Fungi - Morphology and Diagnostic Flashcards
What are fungi?
Eukaryotic organisms that are encased within a rigid cell wall.
What is the cell wall of fungi made up of?
Chitin
Flucan
Mannon
Glycoproteins
Various combinations of these structural components.
Where are fungi found?
Ubiquitous in the enviroment.
Some species of yeast are commensals in humans. (candida in GI tract)
Do fungi cause disease?
Rarely, mostly in immunocompromised patients.
Exception to this is dermatophytes in most dimorphic fungi.
How are fungal diseases commonly classified?
By causative organism
By clinical presentations
What type of organism is yeast?
Unicellular, round/oval shaped fungi
How do yeast reproduce?
Asexually.via budding.
Do yeast form hyphae?
No they form pseudohyphae which are elongated cells that can be confused with true hyphae from mould.
Do yeast form spores?
No
How large are yeasts?
3 - 4 micrometers but can be up to 40 micrometers
What do yeast look like macroscopically?
White and thready
What yeast are clinically significant?
Candida species
Cryptococci (cryptococcus neoformans and cryptococcus gatti)
Other rare fungal diseases also exist.
What are mould?
Multicellular fungi
Do moulds form hyphae?
Yes
What are hyphae?
Chain of multinucleate, tubular, filament-like cells
How do hyphae divide?
Each hypha has a rigid cell wall and elongates via mitosis. May be septated or aseptated
What is a tangled mass of hyphae visible to the naked eye called?
Mycelium
What are the types of mycelium?
Vegetative mycelium (extend into culture media and responsible for absorbing water and nutrients)
Aerial mycelium (extend into the air and is required for reproductions
How do fungi reproduce?
Both sexually and asexually.
They use spores or conidia for asexual reproduction.
How are conidia formed?
From a group of specialised cells called conidiogenous cells.
What are annellides?
Annellides (produces conidia by series of short percurrent proliferations and annellides slowly increase in length)
What is a conidiophore?
Specialized hyphal segment responsible for supporting a conidia bearing head.
What do conidiophores branch into?
Further branch into segments called metulae
What are phalides?
Phalides (produce conidia without increasing in length)
What structures vary between genus and species?
Spore morphology that varies includes:
Conidia shapes
Size of conidia
Arrangement of conidia
Septation (if present)
Colour
Texture
Locations of and shapes of conidiogenous cells (phalides or annellides, shape)
What is fungal morphology used for?
It is used to identify different fungal infections
What is hyphomycosis?
A disease caused by fungus in the class hyphomycetes
What are the classes of hyphomycosis?
Hyaline hyphomycosis (Hyalophyphomycosis)
Demetiaceous Hyphomycosis
What is used for classification of hyphomycosis?
Colour of the mycelium and conidia
What is the most common cause of hyaline hyphomycosis mould infections?
Aspergillus (most common)
Penicillium, fusarium and others may also cause diseases
What does hyphae morphology look like in aspergillus?
Septated hyphae that branch 45 degrees.
Conidiophore is 90 degrees from hyphae
Metulae and phialides grow from the vesicles. Phialides produce conidia.
What does hyphae morphology look like in macroconidia (fusarium)?
Macroconidia
What kind of infection do macroconidia (fusarium) cause?
In normal immune system (keratitis and contact lens associated disease)
In immunocompromised people (disseminated blood stream infection and invasive lung and skin infection)
What kind of infections does scedosporium prolificans cause?
in immunosuppressed hosts causes lung infections, blood stream infections, and rarely can cause keratitis
What do conidial look like in scedosporium prolificans?
Clustered and tear shaped.
Flask-shaped phalides
What kind of infections does dermaticaeous hyphomycosis cause?
Opportunistic infections and cutaneous infections in immunocompromised people.
How diverse is dermaticaeous hyphomycosis?
Widely heterogenous
What group of people does mucomycosis most commonly infect?
Immunosuppressed people primarily people with haematological malignancies
What are mucormyocis structures called?
Sporangia (equivalent to conidia)
Sporangiospores (equivalent to conidiospores)
Sporangiophores (equivalent to conidiophores)
What are the unique features of rhizopus?
Broad branching, aseptate hyphae
Some of the genus have rhizoids. (their location relative to sporangiophore helps in identification)
They have stolons
What are stolons?
Connection between the structures.
What is dermatophytosis?
Infection of skin and nail (tinea or ringworm)
What does dermophytosis require to grow?
Keratin
What is the defining feature of trichophytan?
usually only have microconidia, macroconidia are sometimes seen.
What is the defining feature of Microsporum?
Have macro and microconidia of different shapes and sizes. The microconidia have septation.
What is the defining feature of Epidermophyton floccosum ?
It has only macroconidia
Floccosum is the only species in this genus
What are dimorphic fungi?
Fungi that have different morphologies based on temperature.
Yeast at 36degrees celcius
Mould at 25 degrees celcius
What must be observed in dimorphic fungi?
Culture and microscopy of both yeast and mould morphologies.
What is clinically important about dimorphic fungi?
They cause a variety of clinical diseases and are capable of infecting non-immunosuppressed hosts
What are the species of dimorphic fungi that cause clinically significant disease in humans?
Histoplasma capsulatum
Coccidioides immatis
Blastomyces dermatitidis
Pracoccidiodes brasiliensis
Talaromyces marneffi
SPOROTHRIX SCHENCKII is the only dimorphic fungi endemic to Australia
How are dimorphic fungi distributed?
