AFIP Wednesday Slide Conference - No. 30
May 17, 2000

Conference Moderator:
LTC Thomas P. Lipscomb
Division of Veterinary Pathology
Armed Forces Institute of Pathology
Washington DC 20306-6000
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Case I - 98C10269-53, 54, 55, or 56 (AFIP 2679490)
Signalment: 2-year-old, castrated, 29-kg husky-German shepherd crossbred dog.
History: The dog developed sensory ataxia and conscious proprioceptive deficits in all limbs. The clinical course began abruptly with lameness of the left pelvic limb. The dog walked with a distinctive uncoordinated stumbling gait. Neurological examination revealed loss of proprioceptive and patellar reflexes. Cranial nerves and deep pain reflexes were clinically normal, megaesophagus was absent, and clinical signs were constant. The dog was referred to a veterinary teaching hospital where the tentative diagnosis was diffuse lower motor neuron disease.
Serological test results were "consistent with myasthenia gravis." The dog was treated without improvement with pyridostigmine bromide syrup (Mestinon®). Two attempts were made to corroborate the diagnosis of myasthenia gravis using a short- and rapid-acting cholinergic compound (edrophonium chloride; Tensilon® test). On both occasions the dog's status failed to improve and the clinician concluded the serological finding indicating myasthenia gravis was unrelated to sensory ataxia. The animal's condition continued to deteriorate, in spite of transient remission of some clinical signs. The dog was euthanized after a clinical course of four months.
Case 30-1. Spinal cord, dura. There is a pale white discoloration affecting gray and white matter of the dorsal funiculi.
Gross Pathology: Necropsy revealed a 32.5-kg dog in good flesh. There was no gross evidence of muscular atrophy. A V-shaped area of marked pallor involved dorsal columns at all levels of the spinal cord.
Laboratory Results: Immunoprecipitation radioimmunoassay for AChR antibodies: 2A3 nmol/L. (Reference range in healthy dogs: <0.6 nmol/L)
Contributor's Diagnosis and Comments: Axonal (Wallerian) degeneration, severe, diffuse, chronic, bilaterally symmetrical, dorsal columns and dorsal spinal nerve roots, with associated mild chronic lymphocytic-histiocytic myelitis and neuritis.
The other major change in this dog was multifocal lymphocytic-histiocytic ganglioneuritis of spinal ganglia with neuronal loss and Nageotte nodules. Other lesions were the presence of uncharacterized, PAS-negative spherical inclusions in neurohypophysis, supraoptic hypothalamus and ventromedial periventricular thalamic nuclei; axonal degeneration restricted to selected fascicles in radial and ulnar nerves; moderate diffuse skeletal muscular atrophy in M flexor carpi ulnaris; and mild ganglioneuritis in an unidentified ganglion adjacent to adrenal gland. In addition to the submitted levels of spinal cord, some of the submitted blocks include levels of thalamus and medulla oblongata. Unfortunately the thalamic inclusions are highly localized and are not present in most of the submitted slides.
Clinical signs and lesions were consistent with a diagnosis of sensory neuropathy. The condition has various sobriquets in addition to sensory neuropathy: sensory neuronopathy, chronic idiopathic polyneuritis, and ganglioradiculitis. Degeneration in sensory tracts in dorsal columns is attributable to loss of somata in dorsal root ganglia. The prognosis in the disease is poor. In this case, there was no consistent response to immunosuppression with corticosteroids. The waxing and waning course, which is typical of some cases of this disease, makes it hard to assess the effectiveness of medications used to slow clinical progression.
