Saturday, 22 August 2015

Emmonsia pasteuriana



Emmonsia pasteuriana

Note:  I've held back on uploading this particular post only because I'm not satisfied with the quality of the photographs I've taken.  It is unlikely that I will come across another strain of this fungus in my career so I will post it at this time hoping any readers will understand.

Emmonsia species:  Emmonsia currently consists of three species: E.parva, E.crescens & E.pasteuriana.

Ecology:  Emmonsia is a cosmopolitan soil saprobe (found just about in all temperate climates and lives of decaying organic matter).  It has been isolated from a variety of mammalian species, particularly small rodents.

Pathogenicity:  Emmonsia parva and Emmonsia crescens are the etiologic agents of adiaspiromycosis.  Usually this presents as an asymptomatic pulmonary infection in animals, and more recently in immunocompromised humans after the inhalation of the fungal spores. Lung biopsy may be necessary to diagnose the illness as the organisms may not be present in sputa or bronchial alveolar lavages.  Dissemination of the infection may occur more readily in immunocompromised hosts. E.crescens is more commonly isolated from humans while E.parva is isolated more often from animals.
Reports of E.pasteuriana have increased in recent years, presenting as disseminated cutaneous (skin) mycosis in persons with underlying AIDS infections[i].  Unlike E.parva and E.crescens, E.pasteuriana does not produce adiaconidia and therefore does not cause adiaspiromycosis.  E. pasteuriana appears as yeast in infected tissue.

Emmonsia pasteuriana:
The following description is for an isolate resembling Emmonsia pasteuriana.

Macroscopic Morphology:  E.pasteuriana exhibits slow to moderate growth at 30ᵒC, with a colony diameter of about 25mm after 10 days and 60mm after 21 days.  The colony appeared velvety to powdery in texture with folded, wrinkled, or cerebriform surface contours. The colony spontaneously acquired splits in the surface as it aged (see photo) The colony was primarily white in colour and remained so while some sources state that it may develop a light brown colour as it ages.  The colony produced no diffusible pigment.  The reverse was tan or light brown in colour.

 Emmonsia pasteuriana - Saboraud-Dextrose Agar.  The splits in the colony were created by the growth and not by any prodding of my own.  (SAB), 30ᵒC, 1 Month. (Nikon)

Emmonsia pasteuriana -  Dermasel® agar, 30ᵒC, ~18 Days. (Nikon)

Microscopic Morphology:  Emmonsia pasteuriana is a dimorphic fungus meaning it can exhibit the filamentous fungus form at one temperature and the yeast form at another.

Filamentous form: This isolate produce septate, hyaline (non-pigmented) hyphae of about 1.0 to 1.3 µm diameter.  Thin-walled, slightly verruculose (minutely verrucose or warty), globose to sub-globose (round-ish) conidia (2 – 3 µm X 3 – 4 µm) are formed on slender (0.4 – 0.5 µm) pedicles (stalks) or at the apex of inflated cells.  Initially a conidium may be found at the end of a delicate pedicle which may then develop further to form four to eight pedicles with conidia, establishing a ‘floret’.  Additional sessile or broad-based verrucose (warty) conidia may also be present.

Emmonsia pasteuriana - hyphae with conidia visible throughout.
(400X, LPCB, DMD-108

Emmonsia pasteuriana -  Globose to sub-globose (round-ish) conidia.  They are generally found at the end of a delicate pedicile (stem) which can barely be seen at this magnification.  A few more photos to follow of much the same as this is this really was my first impression of this fungus and what clued me into what it might be.
(400X, LPCB, Nikon)

Emmonsia pasteuriana - ditto.  Numerous conidia formed.
(400X, LPCB, Nikon)

Emmonsia pasteuriana - another as previously, but here in this photo there may be evidence of the conidia occuring in small bunches, each on its own pedicile branching off from a central delicate pedicile.  (400X, LPCB, Nikon)

Emmonsia pasteuriana -small conidia occuring in bunches (as previously)
(400X, LPCB, Nikon)

