Cladosporium species (Revisitied)

Cladosporium species Revisited (Kind Of) -A Black Mould  -Hyphomycetes

This post is a bit of a revisitation to the fungus Cladosporium as I previously included the genera in a previous post on Cladophialophoraspecies.  You can compare the two more thoroughly by checking out my previous post by clicking here.

Ecology: Cladosporium species are cosmopolitan saprophytic fungi found in soil, on plant debris and leaf surfaces. 

Pathogenicity:  Cladosporium species are generally considered to be non-pathogenic although they may be considered as possible opportunists in the severely debilitated host.  Potentially pathogenic species previously included under the Cladosporium genera have recently been reassigned to the genera Cladophialophora (eg. C.bantianum, C.carrionii).  Cladosporium may be encountered in the laboratory as a culture contaminant and must be distinguish from the pathogenic Cladophialophora.

Macroscopic Morphology:  The rate of growth is dependent on the particular Cladosporium species and can vary from slow to moderately rapid.  The isolate discussed in this post expanded in size rather slowly though it matured to produce copious amounts of conidia rather quickly (~7 days).  The colony was velvety to suede-like in texture.   Other sources describe Cladosporium’s texture as ranging from powdery to woolly.  The colony may become slightly heaped and develop gentle folds as it ages.  Colour ranges from greyish-green to olivaceous-green to brownish-black.  The reverse is a dark brown to black in colour.

 Cladosporium species on SAB, 15 Days at 30˚C (Nikon)

 Cladosporium species on SAB, 25 Days at 30˚C (Nikon)

Microscopic Morphology:  Cladosporium produces erect, dark, septate hyphae.  Conidiophores are also darkly pigmented, may be septate and show tree-like branching.  Fragile chains of dematiaceous blastoconidia are produced and exhibit a dark hila or scar at their point of attachment to the conidiophore or other conidia.  The 1-4 celled conidia are round to oval (3 -6 µm X 4 - 12 µm) and may be smooth-walled to verrucose in surface texture.  Cells on the conidiophore which bear the chains of conidia are sometimes septate and appear in the shape of a ‘shield’.  These cells are also conidia but are referred to as shield cells.  Chains of conidia easily disarticulate (break up) and were frustratingly difficult to document using both adhesive tape andslide culture techniques.

Cladosporium species are not thermotollerant and some species may not grow at 37˚C.

 Cladosporium species produce darkly pigmented, septate, branching hyphae,

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

Cladosporium species - Conidiophores are also darkly pigmented, may be septate and also show tree-like branching. (LPCB, DMD-108, 400+10X)

Cladosporium species - The Lactophenol cotton blue stain has taken more deeply in this preparation.  Septate hyphae are clearly visible. (LPCB, DMD-108, 400X)

 

Cladosporium species -1-4 celled conidia are round to oval (3- 6 µm X 4-12 µm) and may be smooth-walled to verrucose in surface texture.

(LPCB, DMD-108, 1000X)

Cladosporium species -the structures are clearly evident here: septate hyphae, darkly pigmented conidiophores and oval conidia in chains.

(LPCB, DMD-108, 1000X)

Cladosporium species -as above.  Younger conidiophores & conidia stain more intensely with the LPCB while the mature structures have developed the pigmentation.

(LPCB, DMD-108, 1000X)

Cladosporium species -the structures easily dis-articulate (break up) and it was difficult to keep them intact regardless of using the adhesive tape technique or slide culture technique.

Short chains of oval, pinmented, conidia are seen.

(LPCB, DMD-108, 1000X)

Cladosporium species - a closer look at the conidiophores bearing round to oval pigmented conidia.

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

Cladosporium species -a single conidia (conidiophore?) seen at the end of a septate hyphae.

(LPCB, DMD-108, 1000X)

Cladosporium species - a structure visible in some of the previous photos is more clearly seen in this picture.  Here you can clearly see what is referred to as a "shield cell", because of its resemblance to a warrior's shield.  These are conidiophores as you can clearly see chains of conidia extending from them. (LPCB, DMD-108, 1000+10X)

Cladosporium species - `Shield` cell conidiophore and conidia

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

Cladosporium species - another view, as above.

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

Cladosporium species - `Shield`cells and chains of conidia

(LPCB. DMD-108, 1000X)

Cladosporium species - Septate hyphae with conidiophore bearing conidia at its apex.  Insert is simply a change of focus.  (LPCB, DMD-108, 1000X)

Cladosporium species - okay, one final photo, just for the heck of it!

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

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Chilomastix mesnili

Chilomastix mesnili  (Flagellate) Intestinal Parasite

Note: I'm always hesitant in adding posts of organisms where the photos don't clearly show the features or structures described.  Such is the case here -fine detail is not evident, but here are the photos for whatever they are worth.

