DIMORPHIC SYSTEMIC FUNGI

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DimorphicSystemicFungi.ppt
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Organisms Causing
Systemic Mycoses


Objectives - List two properties that the systemic dimorphics share.

Discuss conversion of the dimorphic yeasts

Identify from cases using colonial, morphological and geographic information, and diseases, the dimorphic fungi.

Differentiate the systemic dimorphic fungi from similar organisms previously discussed.

Describve the diseases, transmission, causative agents, of blastomycosis, paracocciodes, histoplamosis, and cocciomycosis.

Lab objectives

  • Describe any additional safety precautions needed when working with the systemic dimorphic fungi.

  • Using lactophenol cotton blue or tissue stains – identify Blastomyces dermatitidis, Paracocciodes barasilienses, Histoplasma capsulatum, and Cocciodes immitis.

  • Explain the method from converting these molds to yeast.
  • Describe exoantigen testing

Organisms

  • Histoplasma capsulatum
  • Blastomyces dermatitidis
  • Coccidioides immitis
  • Paracoccidioides brasiliensis
  • Pneumocytis carinii

Safety considerations

  • ALL handling of specimens and cultures MUST be performed in a biological safety cabinet
  • Do not open plates on benchtop
  • Plates should be sealed
  • If C. immitis is suspected, use tubed media instead of plates

Transmission and pathogenesis

  • Transmission is by inhalation of airborne conidia which are normally present in soil
  • Organisms are located in distinct geographical areas
  • Disease severity is dependent upon infective dose and patient’s immune status
  • Competent hosts: asymptomatic to mild respiratory disease
  • Compromised hosts: dissemination is common

Cultural characteristics

  • Very slow growing
  • Up to 6 weeks for growth (except C. immitis)
  • Must do direct examinations
  • Experimental: PCR testing of direct specimens
  • Colonial morphology varies with isolation media
  • Use both enriched (BHIA) and selective media
  • Classic morphology is described on SAB
  • Historically, definitive identification has been made by demonstrating both a yeast or tissue phase (at 370C) and a mold phase (at room temperature)

Methods of identification of
organism grown in culture

  • Exoantigen testing
  • DNA probe
  • Microscopic morphology
  • Conversion to yeast or tissue phase
  • Serological testing
  • Four-fold rise in antibody titer between acute and convalescent paired sera

Histoplasma capsulatum

  • Causes histoplasmosis (Darling’s or spelunker’s disease)
  • Distribution
  • Ohio and Mississippi River valleys, Appalachian mountains
  • Organism is spread by inhalation of airborne conidia
  • Organism multiplies in bird droppings and bat guano

Primary disease

  • 90-95% are asymptomatic or sub-clinical, with a self-limiting mild respiratory infection
  • High percent of population in endemic areas are skin test positive for organism
  • Acute pulmonary disease
  • Night sweats, cough, fever and weight loss
  • Some develop pulmonary cavitary lesions resembling TB
  • Organism is able to multiply in macrophages (observed as pseudoencapsulated yeasts)
  • Dissemination is rare in the immuno-competent

Disseminated disease

  • Immune competent patients
  • Chronic disease of the adrenals, liver, kidneys, skin, oral mucosa, eye and CNS
  • Treatment: amphotericin B (>90% successful)
  • Immunocompromised patients
  • Rapidly fatal disease
  • Organism can be found throughout body, even in peripheral blood monocytes

Direct specimen examination

  • Specimens
  • Sputum, blood, bone marrow, lymph nodes, biopsy of infected tissue
  • Wright Giemsa stain of blood/bone marrow
  • Intracellular clusters of round to oval yeasts with narrow necked budding
  • Seen in macrophages and histiocytes

Culture characteristics

  • 370C
  • Rough, mucoid cream to tan colonies
  • May need several transfers to convert from mold to yeast phase
  • Room temperature
  • Slow growing
  • Starts moist  white to brown aerial mycelium  glabrous, velvety or wooly colonies
  • Original isolates on BHIA should be transferred to SAB to observe characteristic morphologies

04Histoplasm

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mold

Microscopic morphology at 370C

  • Small, single budding yeast
  • Occasional abortive hyphae may be seen

Microscopic morphology at
room temperature

  • Smooth-walled or tuberculate macro-conidia formed at 900 to hyphae
  • Round or tear drop microconidia may be seen
  • Macroconidia may be confused with contaminants:
  • Sepedonium sp.
  • Chrysosporium sp.