Geographically. Only Sporothrix schenckii is endemic to Australia.
What is Histoplasma capsulatum?
Histoplasma capsulatum is mainly found in Ohio, Missouri and Mississippi river basins with isolated cases in Europe and Africa.
Which form of Histoplasma capsulatum has tuberculated macroconidia?
Mould form
What are athroconidia?
When conidia come off the vegetative mycelium itself. The conidia thickens and breaks off.
What is the fungal species that uses athroconidia and what diseases does it cause?
Coccidioides immitis, it causes pneumonia in immunocompetent people but the pneumonia is self-llimiting.
What kind of disease is caused by Blastomyces dermatitidis?
Chronic granulomatous skin disease
Can also cause a primary - self limiting pulmonary disease.
How do Paracoccidioides brasiliensis reproduce?
In the yeast form it forms what is known as a pilot wheel with lots of little yeast cells surrounding a central cell.
In the mould form they form chlamydospores/arthroconidia
What diseases are caused by Paracoccidioides brasiliensis?
Primary pulmonary disease that is self-limiting or progressive.
Rarely, causes disseminated disease
How are fungal diseases identified and diagnosed?
Microscopy and culture is maintay of diagnosis.
Both microscopic and macroscopic morphology are essential for diagnosis.
Mould have quite different growth requirements compared to bacteria.
What is the procedure used for direct microscopy of fungi?
Microscopy of clinical specimens is done with care to avoid crushing hyphae through processing.
Potassium hydroxide is used to dissolve tissues.
Specimens are then stained
What kind of stain is calcoflour white used for?
Immunofluorescence stain
What kind of infection is diagnosed using india ink stain?
cryptococcus infection
What are the stains used for fungal specimens?
Lactophenol cotton blue
Calcoflour white - Immunofluorescence stain
India ink
Gram stain
Why are gram stains commonly used for staining of fungi?
It can pick hyphae and yeast
How do fungal infections stain in a histopathology stain?
Dark purple on a pink background on a H&E stain
Gomori methenamine silver stain has very good specificity for fungal staining.
What are the advantages and disadvantages of direct microscopy?
Advantages:
Quick screening of specimen to diagnose fungal infection. (Culture takes too long)
Disadvantages:
Spores are not usually seen in direct microscopy (this necessitates use of culture to induce sporulation)
What factors are looked for in a culture?
Temperature
Growth rate
Colony morphology
Colony pigmentation
Growth on selective media
Microscopy of colony growth allows identification of spores.
How is temperature’s effect on morphology of fungi measured?
Cultured on non-selective media (Sabaroud agar) at 25 and 36 degrees.
Can yeast grow on normal bacterial agar?
Yes
What is the ideal temperature for growth of mould and yeast?
Mould - 25
Yeast - 36
Some species such as mucormycosis have more specific temperature requirements. (37, 40, 45)
What happens to dimorphic fungi on agar plates?
They grow as mould or yeast based on temperature
How fast do mould usually grow?
Depends on species
How fast do mucormycosis mould grow?
Can grow within 24 - 48 hours
How fast do aspergillus mould grow?
Can take 3 - 5 days
How fast do phaeohyphomycetes mould grow?
Can take up to 2 weeks
How fast do yeast grow?
24 - 48 hours
How long do spores need to start to appear?
Usually only appear a few days after initial growth.
How are yeast and mould different?
They have different morphologies and colours
What shape do yeasts typically have on cultures?
Smooth, round colonies
What colour do yeasts typically have on cultures?
White to creamed colour generally. Selective media may induce different colour.
What agar is selective for cryptococcus?
Bird seed agar.
What agar is used to differentiate between cryptococcus neoformans or cryptococcus gatti?
L-Canavanine Glycine Bromothymol Blue (CGB) Medium
What type of diseases is Rhodoturula mucilaginosa associated with?
Very rare cause of blood stream infections
Known association with peritoneal dialysis infections
How can Rhodoturula mucilaginosa be visualised on SAB agar?
Forms a distinctive Red/Orange colour on SAB agar.
What feature of mould cultures is used for comparison?
Mould cultures vary significantly so it can be unreliable for anything more than having an initial idea.
What do dermatiaceous mould look like on culture?
Darkly pigmented colonies (they also grow slowly)
What colour are mould on cultures?
Surface and reverse have different pigmentation:
White
Cream
Green
Red
Purple
Brown
Grey
What are the possible shapes of a culture?
Seude-like
Waxy
Glabrous
Downy
Can microscopy differentiate between various species of dermatophytes?
Not in a reliable way
What method effectively allows us to differentiate between different species of fungi?
DNA sequencing of the internal transcribed spaces (ITS) region which is relatively conserved within species.
What is the advantage of using molecular methods of diagnosis over microscopy?
Don’t have to wait for fungi to sporulate for identifications and allows for better and more accurate identifications of fungal infection
Where is the ITS region located?
Between the 50S and 23S ribosomes
What is the limitation of using molecular methods of diagnosis?
Sequencing is fairly limited in a clinical setting at the current time.
What are anamorphs?
Sexual form of mould
What we need to know:
Difference between yeast and mould
Understanding the wide heterogeneity between different mould species which is important for identifications.
Macroscopic and microscopic appearance
Growth characteristics
Important to know how to describe them
