Vibrio media

Question 1

What does “TCBS agar” stand for?

Answer 1

thiosulfate-citrate-bile salts-sucrose agar

Question 2

What ingredients in TCBS agar inhibit gram positives?

Answer 2

ox gall, sodium cholate (bile salts)

Question 3

What ingredients in TCBS agar inhibit Enterobacteriaceae?

Answer 3

sodium thiosulfate and sodium citrate

Question 4

What is the pH of TCBS media?

Answer 4

8.6 (very alkaline)

Question 5

What is the purpose of thymol blue and bromothymol blue in TCBS agar?

Answer 5

pH indicators (change color when sucrose fermentation occurs)

Question 6

What color and colony morphology does Vc have on TCBS?

Answer 6

large, yellow colonies

Question 7

What color does Vp have on TCBS?

Answer 7

blue/green colonies (indicates suc-)

Question 8

What color and colony morphology does Va have on TCBS?

Answer 8

large, mucoidal, yellow colonies

Question 9

Vh and Vf should have what color and characteristic on TCBS agar?

Answer 9

blue/green colonies that fluoresce

Question 10

What types of organisms are inhibited on most selective and differntial Vibrio media? (2)

Answer 10

Gram pos

Enterobacteriaceae

Question 11

What are two examples of organisms that may grow on TCBS?

Answer 11

Proteus/enterococci- small yellow colonies
Pseudomonas/aeromonas- small blue colonies

Growth should be partially inhibited!

Question 12

What component of TCBSS agar is diffferent between brands and can cause problems with quality/reproducibility?

Answer 12

bile salts

Question 13

What is the most common culture media for Vibrios?

Answer 13

marine agar/broth

not selective-get lots of background organisms in environmental samples

Question 14

What are some problems with culture methods?

Answer 14

Almost none are specific for a single species and even fewer are selective for pathogenic organisms

Question 15

What species of Vibrio was CHROMagar first developed for?

Answer 15

Vp (Hara-Kudo et al. 2001)

Question 16

What is the recipe for CHROMagar

Answer 16

proprietary

has salts + a “chromogenic mix”

Question 17

What is the pH for CHROMagar

Answer 17

9.0 (very alkaline)

Question 18

What color is Vp on CHROMagar?

Answer 18

purple/violet

Question 19

What color is Vv on CHROMagar?

Answer 19

pale blue

Question 20

What color is Vc on CHROMagar?

Answer 20

pale blue

Question 21

What color is Va on CHROMagar?

Answer 21

white/clear

Question 22

What are some advertised benefits of using CHROMagar instead of TCBS?

Answer 22

fewer false positives, more Vibrio recovered

Question 23

What color do suc+ Vibrios appear on TCBS?

Answer 23

yellow

Question 24

What are two problems associated with growing Vv on TCBS?

Answer 24

Vv has very similar result to Vp– difficult to differentiate

TCBS media is thought to somewhat inhibitory for Vv

Question 25

What is one problem associated with differentiating Vv on CHROMagar?

Answer 25

Vv and Vc have similar result (blue colonies)

Can sometimes get around by adding more salt

Question 26

What media is recommended for the enrichment of Vibrio?

Answer 26

Alkaline peptone water

Question 27

What is the concentration of Peptone in APW?

Answer 27

1-2% (2% more appropriate for marine species)

Question 28

What is the pH of APW?

Answer 28

9.0

Question 29

How does the MPN method work?

Answer 29

10X dilution of samples (usually triplicates), innoculate in nutrient broth to look for growth, use a table to determine MPN based on your results

Must confirm species using colony hybridization or other molecular test

Question 30

What are the biochemical characteristics of Vp O3:K6 (pandemic strain associated with seafood-related illnesses in Asia and the US)

Answer 30

trh-
tdh+
urease-

Question 31

How does colony hybridization work?

Answer 31

Grow colonies on solid media

Blot colonies onto a membrane

Preform cell lysis on membrane

Incubate with probes

Visualize using UV or autoradiography