Emmonsia pasteuriana - and another.
(400X, LPCB, Nikon)

Emmonsia pasteuriana -at a higher magnification, the rather round (globose) or 'round-ish' (sub-globose) conidia can be seen with a couple at the end of a pedicile (stem or stalk) and one (center-left) which may be growing directly from the hypha (sessile).
(1000X, LPCB, DMD-108)

Emmonsia pasteuriana -small clusters of conidia.
(1000X, LPCB, DMD-108)

Emmonsia pasteuriana - as above.  It was my impression of the tiny, round conidia attached by a delicate pedicile which directed me to the identity.
(1000X, LPCB, DMD-108)

Emmonsia pasteuriana - a solitary conidium at the end of a pedicile is seen in this photo (inset)
(1000X, LPCB, DMD-108)

Emmonsia pasteuriana - Initially a conidium may be found at the end of a delicate pedicle (see above) which may then develop further to form four to eight pedicles with conidia, establishing a ‘floret’.  Here we see more distinctly what is described as a 'floret' (inset).
(400+10X, LPCB, DMD-108)

Emmonsia pasteuriana -a very good example of the 'floret', usually composed of between 4 to 8 conidia at the end of the slender delicate and slender (0.4 – 0.5 µm) pedicles.
(1000X, LPCB, Nikon)
 
 Emmonsia pasteuriana -another example of a 'floret' as above.
(1000X, LPCB, DMD-108)

 Emmonsia pasteuriana -a string of florets along a delicate hyaline (non-pigmented) hypha of about of about 1.0 to 1.3 µm diameter running through the photo from lower left to upper right.
(1000X, LPCB, Nikon)

 Emmonsia pasteuriana -a string of florets along delicate hyaline (non-pigmented) hyphae.
(400X, LPCB, Nikon)

 Emmonsia pasteuriana - Additional sessile (directly from hypha) or broad-based verrucose (warty) conidia may also be produced.
(1000X. LPCB, Nikon)

 Emmonsia pasteuriana - once again, single conida on slender stalks.
(400X, LPCB, DMD-108)

 Emmonsia pasteuriana - two conidia can be seen at center left, attached to the delicate pedicile.
(400+10X, LPCB, DMD-108)

 Emmonsia pasteuriana -sessile (directly from hypha) or broad-based verrucose (warty) conidia may be produced.  (1000X, LPCB, DMD-108)

Emmonsia pasteuriana - Once again, thin-walled, slightly verruculose (minutely verrucose or warty), globose to sub-globose (round-ish) conidia (2 – 3 µm X 3 – 4 µm) are formed on slender (0.4 – 0.5 µm) pedicles (stalks) or at the apex of inflated cells.  Here in this photo is a conidium at the apex of an inflated cell.  (1000+10X, LPCB, DMD-108)

Emmonsia pasteuriana - more conidia at the born on delicate pediciles.
(1000X, LPCB, DMD-108)

Yeast form:  The filamentous fungus form can be converted to the yeast form by incubating a freshly inoculated culture at 37ᵒC for 10 to 14 days.  Conversion is enhanced by cultivation on the nutritionally richer Brain-Heat Infusion (BHI) agar.  Smooth, butyrous colonies appear cream to beige in colour which may darken as they age.  Colonies consist of globose to oval yeast colonies which may show narrow-based budding.

 
Emmonsia pasteuriana - isolate was inoculated onto Brain-Heart Infusion (BHI) agar and incubated at 37ᵒC for 14 days.  Emmonsia pasteuriana converted to the yeast form which is the form that is directly recovered from skin lesions.
(400X, LPCB, DMD-108)

 Emmonsia pasteuriana - a more convincing example of the yeast-like phase of E.pasteuriana.
(400+10X, LPCB, DMD-108)

Note:  At various stages of development, Emmonsia species may resemble other fungi such as Blastomyces dermatitidis, Paracoccidioides brasiliensis, Histoplasma capsulatum and Chrysosporium species.  It has been noted that some cross-reaction (false positives) may occur between Emmonsia species and Blastomyces dermatitidis with a both a DNA probe and direct immunofluorescent-antigen tests.

