Harriet the Hitch-hiker

For those interested in the initial story, scroll down to yellow text.

January 28th, 2016: The case of ‘Harriet the Hitch-hiker has been solved!  The culprit in this case was a common house dust mite, most likely Dermatophagoides farina.  

I spoke with a former colleague the other day who informed me that months after I left, there seemed to be a ‘bug’ infestation in the mycology lab.  A few parafilm-sealed plates left at room temperature appeared to show surface movement.  When placed under a plate microscope, tiny creatures could be seen chomping away at the fungal growth.  As my colleague put it “it looked just like tiny buffalo eating grass on the high plains!”  

So where did they come from?  Well, they are considered ‘common’ dust mites and so they may have blown in from the hospital environment.   Hospital beds are full of patients, shedding skin and while proper laundering of bedding and other fomites should destroy these critters, the contents  and mites may be shaken off in the process.  They may have been brought into the lab by the cleaning staff or by laboratory staff such as myself.  They may have come from specimens sent from other labs.  Somehow I was lucky enough to find the very first one most likely caught on the adhesive tape roll that I used to make a mount of the proficiency testing sample.

Well, the completed isolates were quickly discarded.  New specimens were temporarily sent to the Provincial Reference Laboratory for workup and the Mycology laboratory was scrubbed down with disinfectant from top to bottom.  The problem has been solved

It was news to me that mites eat fungi but just Google ‘fungi eating mites’ and you’ll get a number of references and even a YouTube video for those who want to see these mites in action.

Just a few Notes:

Mites are eight-legged, sightless arthropods that live on skin scales and other debris.  The most common house dust mites are:

Dermatophagoides pteronyssinus –known as the European dust mite

Dermatophagoides farina –known as the American dust mite

The common names may be misleading as neither species is restricted to the location in their name.

Euroglyphus maynei - is a third species of dust mite, however, because of the high water content of this organism, it prefers more humid environments to replenish itself.

Once source[i] suggests that fungi are an essential component of the dust mite’s life cycle.  Though not fully understood, it may be that the fungi utilize some of the same nutrients or assist in breaking them down so they may be consumed by the dust mites.  This sounds more like a symbiotic relationship rather than the mites consuming the fungi (fungivore).

 

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[i] http://www.situbiosciences.com/dust-mites-love-fungi/

Another souce;

Featured Creatures –University of Florida

http://entnemdept.ufl.edu/creatures/urban/house_dust_mite.htm

The Story of 'Harriet the Hitch-Hiker

So the laboratory proficiency mycology specimens arrive in the lab and I eagerly have a first look at the direct, teased out, samples.  One specimen is labeled as having been taken from the scalp so I suspect the sample may possibly contain a dermatophyte.  Carefully scanning at low power, I search for any tell-tale clues which may provide a ‘head-start’ (no pun intended). Then, I see something unexpected.  Hey! What’s this?  What are you doing here?  Where did you come from?  That specimen from the scalp has brought along a hitch-hiker.  So who are you anyways?  Follicle Fred? Afro Al? Blow-Dried Billy-Bob?  Hmmm, Hairy Harrison? (Hairy Harry to be less formal).....but wait….what do we have here?  Eggs!  That’s not Harry the hitch-hiker….it’s Harriet the hitch-hiker!           

I suppose it was an unintended addition, but our mould sample from the scalp, also contained a tick(?) of some sort.  The specimen donor has more problems than just a possible dermatophyte!             

‘Googling’ ticks brought up the ‘mug shots’ of a number of possible suspects, however, I’ll leave it up to some entomologist out there to positively identify my freeloader and replace the alias with a proper name.

Below are Hitch-hiking Harriet’s mug shots to add to the unsavory lot of ticks, lice and louses imprisoned in cyberspace.  Sorry, she wouldn’t turn for me to get a profile photo.  No smile either!  One tough character!

Lactophenol Cotton Blue is the background stain as I was looking for moulds in this preparation.

Harriet, my unknown scalp tick which hitch-hiked along on a scalp mycology specimen.

(Didn't keep detailed notes on this one.  X100, I believe -Nikon)

Harriet again.  Female for sure -check out the 5 oval eggs in her abdomen

(?250X, Nikon)

Same 'Harriet' but image reversed due to the difference in the optics between the Nikon and the DMD-108 Digital Micro-imaging Device.

(100X, DMD-108)

And the last mug-shot at a higher magnification.

(?100+10X, DMD-108)

 Links below redirect to the identical post within this blog.

While I like using both photographic devices (Nikon Coolpix microscope mounted camera) and the Leica DMD-108 Digital Micro-imaging Device, I feel that each platform has an advantage in different situations.  I like to try both methods and then chose which gave the best representation of the subject I was attempting to document.  Often it is in the eye of the viewer.

What?  you mean you don't give a familiar name to your organisms?

Fun With Microbiology - as the title of the blog proclaims.