Geographic & Human Habitats:

In nature, Chilomastix mesnili is considered to be cosmopolitan and therefore has worldwide distribution, found in both warm & temperate climates.  In humans, it can be found primarily in the cecal region of the large intestine, but may also occur in the small intestine.  There it feeds on intestinal bacteria and debris.

Pathogenicity:

C.mesnili is considered to be non-pathogenic and treatment is not warranted.  Its presence in stool specimens does indicate that the host has acquired the parasite through the ingestion of its cysts and therefore has come in contact with contaminated material.  Further examination for other more serious parasites might be advised.   Personal and public hygiene are key in the prevention this parasite.

Morphology:

Chilomastix mesnili has both trophozoites and cyst forms.  Diagnosis is made by detecting one or both of these forms in fecal specimens.

Trophozoites:   ‘Trophs’ are elongate, tapering towards the posterior end.  They can measure 6 µm to 24 µm in length but the more common range is 10 µm to 15 µm.   In a freshly obtained stool sample, the trophozoites exhibit a ‘stiff’, rotary motion.  The nucleus is generally not recognizable in unstained samples from lack of contrast.  There are three anterior flagella and a groove running in a spiral manner along the length of the organism.  In stained preparations, the single nucleus can be seen, located near the origin of the anterior flagella.  The cytostome, or oral depression, is bordered by fibrils extending one-third to one-half down the length of the organism.  The most prominent cytostomal fibril curves posteriorly around the cytostome and resembles a ‘shepherd’s crook’.  The flagella stain poorly, if at all.  The nucleus is found at the anterior end of the organism.  Peripheral nuclear chromatin may be present in the form of granules or irregularly distributed as plaques against the nuclear membrane.  A small, central or eccentric karyosome may at times be seen.

 Chilomastix mesnili -trophozoite seen here with a large nucleus in the anterior end with the flagellate tapering posteriorly. The karyosome is seen to be eccentric.  Some inclusions are visible within the cytoplasm.

(Iron-Hematoxylin Stain, 1000X, Nikon)

 Chilomastix mesnili -troph as described in the previous photo.  Karyosome is not obvious.

(Iron-hematoxylin, 1000X, Nikon)

 Chilomastix mesnili -The previous two photos were cropped to better isolate the organism.  This un-cropped photo better shows what the Chilomastix mesnili troph would appear like under the light microscope at 1000X magnification when analyzing a stool specimen.  Parasitology is a science, skill and art combined, making it one of the more challenging disciplines in clinical microbiology to master.

(Iron-hematoxylin, 1000X, Nikon)

Cysts:  The cyst formed by C.mesnili is typically described as lemon or pear shaped, with a prominent knob-like protuberance at the anterior end.  (Appearance always depends on the orientation of the organism when fixed and stained.)   This shape is characteristic of Chilomastix mesnili alone and does not appear in any other intestinal protozoa.  Cysts can range from 6 µm to 10 µm in size but generally average 7 µm to 9 µm in length, with the width being slightly less.  The cyst contains a relatively large nucleus and the chromatin may be condensed to form a large karyosome.  Curved cytostomal fibrils are usually quite prominent and frequently may give the appearance of an open safety pin, alongside the cytostome.

 Chilomastix mesnili -cyst.  Note the slightly distorted shape giving the cyst a lemon or pear-like appearance.  A rather large nucleus is seen with abundant peripheral chromatin.

(Iron-hematoxylin, 1000X, Nikon)

 Chilomastix mesnili -as above.  The skill is to find and identify parasites when often hidden or masked by fecal debris present after processing.

(Iron-hematoxylin, 1000X, DMD-108)

 Chilomastix mesnili - due to the orientation, it is difficult to tell in this photo.  The nucleus appears at one end suggesting a trophozoite, however it may also be a cyst.  Actual size measurement was not recorded for this photo.

(Iron-hematoxylin, 1000X, Nikon)

 Chilomastix mesnili -cyst

(Iron-hematoxylin, 1000X, DMD-108)

 Chilomastix mesnili -cyst (middle of photo) -other structures are artifacts.

(Iron-hematoxylin, 1000X, Nikon)

 Chilomastix mesnili - Chilomastix troph (C.m.) seen above and to the left of an Entamoeba coli cyst (E.c.) -for size comparison.

(Iron-hematoxylin, 1000X, DMD-108)

 Chilomastix mesnili -cyst (Cm) seen.  A Blastocystis hominis (Bch) cell seen at top of photo.  Cell seen near middle of photo shows to few characteristics for definitive identification but may be another Chilomastix organism.

(Iron-hematoxylin, 1000X, DMD-108)

Chilomastix mesnili - cyst in an unstained concentrate.  Can you find it without an arrow pointing to it? 

(Fecal concentrate, 400X, Nikon)

Chilomastix mesnili - trophozoite in an unstained fecal concentrate.