Adioconidia: When the hyphae and conidia are incubated at their maximum temperatures on enriched media, the hyphae become distorted and usually disintegrate while the conidia swell to become round, thick-walled adiaconidia (formerly called adiaospores).  Production of adiaconidia is best achieved by incubation at increased temperatures: 37ᵒC for E.crescens (20 – 14 µm dia) and at 40ᵒC for E.parva (10 -25 µm dia).   As previously mentioned, E.pasteuriana cannot be induced to produce adiaconidia at any temperature.

Physiology:  Emmonsia pasteuriana is not inhibited by cycloheximide and therefore can be grown on Mycosel® or Dermasel® agar.

 

[i] A Dimorphic Fungus Causing Disseminated Infection in South Africa
Chris Kenyon M.D. et al.,
N.Engl. J. Med. 2013, 369 – 1416 - 1424

Saturday, 1 August 2015

Penicillium citrinum



Penicillium citrinum

Ecology:  Penicillium citrinum is a commonly occurring filamentous fungus with worldwide distribution.  It has been isolated from a variety of sources including soils, decaying vegetation, foodstuffs (beans, coffee, cereals & spices) as well as a variety of indoor environments.


Pathology:  While Penicillium species are generally regarded as laboratory contaminants, or at best, opportunists, a number of species have been implicated as being involved in the disease process.  While Penicillium species may be isolated from clinical specimens, it is commonly believed that a true infection can only be established by histological demonstration of tissue invasion.  With that in mind, Penicillium citrinum has been reported in mycotic keratitis (eye), lung infections (pneumonia), a single case of a urinary tract infection (UTI) and one of pericarditis.  Their contribution to the disease process may be secondary an additional underlying illness.  As with all fungi, immunocompromised individuals may be at greater risk of infection including those rarely considered as pathogenic.

Macroscopic Morphology:  Penicillium citrinum exhibits moderately slow growth on Sabouraud-Dextrose agar (SAB) at 30ᵒC.  Surface texture is velutinous (soft, velvety surface) to floccose (woolly tufts of soft “hairs”).  The colonial growth appears radially sulcate (narrow, deep furrows or radial grooves –like spokes on a wheel).  The mature colony has a central greyish-turquoise to greyish-orange colour with a white periphery (outer edge).  Exudates (extrolites) are frequently produced which appear as drops of liquid upon the surface of the colony.  These may appear clear, to pale yellow, to a reddish-brown in colour.  Some strains may also produce a soluble pigment which can diffuse into the surrounding medium.  The reverse is a pale yellow to a light yellow-brown. Colours and growth characteristics are, of course, media and strain dependent.

 
Penicillium citrinum-SAB, 14 days incubation at 30ᵒC (Nikon)
Note the drops of exudate (extrolites) which have formed on the surface.
Colour variation due to maturing of colony but also a difference in my lighting for photography.

 Exudates (or Extrolites): Some fungi can produce exudates as a by-product of their growth, many of which can be collected for commercial use.  Mycotoxins are by-products (secondary metabolites) which are potent poisons.  Penicillium citrinum produces Citrinin, a nephrotoxic mycotoxin which derives its name from the fungus.  It may also produce other extrolites such as tanzowaic acid A, quinolactacins, quinocitrinines, asteric acid and compactin.

Microscopic Morphology:  Penicillium citrinum produces septate, hyaline (clear, not pigmented) hyphae.  Smooth-walled conidiophores stipes are rather long (100 – 300 µm) and is biverticillate (see diagram at end of post).  Metulae are 12 – 15 µm in length which are found in whorls of 3 – 5 divergent structures.  Phialides are ampuliform (flask-shaped) and about 7 – 12 µm in length.  Conidia (2.2 – 3.0 µm dia.) are globose to sub-globose (round to off-round) and are smooth or have a finely roughened surface.  Conidia resist disruption and form rather long chains.  These characteristics: the metulae longer than the phialides and the conidia being both spherical and produced in well-defined chains, are distinguishing features of Penicillium citrinum.

