Nonfermenters and Campylobacter

Question 1

Motility test: wet preparation

Answer 1

Use TSB cultures

• sterile disposable pipette - add one drop onto slide
• cover with cover slip
• examine immediately under low illumination

organisms are pale and difficult to see

Question 2

Principle of O/F (carbohydrate)

Answer 2

OXIDATION or FERMENTATION of GLUCOSE

Aerobic process of creating acid from carbohydrates is OXIDATION
glucose –> gluconic acid (1 molecule of acid:1 molecule of glucose)

Anaerobic process is FERMENTATION; independent from oxygen
Phosporylation of glucose into two triose molecules, furthers into acids such as: lactic, formic, succinic, or acetic.
–> more acid produced, more rapidly

O-F test determines if a sugar is attacked by oxidation or fermentation

Question 3

Principle of NITRATE REDUCTION

Answer 3

Organisms that can reduce nitrate - extract oxygen from nitrates to form nitrites (and other reduction products)

NITRATE –> NITRITE
NO3- + 2e- + 2H+ ——-> NO2 + H20

Used to differentiate Enterbacteriaceae, Haemophilus, Neisseria, Pseudomonas

Question 4

Two ways motility can be tested

Answer 4

Wet preparation (microscopic)
Pattern of growth in motility media (diffusion through semi-solid media)

Question 5

Brownian Movement

Answer 5

Jerky, oscillatory movement in one spot (lacking direction)

–> not true motility

Question 6

True motility

Answer 6

Moving in a direction across the field

- darting, wriggling, tumbling

Question 7

Procedure of O/F (carbohydrate)

Answer 7

Inoculate tubes: stab wire almost to bottom
DO NOT SHAKE (introduces oxygen)
Add 10 drops mineral oil to one tube (close lid tight)
Leave lid loose on “open” tube
Incubate 35°C

Question 8

Interpretation of O/F (carbohydrate)

Answer 8

———————— OPEN CLOSED
Oxidation yellow green
Fermentation yellow green
Glu not utilized blue/green blue/green

yellow = positive (acidic)
blue/green = negative (alkaline)

Question 9

Components of O/F test

Answer 9

Medium:

• high carbohydrate content
• low peptone concentration
• 0.3% agar
• phosphate
• NaCl
• BROMOTHYMOL BLUE INDICATOR

one tube is sealed - anaerobic environment
one tube is open - aerobic environment

Question 10

Procedure of NITRATE REDUCTION

Answer 10

Nitrate broth (Durham tube) - inoculate heavily with organism