Test 3- Pathogenic Yeast

Question 1

Candida

Answer 1

• >200 species
• Candida albicans is the only significant animal pathogen
• Grows as budding yeast cell in culture and tissues • Hyphae or pseudophyphae may also be seen in tissues
• Culture
• No special requirements
• White, shiny, high‐convex colonies in 24‐48 hours • Grows well at 25°C and 37°C

Question 2

Candida albicans generals

Answer 2

Worldwide distribution • Natural Habitat

• Commensal of mucocutaneous areas • Intestinal Tract
• Genital Tract

• Endogenous infections due to:

• Immunosuppression
• Prolonged antibiotic therapy
• Concurrent infection

• Malnutrition

Question 3

Candida albicans- virulence factors

Answer 3

Candida albicans

• Virulence Factors and Pathogenesis
• Neuraminidase- influenza
• Proteases
• Aid in tissue invasion
• Glycoproteins
• Endotoxin‐like activity
• Yeast form colonizes epithelial surfaces
• Transition to (pseudo)hyphal form for tissue invasion
• Inflammatory response is usually neutrophilic (not granulomatous)

—————“Sour crop”

—————-Corrugated yellow‐gray necrotic pseudomembrane

Question 4

Answer 4

Candida albicans

Question 5

candida albicans dx

Answer 5

Candida albicans

• Laboratory Diagnosis

• ———Lesional scrapes
• ——— Centrifuged milk (mastitis)
• ——— Biopsy/Tissue in formalin (histopath)

• Microscopy

• ———Gram stain (appears purple‐blue)
• ———KOH preparation
• ——— Stained tissue sections (or cultured samples)

——————–Thin‐walled,oval,buddingyeastcells+/‐(pseudo)hyphae ———-“Germ tube” formation is characteristic

• Isolation and Identification
———- Colonies visible in 24‐72 hours

• ———White/cream, shiny, convex
• ——— “Beery” smell
• ———Specialized media available
• ———— CHROMagar
• —————— C. albicans, C. tropicalis, C. krusei

————- API‐yeast‐Ident- smell like beer

Question 6

Cryptococcus

Answer 6

>30 species

Cryptococcus neoformans is the only significant

animal (and human) pathogen

Spherical to oval, thin‐walled, budding yeast

Cells are surrounded by a large mucoid polysaccharide capsule

Daughter cells are single and connected to parent by narrow neck

• Four serotypes:
• A: C. neoformans var. grubii (Worldwide; soil/dust)
• B/C: C. neoformans var. gattii * (Associated with decaying eucalyptus wood)

• D: C. neoformans var. neoformans (Worldwide; soil/dust)

Question 7

Cryptococcus Serotypes

Answer 7

Cryptococcus neoformans

• Natural habitat

• Serotypes A (C. n. var. grubii) and D (C. n. var. neoformans):
• Worldwide in soil and dust
• Skin, mucous membranes, and intestinal tract of normal mammals and birds

• Yeasts are concentrated in pigeon feces (rich in creatinine)

• >1 year survivability
• Serotypes B and C (C. n. var. gattii):
• Subtropical and temperate areas
• Eucalyptus and gum trees, also other trees

Question 8

Cryptococcus neoformans pathogenesis and virulence factors

Answer 8

Cryptococcus neoformans- CATS WITH UPPER RESPIRATORY DISEASE

Pathogenesis
• Virulence is mainly associated with the large capsule

• Capsule is diminished in high salt/sugar concentration environments

———- Spores may be aerosolized

Productionofcapsuleinitiatedintissues

Anti‐phagocytic

Immunosuppressive

• Other virulence factors: • Phenoloxidase

• Melanin
• Anti‐oxidant
• Protects yeast in oxidants in phagolysosomes
• Phospholipase
• Mannitol
• Pathogenesis (cont’d)
• Lesions
• Rangefromgranulomastomyxomatousneoplasms
• Containcapsularslime,yeastcells,andinflammatorycells
• Route/Spread of infection
• Usuallyrespiratory
• Localized infection in nasalcavityandsinuses • Eventuallymayinvolvebrainandmeninges
• Subcutaneousgranulomas
• Species affected
• CATS:Rhinitis,nasalgranulomas,CNSinvolvement,

ocular/cutaneous infections

• Dogs:Respiratorytractinfections,someocular/cutaneous manifestations

Cattle:Mastitis(rare)

Horses:Nasalgranulomas

Koalas:Respiratorytractinfections,CNSinvolvement

Humans:Similartocats

Question 9

Answer 9

Cryptococcus neoformans

Question 10

C. neoformans- dx

Answer 10

Cryptococcus neoformans

• Laboratory Diagnosis
• BE CAREFUL! Zoonotic agent!
• Biosafety hood recommended
• Useful samples:CSF,exudates,milk,tissue/biopsies
• Directmicroscopy
• India ink or nigrosin stains (outlines capsule)
• Mayer’s mucicarmine (stains capsule and cell wall) • Isolation/Culture
• Blood or SabDex agar
• Chocolate agar for enhanced capsule production

Streak as for bacteria, 48 hours – 2 weeks

Aerobic, 37°C (other Crypto. species will not tolerate 37°C)

