"

SUBCutaneous mycoses

Subcutaneous mycoses involve the dermis, subcutaneous tissues, muscle and fascia. These infections are chronic causing a pyogranulomatous inflammatory response and can be initiated by piercing trauma to the skin which allows the fungi to enter.

Common sub-cutaneous infections include:

  1. “Kunkers” in horses and dogs caused by the agent of swamp cancer (Pythium insidiosum,  Lagenidium and Paralagenidium species – not a fungus = oomycete), The primitive fungi Basidiobolus ranarum and Conidiobolus coronatus which are found in moist tropical environments also cause similar lesions.
  2. Cantaneous lymphangitis in horses due to the thermal dimorphic fungi Sporothrix schenckii (rare in Australia) and Histoplasma farcinomosum (exotic to Australia)
  3. Mycetomas which are single often expanding fungal granulomas caused by a variety of fungi found in the soil or plants. They can either be caused by dark pigmented or dematiaceous fungi = phaehypomycosis or the unpigmented fungi (hypomycosis).
  4. Protothecosis is not caused by a fungus, but rather algae, belonging to the genus Prototheca,  that is unable to produce cholorophyll.

Oomycota

Learning Objective

Discuss  the cause, diagnosis and treatment “kunkers” as an example of a subcutaneous mycoses in the tropics. Also discuss why it is so common in the Townsville district

The oomycetes are known as water moulds (misnomer) and typically grow on plants in moist environments. These primitive protozoa differ from true fungi in that they have cellulose instead of chitin in their outer cell membrane and the vegetative structures are diploid (2n). They also don’t have septa in their hyphae. Most of the oomycetes are plant and insect pathogens. The name of this group is derived from the presence of an oogonium where eggs are fertilised by another mating type and oospores develop. In animals pathogenic genera include:

  1. Saprolegnia and Aphanomyces: Superficial infections of fish and crustacea. Aphanomyces invadens causes red spot disease, also known as epizootic ulcerative syndrome in fish. It is endemic in Australia and is a notifiable disease. Saprolegnia parasitica is the cause of winter disease in freshwater fish as outbreaks occur when the water temperature is below 16°C. It is characterised by a cotton-like appearance on the gills and body. Both have been introduced to Australia and are present in wild fish populations.
  2. Pythium,  Lagenidium and Paralagenidium: Sub-cutaneous granulomas in horses, dogs and other animals. People can also be infected.

 

The water associated life cycle of an oomycete such as Pythium and Lagenidium
The water associated life cycle of an oomycete such as Pythium and Lagenidium. JP TO REDRAW

Pythiosis (swamp cancer) (KNOW)

Pythiosis is an infectious disease of especially horses and dogs caused by oomycetes; Pythium insidiosum,  Lagenidium and Paralagenidium species. They are endemic to the tropics, including Townsville, and naturally inhabit wetlands, pond, and swamps. The oomycete produces motile flagellate zoospores that invade wounds or skin that is water maceration damaged resulting in extensive ulcerative pyogranulomatous lesions that are rich in eosinophils. Present in these lesions are coral-like structures knowns as “kunkers”. These are usually rich in the hyphal structures of the oomycete.

Townsville horses with granulomatous lesions of the hind limb, chest and ventral abdomen (left picture) caused by P. insidiosum (picture on the right). These were surgically removed
Townsville horses with granulomatous lesions of the left hind limb, chest and ventral abdomen caused by P. insidiosum. These were surgically removed. Images on the left from the collection of James Cook University, used with permission. All Rights Reserved.  Image on the right by Brad Dowling, James Cook University, used with permission. All Rights Reserved.
German Shepherd Dog with an ulcer of the chest caused by Lagenidium species
Townsville German Shepherd Dog with an ulcer of the chest caused by Lagenidium species

A rare gastrointestinal form is also recognised in dogs. Dogs with gastrointestinal pythiosis often have a history of retrieving objects, such as sticks, from water and then chewing on them; young male retriever-type dogs are particularly at risk. Gastrointestinal pythiosis is usually a fatal disease. Pathologically, the organism is highly invasive and angiotropic, so the infection can be complicated by disruption of vascular supply. The infection typically involves the full-thickness of the stomach or intestine and sometimes extends to adjacent organs.

The diagnosis of this disease is usually by cytology of histology of the lesions with the presence of hyphal elements in the affected tissues. These stain well with Gomorri methanamine silver (GMS) stain but not with the fungal stain Periodic Acid- Schiff (PAS). Antibodies to Pythium insidiosum can also be detected using an ELISA.