 Penicillium citrinum-  distinguishing features of Penicillium 'species' can already be made out at low magnification. (250X, LPCB, DMD-108)

 Penicillium citrinum-  distinguishing features of Penicillium 'species' much more evident at 400X.
Typical "fingers" made up of the metulae and phialide structures from which chains of conidia extend. (400X, LPCB, DMD-108)

Penicillium citrinum- a mass of overlapping fruiting structures with copious amounts of conidia.
(1000X, LPCB, DMD-108)

Penicillium citrinum- a little less congested in this photo.  Conidiophores (stipes) seen from which extend the metulae and conidia producing phialides.  Conidia are globose (round) to sub-globose (somewhat off-round) in shape,  (1000X, LPCB, DMD-108)

Penicillium citrinum- long metulae and the somewhat shorter phialides are clearly distinguishable in this photograph.  The conidia are generally smooth or can have a finely roughened surface.
(1000+10X, LPCB, DMD-108)

 Penicillium citrinum- another view.
(1000+10X, LPCB, DMD-108)

 Penicillium citrinum- exhibits biverticillate branching meaning that the conidiophore can branch and the metulae & phialides extend from these branches.  Triverticillate would have the conidia branching and then the branches also branching to finally produce the metulae & phialide fruiting structures.
(1000X, LPCB, DMD-108)

 Penicillium citrinum- Phialides are ampuliform (flask-shaped) and about 7 – 12 µm in length.  Again, conidia (2.2 – 3.0 µm dia.) are globose to sub-globose (round to off-round) and are smooth or have a finely roughened surface. (1000+10X, LPCB, DMD-108)

 Penicillium citrinum- Here we see the proportions of the metulae (M) and the 'flask-shaped' phialides (P) with the metulae being substantially longer than the phialides,  The biverticillate structure is evident in this photo. (ie. each branch extending from the conidiophore (stipe), branches only once and then bears a fruiting structure consisting of the metulae and phialides.
(1000+10X, LPCB, DMD-108)

Penicillium citrinum- another example.
(1000+10X, LPCB, DMD-108)

Penicillium citrinum- a few more photos to finish up.
(1000+10X, LPCB, DMD-108)

Penicillium citrinum- suitable for framing!
(1000X, LPCB, DMD-108)

Penicillium citrinum
(400+10X, LPCB, DMD-108)
Penicillium citrinum
(400X, LPCB, DMD-108)

Penicillium citrinum- another colony showing the exudate (extrolites) which accumulate on the colony surface after extended incubation.  These metabolites may be potent poisonous mycotoxins or might have beneficial uses in industrial or pharmaceutical applications. (Nikon)



Physiology:  The spores of Penicillium citrinum fail to germinate at 5ᵒC and may show restricted growth at 37ᵒC.
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Saturday, 4 July 2015

Stemphylium species



Stemphylium species (Hyphomycetes)

Ecology:
Stemphylium species are widely distributed in nature and can be found in soils as well as plant parasites and as saprophytes on decaying plant material.  As a plant pathogen, Stemphylium species are implicated in the leaf spot of alfalfa (S.botryosum), black rot of carrots (S.radicinum), and grey leaf spot of the tomato plant (S.solani).


Pathogenicity:
Stemphylium species found in the clinical laboratory are generally considered to be contaminants; however, they may contribute to allergic reactions in general and have been reported as agents of phaeohyphomycotic sinusitis. 

Macroscopic Morphology:
Colonies on SAB are velvety to cottony with a rather short ‘nap’ (does not extend very high above the agar surface).  They are olivaceous (grey-green-brown) to a brownish-black in surface colour.  The reverse is black.  Stemphylium species exhibit a moderate growth rate, becoming mature within five days.

