LAB 9 – NON-FERMENTERS AND CAMPYLOBACTER

Question 1

What is the principle of the OF test? What are the components of the media?

Answer 1

• determines if a sugar is attacked by oxidation or fermentation
• one tube is closed (+ mineral oil) for anaerobic conditions while other tube is loosely capped for aerobic
• contains low peptone, 0.3% agar, phosphate, NaCl, and bromothymol blue indicator

Question 2

What does the zinc dust do in the nitrate reduction test?

Answer 2

• zinc is added to “negative” test
  1. It reduces nitrate if still present = red but (-) for nitrate reduction
  2. Identifies denitrification (nitrates reduced beyond nitrites) = no color change; (+) for nitrate reduction

Question 3

Principle of nitrate reduction?

Answer 3

• determines capability of extracting oxygen from nitrates to form nitrites + other reduction products
• addition of sulfanilic acid (reagent A) and dimethyl-alpha-napthylamine (reagent B) to nitrate broth forms p-sulfobenzene-azo-alpha-napthylamine (red diazonium dye)

Question 4

List steps of Nitrate Reduction test and possible results

Answer 4

1. Inoculate heavily into nitrate broth
2. If growth = add drops of reagent A (sulfanilic acid) and reagent B (dimethyl-alpha-napthylamine)
3. If no color change = add sprinkle of zinc

Results:
Red = positive nitrate reduction
Red after zinc = negative reaction
No color change after zinc = denitrification

Question 5

How does Pseudomonas aeruginosa compare to the enteric organisms?
a. Think about gram, oxidase, colony morph, nitrate, TSI, and appearance on
MAC

Answer 5

a. Gram = g-b
b. Oxidase = (+)
c. Colony morph = pyo; lg wht, “fried egg”
d. Nitrate reduction = (+)
e. TSI = K/K; (“aeruginosa” = oxidative)
f. MAC = wh

Question 6

How would you ID C. jejuni ?

Answer 6

1. Colony morph = flt spr
2. Gram = g-b
3. Catalase = (+)
4. Oxidase = (-)
5. Motility = (+)

Question 7

Campylobacter jejuni growth requirements ?

Answer 7

1. 5-10% O2 and 3-10% CO2 (microaerophilic)
2. 42°C
3. 48 hours

Question 8

What pigments does Pseudomonas aeruginosa produce?

Answer 8

1. Pyocyanin**(blue-green)
2. Pyoverdin (yellow-green)
3. Pyorubin (red)
4. Promelanin (brown)

**P. aeruginosa is the only sp. that can produce pyocyanin

Question 9

Give me an example of an organism that is a fermenter, a non-saccharolytic, and an
oxidizer? What are the expected colors of the OF tubes?

Answer 9

Escherichia coli
- fermenter
- non-saccharolytic
- oxidizer
OF Results:
- yellow (+) in both open and closed tubes

Question 10

What are reagents A and B for the nitrate reduction test?

Answer 10

reagent A = sulfanilic acid
reagent B = dimethyl-alpha-napthylamine