in this ppt you will know more about the bacterial causes of meningitis this include its stain ,describtion on culture special media& biochemical reactions

1. Presented By
Christine Efat Tawfeek
(Assistant Lecturer At Microbiology Dep.
Fomscu)

2. Diagnosis:
1. Sample Collection : CSF
2. CSF Analysis
3. Gram Staining
4. Culture
5. Biochemical Reactions
6. Serology

3. CSF Analysis
• Aspect: Turbid
• Glucose Level: < 40 mg/ dl
• Protein Level: >100mg/ dl
• WBCs Count: > 500 count/mm³
• Neutrophils: 70-90% of WBCs count

4. Most Common Bacterial Causes Of Meningitis
1. Neisseria meningitidis
2. Streptococcus pneumoniae
3. Streptococcus agalactiae (Streptococcus Group B)
4. Escherichia coli
5. Hemophilus influenza

5. Basic stain
Acetone-
alcohol

6. There are four reagents used in gram staining:
1-A primary ( basic) stain: crystal violet.
2-a trapping agent (mordant): gram’s iodine.
3-a decolorizing agent: 95% alcohol or acetone-alcohol
4-a counterstain: safranin.
Presentation title 6

7. NEISSERIA MENINGITIDIS
• Gram Stain:
Gram negative diplococci intracellular in
pus cells as well as extracellular

9. CULTURE MEDIA:
• Name : chocolate agar
• Type: enriched medium
• Use: culture of fastidious organisms e.g.
Neisseria spp &
Haemophilus spp.
Presentation title 9

10. Presentation title 10
On chocolate agar : Neisseria meningitidis appear as greyish white ,glistening ,
convex colonies

11. 2- THAYER MARTIN MEDIUM
Presentation title 11

12. Composition:
• Chocolate agar has been modified to be selective for neisseria spp by
adding of antibiotics:
Colistin : inhibition of gram negative bacteria
Vancomycin : inhibition of gram positive bacteria
Nystatin: inhibition of fungi
• Modified Thayer martin : trimethoprim to inhibit proteus
• Nitrogenous nutrients such as casein to neutralize toxic fatty acids .
Presentation title 12

13. Need 5-10% CO2 for growth Candle jar

14. • Identify:
Anaerobic jar used as candle jar
• Use:
Culture of bacteria that need 5-10% CO2 for
growth as neisseria

15. Biochemical Reactions
• Oxidase test : gives positive result ( purple color ) , as neisseria
spp have cytochrome C
Presentation title 15

16. Presentation title 16

17. Oxidase test
Name of the test
Interpret result
Give an example

18. STREPTOCOCCUS PNEUMONIAE
GRAM POSITIVE DIPLOCOCCI

19. Streptococcus pneumoniae
• Microscopic appearance: gram-positive, lancet-shaped, diplococci (arranged
in pairs), but they may also occur in short chains

20. • Culture media :
● Blood agar :
colonies grow on blood agar and
surrounded by αlpha - haemolysis.
.

21. QUELLUNG TEST
(CAPSULE SWELLING TEST)
This method involves testing a pneumococcal cell suspension with specific antisera directed against
the capsular polysaccharide
The antigen-antibody reactions are observed microscopically
A positive quellung reaction is the result of the binding of the capsular polysaccharide of pneumococci
with type specific antibody contained in the typing antiserum (Strept.pneumoniae)
A positive quellung reaction is observed when the capsule appears as a halo (swollen capsule) around
the bacterial cell

23. Streptococcus agalactiae
(streptococcus group B)
GRAM POSITIVE COCCI ARRANGED IN SHORT CHAINS

24. • Culture media:
• Grow on blood agar and
produce complete beta
hemolysis

25. Catalase test
Staphylococci
Streptococci E.g.
streptococcus
agalactiae

26. H2O2 H2O + BUBBLES OF O2
Catalase

29. Escherichia coli
Gram negative bacilli

30. MaCconkey agar
• Type: differential selective medium
• Use: cultivation of enteric bacteria and
differentiation between two major groups of
enteric bacteria:
(1) lactose fermenting bacteria:
• Give pink colonies.
• E.G. E. Coli, klebsiella.
(2) lactose non-fermenting bacteria:
• Give pale colonies.
• E.G. Salmonella, shigella, and proteus.

31. 2) INDOLE TEST
Principle:
The test determines the ability of the
organism to produce indole from the
breakdown of the amino acid tryptophan.
Addition of Kovac’s reagent which acts
with indole gives a red color
Results and interpretation:
A positive result is indicated by the
presence of a red color in the layer on the
surface of the broth. A negative
result appears in yellow.
Example:
E.coli
Example:
Shigella

32. HAEMOPHILUS INFLUENZA
• MICROSCOPIC APPEARANCE:
H. Influenza in a gram stain of CSF sample appears as gram-negative
coccobacilli.

33. • Culture on chocolate agar
• Type: enriched media
• Use: culture of fastidious organisms as
neisseria and haemophilus

34. • CULTURAL CHARACTERS:
• H.influenza is a fastidious, facultative anaerobic organism which grows best at
35-37°C with ~5% CO2 (or in a candle-jar) and requires hemin (X factor) and
nicotinamide-adenine-dinucleotide (NAD, also known as V factor) for growth.
• The standard medium used for growth of h. Influenzae is a chocolate agar plate
which can be prepared with heat-lysed horse blood, a good source of both hemin
and NAD.
• Growth occurs on a chocolate agar because NAD is released from the blood during
the heating process of chocolate agar preparation (the heating process also
inactivates growth inhibitors) and hemin is available from non-hemolyzed as well
as hemolyzed blood cells.

36. • H.INFLUENZA REQUIRES BOTH X AND V FACTORS FOR GROWTH.
• In this culture, H.Influenza has only
Grown around the paper disc that has
been impregnated with both
X and V factors.
• There is no bacterial growth
Around the discs that only
contain either X or V factor

38. MUELLER-HINTON AGAR
• USE:
ANTIBIOTIC SUSCEPTIBILITY
TESTING

39. Zone of inhibition

40. Bacteriological Loop
• Use: inoculation of bacteria on
culture media
• Method of sterilization:
dry heat (red heat) by heating
in the bunsen flame till it turns
red hot.