Stemphylium species - 1 Week on SAB at 30oC (Nikon)


Microscopic Morphology:
Hyphae are septate and develop a pale brown to deeper brownish colour as they age.  Conidiophores also show septations and the structure is simple or occasionally branched.  The cell at the apex of the conidiophore which bears the conidium may show a slight swelling in relation to the rest of the conidiophore.  The conidiophore which generally shows somewhat smooth and parallel walls when young may develop a much more knobby appearance as it ages and produces conidia.  Conidia (12 – 20 µm X 15 – 30 µm) are produced by growing from the tip of the terminal conidiogenous cell (poroconidia).  They can be smooth or rough (echinulate) in texture.  They have been described as oval, oblong, ellipsoidal, obclavate and subspherical.  More simply they may be described as ‘box-like’ with rounded corners.  The conidia may also show a marked constriction around a central septum or division within individual conidia. The conidia are muriform (both transverse and longitudinal septations or divisions), and acquire a dark brown pigment as they mature.  

Caution: Micron scale (µm) may change between 50 or 100 µm at higher magnifications.

Stemphylium species - not much to distinguish between other moulds in this photo.  I've added it just to show the tangled mass of the mycelium that fungi produce.  Conidia are visible as the dark spots but features are indistinguishable at this magnification. 
 (250X, LPCB, DMD-108)

Stemphylium species - edge of a slide culture showing hyphae growing out from the edge of the SAB block from which conidiophores extend and produce pigmented conidia at the ends.
(250X, LPCB, DMD-108)

Stemphylium species -another shot showing numerous pigmented conidia sitting on the ends of the conidiophores which extend at right-angles from the supporting vegetative hyphae.
(250X, LPCB, DMD-108)

Stemphylium species -more detail emerges at this higher magnification.  Septations become visible in the pigmented conidia.
(400X, LPCB, DMD-108)

Stemphylium species -hyphae and conidiophores.
(400+10X, LPCB, DMD-108)

Stemphylium species -rather short conidiophores, extend primarily at right-angles from the parent vegetative hypha.  At the lower left corner there is a branched conidiophore with a rather young, still unpigmented, conidium developing on the left branch. Two more young conidia can be seen in the photo as well.
(400X, LPCB, DMD-108)

Stemphylium species -a few more photos to leave you with an impression of what Stemphylium looks like.  Conidia can be longer and branched which distinguishes it from Pithomyces species.  The shape of the conida also is different from Pithomyces - discussed in later photos.
(400X, LPCB, DMD-108)

Stemphylium species -brown pigmented conidia at the apices of individual conidiophores extending from an 'out of focus' hyphal element.
(400X, LPCB, DMD-108)

Stemphylium species -conidiophores extending from the supporting vegetative hyphae with conidia at various stages of maturity.  Young, blue-stained conidia and brown pigmented mature conidia are present.  (400X, LPCB, DMD-108)

Stemphylium species -ditto. 
(400X, LPCB, DMD-108)

Stemphylium species -self indulgence here.  More photos to demonstrate the same,  Conidiophores with mature pigmented conidia at the tips.  Muriform (longitudinal & transverse) septation is apparent.  (400X, LPCB, DMD-108)

Stemphylium species -the appearance in this photo is very similar to that of Pithomyces species in that there are conidia at the ends of short conidiophores arising at right-angles to the hyphae from which they extend.  Darkly pigmented conidia have internal compartments made by the muriform (both up and down and across) septations.  The pigmented hyphae to the left of the photo is clearly septate and appears to be releasing it's pigment into the medium (brown haze).
(400X, LPCB, DMD-108)

Stemphylium species -as above. 

