Haemophilus, HACEK, Legionella & Other Fastidious Gram-Negative Bacilli Flashcards by Kyra Catedral

(members of the family Pasteurellaceae)

Haemophilus
Actinobacillus
Pasteurella
Aggregatibacter

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• Now dominant etiologies of infective endocarditis

Haemophilus, Actinobacillus, Pasteurella & Aggregatibacter

(members of the family Pasteurellaceae)

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, an emerging cause of brain abscesses

Aggregatibacter aphrophilus

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• Reside in the human oral cavity and nasopharynx;

some with enhanced capacity to cause ENDOCARDITIS.

HACEK - Haemophilus, Aggregatibacter, Cardiobacterium, Eikenella, & Kingella

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• 5 species inhabit human oral cavity,

2 species in oral cavities of dogs & cats

• Capnocytophaga

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• cause zoonoses

Pasteurella
Brucella
Francisella

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• GN pleomorphic, small coccobacilli in direct smears of clinical material to long filaments in stained smears of colony growth

Nonmotile, facultatively anaerobe, carbohydrate fermenters, generally oxidase+, generally catalase+, nitrate reducers

• Obligate parasites of the mucous membranes (URT) of humans & animals

Haemophilus species

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Haemophilus species

8 species:

H. influenzae
H. parainfluenzae
H. haemolyticus
H. paraphrohaemolyticus
H. pittmaniae
H. aegyptius
H. ducreyi
H. segnis (now renamed to Aggregatibacter segnis)

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2 former members,_____ were moved into the genus Aggregatibacter & combined in a single species - Aggregatibacter aphrophilus.

H. aphrophilus & H. paraphrophilus

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Greek word “Blood lover”

Require both or either growth factors present in the blood:

Haemophilus

• Hemin or hematin = X factor
• Nicotinamide adenine dinucleotide (NAD) = V factor

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Do not grow in pure culture on SBA

Haemophilus

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СНОС agar - lysing RBCs by heat releases X & V factors & inactivates NADases in sheep blood

Haemophilus

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Satellitism
a recognized phenomenon by…

H. influenzae

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• On SBA plates, tiny colonies of Haemophilus appears when mixed with Staphylococcus aureus/ Streptococcus pneumoniae/ Neisseria spp.

(exogenous sources of the V factor as by-product of metabolism & obtains X factor from the sheep blood).

Satellitism

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Haemophilus influenzae aka

Pfeiffer’s bacillus

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Haemophilus

• The major pathogen within the genus with wide pathogenic potential.

Haemophilus influenzae (Pfeiffer’s bacillus)

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Haemophilus influenzae (Pfeiffer’s bacillus)

Virulence factors:

• Capsule - most significant; antigenic types a, b, c, d, e, f
• IgA proteases
• Adherence (fimbriae, other structures)
• Outer membrane proteins & LPS

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H. influenzae

Capsule - most significant; antigenic types a, b, c, d, e, f
>__________ - most invasive infections in children before the widespread use of vaccines; now occasionally seen in over age 65 years
>________ - leading cause of meningitis among unvaccinated children

> ________ - composed of polyribitol phosphate
Not all strains are encapsulated -_______

H. influenzae serotype b (Hib)

Hib

Hib capsule

nontypable H. influenzae (NTHi)

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H. influenzae

Encapsulated Strains

Septicemia
Septic arthritis
Pericarditis
Pneumonia
Cellulitis

Meningitis
Osteomyelitis

Epiglottitis
Tracheitis

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H. influenzae

Non-encapsulated Strains

Otitis media with effusion
Conjunctivitis
Sinusitis
Bacteremia
Pneumonia

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Infections Associated with Other Haemophilus Spp.

“pinkeye disease,” an acute, contagious conjunctivitis

• H. aegyptius (Koch-Weeks bacillus)

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Infections Associated with Other Haemophilus Spp.

conjunctivitis in pediatric populations and Brazilian purpuric fever

• H. influenzae biogroup aegyptius

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Infections Associated with Other
Haemophilus Spp.

strictly human pathogen, agent of chancroid (soft chancre), a highly communicable sexually transmitted genital ulcer disease (GUD) [should test also for HIV, syphilis & herpesvirus]

• H. ducreyi

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Infections Associated with Other Haemophilus Spp.

otitis media, acute sinusitis, rarely endocarditis

Study These Flashcards

• H. parainfluenzae

Infections Associated with Other Haemophilus Spp. - pharyngitis

• H. parahaemolyticus

(Koch-Weeks bacillus)

H. aegyptius

Haemophilus: Laboratory Diagnosis Specimen processing & Isolation • Specimens:

blood CSF middle ear exudate joint fluids upper/lower respiratory tract specimens bronchial washings swabs from conjunctivae vaginal swabs abscess drainage

(2) , die rapidly. Prompt transportation and processing are vital.

• Haemophilus spp., H. ducreyi

Haemophilus: Laboratory Diagnosis Specimen processing & Isolation Primary medium: (agar, temperature, CO2); for respiratory specimens, supplement with______

CHOC agar, 33-37°C, 5%-10% CO2 bacitracin (300mg/L)

Haemophilus: Laboratory Diagnosis Specimen processing & Isolation - enriched CHOC medium or Nairobi biplate (half plate: GC agar base with 2% bovine hemoglobin and 5% fetal calf serum; the other half: MHA with 5% chocolatized horse blood; both contains 3mg/L vancomycin); 33°C for 7 days

• H. ducreyi

Haemophilus: Laboratory Diagnosis Specimen processing & Isolation - require special media enriched CHOC agar with 1% Iso VitaleX or Vitox; at least 4 days incubation.