Cytology of Phythium infections. Left picture: Grams stain of a fruiting hyphal element in a kunker showing neutrophils and an eosinophil; middle picture: DiffQuik stain showing a hyphal element surrounded by neutrophils; Right picture: GMS stain of a granuloma showing branching black stained hyphal elements
Cytology of Pythium infections. Left picture: Grams stain of a fruiting hyphal element in a kunker showing neutrophils and an eosinophil; middle picture: DiffQuik stain showing a hyphal element surrounded by neutrophils; Right picture: GMS stain of a granuloma showing branching black stained hyphal elements

 

PAS stain of an intestinal granuloma infected with Lagenidium species in a dog stained with PAS

The oomycota can be cultured on fungal media and identified. Full species identification should be done by genome sequencing.

Left picture: Culture on potato dextrose agar of Pythium. Right picture: Lactophenol cotton blue stain of the hyphal elements and sporangium of Pythium.
Left picture: Culture on potato dextrose agar of Pythium. Right picture: Lactophenol cotton blue stain of the hyphal elements and sporangium of Pythium.

Surgical excision of the lesions with healing by secondary intention combined with topical or systemic antifungal is the treatment of choice.  Spread of the infection to the bone and reoccurrence of lesions after excision are considered to be a poor prognostic indicators. Dogs often require long term treatment with antifungals, antibacterials like doxycycline, and corticosteroids.

True fungal causes of sub-cutaneous granulomas in horses: entomopthoromycosis (KNOW)

Like swamp cancer similar sub-cutaneous granulomatous lesions called entomopthoromycosis can be caused by  two fungi with wide hyphae that are poorly septate. They are known as Basidiobolus ranarum and Conidiobolus coronatus and tend to more often infect horses.  These fungi are also common in wet, tropical environments like Townsville. Clinically and histologically the lesions are difficult to distinguish from oomycete infections as kunkers is present in both and both have wide, branching non-septate hyphae and elicit an eosinophilic response. Interestingly these fungi, unlike the oomycetes, will stain with PAS.

 

Zoopagomycota. A: Growth of Basidiobolus ranaum on Sabouraud's dextrose; B: Sellotape impression of surface of Basidiobolus colony stained with LPCB; C: Colony of conidiobolus coronatus on Sabourauds dextrose agar; D: slide culture on Conidiobolus mounted on LPCB x400.
Zoopagomycota. A: Growth of Basidiobolus ranaum on Sabouraud’s dextrose agar; B: Sellotape impression of surface of Basidiobolus ranaum colony stained with LPCB x400; C: Colony of Conidiobolus coronatus on Sabourauds dextrose agar; D: slide culture on Conidiobolus coronatus mounted on LPCB x400.

Treatment of this disease is reliant on surgical resection of the lesions and antifungal therapy.

cutaneous lymphangitis complex (Know name and disease, No details required)

Traumatic implantation of fungi and bacteria in the skin of horses can lead to infection of the cutaneous lymphatics, with the formation of small pyogranulomas along the length of the lymphatics. Fungi that cause these infections tend to be thermally dimorphic. A yeast at body temperature and a mould at room temperature. The yeast phase is usually ingested by macrophages which allows them to survive and divide by budding in the macrophages. These fungi are slow-growing taking up to 4 weeks to grow. Their identification should always be confirmed by showing growth at both room temperature and 37 C as well as by morphological characteristics. Gene sequencing should also be done.

Two fungal diseases will be discussed:

  1. Sporothricosis
  2. Epizootic lymphangitis

Sporothricosis caused by Sporothrix schenckii  (KNOW)

Sporothrix schenckii (teleomorph:Ophiostoma stenoceras), a decomposing wood and hay dwelling fungus, is a common cause of sub-cutaneous lesions in horses, dog, cats and people in the world. Infections are rare in Australia. In people it is often called “rose gardener’s disease” as the fungus is found on dead woody plants and can be implanted in the skin when it is injured by thorns or bits of wood. Human cases in Australia have occurred after handling mouldy hay. Recorded as a zoonosis in the USA. People with scratch or bite injuries have been infected percutaneously by handling dogs and cats with lesions. After infecting damaged skin, the fungus causes pyogranulomas in the subcutaneous lymphatics causing a chain of lumps as shown in the picture. This fungus is a yeast in tissue often being found intracellularly in macrophages and a dark mould when grown at room temperature.