Stemphylium species -massive amounts of conidia can be produced as seen here.
(400X, LPCB, DMD-108)

Stemphylium species - a slightly closer look.  Muriform septations evident in the mature brown conidia.  The shape has been described as oval, oblong, ellipsoidal, obclavate and subspherical.  More simply they may be described as ‘box-like’ with rounded corners.
(400+10X, LPCB, DMD-108)

Stemphylium species - The conidiophore which generally shows somewhat smooth and parallel walls when young may develop a much more knobby appearance as it ages and produces conidia (lower left).  (400+10X, LPCB, DMD-108)

Stemphylium species - mature brown box-like conidia at the ends of somewhat knobby conidiophores.  (400+10X, LPCB, DMD-108)

Stemphylium species - conidia with muriform septations.
(400+10X, LPCB, DMD-108)

Stemphylium species - ditto, as above.
(400+10X, LPCB, DMD-108)

Stemphylium species -the conidia may also show a marked constriction around a central septum or division within individual conidia (arrows).
(400+10X, LPCB, DMD-108)

Stemphylium species -  conidia (12 – 20 µm X 15 – 30 µm, when mature) are produced by growing from the tip (arrow) of the terminal conidiogenous cell (poroconidia).
(1000X, LPCB, DMD-108)

Stemphylium species - here on another conidiophore you can see the conidium (poroconidium) increasing in size as it develops.  Internal septations have yet to develop.
(1000X, LPCB, DMD-108)

Stemphylium species - branching conidiophore -mature brown conidia and a smaller and younger, pale blue conidium.  To follow along from the last two photos, the pale blue conidium is developing further and starting to develop a transverse septum (light line crossing the inside of the small blue conidium).  (400+10X, LPCB, DMD-108)

 Stemphylium species - Mature brown muriform septate conidia showing central construction along where the transverse septum crosses.  Conidiophores may develop a knobby appearance with maturity and the cell at the apex of the conidiophore which bears the conidium may show a slight swelling in relation to the rest of the conidiophore.
(1000X, LPCB, DMD-108)

 Stemphylium species - as above.
(1000X, LPCB, DMD-108)

Stemphylium species -  As above.  Different stages of maturity.  Too many photos, I know!
(1000X, LPCB, DMD-108)

Stemphylium species - ditto.
(1000X, LPCB, DMD-108)

Stemphylium species - conidia can be smooth or rough (echinulate) in texture.
(1000+10X, LPCB, DMD-108)

Stemphylium species - can be smooth or rough (echinulate) in texture. Conidiogenous cell at tip of conidiophore may be somewhat distended (wider, swollen) in relation to the rest of the conidiophore.
(1000+10X, LPCB, DMD-108)

Stemphylium species - conidiophore shows septation.
(1000+10X, LPCB, DMD-108)

Stemphylium species - again, conidiogenous cell closest to the tip may show a slight swelling in relation to the rest of the conidiophore.
(1000+10X, LPCB, DMD-108)

Stemphylium species - conidia with various surface textures.
(1000+10X, LPCB, DMD-108)

Stemphylium species - yet another photo!  You get the picture by now...
(1000+10X, LPCB, DMD-108)

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

 Stemphylium species

Stemphylium species - poroconidium (doesn't show the annular frill which is seen on somewhat similar looking muriform conidia produced by Pithomyces species.
(1000+10X, LPCB, DMD-108)

 Stemphylium species - looks like a degenerating, rather wilted conidiophore.
(1000+10X, LPCB, DMD-108)

Stemphylium species- branching septate conidiophore.
(1000+10X, LPCB, DMD-108)

 Stemphylium species - branching conidiophore.
(1000+10X, LPCB, DMD-108)

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

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

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

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

Stemphylium species -Last one.  I don't know what I was thinking, uploading some 45 photos to illustrate Stemphylium species.  As with most of the posts in this blog, the photos posted represent only 10% of what I've taken.  Sorry for anyone who I lost with boredom -I find the little critters fascinating.
(1000+10X, LPCB, DMD-108)

Note: May be confused with other darkly pigmented moulds such as Alternaria species, Pithomyces species, and Ulocladium species (see table below).


 Too small to read?  Click on table to get image. Now right click on image and select 'view image'.  In Windows, cursor now has a + sign within it.  Click on image of table to now magnify the table.
Alternatively, just click and download the damn thing...
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