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histo

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Sepedonium

Identification

  • Exoantigen testing
  • Conversion of mold to yeast phase
  • DNA probes and other molecular techniques

Blastomyces dermatitidis

  • Causes North American blastomycosis or Gilchrist’s disease
  • Distribution
  • South of the Ohio and east of the Mississippi Rivers
  • Transmission is by inhalation of conidia or hyphal fragments
  • Organism proliferates near riverbanks surrounded by rotting wood

Disease

  • Men are more often affected than women
  • Associated with outdoor occupations/activities
  • Begins as a respiratory infection
  • 40-50% remain asymptomatic
  • Progresses to chronic, suppurative and granulomatous infection
  • First disseminates to skin (ulcerated and crusted or weepy lesions) and bones (pain and loss of function)
  • Without therapy (amphotericin B or azoles), prognosis is poor

Direct specimen examination

  • Specimens
  • Sputum, skin biopsy, bone
  • Broad-based budding yeast with thick refractile walls
  • Organism does not survive well in clinical specimens
  • Culture immediately

Culture characteristics

  • 370C
  • Waxy, wrinkled, cream to tan colonies
  • Room temperature
  • Initially: yeast-like colony
  • Develops into a white cottony colony that turns tan with age
  • Conversion of mold to yeast phase takes 4-5 days

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Mold phase

08BDermatit

Microscopic morphology at 370C

  • Thick walled, broad based budding yeast
  • Daughter cell separates from mother cell when both are the same size

Microscopic morphology at
room temperature

  • Single, smooth walled conidia at ends of short conid-iophores or directly on hyphae
  • “lollipops”
  • Resembles:
  • Scedosporium sp.
  • Chrysosporium sp.

Identification

  • Exoantigen testing
  • DNA probe
  • Mold to yeast phase conversion
  • Very difficult to perform

Coccidioides immitis

  • Causes coccidiomycosis (San Joachin Valley fever, desert fever)
  • Distribution
  • Southwestern deserts of US
  • Transmission
  • Inhalation of arthroconidia (organism is found in the soil)
  • Arthroconidia are resistant to heat, dryness and salinity

Disease

  • 60% of infected patients remain asymptomatic
  • 40% develop a mild to severe flu-like illness
  • Immune competent individuals fully recover  life long immunity
  • ? Reactivation of disease in the immunosuppressed
  • Good prognosis for primary disease; often fatal if disseminated
  • Treatment: amphotericin B or azoles

Disseminated disease

  • May be gradual or rapid
  • May form cavitary lung lesions
  • Seen in patients with underlying lung disease or diabetes
  • Spread via lymphatics to bone, skin, joints, lymph nodes, adrenals and CNS
  • Risk factors for chronic disease
  • Immunodeficiency
  • Pregnancy
  • Dark skin

Direct specimen examination

  • 10-60 spherule is usually seen in pathology
  • Thick refractile wall
  • Loaded with endospores (2-4 each)
  • Cavitary lesions may also contain arthro-conidia

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spherule

Culture characteristics

  • Grows at both room temperature and 370C in 4-5 days
  • Starts as a moist white colony
  • Later is covered by fluffy white mycellium
  • Grows on routine bacteriological media
  • No conversion to spherule phase in culture

Microscopic morphology at
room temperature

  • Septate hyphae
  • Thick-walled, barrel-shaped arthroconidia
  • Alternating empty disjunctor cells
  • Maybe confused with
  • Geotrichum sp.
  • Trichosporon sp.

Identification

  • Tissue spherules with releasing endospores
  • Exoantigen testing

Paracoccidioides brasiliensis

  • Endemic in South America (esp. Brazil)
  • Primary disease is often asymptomatic
  • 50% of cases disseminate
  • Causes paracoccidiodomycosis
  • Organisms are inhaled  pulmonary disease similar to TB
  • Trauma to oral mucosa
  • Ulcers of the mucous membranes, enlarged cervical lymph nodes
  • Death occurs within 2-3 years due to dissemination or malnourishment

Direct specimen examination

  • Specimens
  • Sputum, lymph node biopsies, mucosal biopsy
  • Multiple budding yeast
  • “ship’s wheel”

Culture characteristics

  • Room temperature
  • Slow growing
  • Starts smooth but later becomes covered with white to tan mycelium
  • 370C
  • Waxy, wrinkled colonies
  • Cream to tan colored
  • Conversion to yeast phase requires 3-7 days

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mold

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yeast

Microscopic morphology
at room temperature

  • Branched septate hyphae
  • Intercalary and terminal chlamydo-spores

Microscopic morphology
at 370 C

  • Thick walled, multiple budding yeast
  • “mariner’s wheel”
  • “Mickey Mouse ears”

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Direct smear

Homework

  • Read module 7 and answer review questions.

  • Do the final exam

  • Describe conversion of mold to yeast phase
  • Explain exo-antigen testing.
  • Make a table for the four systemic dimorphic fungi. Tell the specimen where they can be isolated from, geographically where they are found, what are the most identifying features.
  • What extra precautions should you take when working with systemic dimorphic fungi?

Identify the genus of this mold

Identify this mold and yeast phase from a landscaper clearing brush

Name this species of Actinomycetales group

What is this disease called it reminds you of cauliflower.