Test Name
Purpose
Reagent
Conditions
Type of
Media
Positive
result
Negative
Result
Notes
Carbohydrat
e
fermentation
Detects
production of
acid
Phenol
Red
Glucose
broth
37◦C for 24
hr
Broth
medium
yellow color
at acidic
medium
Stays red in
neutral
medium
Catalase
Detects
production of
catalase enzyme
that neutralizes
H2O2 into O2 and
H2O
hydrogen
peroxide
solution
(3%)
glass slide
rapid
elaboration
of oxygen
bubbles
No Bubbles
Oxidase
Detects the
presence of
cytochrome
oxidase
enzyme.
oxidase
strips
strip
Color
Production
No color
To detect the presence
of gas produced,
Durham’s tube is
placed in the
fermentation broth,
(the evaluation
of gas will be visible as
a bubble)
Anaerobes lack
catalase, in Gram +ve
Cocci; Streptococcus
and Enterococcus are
catalase-negative but
Staphylococcus is
catalase-positive,
the families of
Pseudomonadaceae
(ox+) and
Enterobacteriaceae
(ox-).
From fresh
culture
remove with
plastic loop
not metallic
because
metal may
react with
strip check
only after 20
seconds
Coagulase
Citrate
Urea
Hydrolysis
(urease)
Gelatin
Hydrolysis
Indole
Hemolysis
Detects
Coagulase (an
enzyme that
clots blood
plasma and
protects bacteria
from
phagocytosis)
determine the
ability of a
bacterium to
utilize citrate as
the only source
of carbon.
Detects Urease
(an enzyme that
breaks the
carbon-nitrogen
bond of amides)
detect
gelatinase
(liquefy gelatin)
Detects indole
(product of
tryptophan
hydrolysis by
enzyme
tryptophanase)
Detects
Hemolysins
(bacteria with
Glass tube
Clot
formation
No Clot
differentiate
pathogenic strains of
Staphylococcus
(coag+) from nonpathogenic strains
(coag-).
Bromothy
mol Blue
Simmone’s
Citrate
Media
(Slant green
agar)
Blue color at
basic
conditons
(sodium
carbonate)
Stays green
Positive for klebsiella,
Enterobacter and
Citrobacter, and
negative for E. coli.
Phenol
Red
Urea Broth
Pink in basic
environment
(ammonia
released)
Red in
neutral
conditions
Positive for genus
Proteus
Gelatin
Medium
Medium
Becomes
liquid
Red color
Medium
stays solid
optimally at
25 °C
Kovac’s
Reagent
Tryptophan
broth
(Tryptone
broth)
Blood Agar
Stays yellow
Positive for E. coli. And
Negative for Klebsiella,
Enterobacter, Hafnia,
Serratia

Beta Hemolysins
(lyse blood
completely;
enzymes that
lyse red blood
cells and degrade
hemoglobin)


MR
VP
Nitrate
reduction
Test the ability of
a bacterium to
ferment glucose
and produce
strong acids
to test the ability
of the organisms
to produce
acetoin from
glucose
fermentation.
to determine the
ability of the
organism to
reduce
Methyl
Red
alphanaphthol
(VP(A))
and KOH
(VP(B)).
35°C for 4872 hr.
24 hours at
37°C.
Shake the
tube gently
to expose
the medium
to
atmospheric
oxygen 10-15
min
sulphanilic Add zinc to
acid and
the tube
alpha‐
showing
Staphylococcus
aureus )
Alpha Hemolysins
(lyse blood
incompletely
producing greenish
color around
colony;
Streptococcus
pneumoniae.
Gamma hemolysins
(no breakage)
(MRVP)
broth
Red color
Yellow color
(MRVP)
broth
Red color
Yellow color
KlebsiellaEnterobacter-HafniaSerratia produce
acetoin.
nitrate
broth
Red color
detects
nitrites.
If after zinc
there is
change (red
color) then
Enterobacteriaceae
and Haermophilus and
neisseria genera can
reduce nitrate.
Motility Test
nitrate to nitrites
or free nitrogen
gas.
naphthyla
mine
to differentiate
between motile
and non‐motile
colorless
tetrazoliu
m
absence of
nitrite.
Motility
agar with
tetrazolium
dye.
If after zinc
there is no
change then
positive
result and
the bacteria
has
produced
ammonia or
nitrogen gas.
Red color
negative
result.
colorless
If the bacteria is
motile, the original
stab line will
diffuse out into the
medium as the bacteria
spread throughout. If
bacteria is not motile,
there will only be
growth along the stab
line.
Obligate aerobes will not
grow very deeply into
the medium. It will grow
only on the surface and
thus, it
may not be possible to
interpret its motility.
Amino Acids
tests
Lysine
decarboxylase is
an enzyme that
breaks the bond
between the
bromocre
sol purple
mineral oil is
used for
anaerobic
conditions
Lysine
decarboxyla
se broth
Purple color
in basic
medium
yellow
Kligler Test
carboxylic (COOH) group
leaving basic
amine group
Phenylalanine
deaminase is a
common enzyme
that knocks the
amino
group producing
an acidic
chemical
(phenylpyruvic
acid).
Test ability to: ferment glucose
and lactose into
acids
-produce gas end
products
-possess sulfur
reducers (release
H2S gas)
FeCl3
phenylalani
ne agar
slant
green
avocado
color.
Phenol
red (PH
indicator)
and iron
(H2S
release)
Kligler
Medium
(low Glc,
high Lac,
and
proteins)





rustic
Yellow butt and yellow slant will be detected if
fermentation of Glc and Lac occurred.
yellow butt and red slant will be detected if
fermentation of Glc but not Lac occurred.
Red butt and red slant will be detected if no
fermentation occurred.
If gas is produced as a result of glucose or lactose
fermentation, then fissures will appear in the agar
or the agar will be lifted off the bottom of the
tube.
H2S gas can be also detected as a black
precipitate