Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 1
Diagnostic of streptococci
Table for summary results for tasks 1 - 6
Strain K L M N P Q R S T
Gram staining Task
1
Catalase ­ Task 2a
Slanetz-Bartley Task
2b
Size
Colour
Shape
Profile
Changes
of agar
Culti-
vation
(Blood
agar)
Task
3
Other
Preliminary result
Task 4a: Optochine
test (only str. with
viridation)
Task 4b: Streptotest
16 (only str. with
viridation)
Task 4c: Suscept. to
bacitracin (str. with
viridation)
Task 5a: PYR test
(only haemolytic st.)
Task 5b: CAMP
(only haemolytic st.)
DEFINITIVE
RESULT
Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 2
Task 1: Microscopy of microbial cultures
Stain according to Gram pure cultures of prepared microorganisms. Use immerion oil and
immersion objective. Draw your observations and write results into table. In strains, which are
not Gram-positive cocci cross out the rest of the table.
Task 2: Basic culture and biochemical tests ­ identification of the
genus
a) Catalase test ­ differentiation of staphylococci
Examine strains of Gram-positive cocci by catalase test. Staphylococci are catalase positive, streptococci
and enterococci negative. Write results in your table.
b) Growth on bile-aesculin agar ­ differentiation of enterococci
There are all strains inoculated on the plate. The strain, which is growing on this medium, is enterococcus.
Write results in your table.
Task 3 ­ Cultivation on Blood Agar
Describe morphology of colonies of prepared bacterial strains on Blood Agar. Note colour and
haemolytic properties of observed colonies. Write the strain properties in your table.
Fill in the preliminary result ­ decide which strains are streptococci. To all strains write ,,NO STR"
(no streptococcus), ,,HAEMOL STR" (partial of full haemolysis) or ,,VIRID STR" (streptococcus
with viridation).
Task 4 ­ Species identifiacation of streptococci with viridation
a) Optochine test
Evaluate the optochine test in your two streptococci with viridation. Strain with susceptibility to
optochine is S. pneumoniae, the strain with no zone around optochine disc is ,,oral
streptococcus". Draw the result and write in your table (+ / ­).
Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 3
b) Biochemical identification of ,,oral" streptococcus
In the strain of oral streptococcus read the biochemical profile and evaluate the result with knowledge
from spring term.
VPT 1
H
1
G
1
F
1
E
1
D
1
C
1
B
1
A
2
H
2
G
2
F
2
E
2
D
2
C
2
B
2
A
1 2 4 1 2 4 1 2 4 1 2 4 1 2 4 1 2
Code: Identification
Streptococcus
___________
% of
probability
T index
c) Susceptibility to bacitracin
Streptococcus mutans, streptococcus with high cariogenic potential is resistant to bacitracin.
Evaluate which of your strains is Streptococcus mutans. Write the results in your table.
Strain __ Strain __
Strain __ Strain __
Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 4
Task 5 Identification of streptococci with partial or full haemolysis
Work only with three strain identifies as beta-haemolytic streptococci.
a) PYR test
PYR test is strip test similar to oxidase test, but the reaction is slower, so the result can be read after 5
minutes and addition of PYR-indicator. Positive result is reddish or pinkish colour of the colony. This test
is positive in S. pyogenes (and enterococci). Negative is in Streptococcus agalactiae and non-A-non-B
streptococci.
Draw the result and fill the table.
Strain ____ Strain ____ Strain ____
negative ­ positive negative ­ positive negative ­ positive
b) CAMP test
CAMP test is based on haemolytic synergy between the strain of S. aureus with production of betahaemolysine
and strain of S. agalactiae. Positive result is butterfly-like shape of zones of full haemolysis
in the crux of both strains. Small zone of other shape is considered as negative result. The CAMP test is
negative in S. pyogenes and non-A-non-B streptococci. Draw the result of the test.
Strain ___
Strain ___
Strain ___
Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 5
Task 6 ­ Susceptibility tests in streptococci
Read and evaluate the susceptibility tests (disc diffusion tests) to antibiotics in Streptococcus pyogenes
and Streptococcus agalactiae.
Kmen
Antibiotikum
(celý název)
 zóny
(mm)
Interpr.  zóny
(mm)
Interpr.
Task 7 ­ Diagnostic of late sequellae of streptococcal infection determination
of ASO
On a side table in a wet chamber you will find a microtitration plate. It includes a positive control, sera X. Y and Z.
Determine the titers and interprete the risk for pacients. (For precise instructions, see the spring term ­ J08)
Result: Interpretation of the result:
Medical Oral Microbiology Topic A - 02
Name: Date:
A2, p. 6
Check-up questions:
1. Why it was difficult to see the arrangement of cocci in our task (pairs, clusters, chains)?
2. Which test besides cultivation on Bile-aesculin or Slanetz-Bartley medium might be employed
in Enterococcus differentiation? One disadvantage: not only the enterococci, but one
streptococcus is positive, too. Which one?
3. Is it useful to perform routine detailed diagnostics of oral streptococci? When it has its
importance?
4. Why optochin test and diffusion disc test employs MH agar with blood and not common MH
agar?
5. Can we use CAMP test in Staphylococcus aureus diagnostic? Why?
6. Why in beta-haemolytical streptococci biochemical test is rarely used for detailed diagnostics?
7. In which streptococci late sequellae take part, and what are the late sequellae?
8. Which streptococci cause infections wiht skin symptomas? What is the name of the infections?