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Fonsecaea pedrosoi/monophora

Fonsecaea pedrosoi/monophora

Note 1:  Recent changes in genus Fonsecaea:  Previously the genus Fonsecaea was considered to be comprised of F.pedrosoi and F. compacta.  Recent revisions indicate that F.compacta is simply a morphological variant of F.pedrosoi and therefore considered the same organism.  DNA analysis, however, has added a second species to the genus known as F.monophora.  There are subtle morphological differences between the two; however, they are best differentiated by molecular means.  The fungal disease was first described by Alexandrino Pedroso in 1911, hence the name.

Ecology:

While Fonsecaea can be found worldwide it is more commonly found in tropical and sub-tropical regions where it is found as a saprobe (lives on dead organic matter) in soils and rotting plant materials.  Agricultural workers in Central and South America, India, Africa and Madagascar are more commonly exposed to these soils and exposure to Fonsecaea.  It is usually acquired through traumatic implantation via a splinter or thorn.  Cold blooded animals living near or around swamps may also be infected and carry the fungus.

Pathology:

Fonsecaea is the most common cause of chromoblastomycosis, a chronic subcutaneous infection which is characterized by verrucous lesions and the formation of brown sclerotic fission cells, described as “copper pennies”[i] within the tissue.  Other dematiaceous fungi responsible for chromoblastomycosis are Phialophora verrucosa and Cladophialophora carrionii.  Both F.pedrosoi and F.monophora are recognized agents of human chromoblastomycosis; however, in F.pedrosoi a strict association with this disease is noted, while F.monophora is a more general opportunist.  While prognosis is generally good, the infection itself is difficult to treat and long-term therapy is required.  The presentation of the disease can initially be confused with squamous cell carcinoma.  Systemic (internal) infections have rarely been described however F.pedrosoi has been implicated in a fatal brain infection acquired via haematogenous dissemination.  Keratitis (corneal infection) and a case of paranasal sinusitis have also been reported.

Macroscopic Morphology:

Growth rate for F.pedrosoi/monphora is slow with the colony maturing in about 14 days on Sabouraud Dextrose medium (SAB) at 30ᵒC.

The colony surface may be dark green to olive brown to dark grey or jet-black depending on the strain and medium.  It is covered with a fine, velvety or downy mycelium.  Colonies start of relatively flat however they usually produce a raised convex protrusion at the center where initially inoculated.  The colony becomes somewhat embedded in the agar surface and may break apart when probed.  The colony reverse is black.

Note 2:  I will refer to the organism throughout the remainder of this post as Fonsecaea pedrosoi for my own ease, however, the reader should keep in mind that the organism could be Fonsecaea pedrosoi or Fonsecaea monophora as discussed in Note 1.

 

 Fonsecaea pedrosoi - Sabouraud Dextrose Agar (SAB), 10 Days, 30ᵒC (Nikon)

 Fonsecaea pedrosoi - Sabouraud Dextrose Agar (SAB), ~3 Weeks, 30ᵒC (Nikon)

Microscopic Morphology:

Note 3:  One source (Larone-See Sidebar) suggests that conidiation may be enhanced by growing the organism on Corn Meal Agar (CMA) or Potato Dextrose Agar (PDA).  In fact the isolate presented here did not show any conidiation until grown on CMA.  All microphotographs presented here are from growth on CMA.

Be aware that the scale of the micron bar within photos may vary.

 Fonsecaea pedrosoi - grown on SAB media, this isolate failed to produce any 'fruiting structures'.  It was only after growing the fungus on Corn Meal Agar (CMA) that conidiophores and conidia were observed.  (400X, LPCB, DMD-108)

All further photos are taken from growth on CMA.

Fonsecaea produces dematiaceous[ii] (dark/brown) septate and loosely branching hyphae.  The conidia produced are pale brown or olivaceous in colour.  They are sub-hyaline, smooth textured, thin walled and ovoid or clavate (club-like) in shape.  The conidia (3.5 – 5.0 X 1.5 – 2.0 µm) are produced in short chains at the apex of the conidiophores.

 Fonsecaea pedrosoi - view of mature fungus at lower magnification.

(400X, LPCB, DMD-108)

Fonsecaea pedrosoi -initial 'budding' growth of conidiophores/conidia appear along the hypha.

(1000X, LPCB, DMD-108)

Four types of conidial formation may be observed the same strain of Fonsecaea.

Fonsecaea type:  Conidiophores are septate, erect, and compactly sympodial.  The distal (far) end of the conidiophore develops swollen denticles that bear primary single-celled ovoid conidia.  Denticles on the primary conidia support secondary single-celled conidia that may produce tertiary conidia, but long chains of conidia are not formed.  Elongate conidia often form in verticils at fertile sites along the conidiophore, producing an asterisk-like (*) appearance.

