Bacteriology

HAEMOPHILUS INFLUENZAE

Written by MicroDok

Haemophilus influenzae is a small, Gram-negative, non-sporulating, non-motile, urease positive, indole positive, pleomorphic, rod-like or coccobacillus blood-loving bacterium in the family, Pasteurellaceae. H. influenzae, a non-toxin producing bacterium was first isolated during the 1890 influenza pandemic, and it is often referred to as a “blood-loving” bacterium (i.e. haemophilic bacterium) because it requires growth factors which are present in blood for growth. These growth factors are hemin (X factor) and NAD or NADP (V factor), and they play significant role in the growth of H. influenzae.

Haemin is required by H. influenzae to synthesize cytochromes, catalase and peroxidase while nicotinamide adenine dinucleotide (NAD)/nicotinamide adenine dinucleotide phosphate (NADP) is used as an electron carrier by the bacterium in its oxidation-reduction reaction. In particular, H. influenzae type b (Hib), an encapsulated strain is the chief cause of infections in humans especially children where it causes a variety of infections including bacteraemia, pneumonia, meningitis, and epiglottis. In adults, Hib cause cellulitis, emypyema, septic arthritis and other invasive infections. Hib causes the most severe infections in humans.

PATHOGENESIS OF HAEMOPHILUS INFLUENZAE INFECTION

The respiratory tract is often the major route via which Haemophilus influenzae type b (Hib) enters the body and cause infection. Though the pathogenesis of H. influenzae is not completely explicit, the capsular polysaccharide is believed to be the main driving force behind the microbe’s virulence. The capsular polysaccharide confers protection to H. influenzae from lysis by complement molecules and opsonization by phagocytes. Infections with capsulated H. influenzae can occur in children (under the age of 5 years) who lack capsular antibodies which protect the host against Hib infections.

Generally, Hib, the capsulated strain of Haemophilus causes the most life-threatening forms of infections in children below the age of 5 years old especially when it becomes invasive, affecting various tissues and causing illnesses that include bacteraemia, cellulitis, osteomyeltis, epiglottitis, arthritis and meningitis. However, Hib rarely cause disease in adult individuals and children above the age of 5 years. Pneumonia, Otitis media (ear infection), epiglottitis, bacteraemia, and meningitis are some of the infections caused by Hib in infants and children, and these can occasionally be caused by non-typable H. influenzae strains.

LABORATORY DIAGNOSIS OF HAEMOPHILUS INFLUENZAE INFECTION

H. influenzae requires complex nutritional growth requirements for growth (in particular: blood-containing culture media that supplies haemin and NAD/NADP). Sputum, pus, CSF, blood and nasopharyngeal specimens are often the main specimens required for laboratory investigation for H. influenzae infection. Chocolate agar is the best culture media for the growth of H. influenzae in the laboratory. Growth of H. influenzae occurs aerobically with slight CO2 tension (usually 5 %) at 35-37oC. Gram staining of specimens is also very useful when looking out for H. influenzae because smears demonstrate the pleomorphic and thread-like nature of the Gram-negative rods. Dilute carbol fuchsin instead of safranin (the known and widely used counterstain in Gram staining) is used as the secondary (counter) stain when Gram staining samples suspected to harbour H. influenzae. The demonstration of the requirement for V factor on blood agar is confirmatory for the presence of H. influenzae in a specimen. H. influenzae exhibit satelliting effect on blood agar and this phenomenon demonstrates requirement for V factor.

Image result for Haemophilus influenzae

Illustration of Gram stain of a sputum sample showing H. influenzae appearing as a Gram-negative coccobacilli.

Satellitism (satellite phenomenon) is the observable effect seen when certain growth-factor-requiring microbes (such as H. influenzae) can grow efficiently on a growth media which lacks the required growth factor but which supports the growth of another microorganism which can provide the required growth factor. S. aureus (a feeder organism in this case) is used in performing satellitism test because it supplies V factor into the growth medium, and this phenomenon allows satellite growth of colonies of Haemophilus species that requires NAD/NADP for growth. H. influenzae does not cause haemolysis on blood agar. Requirement for haemin (X) is investigated by the porphyrin test, and the presence of H. influenzae in the sample is confirmed if the result of the test is negative because H. influenzae does not synthesize porphyrin and haeme. Only Haemophilus species that do not require X factor for growth are known to be positive for the porphyrin test because such species synthesize haeme and porphyrins. Serological tests can also be employed to detect the serotypes of Haemophilus. H. influenzae is indole and urease positive when subjected to biochemical testing.  

Satellitism test for detection of  H. influenzae

TREATMENT OF HAEMOPHILUS INFLUENZAE INFECTION

The drugs of choice for the treatment of H. influenzae infections include ampicillin, chloramphenicol, sulphamethoxazoles, aminoglycosides, tetracyclines and some third generation cephalosporins including ceftazidime, cefotaxime, augmentin (amoxicillin-clavulanic acid) and ceftriaxone.

PREVENTION AND CONTROL OF HAEMOPHILUS INFLUENZAE INFECTION

Non-typable species of H. influenzae is a normal flora of the upper respiratory tract of humans. Thus, infections are likely to occur from these species when the immune system becomes weakened. However, Hib can be transmitted from infected persons to non-infected individuals through the respiratory tract as aerosols. Approved Hib conjugate vaccines should be administered to children especially those at higher risk of infection, and booster doses should be included following laid down medical guidelines. Proper vaccination of children from the age of 2 months and above against the infection will go a long way in minimizing mortality especially bacterial meningitis which is associated with H. influenzae infection. 

REFERENCES

Prescott L.M., Harley J.P and Klein D.A (2005). Microbiology. 6th ed. McGraw Hill Publishers, USA.

Madigan M.T., Martinko J.M., Dunlap P.V and Clark D.P (2009). Brock Biology of Microorganisms, 12th edition. Pearson Benjamin Cummings Inc, USA.

Balows A, Hausler W, Herrmann K.L, Isenberg H.D and Shadomy H.J (1991). Manual of clinical microbiology. 5th ed. American Society of Microbiology Press, USA.

Barrett   J.T (1998).  Microbiology and Immunology Concepts.  Philadelphia,   PA:  Lippincott-Raven Publishers. USA.

Basic laboratory procedures in clinical bacteriology. World Health Organization (WHO), 1991. Available from WHO publications, 1211 Geneva, 27-Switzerland.

Murray P.R, Baron E.J, Jorgensen J.H., Pfaller M.A and Yolken R.H (2003). Manual of Clinical Microbiology. 8th edition. Volume 1. American Society of Microbiology (ASM) Press, Washington, D.C, U.S.A.

Murray P.R, Baron E.J, Jorgensen J.H., Pfaller M.A and Yolken R.H (2003). Manual of Clinical Microbiology. 8th edition. Volume 2. American Society of Microbiology (ASM) Press, Washington, D.C, U.S.A.

Murray P.R., Rosenthal K.S., Kobayashi G.S., Pfaller M. A. (2002). Medical Microbiology. 4th edition. Mosby Publishers, Chile.

 

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