Pasteurellaceae: Pasteurellaceae: Haemophilus, PasteurellaHaemophilus, Pasteurella

Fastidious Gram-Negative Fastidious Gram-Negative RodsRods

Pasteurellaceae: Genera • Small, gram(-) bacilli; pleomorphic

(cocci, filamentous rod)• Medically important genera:

– Haemophilus - human oropharynx NF; infection of unimmunized children

– Pasteurella - animal oropharynx NF; bite wound infection, RTI, bacteremia, meningitis

– Aggregatibacter (former Actinobacillus) - human oral NF; periodontitis, endocarditis, bite wound

Haemophilus: Characteristics

• Aerobic, facultative anaerobe• Optimal growth 35-370 C• Atmosphere of 5-10% CO2• Catalase(+)• Oxidase(+)• Nonmotile• Most fastidious, require enriched media• Susceptible to drying and chilling;

autolytic• Clinical specimens should not be

refrigerated, processed immediately

Haemophilus: Characteristics

• Obligate parasites• Mucous membranes of human, animals• “blood-lover”• Most require growth factors

– X (hemin) factor - synthesis of iron containing respiratory enzymes (cytochrome, cytochrome oxidase, catalase, peroxidase)

– V (NAD) factor - coenzyme for oxidation-reduction

Haemophilus: CBA• Grow 1-4 days• Some β-hemolytic• Most grow poorly, or

not at all on CBA• Satellitism - grow as

small colonies on CBA around Staphylococcus aureus (hemolysis secrete NAD into medium, hemin diffuses from RBC)

Haemophilus: Chocolate Agar

• Grow best on highly enriched CHOC • Heat RBC 80°C, 15 min. - destroys

NADase, releases NAD into medium• Can make selective - add bacitracin,

vancomycin, clindamycin to eliminate NF

• Colonies grayish:– Capsulated strains = smooth– Non-encapsulated strains = rough

ID Haemophilus: X & V Factor Requirement

• Only V (NAD) = H. parainfluenzae• Both X (Hemin) and V (NAD) = H.

influenzae

Haemophilus influenzae: Lab ID

• Six serotypes (a-f)– by capsular polysaccharide – Type b most virulent (>95% of all invasive

infections)– Hib ~20,000 pediatric infections/year prior

to vaccine– Capsule contains polyribitol phosphate

(PRP); used for vaccine, elicits host protective antibody response

• Eight biotypes (I-VIII)– Encapsulated type I – pathogens, more

invasive disease– Nonencapsulated type II, III - opportunists

Haemophilus influenzae: Virulence

Factors• PRP capsule – protects against

phagocytosis• LPS – endotoxin; induces inflammation• Protease – IgA specific; facilitates

colonization of mucosal surfaces• Pili and outer membrane proteins –

adhesion, attachment

NonencapsulatedHaemophilus influenzae

• Colonize URT within first few months of newborn

• Opportunistic pathogen - spread locally and cause disease:– Otitis media– Sinusitis– Bronchitis, pneumonia – elderly, patients

with chronic pulmonary disease• Disseminated disease uncommon

EncapsulatedHaemophilus influenzae

• Infrequently found in URT as NF• Infect, able to penetrate nasopharynx

(NP) submucosal, into bloodstream• Common infection of infants and young

children

Encapsulated H. influenzae: Infection and Disease

• Meningitis:– Follow bacteremic spread from NP– Usually infants 3-18 months age– Generally preceded by 1-3 days URT disease– High mortality

• Epiglottitis:– Occurs in 2-4 year olds, mostly boys– Cellulitis, tissue swelling, life-threatening

emergency– Pharyngitis, fever, breathing difficulties– Progress to complete obstruction of airway

leading to death

Encapsulated H. influenzae Infection and Disease

• Cellulitis:– In very young children– Fever, reddish-blue color on cheek,

periorbital area• Arthritis:

– Usually children <2 years age– Infection of large joints, secondary to

bacteremia• Conjunctivitis:

– H. aegyptius– Acute, contagious conjunctivitis– Commonly called “pink eye”

Haemophilus ducreyi• DNA studies – not Haemophilus; but

related to Pasteurellaceae• STD – genital ulcers, soft chancre or

chancroid• Common in hot, tropical countries

(Africa, Asia); less common in Europe, North America

• Diagnosed in men, usually AS in women

• Following 5-7 days exposure, tender papule, progress to painful ulcer, inguinal lymphadenopathy

• Does not spread further• Autoinoculable, resulting in multiple

lesions

Haemophilus: Treatment and Prevention

• Prompt treatment or causes high mortality• Antibiotic resistance:

– ~30% penicillin (β-lactamase)– <1% chloroamphenicol (chloramphenicol

acetyltransferase)• Drug of choice – ceftriaxone (β-lactamase-

resistant cephalosporin), good CSF penetration

• For chancroid – treat with erythromycin• Purified HIB vaccine:

– 3 doses purified PRP-conjugate– Given 2-6 months of age– In USA, dramatically reduced disease in infants

Pasturella multocida: Characteristics

• Bipolar staining• Grows well on CBA, Chocolate agar• Poor growth on Mac and other media

for Gram(-) rods• Fermentative (glucose, sucrose), little

or no gas• TSI confusing because of weak acid

production = “sick” appearance

P. multocida: Infection and Disease

• Three forms of disease• Animal bite wound infection - localized

cellulitis and lymphadenitis• Exacerbation of chronic respiratory

disease of patients with underlying pulmonary dysfunction

• Systemic infection in immunocompromised patient (bacteremia, meningitis)

• Exquisitely sensitive to penicillin– 2 units of penicillin– may be used for presumptive ID)

• Tetracycline, chloramphenicol also used

Class Assignment• Textbook Reading: Chapter 18

Haemophilus and Other Fastdious Gram-Negative Rods - Haemophilus, Pasteurella (Omit HACEK Group, Capnocytophaga)

• Key Terms• Learning Assessment Questions

Case Study – H. influenzae

• A 78-year-old man confined to a nursing home awoke with a severe headache and stiff neck.

• Because he had a high fever and signs of meningitis, the nursing home staff took him to a local emergency department.

• The CSF specimen was cloudy.

Case Study – H. influenzae

• Analysis revealed 400 white blood cells per mm3 (95% polymorphonuclear neutrophils), a protein concentration of 75 mg/dl, and a glucose concentration of 20 mg/dl.

• Small gram-negative rods were seen on Gram stain of the CSF, and cultures of CSF and blood were positive for Haemophilus influenzae.

Case Study - Questions• 1. Discuss the epidemiology of H.

influenzae meningitis,and compare it with the epidemiology of meningitis caused by Streptococcus pneumoniae and by Neiserria meningitidis.

• 2. Compare the biology of H. influenzae strain that is likely to be the cause of this patient’s disease with the strains that historically caused pediatric diseases (prior to vaccination).

Case Study - Questions• 3. What other diseases does this

organism cause? What other Haemophilus species cause disease, and what are the diseases?

• 4. What diseases are caused by Pasteurella multocida? What is the source of this organism?