Exam 2 - Haemophilus and other fastidious GNB

What does fastidious mean?

• The organism has very specific growth requirements

• It does not grow easily on regular lab media

• Fastidious organisms need special nutrients, conditions, or growth factors

HACEK is an acronym representing several bacteria that

• are fastidious Gram-negative organisms

• are part of normal oral flora

• can cause bacterial endocarditis (infection of the heart valves)

  • These bacteria usually live in the mouth and throat, but if they enter the bloodstream (for example during dental work), they can infect the heart valves.

HACEK - H

Haemophilus

HACEK - A

Aggregatibacter

HACEK - C

Cardiobacterium hominis

HACEK - E

Eikenella corrodens

HACEK - K

Kingella

Haemophilus general characteristics

• Small Gram-negative coccobacilli or bacilli

• Small pleomorphic (can vary in shape)

• Non-motile

• Facultative anaerobes

• Oxidase positive, except H. ducreyi

• Catalase positive

• Normal flora of the upper respiratory tract

Haemophilus Family name

Pasteurellaceae

• 13 species: 8 associated with humans

  • e.g. Haemophilus influenzae, H. aegypytius, H. ducreyi, H. parainfluenzae

Virulence Factors of Haemophilus influenzae

The Capsule:

• protects bacteria from phagocytosis

• helps them evade the immune system

• There are 6 capsule types: a, b, c, d, e, f

Most dangerous Haemophilus influenzae virulence factor

Type b (Hib)

• This type causes invasive infections in unvaccinated populations

• has unique anti-phagocytic properties

Non-capsulated strains are

NTHi (non-typeable Haemophilus influenzae)

IgA protease

allows for colonization

• only produced by Haemophilus influenzae

Localized H. influenzae infections

Non-typable H. influenzae (non-capsulated strains)

Invasive H. influenzae disease

• Caused by encapsulated (Hib) strains

• General risk factors

  • Asplenic

  • Sickle cell disease

  • Complement deficiencies

  • Other immunocompromised states

Localized infections or Invasive disease transmission is from

colonizing strains or through respiratory droplets

Haemophilus influenzae Clinical infections (Invasive)

• Meningitis (usually caused by HiB)

• Acute epiglottitis (usually caused by HiB)

• Pneumonia

• Bacteremia

• Septic arthritis

Haemophilus influenzae - Meningitis

• Common in children 6 months – 2 years

• Symptoms: headache, stiff neck, sensitivity to light

• Complications can include: hearing loss, paralysis, brain damage, retardation, hydrocephalus, ataxia.

• Decreased Incidence with vaccine

Haemophilus influenzae - Acute epiglottitis

• Common in Children 2-5 years old

• Symptoms: Fever, sore throat, hoarse, “barking” cough

• Rapid disease progression

• This condition can block the airway, making it a medical emergency.

Haemophilus influenzae - Pnemonia

• Reduction of mucosal clearing of organism

• Smoking, COPD, viral infection

Haemophilus influenza - Non-invasive clinical infections

These are localized infections usually caused by non-encapsulated strains

• Otitis media (ear infection)

• Sinusitis

• Bronchitis

Treatment for Haemophilus influenzae

• Invasive: 3rd generation cephalosporin

• Non-invasive: 25% produce beta-lactamase, Beta-lactam/beta-lactmase inhibitor

H. aegyptius

Acute, contagious conjunctivitis (“Pink Eye”)

• Closely related to H. influenzae

H. ducreyi

• Causes agent of Chancroid

• This is a STD - genital ulcer disease

• Soft swollen chancre lymph nodes (buboes) common

• Rare in the US but common in: Africa, Asia, Latin America

• Women more asymptomatic then men

H. parainfluenzae

• Normal flora of the mouth and throat

• rare but can cause otitis media (ear infection) and sinusitis

• Rarely associated with bacterial endocarditis

Growth Requirements of Haemophilus

• X Factor (Hemin)

• V Factor (NAD)

• Chocolate agar

• Satellitism

X factor

Hemin

• Directly available in blood agar

V factor

Nicotinamide adenine dinucleotide (NAD)

• Within intact red blood cells

• V factor dependent organism will NOT grow on blood agar

• Will GROW on chocolate agar due to the lysed RBCs releasing V factor

Haemophilus - Satellitism

small colonies around colony of other organism, S. aureus

• S.aureus produces V factor as by product alpha-toxin, this causes satellite colonies around the S. aureus colony

