Microbiology Lab

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Microbiology Lab
2011-04-21 23:29:09

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  1. Selective Mediums
    • Either inhibits the growth of a certain group of microbes or supports the growth of very few microbes
    • Antibiotics
    • Salts
    • Toxic Dyes
    • Limit carbon/energy source
    • Either Gram + OR Gram -
  2. Differential Mediums
    • Distinguish closely related organisms from one another (usually biochemical abilities)
    • Typically a visual difference (pH indicator)
  3. Staphylococcus 110
    • A type of selective medium
    • Contains NaCL and mannitol
    • Lacks phenol red
    • Favors colony pigmentation by different strains of S. aureus
    • No color change as mannitol is fermented
  4. Blood Agar Plate
    A type of differential medium
  5. Alpha Hemolysis
    • A partial breakdown of hemoglobin to methemoglobin
    • Red to green
  6. Beta Hemolysis
    Capable of completely hydrolyzing hemoglobin
  7. Gamma Hemolysis
    Possess no hemolytic actions whatsoever
  8. MacConkey Agar (MAC)
    • Indicator: Neutral red, pH
    • Inhibitor: Crystal violet and bile salts are toxic to gram +
    • Results:
    • -Gram += inhibited growth
    • -Gram - fermenting lactose= pink colonies
    • -Gram - not fermenting lactose= colorless colonies
  9. Eosin Methylene Blue Agar (EMB)
    • Indicator: Methylene blue and eosin Y
    • Inhibitor: Methylene blue is toxic to gram +
    • Results:
    • -Gram += inhibited growth
    • -Gram - fermenting lactose= pink or purple colonies with a metallic green sheen
    • -Gram - not fermenting lactose= colorless colonies
  10. Salmonella Shigella Agar (SS)
    • Indicator: Neutral red for lactose fermentation, FeS for H2S production
    • Inhibitor: Sodium citrate and brilliant green dye are toxic to most bacteria except these
    • Results:
    • -Gram += inhibited growth
    • -Gram - fermenting lactose= pink colonies; if H2S is produced, the center of the colony will be black
    • -Gram - not fermenting lactose= colorless colonies
  11. Mannitol Salt Agar (MSA)
    • Indicator: Phenol red, pH
    • Inhibitor: 7.5% NaCl
    • Results:
    • -Gram + staphylococci fermenting mannitol= yellow media, neutral pH
    • -Gram + staphylococci not fermenting mannitol= red media, basic pH
    • -Gram + streptococci= inhibited growth
    • -Gram -= inhibited growth
    • NaCl inhibits other organisms than staphylococci
  12. Use a drop of isopropanol on lens paper
    How to Remove Immersion Oil
  13. Oil Immersion
    • Ensures most light will travel straight into the objective lens
    • Prevents light from being lost to the surrounding air
  14. Simple Staining
    • Two types of dye- acidic and basic
    • Used to see the morphology and shape of bacteria
  15. Positive Simple Stain
    Colors the cell, leaving the background colorless
  16. Negative Simple Stains
    Cell remains colorless, the background retains the stain
  17. Gram Negative
    • Any organism that takes up the red safranin in gram staining
    • Red
    • Have a higher lipid content in the cell wall
    • Lipids are dissolved by the alcohol causing the crystal violet to leak from the cell
  18. Gram Positive
    • Any organism that retains the primary dye in gram staining
    • Purple
    • Have a thick layer of peptidoglycan
    • Pores in the cell wall are closed and prevent the loss of the crystal violet
  19. Differential Staining
    A method to distinguish between different types of bacteria
  20. Bacillus, Clostridium and Sporosarcina
    Common species that form endospores
  21. Defined Media
    • Composed of designated amounts of specific compounds
    • Exact contents are known
    • Amounts are known
  22. Complex Media
    • Contain nutrient-rich substances and precise chemical constituents are unknown
    • Amounts of chemicals are not known
    • Yeast extract, peptone, and tryptone are some substances are used
  23. Different nutrient needs
    Different metabolic needs
    Why can't all bacteria be cultured?
  24. Streak Plate
    • Useful in isolating colonies
    • Can be made from broth and/or plate cultures
  25. Spread Plate
    • Useful in quantifying bacteria
    • Also isolate colonies (if proper dilution is used)
    • Must be done with a liquid culture
  26. Zone of Inhibition
    • A function of molecular structure weight, charge, solubility, and diffusion within the medium
    • Indicates some chemical activity occurred in vitro
  27. Spoilage
    When microbes grow within or on the surface of foods
  28. Pathogenic Spoilage
    A potential major health hazard
  29. Cured
    Types of fermenting
  30. Microbial growth is slowed
    Metabolism is delayed due to the slower responses of bacterial enzymes
    Why store food at low temperatures
  31. Thermophiles and Hyperthermophiles
    • Live in very hot conditions (above 45C)
    • Compost heaps, hot springs
  32. Mesophiles
    • Live in conditions like body temperature (~37C)
    • Most human pathogens
  33. Psychrophiles
    • Live in cooler climates
    • Refrigerators (4C)
    • Causes spoilage of refrigerated foods
  34. Adding salt, sugar, and/or some spice
    Can lead to bacterial cell destruction
    What changes the osmotic conditions of food environments?