A. Sporothricosis in a horse. Note the sub-cutaneous nodules and ulcerated lesions. B: A culture of Sporothrix schenckii incubated at 37°C. Note that it is a yeast with a similar morphology to what it is intracellularly in macrophages. C: Culture of Sporothrix schenckii at room temperature. It is a mould at this temperature
A. Sporothricosis in a horse. Note the sub-cutaneous nodules and ulcerated lesions. They can form a chain along the skin lymphatics; B: A culture of Sporothrix schenckii incubated at 37°C. Note that it is a yeast with a similar morphology to what it is intracellularly in macrophages. C: Culture of Sporothrix schenckii at room temperature. It is a mould at this temperature

Epizootic lymphangitis caused by Histoplasma farcinomosum (brief no details)

Absent from Australia, this thermal dimorphic fungus known as Histoplasma farcinomosum (related to Histoplasma capsulatum) causes similar lesions in horses, but more dramatic to that of sporotrichosis. It is carried by horses. This is a Notifiable Disease and cases are reported to the WOAH.  The pictures below show the disease in horses in Ethiopia, a country in Africa that is endemic for this fungus. Cytology and histology of affected tissues reveal, signet-ring shaped yeasts swelling the cytoplasm of macrophages.

A: Subcutaneous nodules along the leg of a horse in Ethiopia infected with H. farcinomosum.; B: Ulcerated lesions in a horse in Ethiopia due to H. farcinomosum. These lesions heal with scarring.
A: Subcutaneous nodules along the leg of a horse in Ethiopia infected with H. farcinomosum; B: Ulcerated lesions in a horse in Ethiopia due to H. farcinomosum. These lesions heal with scarring. Image author unknown. All Rights Reserved. Contact the JCU Library OER team if you know the author.
Histoplasma farcinomosum cytology. A: H. farcinomosum yeasts in the cytoplasm of macrophages. DiffQuik stain; B: H. farcinomosum yeasts in the exduate of a sub-cutaneous granuloma from a horse. Gram's stain.
Histoplasma farcinomosum cytology. A: H. farcinomosum yeasts in the cytoplasm of macrophages. DiffTMQuik stain; B: H. farcinomosum yeasts in the exduate of a sub-cutaneous granuloma from a horse. Gram’s stain

Diagnosis of the disease can also be made using the eye intradermal test using proteins purified from Histoplasma farcinomosum and serology using the Complement-fixation test.

Protothecosis

Just know name and diseases it causes. It is a rare infection.

Protothecosis is a rare cutaneous, sub-cutaneous and systemic infection of primarily dogs, sometimes cattle and rarely cats caused by  non-chlorophyll producing algae known as Prototheca. This algae is common in the environment and is found globally. Disease is more common in warm climates and for its size, Townsville has an unusually high number of cases. Common clinical signs in cattle are chronic mastitis with infection via the teat canal and in cats a cutaneous infection. It is in dogs where the disease often becomes disseminated. Dependent on the portal of entry, it is initially a gastrointestinal tract or cutaneous infection. Thereafter the algae disseminate to the rest of the body, infection becoming established in a number of different organs including the lymph nodes, brain, eyes and kidneys. Being rare and unspecific in its clinical signs, protothecosis is rarely suspected as a cause of infection.

Diagnosis is based on cytology of fine needle aspirates or biopsies of lesions, cerebral spinal fluid or urine. Whilst they may be observed with most stains including DiffTMQuik, Periodic-Acid Schiff (PAS) stains reveal them the best. Algae are distinguished cytologically from yeasts by being oval or round in shape with a thick, refractile cell wall. They do not divide by budding as yeasts do but by produce sporangia with 2 to 20 endospores. They are easily cultured on Sabouraud’s dextrose medium producing white yeast-like colonies. Two species are known to be pathogenic, the most common is Prototheca zopfii and then Prototheca wickermanii.

Cutaneous lesions should be surgically removed with a wide margin. Their prognosis is much better than animals with systemic disease. Protothecosis like most systemic fungal infections should be treated with amphotericin B short-term following by an azole long-term therapy.

Left - PAS staining subcutaneous granuloma in a dog (Tessa Mackie, 2020); Right - Trypan blue wet mount of cultured Prototheca zopfii. The blue arrows point to sporangia with endospores
Left – PAS staining subcutaneous granuloma in a dog.  (Tessa Mackie, 2020); Right – Trypan blue wet mount of cultured Prototheca zopfii. The blue arrows in both pictures point to sporangia with endospores

 

END OF CHAPTER

License

Bacterial and Fungal Diseases of Animals Copyright © by Jackie Picard. All Rights Reserved.

Share This Book