Smooth, moist, shiny colonies; white to yellowish

Mucoid growth with age

Capsule is less pronounced from culture

Melanin is produced on birdseed agar

• Immunologicalandmoleculardiagnosticsexist

• Treatment
• Fluconazole,itraconazole • 5‐fluorocytosine
• AmphotericinB

Question 11

Malassezia generals

Answer 11

Malassezia

20 species

Commensal organisms on oily areas of skin and

ears of dogs/cats

• Malassezia pachydermatis is the most significant species in animals

• Otitis externa
• Seborrheic dermatitis

• Feline chin acne
• Small, bottle‐ or peanut‐shaped yeast (broad‐ based bud)
• Reproduce by bud fission (septum detachment)

Question 12

Malassezia pachydermatis

pathogenesis and dx

Answer 12

Malassezia pachydermatis

• Pathogenesis

Predisposing factors
• Immunosuppression
• Allergic dermatitis
• Hairyand/or pendulous ears

• Virulence factors
•Zymogen(activates complement, damages keratinocytes, inflammation/pruritis)
• Proteases(damageearcanalmucosa)
Lipases(adjust the sebum consistency to a more favorable environment for yeast)- why it can live in ears

• Leads to more sebaceous secretions

Laboratory diagnosis

• Interpretcarefully(normalflora?)
• Consider clinical signs and number of organisms
• Directmicroscopy(Gramstainormethyleneblue) • Isolation
• SabDex agar, 35‐37°C
• Colonies are small, smooth, “wet‐dog” odor
• Moleculartechniques(PCR)allowspeciesdifferentiation(ifneeded)

Question 13

Dimorphic Fungi

Answer 13

Sporothrix, Blastomyces, Coccidioides, Histoplasma

Question 14

Dimorphic Fungi growth forms

Answer 14

• Present two growth forms
• Appear as a mold when growing saprophytically in the environment
• Or in culture at 25‐30°C
• Usually the more stable form
• Appear as a yeast in animal tissues
• Or in culture/enriched media at 37°C
• May cause deep or systemic mycosis in animals and humans
• Cultures are biohazardous to laboratory workers. Be careful!

Question 15

Sporothrix schenkii

Answer 15

• Disease
• Sporotrichosis
• “Rose‐handler’s disease”
• Distribution
• Worldwide (especially tropics and subtropics)
• Hosts
• Horses, dogs, cats, humans
• Natural Habitat
• Wooden posts, rose thorns, dead vegetation, soil

• Site of Lesions
• Subcutaneous nodules, Lymphocutaneous ulcerations

• Lymphatics (especially horses)

Question 16

Answer 16

Sporothrix schenkii

Question 17

Answer 17

Sporothrix schenkii

Question 18

Sporothrix schenkii morphology

Answer 18

• Colonial morphology
• Grows well on SabDex agar
• White to cream colonies, wrinkled with aerial hyphae
• Becomes dark and leathery
• Microscopic appearance

37°C Culture- CIGAR SHAPED

Cytology

• Plentiful yeast seen with methylene blue stain (especially samples from cats)
• Appearance varies depending on culture conditions (cigar‐shapes)

Question 19

Answer 19

Sporothrix schenkii

Plentiful yeast seen with methylene blue stain (especially samples from cats)

Question 20

Disease Patterns of S. schenkii

Answer 20

• Dogs, Horses
• Cutaneous/subcutaneous pyogranulomatous inflammation and ulceration at site of inoculation

Infectious process follows subcutaneous lymph channels with suppurative ulcers at intervals

• Disseminated disease is rare in immunocompetent
• Cats
• Similar to dogs initially, but disseminated disease may develop regardless of immune status
• Dissemination to viscera, joints, bones, CNS

Question 21

• Epidemiology of S. schenkii

Answer 21

Typically acquired from the non‐living environment- mechanical transmission

• Zoonotic transmission from cats to humans occurs

Question 22

Laboratory Diagnosis of S. schenkii

Answer 22

Direct examination of exudates

May be difficult to distinguish from Histoplasma or Cryptococcus

Culture

Immunological tests (AGID, latex agglutination)

Molecular Tests (PCR)

Question 23

Treatment of S. schenkii

Answer 23

Sodium or Potassium Iodides

Amphotericin B, flucytosine (disseminated forms)

Ketoconazole, itraconazole

Terbinafine (cutaneous forms)

No vaccines available

Question 24

Blastomyces dermatitidis

Answer 24

• Disease
• Blastomycosis
• Distribution
• Eastern North America (sporadically Europe, India, Middle East)

• Hosts
• Dogs, cats, horses, humans

• Natural Habitat
• Acidic soil rich in organic material
• Site of Lesions
• Primary lesions in lungs
• Metastasis to skin and other organs

Question 25

Answer 25

Blastomyces dermatitidis

Question 26

Blastomyces dermatitidis microscopic appearance

Answer 26

Blastomyces dermatitidis * Colonial morphology * Grows well on SabDex agar (2‐4 weeks) * Small colonies, white cottony aerial hyphae • Becomes gray or dark brown • Microscopic appearance 37°C Culture 25°C Culture * Large round/oval, thick‐walled yeast cells; broad‐based buds * Appearance varies depending on culture conditions

Answer 27

Disease Patterns