Morphology and cultural characteristics
Haemophilus influenzae belongs to the family Pasteurellaceae. These are small, Gram-negative, non-motile, non-spore-forming bacteria possessing pili. They exhibit pleomorphism: smears may contain spherical, ovoid, and rod-shaped forms, as well as short filaments. A crucial, though non-constant, structural element is the capsule.
In terms of respiration, they are facultative anaerobes. Their key metabolic feature is the inability to synthesize respiratory enzymes. They require preformed blood factors for growth:
- X-factor (protoporphyrin X) — required for the synthesis of respiratory chain enzymes.
- V-factor (NAD or NADP) — a cofactor for oxidation-reduction reactions.
On ordinary blood agar, growth is poor because the V-factor is destroyed by NADases of intact erythrocytes. The medium of choice is chocolate agar. During its preparation, blood agar is heated to 80 °C, which causes hemolysis (releasing factors X and V) and inactivates the enzymes destroying them. Also characteristic is the satellite phenomenon («feeder» effect): Haemophilus grows abundantly around colonies of staphylococci that secrete NAD into the medium and lyse erythrocytes.
Antigens and pathogenicity factors
The bacterium possesses a somatic O-antigen and a capsular polysaccharide K-antigen. Based on the capsule, 6 serotypes (a–f) are distinguished. Of greatest clinical significance is type b (Hib), whose capsule consists of polyribosylribitol phosphate (PRP).
Main pathogenicity factors:
- Capsule — the leading factor. It has anti-complementary action, protects against phagocytes, and helps the bacterium survive and invade tissues.
- Endotoxin (lipooligosaccharide) — increases the permeability of tissue barriers (including the blood-brain barrier) and causes paralysis of the cilia of the ciliated epithelium.
- IgA protease — inactivates mucosal protective antibodies.
- Pili — provide adhesion to the epithelium. Interestingly, unencapsulated strains possess greater adhesiveness, which is why they remain at the portals of entry and cause local inflammation.
Epidemiology and pathogenesis
Haemophilus influenzae — a strict anthroponosis. The source of infection is a sick person or a bacterial carrier (unencapsulated strains are more frequently carriers). The transmission mechanism is airborne.
The incidence rate is closely tied to age-related immunology. Up to 3–6 months, infants are protected by maternal IgG. Then their level drops, and the child's own production of antibodies to the T-independent capsular antigen is physiologically reduced in children under 2 years of age. A peak of severe infections occurs during this period.
Clinical courses of infection:
- Unencapsulated strains: cause sinusitis, otitis media, bronchitis, and pneumonia (frequently in the elderly, smokers, and immunocompromised individuals).
- Encapsulated strains (Hib): cause generalized forms due to hematogenous dissemination. These include purulent meningitis (with the risk of vision loss, deafness, and hydrocephalus) and acute bacterial epiglottitis (fulminant laryngeal edema with the risk of asphyxia). The mortality rate of severe forms without treatment reaches 90%.
Diagnostics and treatment
The diagnosis of meningitis can be preliminarily established by microscopy of the cerebrospinal fluid. For rapid detection of the capsular antigen, PCR, DFA, and latex agglutination are used. The primary method is bacteriological: inoculation on chocolate agar with confirmation of the requirement for X- and V-factors.
In generalized forms, therapy is initiated immediately (empirically) with 3rd-generation cephalosporins (ceftriaxone). About 25% of strains produce $\beta$-lactamases; therefore, protected penicillins (amoxicillin + clavulanic acid) are used to treat local forms.
Prevention: conjugated vaccines are used (where the PRP capsular antigen is bound to a carrier protein, such as diphtheria toxoid). This converts the antigen into a T-dependent one and successfully induces immunity in children from an early age.
Briefly: answers to topical questions
What nutrient media are used to isolate Haemophilus influenzae?
To isolate Haemophilus influenzae, media that provide the bacterium with growth factors X and V are used.
- Chocolate agar — is the medium of choice; it is prepared by heating blood agar to 80 °C, which causes hemolysis and the release of essential factors.
- Blood agar supplemented with sheep blood — is used in the bacteriological method, although independent growth on it is poor or absent.
- Blood agar with colonies of other bacteria — growth is possible due to the "satellite phenomenon", where "feeder" bacteria (e.g., staphylococci) secrete the V-factor.
What biochemical properties are characteristic of Haemophilus influenzae?
Haemophilus influenzae is characterized by a chemoorganotrophic type of nutrition combining respiratory and fermentative metabolism.
- It utilizes glucose with acid production; other carbohydrates are fermented weakly.
- It reduces nitrates to nitrites.
- For identification, indole production and urease activity are additionally tested.
- Hemolysis on blood agar is usually absent.
What antigens are part of the antigenic structure of Haemophilus influenzae?
The antigenic structure of Haemophilus influenzae includes:
- O-antigen — somatic.
- K-antigen — capsular polysaccharide antigen, serving as the basis for serotyping; 6 serotypes are distinguished: a, b, c, d, e, f.
Of greatest clinical significance is the serotype b capsular antigen (Hib), which is a polymer of ribose and ribitol — polyribosylribitol phosphate (PRP).
What groups of antibiotics are used to treat infections caused by Haemophilus influenzae?
To treat infections caused by Haemophilus influenzae, the following are used:
- 3rd-generation cephalosporins — ceftriaxone and cefotaxime; drugs of choice for severe generalized forms.
- Penicillins: ampicillin; for localized forms — protected penicillins, e.g., amoxicillin/clavulanate.
- Co-trimoxazole (biseptol) — an alternative for localized forms.
- In purulent meningitis caused by Haemophilus influenzae, chloramphenicol is also indicated as a source.
Mnemonic for cultural properties: «Chocolate XV». Haemophilus prefers Chocolate agar because it contains factors X and V.
Class and exam questions
Why doesn't Haemophilus influenzae grow on ordinary blood agar?
Intact erythrocytes contain NADase enzymes. They destroy the V-factor (NAD), which is vital for Haemophilus influenzae to operate its respiratory enzymes.
What is the fundamental difference between encapsulated and non-encapsulated strains?
Unencapsulated strains have high adhesion, colonize the nasopharynx, and cause only local inflammation (otitis, sinusitis). Encapsulated strains (primarily type b) are capable of overcoming barriers, entering the bloodstream, and causing generalized infections (meningitis, sepsis).
Why is a carrier protein used in the Haemophilus influenzae type b vaccine?
The capsular antigen of Haemophilus is T-independent, and the immune system of young children barely responds to it. Binding the antigen to a carrier protein makes it T-dependent, allowing for the generation of robust immunity even in infants.
What else to study on this topic
- Biochemical activity: carbohydrate fermentation and nitrate reduction
- Immunological paradox: pneumonia and arthritis in adults in the presence of antibodies to PRP
- Mechanism of the satellite phenomenon during co-cultivation with Staphylococcus
- Pathomorphology of purulent meningitis: subdural edema and damage to the cerebral meninges' vessels
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