  35. Halophiles
    Live in high salt conditions
  36. Saccrophiles
    Live in high sugar concentrations
  37. Neutrophiles
    Live in neutral pH environments
  38. Acidophiles
    Live in more acidic pH's
  39. Alkalophiles
    • Live in higher pH environments
    • Degrade proteins for their energy
    • Produce amines and ammonia
  40. Refrigeration
    pH changes
    Addition of salt, sugar, vinegar, or some spices
    What can prevent food spoilage?
  41. They are born
    Bacteria begin the colonization process
    Babies are sterile until...
  42. Commensal Organisms
    • Live with humans in a harmonious relationship
    • Normal flora
  43. Saprophytes
    Live off the nutrients of dead cells and tissues
  44. Metabolic end products which are absorbed by their human host
    Some bacteria produce...
  45. Environment
    Body chemistry
    Body secretions
    Physical activities
    Antibacterial therapy
    Specific growth environments
    What changes the structure of the normal flora community in a host?
  46. Staphylococcus
    • Normal flora for the skins
    • Deadly pathogen if introduced in the bloodstream
  47. Skin
    • Normal flora:
    • Staphylococcus
    • Corynebacterium
    • Proprionibacterium
    • Streptococcus
  48. Throat
    • Normal flora:
    • Streptococcus
    • Corynebacterium
    • Neisseria
    • Branhamella
  49. Large Intestine
    • Normal flora:
    • Escherichia
    • Bacteriodes
    • Lactobacillus
    • Candida
    • Clostridium
    • Enterococcus
  50. Urethral/Anal Openings
    • Normal flora:
    • Escherichia
    • Lactobacillus
    • Enterococcus
    • Bacteriodes
    • Streptococcus
  51. Vaginal Openings
    • Normal flora:
    • Lactobacillus
    • Candida
    • Stretococcus
  52. Skin
    • Dry environment
    • Acidic pH
    • Inhospitable for many microbes
    • Microflora are gram-positive
    • Anaerobic bacteria living in pores
  53. Propionibacterium acnes
    • Metabolizes sebum
    • Releases fatty acids
    • Enhance anti-microbial environment provided by the skin
  54. Mannitol Salt Agar (MSA)
    Staphylococcus Medium 110 (SM110)
    Two medias used for primary isolation on skin
  55. Staph and strep
    Gram stains help differentiate between what two bacterias?
  56. Catalase Test
    • Checks for the presence of the enzyme catalyse
    • Splits hydrogen peroxide into hydrogen and water
    • Bubbles= positive
    • Lack of bubbles= negative
  57. Catalase Positive
    • Gram positive cocci
    • Either micrococci or staphylococcus
  58. Catalase Negative
    • Gram positive cocci
    • Streptococcus
  59. Beta-Hemolytic Streptococci
    • Belong to Lancefield groups A, B, and D
    • Most important pathogens
  60. Hemolysins
    • Uses blood agar
    • BAP distinguishes between 3 different types of hemolysis
  61. Alpha Hemolysis
    • Greenish zone around the colony
    • Due to partial breakdown of red blood cells
    • Most bacillus species
  62. Beta Hemolysis
    • Clear around the colonies
    • Complete lysis of the red blood cells
    • Strep pyogenes, Staph aureus, and all gram negative enteric bacteria
  63. Gamma Hemolysis
    • No hemolysis
    • Staph epidermis and Strep faecalis
  64. Blood degeneration
    Over incubation in the hemolysins' test causes
  65. Coagulase Test
    • Confirms Staphylococcus aureus
    • 97% of strains are positive
    • Others are negative
    • Test only valid for gram-positive
    • Some gram-negative rods cause false-positive reations
  66. Positive Coagulation
    • A loose clot
    • Solid immovable clot
  67. Negative Coagulation
    Slanted agar
  68. Hydrolases
    Split complex organic compounds into smaller units in the presence of water
  69. Amylase and Gelatinase Hydrolysis Test
    Identify bacteria
  70. Amylase
    • Bacteria that hydrolyze starch produce..
    • Clear
  71. Molecules of maltose, glucose, and dextrin
    The breakdown of starch produces...
  72. Dyes starch blue
    Indicates a lack of starch hydrolysis
    Iodine and Starch
  73. Gelatinase
    • Gelatin is a protein that can be broken down by the enzyme..
    • Indicated by the liquid in the tube
  74. No Starch Hydrolysis
    Medium has a clear zone
  75. Mannitol Salt Agar (MSA)
    Staphylococcus Medium 110 (SM110)
    Two medias used for primary isolation in the nose
  76. Fastiduous
    Fussy eaters
  77. Blood Agar Plates (BAP)
    Enriched media loaded with vitamins and nutrients
    Allows detection of alpha and beta hemolytic streptococci
    What type of agar is used for the throat?
  78. CAMP Test
    Help in the identification of Streptococci that produce beta hemolysis
    What type of test is designed for the throat?