In this case no attempt was made to isolate infectious agents, including viruses. Nothing in the animal's history indicated recent exposure to toxic compounds, including medications. Huskies are over-represented in case reports and it may be significant the dog was part husky. The submitting veterinarian was unable to obtain a history about the fate of this dog's littermates. Summers et al speculate this disease is an autoimmune T-lymphocyte mediated reaction directed against spinal ganglia, and note its similarity to Sjögren's syndrome in people. I was unsure of the significance of positive results indicating myasthenia gravis, an autoimmune disease that leads to loss of acetylcholine receptors in neuromuscular junctions. Clinical signs and response to treatment were inconsistent with generalized myasthenia gravis. A recent large-scale retrospective study of acquired myasthenia gravis in dogs did not report an association with sensory neuropathy, and it has not been noted in case reports of this disease.
AFIP Diagnoses:
1. Spinal cord, dorsal columns: Axonal (Wallerian) degeneration, diffuse, severe, with numerous gitter cells, mild lymphocytic meningomyelitis and mild astrocytosis, husky X German shepherd dog cross, canine.
2. Dorsal spinal nerve roots: Axonal (Wallerian) degeneration, diffuse, severe, with Schwann cell proliferation, gitter cells and mild lymphocytic neuritis.
Conference Note: Conference participants agreed that the lesions suggested sensory neuropathy. Much of the discussion centered on Wallerian degeneration. Classically, Wallerian degeneration includes the changes that take place in the distal segment of a transected axon in the peripheral nervous system. There are significant differences between axonal injury in the peripheral and central nervous systems. Thus, some refer to similar changes in the CNS as Wallerian-type degeneration. In either location, the process basically consists of the breakdown and removal of the distal axon and the myelin tube.
Differences in axonal repair have major clinical significance. In the PNS, axonal sprouts growing from the stump of the proximal axon are guided by the preexisting basal lamina into a newly formed column of Schwann cells. In the CNS, there is no basal lamina and oligodendrocytes don't form columns to guide reinnervation. Additionally, oligodendrocyte myelin proteins inhibit axonal sprouting. Thus, CNS axonal repair is much less effective than that in the PNS.
Contributor: Wyoming State Veterinary Laboratory, 1174 Snowy Range Road, Laramie, WY 82070.
1. Shelton GD, Schule A, Kass PH: Risk factors for acquired myasthenia gravis in dogs: 1,154 cases (1991-1995). J Am Vet Med Assoc 211(11):1428-31, 1997
2. Summers BA, Cummings YF, de Lahunta A: Veterinary Neuropathology, pp. 428-431. Mosby-Year Book Inc., Baltimore, MD, 1995
Case II - HB2418 (AFIP 2602959)
Signalment: A nine-year-old female mixed breed dog.
History: This dog had large subcutaneous mass, located on the right side of the body, close to the diaphragm. Initially, a needle biopsy was performed, but the specimen was inadequate for diagnosis. Then the mass was incompletely resected surgically.
Gross Pathology: This large mass, 8.8 x 7.9 x 7.4 cm, was not associated with bone (by X-ray). It was adhered to diaphragm, pink and granular on cut surface.
Contributor's Diagnosis and Comments: The neoplastic cells are arranged in solid sheets. The cells vary in size, have eosinophilic cytoplasm, are round to polyhedral, and are highly pleomorphic. The cells have large vesicular round to ovoid nuclei that contain one or more prominent, irregular-shaped nucleoli. Other characteristic features include vacuolated cells, moderate fibroplasia, osteoclast--like giant cells, and giant cells containing a single large nucleus. Mitotic figures were frequently seen.
The neoplasm was diagnosed as rhabdomyosarcoma (pleomorphic type), originating from rhabdomyoblasts. Although special staining revealed intracellular glycogen (PAS positive), striations were not seen after phosphotungstic acid hematoxylin (PTAH) staining. Immunohistochemical procedures demonstrated the presence of desmin, vimentin and myoglobin type of intermediate filaments in the neoplastic cells.
Rhabdomyosarcomas vary widely in histological appearance, depending on the growth pattern, cellularity, degree of differentiation, and configuration of the individual tumor cells. These tumors are usually highly malignant and metastasize either via the lymphatic or venous routes. Metastatic sites include lymph nodes, lung, spleen, heart and skeletal muscle.
