Genus Haemophilus

Non-Motile; Gram-Negative Bacilli ranging from coccobacilli to filamentous rods; the more mature they are the more pleomorphic they are.

DO NOT GROW ON SHEEP BLOOD AGAR; only species that does this

Fastidious: require one or both of the accessory growth factors, called X Factor (Heme; heat stable) and V Factor (NAD: nicotinamide adenine dinucleotide)

Nitrate Reduction Positive

Members of indigenous microflora of oral cavity and upper respiratory tract

HACEK

Acronym of the first initial of each genus that belong in the group:
Haemophilus aphrophilus

Actinobacillus actinomycetemcomitans

Cardiobacterium hominis

Eikenella corrodens - pitts the agar; smells like bleach

Kingella species

Found as indigenous microflora of the oral cavity and can cause a certain type of endocarditis

Haemophilus influenzae

Small, Gram-Negative Bacilli or coccobacilli

One of the leading causes of meningitis, ear infections, and sinus infections.

Satellites around S. aureus and some other colonies on sheep agar

Porphyrin synthesis (δ-aminolevulinic acid test) NEGATIVE

Requires BOTH X AND V FACTORS

Grows on Chocolate agar in 5% CO₂; DOESN'T GROW ON SHEEP AGAR

Haemophilus parainfluenzae

Most common Haemophilus spp. in the indigenous microflora of the upper respiratory tract

Colonies are flat, grayish, semiopaque, and either smooth or rough and wrinkled after 24 hours incubation

X Factor Negative; Requires V Factor (Positive)

ALA - Porphryrin Test Positive

Beta-Hemolysis Negative

H. influenzae biogroup aegyptius

(H. aegyptius)

Indigenous microflora of the upper respiratory tract

Associated with acute purulent & contagious form of conuuntivitis (pink eye)

Especially fastidious

Very small, smooth colonies

Requires BOTH X and V FACTORS

ALA - Porphryrin Test Variable (+/-)

Beta-Hemolysis Negative

H. haemolyticus

Indigenous microflora of upper respiratory tract

Not pathogenic

REQUIRES BOTH X and V FACTORS

ALA-Porphryrin Test Negative

Beta-Hemolytic POSITIVE

H. parahaemolyticus

Indigenous microflora of upper respiratory tract

Not pathogenic; associated with pharyngitis, lower respiratory tract infections, and oral abscesses

REQUIRES X FACTOR; V Factor Negative

ALA-Porphryrin Test Positive

Strongly Hemolytic; large zones of hemolysis

H. aphrophilus

Long, slender Gram-Negative bacillus

Microflora of oral cavity (found in dental plaque), blood and Meninges

Associated with brain abscesses, postoperative infections, arthritis

Rough, raised colonies rarely exceeding 1 mm in diameter

REQUIRES NEITHER X and/or V FACTORS

ALA-Porphryrin Test Positive

Beta-Hemolysis Negative

H. ducreyi

Causes CHANCROID a sexually transmitted disease in Asia, Africa, Latin America

Especially Fastidious

Grows best at 33°C

Produces long parallel rows of small bacilli in chains; school of fish, railroad track or fingerprint appearance)

Smooth, semitranslucent, gray

Cohesive colonies can be pushed across the agar

REQUIRES X FACTOR; V Factor Negative

ALA-Porphryrin Test Negative

Beta-Hemolysis Negative

Capnocytophaga

Gram-Negative bacilli

Require an increased CO₂ environment (5-10%)

Flora of oral cavity; significant cause of endocarditis; opportunists in immunocompromised patients

Colonies have irregular edges and finger-like projections (gliding or sliding motility) that appear as a thin film surrounding the central part of the colony.

Colonies may have a moist, mottled, or "sweaty" appearance

Colonies adhere to agar surface and most are non-hemolytic

Capnocytophaga

(Continued)

Catalase Negative

Oxidase Negative

Indole Negative

Urease Negative

Produce Negative decarboxylase tests

Capnocytophaga spp. associated with infected dog/cat bite wounds are: C. canimorsus and C. cynodegmi;

Oxidase Positive                                  Catalase Positive

Bordetella pertussis

 Small Gram-Negative coccobacillus that appears singly or in pairs

Etiologic agent (cause) of whooping cough (very contagious)

Preferred Media: Bordet-Gengou and Regan-Lowe (CHB) mediums

Colonies look like mercury droplets or partially imbedded pearls

SHOULD BE INCUBATED AT 35°c IN ROOM AIR FOR MIN. OF 7 DAYS

Catalase Positive                              Oxidase Positive

Nitrate Negative                               Urease Negative

Non-Motile               NO GROWTH ON BAP OR MAC

Erythromycin = drug of choice for treatment & prophylaxis of pertussis

Pasteurella Species

General Characteristics:

Colonizes mucous membranes of the upper respiratory tract and gastrointestinal tracts of mammals or birds

Human infections occur from bites and scratches infected by animals (mostly dog and cat bites

Penicillin is the drug of choice for treating Pasteurella infections       Most common isolated species is Pasteurella multocida

Pasteurella multocida

Blood/Chocolate Agar: small, smooth, convex colonies; "Musty Odor"

May be mucoid and watery when isolated from respiratory secretions

NO GROWTH ON MAC

Very small, Gram-Negative Rods with "safety pin" (bipolar) appearance

Bipolar staining with Giemsa or methylene blue

Oxidase Positive                               Catalase Positive

Indole Positive                                 Ferments Glucose

Quick screening is their penicillin susceptibility (unusual for GNB)

Legionella Species

Gram-Negative Rods found, appearing, or present everywhere

Fastidious, aerobic bacteria:

* Unable to grow on blood agar and require the amino acid L-cysteine for growth.

Buffered Charcoal Yeast Extract (BCYE) = preferred agar

Acquired through inhalations of aerosols by humans

Febrile disease w/pneumonia (Legionnaire's disease) and extrapulmonary involvement

Pontiac Fever (without pulmonary involvement)

Asymptomatic infection

L. pneumophilia = the species most commonly associated with human disease

Legionella pneumophila

Motile, faintly staining Gram-Negative Bacilli; range in appearance from small coccobacilli to long filaments

One of the etiologic agents (causes) of legionellosis (Legionnaires disease)

Requires cysteine for in vitro growth

Will grow on Chocolate Agar; will not grow on standard culture media

Buffered Charcoal Yeast Extract (BCYE) is primary agar medium used; usually supplemented with α-ketoglutarate, L-cysteine, and ferric pyrophosphate

Colonies are whitish green

L. pneumophila

Identification Methods

3 Rapid methods for identifying Legionella isolates approved by FDA:

* Direct fluorescent antibody (DFA)

* DNA detection

*Urine antigen Testing

Usual basis for Identification Include:

1) cellular morphology (thin, weakly staining Gram-Negative bacilli or filaments)

2) their requirement for cysteine

3) their non-fermentative metabolism

4) serotyping by slide agglutination or direct fluorescent antibody stain

Francisella tularensis

Tiny, faintly staining Gram-Negative coccobacilli

Non-Motile

Etiologic agent (cause) of tularemia (aka: tick fever, deerfly fever, and rabbit fever) Hunters especially with contact w/infected animals

Potential bioterrorism land biological warfare agent

Grows poorly, if at all, on ordinary media

Optimal growth with: media supplemented with sulfhydral compounds, such as cysteine, cystine, thio-sulfate, and Iso Vitalex

Causes acute febrile granulomatous disease with rapid onset and flu-like symptoms