Biology 220 - Microbiology

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Biology 220 - Microbiology

• Kathleen Devlin
• kdevlin_at_csusb.edu

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General Lab Info

• Lab Midterm 100 pts
• Lab Final 100 pts
• Lab Assignments 32 pts
• This may include
• weekly in-lab assignments
• quizzes
• Lab Participation 18 pts
• 1 pt each week for signing in on time
• 1 pt each week for proper cleanup

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Lab Protocols

• Be on time
• Read your lab prior to lab
• Clean up after yourselves

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Biohazard Disposal

• Put tubes in tube rack
• Put plates, pipet tips, microtubes, etc in
  biohazard bucket
• Put dirty slides in Dirty Slides bucket
• Put broken glass in Broken Glass bucket

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Lab Safety

• No food or drinks
• Clean your lab bench with disinfectant before and
  after lab
• Report any broken glass or injuries immediately
• Be aware when using bunsen burners

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Exercise 1 - Scavenger Hunt

• Objective Find items commonly used in lab

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Exercise 2 The Unseen Organism

• Bacteria and some Fungi are too small to be seen,
  but are everywhere (ubiquitous)
• Objective Demonstrate that microorganisms exist
  in the air and on surfaces

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Types of Cultures/Media

• Broth liquid
• Slant solid, agar containing tube, dried at an
  angle
• Plate solid, agar containing petri dish
• We will use many types of media throughout the
  quarter, each having different nutrients/propertie
  s

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Aseptic Transfers

• Movement of bacteria from one media culture to
  another, without introducing contamination
• Tools
• Loop
• Needle
• Flame
• Sharpie (label everything!!!)

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Aseptic Transfer, cont.

• Label the new, sterile media (initials and
  bacteria being transferred)
• Label tube glass or bottom of plates
• Hold both tubes in hand not holding inoculating
  loop/needle
• Flame inoculating loop/needle until red hot
• Cool 15 seconds
• Remove both caps with little finger
• Pass tube opening through flame
• Take inoculum from tube and quickly place in/on
  new medium
• Flame tube openings
• Recap

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Exercise 3 - Culture Transfer Techniques

• Objective Transfer a bacterium from a culture to
  sterile media without introducing contamination
• Each group of 4 students will need
• 2 broth tube
• 2 slant tube
• 2 agar plate
• Transfer Micrococcus luteus broth to each type of
  media
• Transfer Micrococcus luteus slant to each type of
  media
• Place tubes in incubation rack and plates in tub
  (always incubate plates upside down!!!!)

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Microscopy
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Microscope Care

• Make sure lenses are clean (w/lens paper)
• Store with lowest power objective lens in
  position
• Stage should be all the way down
• Turn power off

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Tips for Using the Microscope

• Always start on the lowest power (4x)
• Use course focus to bring object into view
• Use fine focus to make the image sharp
• Move to the next power
• Use only the fine focus from here on out
• Before you get to the 100x lens, put a drop of
  oil on the slide
• Use the iris diaphram and light intensity to
  adjust light levels

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Exercise 4 ID of Cells

• Objectives Recognize visual differences between
  prokaryotic and eukaryotic cells Identify three
  basic morphologies of prokaryotic cells.
• Eukaryotic cells of animals, plants, fungi, and
  protists. Contain membrane bound organelles
• Prokaryotic cells of bacteria. No membrane
  bound organelles. 3 basic shapes
• Cocci sphere
• Bacilli rod
• Spirilla spiral

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Staphylococcus aureus

• Gram-positive, cluster forming (like grapes)
  cocci
• Normally found in nose and on skin
• Can cause pneumonia, nosocomial infections, food
  poisoning, TSS

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Bacillus cereus

• Gram-positive rod
• Causes gastrointestinal intoxication
• Found in a wide variety of foods

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Treponema pallidum

• Spirochaete bacterium
• Causes syphilis

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Yeast

• Eukaryotic
• Fungus
• Used in making beer, wine, and cheese
• Can form symbiotic relationship with bacteria
• Note These cells are round, but we do not use
  the term cocci for eukaryotic cells.

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Human Blood Smear

• Eukaryotic cell
• Note These cells are round, but we do not use
  the term cocci for eukaryotic cells.

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Note the size difference between prokaryotic and
eukaryotic cells
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Exercise 5 - Motility

• Objective Determine if a bacterium is motile
• Some bacteria have the ability to move toward or
  away from a stimulus
• TTC will turn red along the path traveled by the
  bacterium