In humans, this neoplasm is the most frequent soft tissue tumor in children but is uncommon in adults. Numerous reports have documented the occurrence of rhabdo-myosarcomas in the head and neck, the genitourinary tract and retrope-ritoneum, and the upper and lower extremities.
The age distribution is not as well documented in domestic animals, but the tumor is considered to be more frequent in younger animals. The three major types are alveolar, embryonal, and pleomorphic. Although osteoclast--like giant cells are prominent in this case, the most useful diagnostic charac-teristic of tumor cells in embryonal and pleomorphic types of rhabdomyosarcoma is the irregular angularity of the cells, including an extreme range m the size of the nuclei.
AFIP Diagnosis: Skeletal muscle: Pleomorphic rhabdomyosarcoma, mixed breed, canine.
Conference Note: Among animals, rhabdomyosarcomas are most often encountered in dogs. Pleomorphic and embryonal types are described. Most of the embryonal types have been found in the urinary bladders of young, large breed dogs. The diagnostic criteria for rhabdomyosarcoma have changed over time. When cross-striations were required for diagnosis, the tumors were very rare. With the advent of immunohistochemistry, diagnosis rests on compatible histomorphology combined with positive staining for muscle markers such as desmin, myoglobin and muscle specific actin (HHF35). The main diagnostic problem is distinguishing rhabdomyosarcoma from other pleomorphic sarcomas, principally malignant fibrous histiocytoma.
In the absence of cross striations, the most specific histomorphologic feature of rhabdomyosarcoma is the presence of rhabdomyoblasts. These cells range from slender spindle-shaped cells with a small number of peripherally placed myofibrils, to large, eosinophilic cells with a strap, ribbon, tadpole or racquet shape and one or two central nuclei and prominent nucleoli. Rhabdomyoblasts often appear as round, eosinophilic cells in which the nucleus is surrounded by filamentous stringy material. Cells with similar morphology and multiple peripheral cytoplasmic vacuoles (so called "spiderweb" cells) are also highly characteristic. Good examples of rhabdomyoblasts and spiderweb cells are present in this case.
Contributor: Laboratory of Comparative Pathology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060, Japan.
1. Enzinger FM, Weiss SW: Soft Tissue Tumors, 2nd ed, pp.448-488. Mosby, ST. Louis, MO, 1988
2. Hulland TJ: Tumors of the muscle. In: Tumors in Domestic Animals, ed. Moulton JE, pp.88-101. University of California Press, Berkeley, CA. 1990
3. Martin de las Mulas I, Vos JH, Van Mil FN: Desmin and vimentin immunocha-racterization of feline muscle tumor. Vet Pathol 29:260-262, 1992
Case III - 99-19676 (AFIP 2675234)
Signalment: 23-month-old, intact male, Italian greyhound
History: Acute onset of anorexia followed by seizures of 1-2 days duration. The dog presented to the referring veterinarian stuporous with extreme neck pain. Analysis of cerebrospinal fluid revealed a marked pleocytosis and numerous fungal organisms (morphologically consistent with Cryptococcus). Ophthalmologic examination showed papilledema. The owners elected euthanasia.
Gross Pathology: Small quantities of exudate were noted in the left nasal passage and right retrobulbar space. The cerebrospinal fluid contained white flocculent material. The brain was grossly unremarkable prior to formalin fixation. Following adequate fixation, the meninges covering virtually all of the brain and brainstem were faintly cloudy to opaque. Also noted was megaesophagus of the intrathoracic esophagus.
Contributor's Diagnosis and Comments: Severe, chronic, diffuse granulomatous meningitis with numerous intralesional yeasts (Cryptococcus) and rare multinucleated giant cells.
