Title: Haemophilus and the HACEK Organisms
1Haemophilus and the HACEK Organisms
- Dr. John R. Warren
- Department of Pathology
- Northwestern University
- Feinberg School of Medicine
- June 2007
2Characteristics 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)
3Characteristics 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)
4Characteristics 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
5Most Common Species of Haemophilus
- Haemophilus influenzae
- Haemophilus parainfluenzae
- Haemophilus aphrophilus
- Haemophilus ducreyi
6Haemophilus 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)
7Haemophilus 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
8Haemophilus 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
9Haemophilus Modes of Infection
- Endogenous non-typeable strains of Haemophilus
influenzae associated with invasive infection in
debilitated adults - Haemophilus ducreyi sexually transmitted
10Haemophilus 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
11Haemophilus 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)
12Haemophilus 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
13Haemophilus 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)
14Haemophilus Types of Infectious Disease
- Haemophilus haemolyticus, H.
- parahaemolyticus, H. segnis,
- and H. paraphrophilus rarely
- produce human infection.
15Haemophilus 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
16Haemophilus 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
17Haemophilus 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
18Haemophilus Species Identification
- Growth requirement for X factor ? Haemophilus
aphrophilus (requirement lost upon
subcultivation)
19(No Transcript)
20Haemophilus 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
21Haemophilus 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
22Haemophilus 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
23Haemophilus influenzae Biotypes
- Indole Urease Ornithine
- I
- II
- III
- IV
- V
- VI
- VII
- VIII
24HACEK Organisms
- HHaemophilus aphrophilus
- AActinobacillus actinomycetemcomitans
- CCardiobacterium hominis
- EEikenella corrodens
- KKingella kingae
25HACEK 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)
26HACEK 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
27Recommended 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
28Recommended 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