  79. Optochin Susceptibility
    Differentiates alpha-hemolytic viridans group streptococci from the pneumococci
  80. Camp Factor
    An enlarged arrowhead-shaped hemolytic zone at the juncture where the unknown meets the S. aureus means it is positive
  81. Cultivating fecal samples of selective and differential media which contain lactose
    Enteric bacteria may be isolated by...
  82. MacConkey agar
    Levine eosin methylene blue (EMB)
    Two types of agar used for normal flora of the intestinal tract
  83. MacConkey Agar
    Contains bile salts to inhibit non-enteric bacteria and two dyes, neutral red and crystal violet, which are taken up by lactose-fermenting bacteria
  84. EMB Agar
    Inhibitory to gram-positive bacteria
  85. Lactose-Fermenting Bacteria on MAC
    • Appears red
    • E. coli is brick red
    • Enterobacter, Klebsiella and other lactose fermenters are pink to red with a mucoid texture
  86. Gram Stains in Intestinal Tract
    Isolates to confirm the presence of gram-negative rods
  87. Results of EMB of Intestinal Tract
    • E. coli- blue-black with a green metallic sheen in reflected light
    • Enterobacter aerogenes- convex and mucoid with dark centers and creamy pink borders
  88. Triple Sugar Iron (TSI) Agar
    • Valuable medium for the further differentiation of enteric bacteria
    • Contains three carbohydrates: glucose, lactose, and sucrose
    • pH indicator phenol red
    • Anaerobic conditions
    • Contains iron ions and sodium thiosulfate
  89. Gas has been produced
    Large amounts of gas may push the medium up the tube
    Cracks in the agar of TSI means..
  90. There is H2S reacting with iron making iron sulfide
    If the TSI medium is black, that means...
  91. TSI medium
    Bacteria isolated on MAC and EMB plates will be differentiated by their reactions in...
  92. Indole production
    Methyl red test
    Voges-Proskauer test
    Citrate test
    IMViC Series
  93. Indole Test
    • Amino acids are produced as a result of the hydrolytic degradation of proteins, peptones, and peptides
    • Tryptophan
  94. Methyl Red Test
    • Depends on an organism's ability to ferment glucose to acid
    • Red in acid solutions
    • Yellow in alkaline or neutral solutions
  95. Voges-Proskauer Test
    • Depends on the digestion of glucose to acetylmethylcarbinol
    • If present it will form a red color
  96. Citrate Test
    • Based on the ability of certain bacteria to utilize a salt of a citric acid as a sole carbon source in growth
    • Indicator bromthymol blue becomes deep blue when pH of medium rises
    • If utilization does not occur, then appears green
  97. Indole Production Results
    • Positive- presence of a red surface layer
    • Negative- no production
  98. Methyl Red Results
    • Positive- acid is produced- red
    • Negative- yellow
  99. Voges-Proskauer Results
    • Positive- red
    • Negative- lack of color change
  100. Citrate Test Results
    • Positive- ability to utilize citrate- blue
    • Negative- green
  101. E. coil, followed by Pseudomonas aeruginosa, and Staphylococcus aureus and s. epidermis
    Common cause of UTIs
  102. Inflammation of the bladder (cystitis)
    Inflammation of one or both kidneys (pyelonephritis)
    Two major infections of the urinary tract
  103. Blood Agar Plates (BAP)
    Eosin-Methylene Blue (EMB)
    Two types of agar used for primary detection in urinary tract
  104. 30-300
    Fewer than means could not be statistically validated
    More than means there are colonies touching
    Range of countable colonies
  105. Dilution
    Transfer of a known volume of a sample into a known volume of diluent
  106. volume of sample added
    volume of sample added + volume of diluent
  107. Dilution 1 x dilution 2 x dilution 3...
    Total dilution=
  108. 1
    total dilution
    Dilution Factor=
  109. # of colonies x dilution factor on a countable plate
    = # of bacteria in the original sample
  110. Compare to chart
    If positive, circle number
    How do you score an enterotube test?
  111. Positive- indole and methyl red
    Negative- citrate and voges-proskauer
    What tests are E. coli positive and negative in IMViC?
  112. Citrate- bromethymol blue
    Methyl red- methyl red
    What is the indicator for citrate and methyl red test?
  113. Fermentation
    Gas production
    H2S production
    What are 3 general outcomes of TSI slant?
  114. Nosocomials
    Hospital acquired infections
  115. MAC- methylene blue
    EMB- neutral red
    What are the indicators in MAC and EMB agar?
  116. Positive
    Bacteria that grows on the outside of the body is...
  117. Negative
    Bacteria that grows on the inside of the body is...
  118. E. auregenus and E. coli
    What are the two bacteria used in IMViC?
  119. Diplo-
    Pair of cocci or bacilli
  120. Strepto-
    chain of three or more cocci or bacilli
  121. Staphylo-
    Cluster of cocci
  122. Tetrad
    Packet of four cocci in a square
  123. Sarcina
    Packet of eight cocci in a cube
  124. The inside
    What part of your body is sterile?
  125. Enterococci and E. coli
    What two types of bacteria are used in water?
  126. Protein and starch hydrolysis
    What are two types of hydrolysis reactions?