Although several different species exist, Cryptococcus neoformans is primarily responsible for disease in animals and man. Two variants (neoformans and gattii) and five serotypes (A, B, C, D, AD) of C. neoformans have been identified to date. In the environment, the chief reservoir of C. neoformans is avian excrement (particularly pigeon excreta). The organism is also found in the soil, grass and milk. Barring direct sunlight or desiccation, organisms may remain viable in pigeon droppings for at least 2 years. The primary route of infection appears to be inhalation of unencapsulated organisms within the environment. Following colonization of the respiratory tract, cryptococci regenerate their capsules. Infection of the CNS may result from hematogenous spread or local extension from the nasal cavity.
While this case does not pose a diagnostic challenge, it does raise questions regarding the pathogenesis of this disease, specifically: Why is there such a massive proliferation of organisms, yet a minimal inflammatory response by the host? Why is there such variation in inflammatory responses between infected individuals?
Factors governing the establishment and spread of infection include the virulence of the invading species and the status of the host's immune system. The cryptococcal capsule may be absent to small during the initial phases of infection (as in organisms isolated from the environment). In general, as in this case, organisms recovered from cerebrospinal fluid are heavily encapsulated. Many strains of C. neoformans are covered by a thick polysaccharide capsule (up to 35 microns thick) which serves as a diagnostic hallmark of cryptococcosis.
It is well established that the capsule is a prominent virulence factor and possesses antiphagocytic and tolerogenic properties. While most strains are heavily encapsulated, an acapsular strain has been described in a dog with cryptococcal lymphadenitis. In that dog, the infection was characterized by the presence of many intracellular (within macrophages) organisms and intense granulomatous inflammation. In addition to inhibition of phagocytosis, the capsule suppresses cellular and humoral immunity.
Capsular antigens may also suppress cytokine production. Purified capsular polysaccharide has been shown to inhibit TNF-a secretion induced by LPS in human monocytes. Acapsular strains of C. neoformans stimulate higher levels of cytokine production than do thinly encapsulated strains. In contrast, organisms with thick capsules are poor inducers of TNF production. Recently, an acapsular strain of C. neoformans was shown to induce higher levels of CD4, a T cell associated transmembrane glycoprotein, than encapsulated strains on human monocytes.
The exact contribution of the host's immune system is not clearly known in dogs. In humans, most cases (up to 85%) of cryptococcal meningitis have an underlying condition, which is inhibitory to the function and/or number of lymphocytes. Such conditions include immunosuppressive therapy (glucocorticoids, chemotherapeutics, or other agents), lymphoid tumors, diabetes mellitus, tuberculosis or AIDS. Five to 10% of all patients with AIDS are reported to develop cryptococcosis. While immunosuppressive disease or the use of immunosuppressive agents has been associated with cryptococcosis in dogs, one study could identify underlying immunosuppressive factors in less than 6% of affected dogs.
Most dogs with cryptococcosis are young adults with the average age of 3.5 years. The dog in the present case was slightly less than 2 years of age. Histopathology confirmed the presence of Cryptococcus in the nasal cavity and retrobulbar region (forming a dense cuff around the optic nerve and causing the papilledema noted clinically). In light of these findings, a direct extension from a rhinosinusitis is the presumptive source of the CNS infection.
AFIP Diagnosis: Cerebellum: Meningitis, granulomatous, multifocal, moderate, with numerous yeasts, Italian greyhound, canine, etiology consistent with Cryptococcus sp.
Conference Note: Cryptococcosis is the most common systemic fungal infection of cats. The nasal cavity is affected in over 80% of cases, and typical signs are those of upper respiratory tract disease. Often, a firm swelling over the bridge of the nose is evident. Skin lesions are also common and generally result from systemic dissemination.
In humans, cryptococcal infection of the CNS is associated with increased intracranial pressure. The pathogenesis is uncertain, but increased CSF osmolality caused by the presence of high molecular weight cryptococcal polysaccharide and the production of D-mannitol by the fungus are possible contributing factors.