Rhinocladiella type:  Conidiophores are septate, erect, and sympodial; swollen denticles bear ovoid conidia at the tip and along the side of the conidiophore.  Usually only primary conidia develop.

Cladosporium type:  Conidiophores are erect and give rise to large primary shield-shaped conidia that in turn produce short, branching chains of oval conidia having small dark hila (scars of attachment)

Phialophora type:  Phialides are vase shaped with terminal cup-like collarettes.  Round to oval conidia accumulate at the apex of the phialide.  This type of conidiation is often scant or lacking.

 Fonsecaea pedrosoi - as the colony matured, the hyphae darkened with the production of melanin pigment.  Initially only primary, or a single tier of conidia were seen to be produced which suggested that this organism might be Rhinocladiella species.  Therefore, this is what I believe sources refer to as Rhinoladiella conidiation as described in the previous text.

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi -again, only primary conidia (each conidia attached to the conidiophore) is seen suggesting the Rhinocladiella type conidiation.

(1000+10X, LPCB, DMD-108)

 Fonsecaea pedrosoi -an interesting feature observed is the sympodial growth of the hyphae/conidiophore (arrow).

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi -Conidia have dispersed revealing the sympoidal growth pattern (arrows)

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi -a final photo showing this feature.

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi -secondary conidia appear which are attached to the primary by a delicate denticle.  Rhinocladiella is not known for producing other than primary conidia therefore the evidence began to suggest that this was, indeed, Fonsecaea.

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi - still mostly primary conidiation however a typical example of Fonsecaea.

(1000X, LPCB, DMD-108)

 Fonsecaea pedrosoi - typical growth.  Branching conidiophores present.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - from edge of slide culture on CMA.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - compactly sympodial conidiation of Fonsecaea.

(1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - another photo (sometimes I post a photo just 'cause it looks cool!)

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - more typical of mature Fonsecaea as you can see that there are several tiers to the conidiphore & conidia fruiting structure.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - conidiophores bearing conidia along the septate hyphae.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - typical complex fruiting structure -center left of photo.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - another typical example.

(1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - more examples.  Conidiophores of varying lengths along the hyphae. Primary and more mature central hypha has developed dark pigmentation.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - a typical example of the conidiphore with compactly sympodial growth of conidia.  (1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi -  Fonsecaea type conidiation:  Conidiophores are septate, erect, and compactly sympodial.  The distal (far) end of the conidiophore develops swollen denticles that bear primary single-celled ovoid conidia.  Denticles on the primary conidia support secondary single-celled conidia that may produce tertiary conidia, but long chains of conidia are not formed.

(1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - Elongate conidia often form in verticils at fertile sites along the conidiophore.

(1000X, LPCB, DMD-108) 

Fonsecaea pedrosoi - Elongate conidia often form in verticils at fertile sites along the conidiophore.

(1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - Elongate conidia often form in verticils at fertile sites along the conidiophore, producing an asterisk-like (*) appearance. (Below)

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - Elongate conidia often form in verticils at fertile sites along the conidiophore, producing an asterisk-like (*) appearance.

(500X, LPCB, Nikon)

Fonsecaea pedrosoi - Cladosporium type conidiation: conidiophores are erect and give rise to large primary shield-shaped conidia (inset) that in turn produce short, branching chains of oval conidia having small dark hila (scars of attachment).  Conidia were easily disrupted and I have no photos for chaining of the conidia)  (1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - remnants of conidial attachment on a septate conidiophore.

(1000X, LPCB, DMD-108)

Fonsecaea pedrosoi - Septate conidiophore with conidia shown.  The conidia produced are pale brown or olivaceous in colour.  They are sub-hyaline, smooth textured, thin walled and ovoid or clavate (club-like) in shape.  The conidia (3.5 – 5.0 X 1.5 – 2.0 µm) are produced in short chains at the apex of the conidiophores.

(1000+10X, LPCB, DMD-108)

Fonsecaea pedrosoi - Phialophora type conidiation (?): Phialides are vase shaped with terminal cup-like collarettes (inset - arrows).  Round to oval conidia accumulate at the apex of the phialide.  This type of conidiation is often scant or lacking.  The 'staggered' sympodial growth pattern appears evident at base of inset photo)

(1000+10X, LPCB, DMD-108)

 Fonsecaea pedrosoi

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[i] In tissues, this fungus, as well as other etiologic agents of chromoblastomycosis appears as large (5 – 12 µm diameter), round, brownish and thick-walled bodies, hence the resemblance to the coin and common description of “copper pennies”.  When the fungus is cultured on laboratory media at 25, 30, or 37ᵒC, the fungus is filamentous.

[ii] Dematiaceous fungi represent a large and heterogeneous group of filamentous moulds containing melanin in their cell walls. The term phaeohyphomycosis was proposed by Ajello and Georg in 1974 as “a collective name for a group of mycosis caused by diverse genera and species of dematiaceous fungi”

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