Related Material for Lab 4 Tuesday October 1 Group A, Thursday October 3 Group B
4A. Id of GNR:
Here is a copy of the lab wallchart
showing typical color reactions in the API 20 E assay and the
calculation of the biotype number; and the interpretation of the
reactions (which also appears on p 30 of your manual).
(1) Fermentation of the carbohydrates begins in the most anaerobic portion (bottom) of
the tube. Therefore, these reactions should be read from the bottom of the tube
to the top. (2) A yellow color at the bottom of the tube only indicates a weak or delayed
positive reaction.
Tube
Incubation
Positive
Negative
Comments
ONPG
Yellow
Colorless
(1) Any shade of yellow is a positive reaction.
(2) VP tube, before the addition of reagents, can be used as a negative control.
ADH
18-24 hrs
36-48 hrsRed or Orange
Red Yellow
Yellow or Orange Orange reactions occurring at 36-48 hours should be interpreted as negative.
LDC
18-24 hrs
36-48 hrsRed or Orange
RedYellow
Yellow or Orange Any shade of orange within 18-24 hours is
positive reaction. At 36-48 hours, orange decarboxylase reactions should be
interpreted as negative.
ODC
18-24 hrs
36-48 hrsRed or Orange
RedYellow
Yellow or Orange
Orange reactions occurring at 36-48 hours should be interpreted as negative.
CIT
Turquoise or
Dark BlueLight Green
or Yellow(1) Both the tube and cupule should be filled. (2) Reaction is read in t he aerobic (cupule) area.
H2S
Black
DepositNo Black
Deposit(1) H2S production may range from a heavy black deposit to a
very thin black line around the tube bottom. Carefully examine the bottom of the tube before
considering the reaction negative. (2) A "browning" of the medium is a negative
reaction unless a black deposit is present. 'Browning occurs with TDA positive
organisms.
URE
18-24 hrs
36-48 hrsRed or Orange
RedYellow
Yellow or Orange
Klebsiella, Proteusand Yersinia, routinely give positive reactions.
TDA
Add 1 drop 10% Ferric chlorideBrown-Red
Yellow
(1) Immediate reaction. (2) Indole positive organisms
may produce a golden orange color due to indole production. This is a negative reaction.
IND
Add 1 drop Kovacs' ReagentRed Ring
Yellow
(1) The reaction should be read within 2 minutes after the
addition of the Kovacs' reagent and the results recorded. (2) After several minutes,
the HCl present in Kovacs' reagent may react with the plastic of the cupule resulting
in a change from a negative (yellow) color to a brownish-red. This is a negative
reaction.
VP
Add 1 drop of 40% Potassium hydroxide, then 1 drop of alpha-naphtol
Red
Colorless
(1) Wait 10 minutes before considering the reaction negative.
(2) A pale pink color (after 10 min.) Should be interpreted as negative. A pale pink
color which appears immediately after the addition of reagents but which turns dark pink
or red after 10 min. should be interpreted as positive.
Motility may be observed by hanging drop or wet mount preparation.
GEL
Diffusion of the Pigment
No Diffusion
(1) The solid gelatin particles may spread throughout the
tube after inoculation. Unless diffusion occurs, the reaction is negative.
GLU
Yellow or
GrayBlue or
Blue-GreenFermentation(Enterobacteriaceae, Aeromonas, Vibrio)
MAN
INO
SOR
RHA
SAC
MEL
AMY
ARA
Yellow
Blue or
Blue-GreenOxidation (Other Gram-negative)
(1) Oxidative utilization of the carbohydrates begins in the most aerobic
portion (top) of the tube. Therefore, these reaction should be read from the top to the
bottom of the tube. (2) A yellow color in the upper portion of the tube and a blue in the
bottom of the tube indicates oxidative utilization of the sugar. This reaction
should be considered positive only for non-Enterobacteriaceae gram-negative rods.
This is a negative reaction for fermentative organisms such as Enterobacteriaceae.
4C. Cryptococcus neoformans
Cryptococcus neoformans yeast cell in the process of budding; the specimen of spinal
fluid was centrifuged and the pellet examined in a suspension of India ink.
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