu/mathsci/reynolds/micro/lab_manual/Coloniy_morph.jpg

  18. How to open a tube • Hold the loop like a pencil • Curl the little finger of the same hand around the cap of the tube • Turn the tube with the other hand • Remove the cap (keep in your hand) • Flame the opening of the tube • Remove samples with loop • Flame the opening of the tube & replace the cap

  19. Objective 3:b How to inoculate a deep • Semi-solid media (0.5% agar) • Oxygen gradient in the tube • Can be used to look at bacteria motility • Sterilize the needle (until red hot)  wait a few seconds  pick your sample stab the needle in the middle of the deep and remove it through the same stab • Do not use a loop to inoculate the deep*

  20. Bacteria motility • Non motile bacteria will only be found at the site of inoculation • Motile bacteria  swim around go everywhere http://www.bact.wisc.edu/bact100/Motility.jpg

  21. Objective 3:c How to inoculate a slant • Provide a solid growth surface in a tube format (take less space) • Inoculate as you did for the petri plate • One streak in the middle of the surface do not dig/ nor stab. on the surface. http://www.liddil.com/beer/culture/slant.gif

  22. Slant observation http://www.rlc.dcccd.edu/MATHSCI/reynolds/MICRO/lab_manual/slant_patterns.jpg

  23. Broth observation http://www.rlc.dcccd.edu/MATHSCI/reynolds/MICRO/lab_manual/broth_patterns.jpg

  24. Broth High concentration of bacteria Slant Space saving solid culture Plate Individual colonies Can be used to count bacteria Deep Look at motility & oxygen requirement Uses

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