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Haemophilus and the HACEK Organisms

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Title: Haemophilus and the HACEK Organisms


1
Haemophilus and the HACEK Organisms
  • Dr. John R. Warren
  • Department of Pathology
  • Northwestern University
  • Feinberg School of Medicine
  • June 2007

2
Characteristics of Haemophilus
  • Small, pleomorphic gram-negative coccobacilli
  • Positive for cytochrome oxidase
  • No growth on MacConkey agar
  • Growth in culture requires exogenous hemin
    (oxidized ferroprotoporphyrin) (X factor) and/or
    nicotinamide adenine dinucleotide (NAD) (V
    factor)

3
Characteristics of Haemophilus
  • Sheep blood contains NADase, a V
    factor-destroying enzyme, and consequently
    clinical strains of Haemophilus do not grow on
    standard 5 sheep blood agar
  • Heating sheep blood agar at 80oC denatures NADase
    and heat-lysis of red cells releases free NAD
    (chocolate agar)

4
Characteristics of Haemophilus
  • Growth of Haemophilus on chocolate agar in the
    presence of 5-10 CO2 (capnophilia)
  • Contemporary chocolate agar is a synthetic mix
    of NAD, hemoglobin, vitamins (cobalamin, thiamine
    hydrochloride), minerals (iron, magnesium),
    cysteine, glutamine, and glucose

5
Most Common Species of Haemophilus
  • Haemophilus influenzae
  • Haemophilus parainfluenzae
  • Haemophilus aphrophilus
  • Haemophilus ducreyi

6
Haemophilus Natural Habitats
  • Normal inhabitant of the upper respiratory,
    gastrointestinal, and genital tracts of humans
    except Haemophilus ducreyi
  • Haemophilus ducreyi found only in humans during
    disease (not normal microbial flora)

7
Haemophilus Modes of Infection
  • Encapsulated strains of Haemophilus influenzae
    associated with invasive infection caused by
    person-to-person spread of H. influenzae due to
    inhalation of infectious respiratory droplets
  • Type b H. influenzae was most commonly associated
    with disease prior to conjugate vaccine but
    prevalence has declined with advent of vaccination

8
Haemophilus Modes of Infection
  • Prevalence of invasive type b H. influenzae
  • disease (meningitis, epiglottitis, pneumonia,
  • septic arthritis, osteomyelitits, facial
  • cellulitis) in children less than 5 years of
  • age decreased 95 in the United States
  • from 1987 to 1993 (41 cases per 100,000 to
  • 2 cases per 100,000) due to childhood
  • vaccination

9
Haemophilus Modes of Infection
  • Endogenous non-typeable strains of Haemophilus
    influenzae associated with invasive infection in
    debilitated adults
  • Haemophilus ducreyi sexually transmitted

10
Haemophilus Types of Infectious Disease
  • Encapsulated (types a-f) strains of H. influenzae
    produce invasive infection (pneumonia,
    meningitis, epiglottitis, and bacteremia)
  • Unencapsulated (non-typeable) strains of H.
    influenzae cause otitis media in children, and
    lower respiratory tract infections (acute
    tracheobronchitis, pneumonia) in children and
    adults

11
Haemophilus Types of Infectious Disease
  • Haemophilus influenzae biogroup aegyptius
  • (formerly H. aegyptius) lacks a capsule but
  • capable of invasive infection, including
  • Purulent conjunctivitis
  • Sepsis syndrome in children (1-4 years of age)
    following purulent conjunctivitis with mortality
    as high as 70 (Brazil and Australia)

12
Haemophilus Types of Infectious Disease
  • Respiratory infection by non-typeable H.
    influenzae associated with underlying conditions
    (cystic fibrosis, chronic obstructive pulmonary
    disease, malignancy)
  • H. parainfluenzae of low virulence with sporadic
    cases of endocarditis and bacteremia

13
Haemophilus Types of Infectious Disease
  • Haemophilus aphrophilus an uncommon cause of
    slowly progressive (subacute) endocaridits and
    brain abscess (member of the HACEK group)
  • Haemophilus ducreyi causes chancroid, a sexually
    transmitted disease (400 cases annually in the
    US) with shallow and painful genital ulcers
    associated with inguinal lymphadenitis (bubo
    formation)

14
Haemophilus Types of Infectious Disease
  • Haemophilus haemolyticus, H.
  • parahaemolyticus, H. segnis,
  • and H. paraphrophilus rarely
  • produce human infection.

