Signalment:  

5-year-old, female Quarter horseLarge (15 cm diameter), ulcerative skin lesion on ventral midline

Gross Description:  

This 15 cm diameter, ulcerated, crater-like mass in the skin has a 3 cm thick fibrous wall extending through the subcutis. Within the ulcerated surface, 1 mm yellow granules are seen (kunkers) (Fig. 1)

Histopathologic Description:

The mass is mostly fibrovascular connective tissue containing a diffuse inflammation of eosinophils, lymphocytes and plasma cells. Distinct foci of coagulative necrosis are present circumscribed by a thin rim of macrophages (Fig. 2). Within the necrotic foci are hyphae that are negatively stained and show only a clear space outlined by the hyphal wall (Fig. 3). The hyphae are 10 microns in width with non-parallel sides and rare septation. A Gomoris methenamine silver stain outlines the hyphal wall (Fig. 4). 

Morphologic Diagnosis:  

Skin: chronic, ulcerative and fibrotic dermatitis with multifocal caseous granulomas and intralesional hyphae

Lab Results:  

Pythium insidiosum was cultured from the cutaneous wound and the presence of P. insidiosum DNA was confirmed by PCR

Condition:  

Pythium insidiosum

Contributor Comment:  

The gross appearance, location of the lesion, and the hyphae within the lesion are most consistent with an infection by Pythium insidiosum. The organism was cultured from the lesion and confirmed with PCR. Pythium is a water-born organism classified as an Oomycte in the kingdom Protista. The organism requires a water environment with decaying vegetation to maintain its life cycle and temperatures between 30 C and 40 C for reproduction. Infective zoospores are produced and invade damaged plant or animal tissue. Animals acquire the infection with prolonged exposure to freestanding water containing the Pythium organism.

Cutaneous disease in horses is the most common infection seen in animals, but infection is reported in many other animal species and in other locations, such as the intestinal and respiratory tracts (1). Cutaneous lesions usually occur on the distal extremities or ventrum, areas most likely to be in contact with stagnant water. 

The zoospores produce an ulcerative lesion that rapidly expands to a large mass of granulation tissue containing draining sinus tracts. Distinct yellow to tan masses, 2-10 mm in diameter, form within the sinus tracts and are known as kunkers. Kunkers represent areas of tissue necrosis containing hyphae and necrotic eosinophilic inflammation.

JPC Diagnosis:  

Haired skin: Dermatitis and panniculitis, pyogranulomatous and eosinophilic, focally extensive, severe, with ulceration, vasculitis, and few hyphal structures, Quarter Horse (Equus caballus), equine.

Conference Comment:  

There is significant slide variation with some sections having prominent vasculitis with fibrin thrombi.

Pythium insidiosum and Lagenidium sp. are aquatic dimorphic water molds of the kingdom Protista. Lagenidium sp. have only been described in dogs.(4) The infective stage of pythium is a biflagellate zoospore that is attracted chemotactically to injured tissue. Upon contact with host tissue the zoospores lose their flagellae and form germ tubes to allow penetration and invasion of tissue.(7)

Histologic features of pythiosis are granulomatous inflammation with foci of liquefactive necrosis and extensive fibrosis with eosinophilic coagula. Circulating monocytes are recruited from the circulation by several chemotactic stimuli such as C5a, TGF-α, and platelet derived growth factor. The tissue macrophages are then activated primarily by IFN-γ, or by endotoxin.(5)

Pythium sp. stain well with GMS but do not stain well with PAS.(7) Fungal hyphae or hyphal-like structures are distinctive between various entities and are often described according to the following structural features.(6)

Hyphae with conidiophores

Species	Hyphael type	Septae	Width	Branching	Walls	Misc
Aspergillus sp	Hyphae, +/- fruiting bodies	Regularly septate	2.5-4.5 um	Acute angle, dichotomous	Parallel walls	Conidia not seen in tissue
Candida sp.	Hyphae, pseudohyphae & budding yeast	septate	3-6 um	Irregular branching
Pseudallescheria boydii	septate	5 um	Less acute, highly branching, intertwined	15-20um intercalary chlamydoconidia (swollen cells)
Pythium insidiosum	Hyphal like structures	Rarely septate	2-9 um	Irregularly branching	Non-parallel	Kunkers or leeches
Zygomycetes	May occur as 5-30um chlamydoconidia	Few septae	3-25 um	Non-dichotomous irregular branching	Non-parallel	Mucor sp., Basidiobolus sp., Rhizopus sp.

Eosinophils are a prominent feature of equine pythiosis. Residents discussed the four primary differentials for eosinophilic and granulomatous dermatitis in a horse:
1. Pythiosis Hyphal-like structures, negative images on H&E that are highlighted with GMS stain, necrotizing vasculitis
2. Habronema Nematode larvae within eosinophilic granulomas
3. Mast cell tumors monomorphic population of mast cells surrounding eosinophilic granulomas
4. Eosinophilic collegenolytic granulomas Usually centered on collagen bundles

References:

1. Chaffin MK, Schumacher J, McMullan WC: Cutaneous pythiosis in the horse. Vet Clin N Amer:Eq Prac 11: 91-103, 1995
2. Ginn PE, Mansell JEKL, Rakich PM: Skin and appendages. In: Jubb, Kennedy and Palmers Pathology of Domestic Animals, ed. Maxie MG, 5th ed., pp 704-707. Elsevier Ltd, Philadelphia, PA 2007
3. Gross TL, Ihrke PJ, Walder EJ, Affolter VK: Skin diseases of the dog and cat, 2nd ed., pp. 303-309. Blackwell Science, Oxford, UK, 2005
4. Hargis AM, Ginn PE: The integument. In: Pathologic Basis of Veterinary Disease, eds. McGavin MD, Zachary JF, 4th ed., pp. 1196. Elsevier, St. Louis, MO, 2007
5. Kumar V, Abbas AK, Fausto N: Acute and chronic inflammation. In: Robbins and Cotran Pathologic Basis of Disease, 7th ed., pp. 877-937. Elsevier Saunders, Philadelphia, PA, 1999
6. Rippon JW: Medical Mycology: The Pathogenic Fungi and the Pathogenic Actinomycetes, pp. 569, 638, 653, 687, 739. W.B. Saunders Company, Philadelphia, PA, 1988
7. Scott DW, Miller WH: Equine Dermatolgy, pp. 287-293. Elsevier Science, St. Louis, MO, 2003

A virtual slide is not available for this case.

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