Contributor: University of Illinois, 2001 S. Lincoln Ave, Urbana, IL 61801.
1. Jacobs GJ, Medleau L: Cryptococcosis. In Infectious Diseases of the Dog and Cat, ed. Greene CE, pp 383-390. WB Saunders Company, Philadelphia, PA, 1998
2. Lichtensteiger CA, Hilf LE: Atypical cryptococcal lymphadenitis in a dog. Vet Pathol 31:493-496, 1994
3. Pietrella D, Monari C, Retini C, Palazzetti B, Kozel TR, Vecchiarelli A: Cryptococcus neoformans and Candida albicans regulate CD4 expression on human monocytes. J Infect Dis 178:1464-1471, 1998
4. Vecchiarelli A, Retini C, Pietrella D, Monari C, Tascini C, Beccari T, Kozel TR: Downregulation by cryptococcal polysaccharide of tumor necrosis factor alpha and interleukin-1ß secretion from human monocytes. Infect and Immun 63:2919-2923, 1995
Case IV - 96-1273 (AFIP 2593977)
Signalment: 15-week-old, 3.4 kilogram, male dog (toy poodle)
History: The dog was initially vaccinated by the breeder and then two weeks later by the referring veterinarian with DHLP-P. One day after the second vaccination the dog developed corneal edema. A veterinary ophthalmologist performed a conjunctival flap for a bulla which developed on the right eye. The dog was treated postoperatively with topical and systemic corticosteroids. Three weeks after the second vaccination the dog developed a non-productive cough with increased abdominal effort. A few days later the dog had difficulty breathing and became cyanotic. A severe interstitial pulmonary pattern was present on thoracic radiographs. The dog was euthanized due to poor prognosis.
Gross Pathology: The lungs were deep red and heavy (edema) and had multiple, locally extensive, irregularly shaped white regions (approximately 2 - 5 mm) randomly distributed throughout all of the lobes. The liver was mottled tan and red on the capsular and cut surfaces of all of the lobes. Bilaterally the corneas were light blue (corneal edema).
Laboratory Results:
Canine arterial blood gas:
 Test  Result  Expected  Units
 pH  7.324  7.370 - 7.510  
 pCO2  21.0  21.5 - 35.5  MM/HG
 pO2  36  85 - 100  MM/HG
 Bicarb  11.0  15.5 - 23.5  MMOL/L
 TCO2  11.7    MMOL/L
 Base excess  -11.9  -2.0 - 2.0  
Anti-Toxoplasma gondii antibody:
Structures morphologically consistent with Toxoplasma gondii stain.
Contributor's Diagnosis and Comments: Lung: Interstitial pneumonia, severe, diffuse, with multifocal to locally extensive necrosis, intralesional protozoal organisms, and rare intracytoplasmic viral inclusion bodies.
Toxoplasma gondii and canine distemper virus (CDV).
In addition to the pulmonary lesions observed on the slide submitted, clinically significant lesions were observed in the liver (severe hepatic necrosis with intralesional protozoal organisms) and brain (encephalitis with intralesional protozoal organisms and intracytoplasmic viral inclusion bodies). Viral inclusion bodies were also observed in the gastric epithelial and superficial chief cells. The low pO2 and mild respiratory acidosis were interpreted to be due to decreased gas exchange secondary to the thickening of the alveolar septa and multifocal obliteration of alveolar lumina.
Toxoplasma gondii and Neospora caninum can be very difficult to distinguish histologically. In fact, prior to 1988, Neospora caninum was misdiagnosed as Toxoplasma gondii. However, Neospora caninum is not a new disease as retrospective studies using monoclonal antibodies to Neospora caninum on formalin fixed, paraffin embedded tissue indicate that prior cases, as early as 1957, of Toxoplasma gondii were actually Neospora caninum. The protozoal organisms in this case are most likely Toxoplasma gondii as Neospora caninum tissue cysts are only found in the central nervous system (CNS). Tissue cysts in this dog were found in the CNS as well as the lungs, liver, and an adrenal gland. Toxoplasmosis is further supported, in addition to the extensive extraneural tissue cyst distribution, by the specific staining of the organisms using immunohistochemistry with a polyclonal antibody against Toxoplasma gondii (Biogenex Laboratories - San Ramon, California).