15
Haemophilus Species Identification
  • Preliminary findings small faintly-staining
    (with safranin) gram-negative coccobacillary to
    filamentous rods that grow on chocolate agar but
    not sheep blood agar (except H. aphrophilus that
    with passage in culture grows on both)
  • X and V factor requirements determined using X,
    V, and XV factor impregnated paper strips on
    Mueller-Hinton agar

16
Haemophilus Species Identification
  • Growth requirement for X and V factors, no ?
    hemolysis on horse blood agar ? Haemophilus
    influenzae
  • Growth requirement for V factor, no ? hemolysis
    on horse blood agar ?Haemophilus parainfluenzae

17
Haemophilus Species Identification
  • Growth requirement for X and V factors, ?
    hemolysis on horse blood agar ? Haemophilus
    haemolyticus
  • Growth requirement for V factor, ? hemolysis on
    horse blood agar ?Haemophilus parahaemolyticus

18
Haemophilus Species Identification
  • Growth requirement for X factor ? Haemophilus
    aphrophilus (requirement lost upon
    subcultivation)

19
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20
Haemophilus Species Identification Using the
Quad Plate (Remel)1
  • Quad 1-Horse Blood Agar
  • Quad 2-X Factor 1ßß-hemolysis, Noßno
  • Quad 3-V Factor ß-hemolysis, ggrowth,
  • Quad 4-XV Factor ngno growth
  • 1 2 3 4
  • H. influenzae Noß ng ng g
  • H. haemolyticus ß ng ng g
  • H. parainfluenzae Noß ng g g
  • H. parahaemolyticus ß ng g g

21
Haemophilus Species Identification Using
Biochemicals1
  • X V Hem Glu2 Suc2 Lac2 Man2
  • H. inf
  • H. para
  • H. haem
  • H. aph3
  • 1XX factor, VV factor, Hemß-hemolysis on horse
    blood agar
  • 2Glucose (Glu), sucrose (Suc), lactose (Lac), and
    mannose (Man) broth supplemented with X and V
    factors
  • 3X factor requirement often lost upon
    subcultivation

22
Haemophilus influenzae Biotypes
  • Biotypes II and III associated with respiratory
    tract, middle ear, and eye infections
  • Biotype III includes Haemophilus influenzae
    biogroup aegyptius (Koch-Weeks bacillus)
  • Biotype IV associated with genital infections,
    neonatal sepsis (mortality 50-90) and postpartum
    sepsis associated with endometritis
  • Most serotypes a, b, and f strains biotype I
  • Serotype c strains usually biotype II
  • Serotype d and e are biotype IV

23
Haemophilus influenzae Biotypes
  • Indole Urease Ornithine
  • I
  • II
  • III
  • IV
  • V
  • VI
  • VII
  • VIII

24
HACEK Organisms
  • HHaemophilus aphrophilus
  • AActinobacillus actinomycetemcomitans
  • CCardiobacterium hominis
  • EEikenella corrodens
  • KKingella kingae

25
HACEK Organisms
  • Slow growing, capnophilic, gram-negative bacteria
    that cause endocarditis
  • Clinical suspicion of endocarditis with negative
    blood cultures after standard 5 days of
    incubation (incubate blood cultures for 2 weeks)

26
HACEK Organisms
  • H A1 C2 E3 K4
  • Mac /5 /
  • Oxidase /
  • Catalase
  • Indole
  • Nitrate
  • Glucose
  • Mannitol /
  • Sucrose
  • 1Colonies show central opaque dot that with
    incubation forms a star-like configuration like
    crossed cigars visible on clear agar medium
    such as brain heart infusion (supplemented with
    serum) at 100X magnification may show light
    growth on MacConkey agar
  • 2Irregularly-staining gram-negative rods with
    bulbous (swollen) ends indole detected by xylene
    extraction
  • 3Cultures smell of hypochlorite (bleach)
  • 4Colonies show small but distinct zones of
    ß-hemolysis
  • 5X-factor requirement lost with passage in culture

27
Recommended Reading
  • Winn, W., Jr., Allen, S., Janda, W., Koneman,
  • E., Procop, G., Schreckenberger, P., Woods,
  • G.
  • Konemans Color Atlas and Textbook of
  • Diagnostic Microbiology, Sixth Edition,
  • Lippincott Williams Wilkins, 2006
  • Chapter 9. Miscellaneous Fastidious
    Gram-Negative
  • Bacilli

28
Recommended Reading
  • Murray, P., Baron, E., Jorgensen, J., Landry,
  • M., Pfaller, M.
  • Manual of Clinical Microbiology, 9th
  • Edition, ASM Press, 2007
  • Kilian, M. Chapter 41. Haemophilus.
  • von Graevenitz, A., Zbinden, R., and Mutters, R.
    Chapter 40. Actinobacillus, Capnocytophaga,
    Eikenella, Kingella, Pasteurella, and Other
    Fastidious or Rarely Encountered Gram-Negative
    Rods
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