1. Haemophilus influenzae

2. Scientific Classification
Kingdom: Bacteria
Phylum: Proteobacteria
Class: Gamma Proteobacteria
Order: Pasteurellales
Family: Pasteurellaceae
Genus: Haemophilus
Species: H. influenzae
Binomial Name: Haemophilus influenzae

3. History
H. lnfluenzae is also called Pfeiffer's bacillus as it was discovered by Pfeiffer ( 1892)
Was mistaken as the causative agent of Human Influenza
Isolated in 1933 by Smith, Laidlaw and Andrewes

4. Overview Haemophilus means "blood loving."
infiuenzae (the flu-an upper respiratory illness)
Haemophilae are small
Coccobascillus
Pleomorphic
Gram negative bacilli
Are non-motile
Non-sporing
Catalse and oxidase positive

6. Overview Present on the mucous membranes of humans
Normal flora of the human respiratory tract and the oral cavity
Transmitted via droplets discharged from upper respiratory tract during the infectious period
or contact with contaminated secretions

7. Overview Infectious Period
As long as the organism is present even if there is no nasal discharge. Organism is noninfectious within 24 to 48 hours after the start of antibiotics.
There are two major categories of H. influenzae;
Encapsulated strain(typeable)
Unencapsulated stains(nontypeable)

8. Overview
The Encapsulated strains are classified into six serotypes based on their capsular antigens (designated a to f )
Unencapsulated strains are called nontypeable because they lack the polysaccharide capsule and consequently capsular antigens
Type b (Hib) causes about 95% of the invasive diseases. Important human pathogen

9. Epidemiology
Encapsulated strains colonize approximately 40-80% of children and adults
Capsulated(Hib)Colonizes approximately 3-5% of children 2-5yrs
By the age of 5–6 years more than 50 % of children will be colonized with this bacterium and most healthy adults (at least 75 %) will be
Neonates – protected by maternal antibodies. Older children and adults develop bacteriocidal antibodies during carriage
Closed communities e.g. nurseries – increased risk of infection
Incidence has reduced considerably since introduction of conjugate vaccine – by about 95%.
Cases exclusively seen in unimmunized children

10. Cultural Characteristics
H. influenzae requires two accessory growth factors present in blood; Factor X and Factor V
Factor X consists of group of heat-stable compounds such as hemin or other porphyrins required for the synthesis of enzymes such as cytochrome, catalase and peroxidase.
It is involved in the aerobic respiration. It is not required when H. influenzae grows anaerobically.

11. Cultural Characteristics
Factor V: It is a heat-labile, nicotinamide adenine dinucleotide (NAD), which is also produced by some animals, plant cells and other bacteria, such as Staphylococcus aureus.
It gets inactivated by NADase present in sheep blood.

12. Cultural Characteristics
. In the laboratory it is classically grown on chocolate agar
Another way to grow it, is to grow with staphylococcus colonies, on blood agar which provides factor V via red blood cell hemolysis
On both chocolate agar and blood agar, H.influenzae grows into convex, smooth, gray or transparent colonies

13. Visible growth of H.influenzae on a chocolate agar

14. Cultural Characteristics
The growth of H.influenzae vary in different media depending on the availability of X and V factors.
Does not grow on ordinary media: Nutrient agar or peptone water lack X and V factors, hence does not support Haemophilus growth

15. Cultural Characteristics
Growth is best on chocolate agar because it has both factor X and V
While preparing chocolate agar, blood is poured into molten agar at 75°C which inactivates NADase and lyses RBCs releasing excess of factor V.
Hence it supports the growth of H.influenzae.

16. Cultural Characteristics
Growth is scanty on blood agar because only Factor X is available in this medium and Factor V is largely intracellular, present only inside the RBCs.
Factor V is available in very minute quantities freely in the medium especially in sheep blood . More so, blood contains NADase that destroys factor V.
Growth can be enhanced if medium is supplemented with an extra source of NAD:
H.influenzae does not grow on sheep blood agar except around colonies of staphylococci (“satellite phenomenon”)

17. Cultural Characteristics
Streaking S. aureus across surface of BA produces growth stimulation in its vicinity or
Use of paper discs impregnated with X and V factors alone, and in combination
The NADase in the blood may be in activated by heating to 70 – 80°C
The enhanced growth around V factor – called SATELLITISM.

18. Satellitism
When Staph aureus is streaked across plate of blood agar with species containing H influenza
The large colonies of H influenza develop along the streak of Staphylococcus aureus and the small colonies further away

19. Satellitism

20. Cultural Characteristics
In cultures, the morphology depends both on the length of incubation and on the medium.
At 6–8 hours in rich medium, the small coccobacillary forms predominate. Later, there are longer rods and very pleomorphic forms.
Facultative anaerobes but do not grow at a temperature less than 22°C
Optimal growth is seen at 35-37°C. Better growth is obtained under aerobic conditions as compared to anaerobic environment. Presence of CO2 improves the growth.

21. Pathogenesis Virulence Factors
Sword-like structure that helps it attack and destroy the host cell.
Encapsulated strains of Haemophilus influenzae are covered by a polysaccharide layer called the capsule
They also have pili and adhesion proteins (HMW1 and HMW2) for attachment to host cells.
Both encapsulated and unencapsulated have an outer membrane which consists of lipooligosaccharide which inhibits self-clearing mechanism of the bronchi.
Encapsulated and unencapsulated strains make IgA protease which destroys IgA.
IgA protease neutralizes the first line of mucosa defense
Unencapsulated strains have two other abilities to help invade the immune system
1 Phase variation where the strains consist of oligosaccharide
2 Ability to produce biofilms (Made of exopolysaccharides) within which H.I live and produce.

22. Invasive diseases
Epiglottitis-fever, sore throat, difficulty speaking and dyspnea
Cellulitis-fever, warm and tender area of erythema or cheek or periorbital area.
Bacteremia-fever, chills, hypotension and tachycardia
Meningitis- fever, lethargy, irritability, vomiting, sore neck and altered mental status
Osteomyelitis-fever, bone pain and weakness
Septic arthritis- fever, pain, swelling and tenderness of the affected joint

23. LABORATORY DIAGNOSIS:
SEROLOGICAL METHODS
BIOLOGICAL SAMPLES
Blood
CSF
Synovial fluid
Pleural fluid
Fluids retained by sinus aspiration
Tympanocentesis
Tracheal or lung aspiration
Bronchoscopy
Bronchoalveolar lavage
Latex agglutination
Enzyme immunoassay
Coagglutination
For epiglottitis, a laryngoscopy can be done which shows a cherry red and swollen epiglottis and an X-ray which shows a thumbprint sign on the epiglottis

25. Treatment Infections with H.influenza type b are treated with;
1st choice (ceftriaxone)
Chloramphenicol

26. PREVENTION & CONTROL Vaccination( for H.inflenzae type b)
Recommended between 2 and 18 months of age
Contains type b capsular polysaccharide conjugated to a Diphtheria toxoid component
Used to greatly decrease H.influenzae meningitis
The use of antibiotics.
Contacts of children with H.inflenzae type b may be treated with chemoprophylaxis by using RIFAMPIN
Avoid close contact.
Practice good hygiene.
Breathing support.
Medication to treat low blood pressure.
Wound care for parts of the body with damaged skin.

27. REFERENCES
“Haemophilus influenza Disease (Including Hib)”. Centre For Disease Control And Prevention. 18th March, 2021.