BIOCHEMICAL TEST
Biochemical test is very
important for specific microorganism identification.
a)STAPHYLASE TEST
Purpose: To differentiate
species within the genera Staphylococcus (S. aureus usually positive from other
Staphylococcus spp.
Principle: S.aureus able
to produce free and bounce coagulase(clumping factor). Staphylase test will
detect the presence of clumping factor through clumping fibrinogen sensitized
sheep red blood cell.
Reagent: Staphylase Rest
Reagent
Result
Positive– Clumping of test cell (Staphylococcus
aureus)
Negative– No Clumping (Other
Staphylococcus sp)
b)STREPTOCOCCAL GROUPING
Purpose: For the
qualitative detection and identification of the Lancefield group of
Streptococci
Principle: Latex
agglutination test for the identification of streptococcal group, and the
reagent are provided for groups A, B, C, D, F and G that consist of specific
antigen for ach group.
Result: Test is considered
positive when agglutination occurs with one grouping reagent or when one
grouping reagent gives a substantially stronger reaction tan the other five.
c)P.Y.R. HYDROLYSIS TEST
Purpose: The PYR is a
rapid colorimetric test for the determination of PYRase activity in
Streptococci ad Citrobacter spp
Principle: PYR test usus
test cards impregmenated with L-pyroglutamic acid
&-amino-4-methylcoumarin(7AMC) and dimethylamino-cinnamaldehyde for the detection
of PYRase activity. The enzymatic hydrolysis of this substrate by Enterococci,
Group A Streptococci and Citrobacter spp. produces a purple color following the
addition of the developing solutions.
Result
Reactivity with
|
||
Organism
|
Lancerfield Group
|
P.Y.R
|
Streptococcus pyogenes
|
A
|
+
|
Streptococcus agalactase
|
B
|
-
|
Streptococcus Group C
|
C
|
-
|
Streptococcus bovis
|
D
|
-
|
Streptococcus feacalis
|
D
|
+
|
Streptococcus feacium
|
D
|
+
|
Citrobacter
|
F
|
-
|
Salmonellae
|
G
|
-
|
Escherichia coli
|
+
|
|
d)SLIDEX PNEUMO KIT TEST e)BACTERIAL ANTIGEN TEST f)X,V, XV GROWTH FACTOR g)OPTOCHIN
Purpose:
It is used to determine susceptibility organism to Optochin chemical and
fragility of cell membrane. It is for presumptive identification of S.pneumonia
from alpha-streptococcus spp.
Principle:
Optochin disc are impregmented with ethylhydrocupreine hydrochloride
(optochin),a drug used for pneumonia theraphy before sulfonamide became
available. The growth of pneumococci, but not other streptococci, is markedly
inhibited by this chemical. Pneumococci may, therefore be differentiated from
other alpha-hemolytic streptococci by the formation of a zone of inhibition
around a optochin disc placed on a blood agar plate heavily inoculated with
pure culture suspected to be Streptococcus pneumoniae.
Result And Interpretation
Positive- zone
of inhibition of 14mm are formed.
h)TRIPLE SUGAR IRON (TSI)
For
Determine whether a Gram negative bacillus utilize glucose, lactose or sucrose
fermentatively and forms hydrogen sulphide
Results
- Alk slant/alk butt (K/K)= Non fermenter
- Alk slant/acid butt(K/A)=Glucose fermenter only
- Acid slunt/acid butt (A/A)=Glucose, sucrose and/of lactose fermenter
- Bubble and crack=Gas formation
- Black precipitate=Produce hydrogen sulphide
Quality
Control
- Citrobacter=A/A with gas and H2S production
- Providencia= K/A
- Pseudomonas=K/K
i)MOTILITY TEST
Detection
of motility of gram-negative bacilli
Result
- Motile Organism= spread growth from the line of inoculation. Eg: Enterobacter aerogenes, E.coli, Proteus vulgaris, etc
- Non-motile organism= Growth only along the inoculation line. Eg:Klebsiella pneumoniae, Shigella flexneri, etc
j)UREASE TEST
Purpose
The test is to determine the
ability of an organism to split urea into ammonia by the action of the enzyme,
urease. When organism hydrolyses urea, ammonia is formed during incubation
making the media alkaline. The phenol red indicator incorporated in the medium
turns from yellow (acid) to pinkish red (alkaline).
Reading And Interpretation
- Positive test-Intense pink red to red violet color
- Negative test-No color change
k)OXIDATIVE FERMENTATIVE TEST
Oxidation fermentation media is
used to determination of oqidative and fermentative metabolism of carbohydrates
by gram-negative bacilli on the basis of acid reaction in either the open or
closed system
Result
|
Open system
|
Closed system
|
Oxidation
|
Yellow
|
Green
|
Fermentation
|
Yellow
|
Yellow
|
No reaction
|
Blue/green
|
green
|
l)COAGULASE TEST
Shows ability of organism to
agglutinate or plasma to clot with the action of coagulase enzyme. It can
differentite Staphylococcus aureus from other Staphylococcus sp because of
Staphylococcus aureus ability to produse coagulase enzyme.
Result
- Coagulse positive- Staphylococcus aureus
- Coagulase negative- Staphylococcus sp.
m)GERM TUBE TEST
To identify and differentiate
between Candida albican and non-Candida albican.
Method
- Place few yeast colny in 50ul of human serum and incubate for 2-3 hours.
- Transfer 1 drop of serum yeast culture to a glass slide and cever with coverglass
- Examine and search for sprouting yeast cell
Reading And Interpretation
- Presence of sprouting yeast cell- Candida albicans
- No sprouting yeast cell- Candida spp.
n)CATALASE TEST
Determine the presence of
catalase enzyme produced by bacteria
Reading And Interpretation
- Production of bubble- positive catalase
- No bubble- negative catalase
o)OXIDASE TEST
- Oxidase test are used in the Kovac oxidase test as a qualitative reaction in the identification of non-fermenters and miscellaneous Gram negative bacilli
- Production of indophenol oxidase enzyme oxidize a redox dye(present in reagent) which results in color change of yellow to dark purple. Indophenol oxidase in the presence of atmospheric oxygen, oxidizes the phenylenediamine oxidase reagent to form a dark purple compound, indophenol
Reading And Interpretation
Positive reaction turns the
bacteria streak to purple immediately or within 10-30 seconds
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