(Fecal concentrate, 400X, Nikon)

Again, the morphology of Chilomastix mesnili makes it rather easy to identify in well stained preparations.  However, both trophs and cysts may stain lightly making them difficult to distinguish when mixed in with fecal material.  It must be distinguished from other small parasites that may be present such as Enteromonas hominis and Retortamonas intestinalis.  

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Hortaea werneckii

Hortaea werneckii -Mould               (Phaeoannellomyces werneckii)

Ecology:

Hortaea werneckii is a saprophytic fungus which can be found in tropical and sub-tropical soils, compost, and decaying wood.  This fungus is also halophilic as it has been isolated from salted foods, saline waters and natural saltpans, growing in salt concentrations of up to 10%.

Pathogenicity:

H.werneckii is the etiological agent of superficial skin infections known as tinea nigra.  This infection usually manifests itself as brown to black macules which are most often found on the palms of the hands.  The appearance may initially resemble that of malignant melanoma.

Macroscopic Morphology:

Hortaea werneckii is a rather slow growing fungus which initially appears as shiny black, slimy or mucoid yeast.  As the colony matures, development of aerial hyphae may give the colony a velvety texture.  The reverse also has a rather non-nondescript brown-black colour.

 Hortaea werneckii SAB 16 days at 30˚C (Nikon)

Reverse: (not shown) appears much like the surface, perhaps slightly more olivaceous in colour.

Right: single colony incubated for approximately one month shows a slight velvety texture.

Microscopic Morphology:

Aerial mycelia develop and acquire the olivaceous black colour as they age.  Septate hyphae may be rather wide, reaching up to 6 µm in width.

Intercalary or lateral conidiogenous (annellide[i]) cells develop along the hyphae which produce the conidia (annelloconidia).  When released the annelloconidia show a prominent annellated ring, 1 -2 µm wide where once attached. 

Annelloconidia are also hyaline (clear) becoming translucent olivaceous brown-black in maturity.  They are smooth-walled and have a broad ellipsoidal appearance (7.0 – 9.5 µm X 3.5 – 4.5 µm).  Annelloconidia are 1 to 2 celled and the internal cell wall or septum is usually deeply pigmented.  With aging the annelloconidia may develop into chlamydoconidia-like aggregates (chlamydospores).

 Hortaea werneckii - At low magnification there is not much to see other that a sea of cells (conidia)

(KOH, 250X, DMD-108)

 

Hortaea werneckii -brown or olivaceous pigmented conidia are visible

(KOH, 400X,  DMD-108)

Hortaea werneckii - two celled conidia can now be distinguished.

(KOH, 400+10X, DMD-108)

Hortaea werneckii -Pigmented, septate hyphae with lateral annellides.

(LPCB, 1000X, DMD-108)

Hortaea werneckii -Here we can see the broadly ellipsoidal, two-celled annelloconidia.  Mature conidia have the brown pigmentation while younger cells stain more intensely with the Lactophenol Cotton Blue (LPCB).  One end of the annelloconidia usually stains darker indicating the location of the annellated ring or the point where it was previously attached to the conidogenous annellide.

(1000X, LPCB, DMD-108)

 

Hortaea werneckii -Another view of a curving septate hyphae surrounded by numerous free conidia.  Annellides can be seen at various points along the hyphae from where the conidia are produced.

(1000X, LPCB, DMD-108)

Hortaea werneckii -The arrow points to an annelide with its developing annelloconidium still attached.  Also seen is a pigmented and septate hyphal element near the top left as well as annelloconidia in various states of maturity.

(1000X, LPCB, DMD-108)

Hortaea werneckii -Another view of a developing annelloconidium attached to the parent annellide.

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

Hortaea werneckii -and one more.  Annelloconidium developing in the center of the photo.

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

Hortaea werneckii -The primarily two-celled conidia, or more precisely annelloconidia are seen here in various states of pigmentation.  Note that the septum dividing the two cells is darkly stained, as is one end, where the annelloconidium was once attached to its parent annnellide.

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

Hortaea werneckii - With aging the annelloconidia may develop into chlamydoconidia-like aggregates (chlamydospores) as seen here in the center of the photograph.

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

Hortaea werneckii -The annelloconidia can themselves "bud" and act as annellides, producing annelloconidia as seen here.  

(400X, LPCB, 400X -adhesive tape mount)

Hortaea werneckii -germination of an annelloconidium at top.  Not also the bud developing at the bottom of the two celled annelloconidium.

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

Physiology:

-Does not grow at 37˚C

-Nitrate +ve

-Urease +ve

-Halophilic up to 10%

Differentiation:

The annellated zones on Hortaea werneckii are much broader than those of Exophiala species.  Lack of growth at 37˚C also distinguishes the two.

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[i] Annellide –A specific conidiogenous cell that produces conidia in succession, each leaving a ring-like collar on the cell wall when released.  ‘Annello’ prefix simply specifies the type of conidiogenous cell or conidia which is produced.

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