Haemophilus influenzae is a Gram-negative coccobacillus first isolated in 1892 during an influenza outbreak. It is classified within the phylum Proteobacteria and can cause several diseases in humans like meningitis, pneumonia, epiglottitis and cellulitis. H. influenzae possesses several virulence factors like a capsule, lipopolysaccharides and IgA1 protease that help it evade the immune system and cause disease. Diagnosis involves culture and identification of the bacteria from clinical specimens as well as antigen and molecular detection techniques. Vaccination against H. influenzae type b is available to prevent disease.

1. HAEMOPHILUS INFLUENZAE

2. HISTORY
 Hib was found in
a group of
patient during an
influenza in
1892.
 Haemophilus
Influenzae was
first isolated in
1890 by Richard
Pfeiffer.

3. SCIENTIFIC CLASSIFICATION
 Kingdom : Bacteria
 Phylum: Proteobacteria
 Class: Gamma Proteobacteria
 Family: Pasteurellaceae
 Genus : Haemophilus
 Species : Influenzae

4. SPECIES
 Haemophilus Influenzae
 Haemophilus Influenzae biotype aegyptius
 Haemophilus haemolyticus
 Haemophilus ducreyi
 Haemophilus aprophilus
 Haemophilus parainfluenzae
 Haemophilus parahaemolyticus
 Haemophilus paraphrophilus

5. Species X Factor Y Factor Haemoly
sis
CO2
Require
ment(5%
)
Lactos
e
Sucrose Glucose Mannos
e
catalase Xylose
H.influen
zae
+ + _ + _ _ + _ + +
H.Influen
zae(aegy
ptius)
+ + _ _ _ _ + _ + +
H.haemo
lyticus
+ + + _ _ _ + _ + _
H.ducreyi + _ +w _ _ _ _ _ _ _
H.aphrop
hilus
_ _ _ + + + + + _ +
H.parainf
luenzae
_ + _ _ _ + + + V _
H.paraha
emolyticu
s
_ + + _ _ + + _ + _
H.paraph
rophilus
_ + _ + + + + + _ +

6. CHARACTERISTICS OF SIX BIOTYPES
character I II III
IV V VI VII
VIII
Indole production + + _ _ + _ + _
Urease Activity + + + + _ _ _ _
Ornithine decarboxylase + _ _ + + + _ _

7. MORPHOLOGY

9. VIRULANCE FACTOR
 Outer membrane protein
 Pili
 Immunoglobulin A1 Protease
 Lipopolysaccharides
 Capsule

10. VIRULANCE FACTOR BIOCHEMICAL FUNCTION
Capsular polysaccharides PRP of the capsule is antiphagocytic. It
resists phagocytosis of the bacteria
Lipopolysaccharides Causes meningococcal inflammation.
IgA1 protease Causes breakdown of IgA, facilitate
colonization of H.influenzae on the
mucosal surface
Pili Helps in adherence of H.influenzae to
epithelial cells.

11. MODE OF TRANSMISSION

12. PATHOGENESIS
H.Influenzae enters the human host by respiratory route.
Pilus and non pilus adhesions of the bacteria mediate colonization in
the nasopharynx and oropharynx
Lipid A lipopolysaccharides impairs ciliary function and IgA1 protease
breakdown IgA1
Cause damage of the respiratory mucosa
A large bacterial load or the viral infection potentiate the bacterial
infection that invade mucosa and enter the blood stream.

13. The presence of antibodies, complement components and phagocytes
cause clearance of bacteremia
The absence of anti PRP antibodies contributes to bacterial infection
High grade bacteremia leads to disseminate to various sites including
meninges, subcutaneous tissue,joints,pleura and pericardium
Responsible for causing Meningitis, Arthritis, pneumonia and endocarditis
Colonization of noncapsulated strains will direct extend to sinuses,
eustachian tube etc cause sinusitis and otitis media

14. CLINICAL SYNDROME

16. Cellulitis cause by H.influenzae in foot

17. Cellulitis caused by H.influenzae in cheek

18. H.Influenzae causing Otitis Media

19. Clinical Syndrome of H.influenzae

20. MATERIALS AND METHODOLOGY

21. PREANALYTICAL PHASE
 Sample Collection
 Sample Transport

22. SPECIMEN
 CSF
 Throat Swab
 Sputum
 Pus
 Joint Aspirates
 Pleural Aspirates
 Bronchial Aspirates
 Pericardial Aspirates

23. COLLECTION
Lumber Puncture for CSF Collection

24. Sputum Collection

25. specimen collection for throat swab

27. ANALYTICAL PHASE
 Direct Examination :
Microscopic Observation
H.Influenzae in Gram Stain

28. H.Influenzae in coccobacilli form in Gram stain

29. Direct Smear Of H.influenzae in CSF in a case of meningitis

30. H.Influenzae in immunofluorescence microscopy

31. ANTIGEN DETECTION
Latex Agglutination Test For H. influenzae

32. CULTURAL CHARACTERISTICS
H.Influenzae in Chocolate Agar

33. SATELLITISM
Satellitism Test for H.Influenzae

34. SELECTIVE MEDIA
 Levinthal’s Agar:
Prepared by boiling and filtering a mixture of blood
and nutrient broth.
Fildes Agar : By adding peptic digest of blood to
nutrient broth

35. H.Influenzae in Fildes Agar

36. ENRICHMENT MEDIA
 Blood Culture:
Brain Heart Infusion Broth (BHI Broth) :

37. BIOCHEMICAL TEST
CATALASE TEST OXIDASE TEST

38. NITRATE REDUCTION TEST

39. NITRATE REDUCTION TEST

40. ISOLATION FROM CLINICAL SPECIMEN

41. IDENTIFICATION
X and V Test For H.influenzae

42. H.Influenzae in X And V Test

43. H.Influenzae in Latex Agglutination Test

44. Flowchart for isolation of H.influenzae

45. POST ANALYTICAL PROCEDURE
 Interpretation of Laboratory Results:
Antigen Detection :
Culture :

46. ANTIBIOTIC SENSITIVITY TEST
Kirby Bauer Disk diffusion Method For H.influenzae
performing in Chocolate Agar

47. Acidometric Paper Strip Method For Penicillin

48. Nitrocefin Test For Cephalosporin

49. PREVENTION AND CONTROL
 Vaccination :
Recently,three types of haemophilus influenzae
type b vaccines are available. These vary in
following properties:
1.The protein carrier used.
2.The molecular size of saccharides.
3.The method of conjugating the protein to the
Saccharides.

50.  Three types of Vaccines are as follows:
 HbOC (Mutant Diphtheria toxin as a carrier)
 PRP-T (Tetanus Toxoid as a carrier Protein)
 PRP-OMP (Major OMP Of Neisseria meningitidis
serotype B as a carrier protein )

51. Child Vaccinated with Haemophilus Type b Vaccine

52. THANK YOU
ANY ?????????

53. REFERANCES
 Practical Medical Microbiology(Mackie And
McCartney)
 Diagnostic Microbiology (Bailey And Scott’s)
 Textbook Of Microbiology And Immunology (Parija)
 A Textbook Of Microbiology (Chakraborty)
 Textbook Of Microbiology (Dr. Prof. CP Baveja)
 Textbook Of Microbiology (Anantanarayan And
Paniker’s)