Name: _________________________

Lab Module 2 Question Set

56. Fermenters in this test ___________________ the medium more that do oxidizers. (____________ the pH)

57. Why does this medium have a high sugar-to-peptone ratio?

58. The pH indicator in this medium is bromothymol blue, which is _____________ at pH 6.0 and ______________ at
pH 7.1.

59. The lower-than-usual concentration of agar in this medium allows the student to determine
____________________ of the organism as well as its oxidative or fermentative abilities.

60. The fermentable carbohydrate that we have included in this medium is _______________________.

61. After inoculation, one tube of O-F medium for each organism is sealed with a layer of mineral oil to promote
__________________________ and __________________________; the other tube for this organism does not
get a layer of mineral oil, in order to promote ___________________________ and _______________________.

62. If an organism grows in this medium, either with or without the mineral oil, and turns the medium blue, why did
this change occur? What did the organism use as a food source?* Is the appearance of a blue color in the
medium a positive or negative test for the oxidation or fermentation of carbohydrate and why?*

63. What do you call an organism (oxygen requirements) that can ferment and oxidize the carbohydrate in this
experiment (and which bacteria that we looked at fell in this category)? What do you call an organism that can
oxidize the carbohydrate but not ferment it (and which bacteria that we looked at fell in this category)?

64. Our OF medium contains glucose, but it could contain the carbohydrate __________________ or the
carbohydrate ________________________.

5-3: Phenol Red Broth (some of these questions also go with MR and VP)

65. A “lactose fermenter” is an organism that splits the ____________________ lactose into _________________ and
____________________, and then ferments the ___________________________.
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 66. The Methyl-Red (MR) test detects bacteria capable of performing a ______________________ fermentation.

67. The Voges-Proskauer (VP) test identifies bacteria able to produce ________________________ as part of a
___________________________ fermentation.

68. Phenol red broth contains a fermentable ________________, ________________, and the pH indicator
__________________________.

69. Phenol red is ____________________ below pH 6.8 and _____________________ above pH 7.4.

70. ____________________ of amino acids in the peptone in this broth produces ____________________, which
__________________ the pH and turns the broth _________________ or _______________ (colors); the color of
the broth may also remain the same. Do these results indicate a positive or negative test for the fermentation of
the carbohydrate and why?*

71. An inverted Durham tube (small test tube) is placed inside the culture tube to:

72. If the medium is examined after 18-24 hours of incubation, what does a yellow color indicate? Is this a positive or
negative test for fermentation of the carbohydrate and why?

73. To state that a particular organism is not capable of fermenting a particular carbohydrate (negative test), in
addition to a color change, the medium must be _________________. If this last criterion is not met, why is it
impossible to determine if that organism could ferment that carbohydrate?*

74. Why is it important to read these tubes after 24-48 hours after incubation?* What color would the tube be after
48 hours? Why?

5-4: Methyl Red & Voges-Proskauer Tests

75. Glucose is included in MR-VP medium as a __________________________; the purpose of the phosphate buffer
is:

76. The MR test is designed to detect organisms capable of _______________________ the buffer and
________________________________________.

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 77. Bacteria that are MR+ carry out what is called a ______________________________________. The acids
produced by these organisms tend to be ________________, whereas acids produced by other organisms tend to
be:

78. The pH indicator methyl red is _______________ at pH 4.4 and _______________ at pH 6.2 (note that this is
exactly the opposite of the color range of the pH indicator __________________). Between pH 4.4 & pH 6.2,
methyl red is various shades of _____________________.
79. ____________ color is considered the only true indication of a + result in the MR test; orange broth is considered
________________ or_________________. _______________________ is negative.

80. The VP test is designed to identify organisms that ferment glucose but quickly convert their acid products to
___________________ and ___________________________.

81. A positive VP test is ________________ (color); no color change is _______________________.

82. A copper color in the VP test is the result of interaction between the reagents and should not be confused with the
true __________________ color of a + result.

83. If an organism that cannot ferment glucose, grows in this both, what did it use for a food source?*

84. Why are organisms that are MR+ usually VP-, and VP+ organisms usually MR-?*

85. The MR and VP tests are components of a battery of tests known by the initials ____________________. Indicate
what each of these letters stands for:

86. All fermentation pathways start with the compound ____________________, produced by
_______________________. (process)

5-9: Citrate Test

87. The only carbon-containing compound in citrate medium is ______________________. Why do organisms need a
carbon source?*

88. The only nitrogen source in citrate medium is ________________________. Why do organisms need a nitrogen
source (to make which 2 macromolecules)?*

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 89. In a medium where citrate is the only available carbon source, bacteria that possess the enzyme
______________________________ can transport the citrate molecules into the cell and metabolize them.

90. The pH indicator in citrate medium, bromothymol blue, is _________________ at pH 6.9 and ________________ at
pH 7.6.

91. Bacteria that survive in this medium and utilize the citrate create an _____________________ (increase or
decrease?) in the pH of the medium.

92. Describe a positive result in this medium. Why does the pH indicator in the medium change color in a positive test?
Indicate the name of the product and the relative pH (acidic or basic) that caused the color change.

93. Describe a negative result in this medium. Why is there no growth in a negative result?*

94. If there is growth on citrate medium after incubation but no color change, this is considered a
__________________________ result.

95. Citrate utilization is one part of a test series referred to as IMViC series; these letters stand for the 4 tests:

96. The presence of ammonium hydroxide in the medium is INDIRECT evidence that the bacterium has broken down
_______________________.

5-13: Starch Hydrolysis

97. What is a polysaccharide? What type of macromolecule is a polysaccharide?

98. Why will starch not pass through the bacterial cell (plasma) membrane?

99. Bacterial enzymes that can hydrolyze starch are _______________________ enzymes, namely
___________________________ and _________________________.

100. The overall reaction for the hydrolysis of starch involves breaking the polysaccharide down to
________________________________________.

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101. Why is it not possible to see either starch or its sugar subunits in starch agar?

102. What is the purpose of adding gram’s iodine to starch agar plates after incubation?

103. Describe a positive result in this experiment. What does this result mean?

104. Describe a negative result in this experiment. What does this result mean?

105. After adding iodine, what would an uninoculated plate of starch agar look like? Why?

106. Starch hydrolysis is one of the tests that aids in the differentiation of species in the genus Clostridium, members of
which cause the diseases ____________________, and ______________________, and species in the genus
Bacillus, which causes ______________________.

107. What do the 2 bacterial genera mentioned in #106 have in common?

108. What conclusion can you draw from the fact that after iodine has been added, the clear area on starch agar
around the colonies of the organism that is + for starch hydrolysis is only found a short distance from the
colonies?* If the organisms is + for starch hydrolysis, why isn’t the whole plate clear after iodine has been added?*

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