Haemophilus species - Specimen collection

• Blood, CSF, swabs (ears and eyes)

• These organisms die quickly, so samples must be processed fast

Haemophilus species - Colony morphology

• Chocolate agar: Grayish/tan, smooth, moist

• Blood agar: No growth, or satellite colonies

Haemophilus species - Culture requirements

• Chocolate agar is media of choice

• 35 degrees C + increased CO2 content

Haemophilus species - Microscopic morphology

• Small pleomorphic Gram-negative coccobacilli

• They often stain faintly, so sometimes you must examine the edge of the smear

• May see capsule in primary smear

Requires both X and V factor

H. influenzae

Requires only V factor

H. parainfluenzae (V and XV)

Porphyrin (ALA Test)

• Detects ability of organism to convert ALA into porphyrins

• Helps identify Haemophilus species

• If porphyrins are detected, they glow under UV light.

• This indicates the organism does not require X factor (hemin) to grow

  • Advantage: This test is quicker than growth tests (about 4 hours).

  • Disadvantage: Identification of H. influenzae based off a negative result

Aggregatibacter aphrophilus

• Most prevalent HACEK organism

• Found in dental plaque

• Does not require CO2, but grows better with it

• Endocarditis

Aggregatibacter actinomycetemcomitans

• Oral flora

• Does not require CO2, but grows better with it

• Slow growing, >24 hours to observe

• Star shaped in center of colony after 48 hours

• Typical animal pathogen

• Periodonitis

Cardiobacterium hominis

• “Rosettes” on Gram stain

• Normal flora in nose, throat, and mouth

• Grow slowing on blood and chocolate

Eikenella corrodens

• Wound infections from human bites or fights

• Normal flora of oral and bowel

• Endocarditis associated with poor dental hygiene or oral surgery

• Cellulitis in IV drug users

• Growth

  • Colonies “pit” agar, Bleach-like odor, greening around colonies

Kingella

• Colonize upper respiratory tract/tonsils

• Poor dental hygiene or oral surgery associate with infection

• Bone and joint infections in children (<3 years old)

  • Associated with daycare disease outbreaks

Kingella morphology

Rod with square ends, resist decolonization and may stain purple

Capnocytophaga spp.

• Normal oral flora in humans or animals

• Infections subsequent to dog/cat bites

• Long thin gram-negative bacilli with tapered ends

• Common in Neutropenic patients

Most common human pathogen

Pasteruella multocida

Pasteurella spp.

• Normal oral flora of birds and mammals

  • Infections usually result of an animal bite

• Growth on blood and chocolate agar

• Does NOT grow on MacConkey

• Small gram-negative coccobacilli

• Oxidase and Catalase Positive

Brucella spp.

Normal flora in animals

Brucella Transmission

• Direct via work with animals or animal products

• Indirect via consumption of contaminated food

• Laboratory acquired

Brucella Disease

Brucellosis

• 7-21 days after exposure

• Malaise, fever chills, body aches, headache

• Severe: arthritis, orchitis, depression, endocarditis

4 common species of Brucella are

human pathogens

B. melitensis

sheep and goats

B. abortus

cattle

B. suis

swine

B. canis

dogs

Brucella laboratory diagnosis

• Considered to be an agent of bioterrorism

• Rule out testing

  • Oxidase and Catalase positive

  • Urea positive

Francisella

Normal flora in wild animals (rabbits most common)

• F. tularensis is most common human pathogen

Francisella Transmission

• Direct via work with animals or animal products

• Bite or scratch from infected animal

• Tick bites

• Deer files

• Laboratory acquired

Francisella Disease

Tularemia

• Clinical presentation depends on route of infection

  • Most common form is ulceroflandular

    • Ulcer at site of inoculation, followed by swelling of regional lymph nodes

Francisella laboratory diagnosis

• Blood most common specimen

• Slow growing

• Growth on chocolate

• Small pleomorphic Gram-negative coccobacilli

• Considered to be an agent of bioterrorism

Francisella Rule out testing

• Oxidase negative

• Catalase weakly positive

• Urea negative

Bordetella has 2 clinically significant human isolates

Bordetella pertussis and Bordetella parapertussis

Bordetella spp.

• Obligate aerobe

• Small Gram-negative bacilli or coccobacilli

Bordetella pertussis virulence factor

Slider 34