• H. aegyptius

half plate: GC agar base with 2% bovine hemoglobin and 5% fetal calf serum; the other half: MHA with 5% chocolatized horse blood; both contains 3mg/L vancomycin

Nairobi biplate

Haemophilus: Laboratory Diagnosis Colony Morphology TRANSLUCENT, TANNISH, MOIST, SMOOTH, CONVEX, with a distinct "MOUSY" or BLEACHLIKE odor. Encapsulated strains larger, more mucoid than NTHi strains.

H. influenzae (and H. aegyptius) on CHOC:

Haemophilus: Laboratory Diagnosis Colony Morphology : tannish, drier, medium to large size; are non-hemolytic

• H. parainfluenzae

Haemophilus: Laboratory Diagnosis Colony Morphology : similar to H. parainfluenzae but beta hemolytic on horse or rabbit blood agar

• H. parahaemolyticus

Haemophilus: Laboratory Diagnosis Colony Morphology : tan or yellow, small, flat, smooth, transparent to opaque colonies

• H. ducreyi

Haemophilus: Laboratory Diagnosis Colony Morphology distinct "MOUSY" or BLEACHLIKE odor.

H influenczae

Haemophilus: Laboratory Diagnosis Microscopic Morphology • Often stains faint pink, with Gram stain: small, gram-negative coccobacilli in clinical specimens to long filaments in colonies • Capsules of_______ - clear, nonstaining halos in purulent secretions • (2) stains may help detect Haemophilus.

H. influenzae Acridine orange or methylene blue

Haemophilus: Laboratory Diagnosis Microscopic Morphology - Gram stain: ***school of fish or railroad tracks,*** fingerprints, pale-staining, small GN coccobacilli in chains

H. ducreyi

Haemophilus: Laboratory Diagnosis Laboratory Identification FIRST CLUE:

pleomorphic GN coccobacilli on CHOC, NO GROWTH ON SBA & MAC in pure culture.

• X and V growth factor test, porphyrin, oxidase, catalase, biochemicals, hemolysis • CSF detection for capsular antigens (type b capsular Ag, i.e., Directigen Meningitis Combo Test Kit); also test for beta lactamase • Automated commercial systems, ie, Haemophilus biotyping within 4 hours & HACEK identification by MALDI-TOF MS (matrix-assisted laser desorption/ionization-time-of-flight mass spectrophotometry

Haemophilus: Laboratory Diagnosis Laboratory Identification

Haemophilus: Laboratory Diagnosis Laboratory Identification X Factor and V Factor Requirements • Use of impregnated strips or disks • Care not to use X-factor containing medium to prevent erroneous results • H. influenzae may be mistaken for______ • H. influenzae do not require_____ when grown anaerobically but still require_____

H. parainfluenzae heme; NAD

Haemophilus: Laboratory Diagnosis X Factor and V Factor Requirements • Isolates identified based on the factors required for growth & presence of____ • Use of______ or _____ media, ______ and ______impregnated strips/disks. Incubate at_____ with CO2 for_____. Check for growth.

hemolysis MHA or TSA X and V 35C 24H

•__________ ATCC 35056 grows around ***XV-disk only.*** •__________ ATCC 7901 grows around ***V-disk and XV-disk.*** • __________ATCC 33940 grows around ***X-disk and XV-disk.***

Haemophilus influenzae Haemophilus parainfluenzae Haemophilus ducreyi

Haemophilus: Laboratory Diagnosis : XV no hemolysis : V no hemolysis : XV with hemolysis : V with hemolysis : X only no hemolysis

Hi Hpi Hh Hph Hd

Haemophilus: Laboratory Diagnosis Laboratory Identification • Alternative method for heme-producing species

Porphyrin Test

Haemophilus: Laboratory Diagnosis Laboratory Identification Porphyrin Test • Principle: Inoculate isolates on medium with_________ → incubate 35°C, 4 hours → organisms are able to convert ALA into_____________ (intermediates of X factor) → add _______________→ check under UV lignt (Wood lamp, 360nm) for__________color

S-aminolevulinic acid (ALA) porphyrins or porphobilinogen Kovac reagent (p-dimethylaminobenzaldehyde) reddish orange color

Positive porphyrin reaction

reddish orange color

CLSI abbreviated test guidelines for ID of commonly isolated bacteria. 1. GNB, coccobacilli from ***RT specimens*** 2. CS with ***larger than 1 mm*** colonies on CHOC 3. ***No growth or satellitism on SBA*** 4. ***NEGATIVE PORPHYRIN*** test → identified as

H. influenzae

Factor X +, V +, Porphyrin-

H. influenzae H. haemolyticus

Factor X-, V +, Porphyrin +

H. parainfluenzae H. parahaemolyticus H. paraphrohaemolyticus A. segnis

Factor X +, V-, Porphyrin-

H. ducreyi

Factor X-, V-, Porphyrin +

A. aphrophilus

Factor X +, V +, Porphyrin- HEMOLYSIS (Horse/ Rabbit)

H. influenzae (-) H. haemolyticus (+)

Factor X-, V +, Porphyrin + Hemolysis (Horse/ Rabbit)

H. parainfluenzae H. parahaemolyticus (+) H. paraphrohaemolyticus (+) A. segnis

Factor X +, V-, Porphyrin- Hemolysis

H. ducreyi (+/-)

Factor X-, V-, Porphyrin + Hemolysis

A. aphrophilus (-)

Haemophilus influenzae type b (Hib) vaccination recommended for…

children younger than 5 years old

Haemophilus, HACEK, Legionella & Other Fastidious Gram-Negative Bacilli Flashcards

(58 cards)