Neospora caninum is often a primary disease in dogs whereas Toxoplasma gondii typically occurs concurrently with CDV. CDV is known to cause immunosuppression in the dog. Immunocompetent dogs can become infected with Toxoplasma gondii and elicit an immune response which precludes clinical disease. However, in the face of an immunosuppressive disease such as CDV, the host immune system can no longer keep the Toxoplasma gondii in check. It is not known in this case if steroids used to treat this animal clinically also contributed to this dog's inability to mount an immune response against Toxoplasma gondii. The corneal edema may have been vaccine-induced or secondary to the Toxoplasma gondii, but there were no specific lesions to suggest an etiology.
Case 30-4. Lung. There are many macrophages in alveoli and expanding alveolar septa. Note syncytial cell (upper right) and a cell containing tachyzoites (lower left).
Case 30-4. Lung. Immunohistochemical staining directed against Toxoplasma gondii antigens renders zoites dark red-brown.
AFIP Diagnosis: Lung: Pneumonia, interstitial, necrotizing, subacute, diffuse, severe, with type 2 pneumocyte hyperplasia, bronchiolitis, hemorrhage, numerous protozoa, few syncytia cells and few intracytoplasmic inclusion bodies, toy poodle, canine.

Conference Note: Immunohistochemistry performed at the AFIP demonstrated the presence of morbilliviral antigen confirming the diagnosis of canine distemper. Immunohistochemistry for Toxoplasma gondii was also positive.
The public health significance of toxoplasmosis was discussed in conference. Human seroprevalence approaches 100% in tropical climates. In the United States, 25 to 50% of people are antibody-positive. The vast majority of people infected after birth have no clinical illness. The most significant forms of human toxoplasmosis are transplacental infection and infection of immunosuppressed individuals. Cats are the definitive hosts of Toxoplasma gondii and contaminate the environment with oocysts; however, there is no correlation between toxoplasmosis in adults and cat ownership. Herbivores become infected by consuming oocysts. Most people become infected by consuming undercooked bradyzoite-containing meat. Most cats are infected shortly after weaning and shed oocysts for only a few weeks. Seropositive cats pose little risk to humans since they are unlikely to shed oocysts. Seronegative cats are of greater risk because they may become infected and shed oocysts.
Contributor: North Carolina State University, College of Veterinary Medicine, Department of Microbiology, Pathology, and Parasitology, 4700 Hillsborough Street, Raleigh, North Carolina 27606.
1. Dubey JP: Toxoplasmosis in dogs. Canine Pract 12:7-25, 1985
2. Dubey JP, Lindsay DS: Neosporosis. Parasit Today 9:452-458, 1993
3. Rhyan J, Dubey JP: Toxoplasmosis in an adult dog with hepatic necrosis and associated tissue cysts and tachyzoites. Canine Pract 17:6-10, 1992
4. Dubey JP, Lappin MR: Toxoplasmosis and Neosporosis. In: Infectious Diseases of the Dog and Cat, ed. Greene CE, pp 493-509. WB Saunders Company, Philadelphia, PA, 1998
J Scot Estep, DVM
Captain, United States Army
Registry of Veterinary Pathology*
Department of Veterinary Pathology
Armed Forces Institute of Pathology
(202) 782-2615; DSN: 662-2615
* The American Veterinary Medical Association and the American College of Veterinary Pathologists are co-sponsors of the Registry of Veterinary Pathology. The C.L. Davis Foundation also provides substantial support for the Registry.
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