AFIP Wednesday Slide Conference - No. 8
November 06 1996

Conference Moderator: LTC Kelly J. Davis
Diplomate, ACVP
U. S. Army Medical Research Institute of Infectious Disease
Bldg. 1425
Frederick, MD 21702-5011

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Case I - 96010873 (AFIP 2551107)

Signalment: 6-year-old male mixed breed dog.

History: Dog was vaguely ill with depression, stiffness and intermittent fever for 4 months. The dog then became inactive, weak, developed conjunctivitis and muscle wasting. It died one month later.

Gross Pathology: A cosmetic necropsy was requested. The carcass was emaciated with gray streaks in the myocardium, an enlarged friable liver with capsular rupture and hemorrhage, hemoperitoneum, and mild splenomegaly. Skeletal muscle was pale with reduced mass. The temporal muscles were especially pale and atrophied.

Laboratory Results: WBC = 136,000, 83% segmented neutrophils and 12% bands.

Contributor's Diagnosis and Comments: Skeletal muscle, myositis, granulomatous with extensive atrophy due to Hepatozoon canis.

Parasitic forms of H. canis occur in many organs and tissues of the body but are most frequently present in skeletal and cardiac muscle. Inflammatory changes associated with encysted forms of the organism tend to be minimal. Granulomatous responses appear to occur when excystment and gamont development occur.

AFIP Diagnosis: Skeletal muscle: Degeneration and atrophy, diffuse, moderate, with multifocal chronic-active myositis, vascular proliferation, and multiple protozoan cysts, mixed-breed, canine, etiology consistent with Hepatozoon canis..

Conference Note: In the United States, Hepatozoon canis infection has only been reported along the Texas gulf coast, where potential for spread is limited to the ideal habitat for its vector, Rhipicephalus sanguineous, the brown dog tick. Hepatozoon canis is a protozoan in the family Haemogregarinidae, phylum Apicomplexa. Distribution of Hepatozoon canis is worldwide and its life cycle is similar to other Hepatozoon species. The infected nymphal or adult brown dog tick contains the infective parasitic oocyst within its hemocoel. Ingestion of the tick is the only known mode of natural transmission. Sporozoites are released, penetrate the intestinal wall, and are then transported by the blood or lymph to mononuclear phagocytes especially in the spleen and bone marrow but also multiple other tissues. Schizonts have been reported in many tissues but are most common in the lung, myocardium, skeletal and smooth muscle; less frequent sites include liver, spleen, and lymph nodes. Mature schizonts produce merozoites which invade neutrophils and monocytes and form gametocytes. Ticks are infected by ingesting gametocytes. The gametocytes undergo syngamy in the tick gut and produce a zygote. The zygote penetrates the gut wall and undergoes sporogony in the hemocoel, completing the life cycle.

Clinically, Hepatozoon canis infections are associated with muscular pain and atrophy, lumbar pain, emaciation, lethargy, anemia, neutrophilic leukocytosis and periosteal new bone proliferation at sites of muscle attachment. Occasional lesions include pulmonary congestion, lymphadenopathy, gastric congestion, and pale kidneys. Microscopically, schizonts are classically found in skeletal and cardiac muscle, lymph nodes, spleen, liver, kidney, or skin predominantly in the form of microschizonts with a characteristic "cart wheel" appearance or cysts containing fine lamellar membranes with a foamy or onion-skin appearance. When mature schizonts rupture in muscle, there is an intense pyogranulomatous response characterized by about equal numbers of neutrophils and macrophages with varying numbers of eosinophils and few lymphocytes or plasma cells. Bony proliferation is probably due to local muscle inflammation. As demonstrated in this case, there is also vascular proliferation into the area of schizont rupture.

Contributor: Department of Anatomy, Pathology, & Pharmacology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.

1. Barton CL, Russo EA, Craig TM, Green RW. Canine hepatozoonosis: a retrospective study of 15 naturally occurring cases. J Am Anim Hosp Assoc 1985;21:125- 134.

2. Craig TM, Jones LP, Nordgren RM. Diagnosis of Hepatozoon canis by muscle biopsy, J Am Anim Hosp Assoc 1984;50:83-99.

3. Droleckey RE, Mercer SH, DeLoach JR, Graig TM. Ultrastructure of Hepatozoon canis in the dog, Vet Parasitol 1993;50:83-99.

4. Greene GE, Hall HF, Dreesen DW, Chandler FW (eds). Clinical Microbiology and Infectious Diseases of the Dog and Cat, W.B. Saunders Company, 1984, pp. 771- 780.

5. Baneth G, Harmelin A, Presentey B. Hepatozoon canis infection in two dogs. J Am Vet Med Assoc 1995;206(12):1891-94.

International Veterinary Pathology Slide Bank:
Laser disc frame #11088, 14304, 14305.


Case II - 23268 (AFIP 2549332)

Signalment: Adult female Thoroughbred horse.

History: This was a "redbag" presentation. The placenta was delivered with a live full term foal.

Gross Pathology: The allantochorion weighed 27 pounds and the amnion weighed 7 pounds. The chorionic surface of the placenta was diffusely granular, thickened, and slightly pale. The amnion was also edematous.

Laboratory Results: Cultures of the placenta yielded Cryptococcus neoformansin numerous quantities along with nonpathogenic bacteria. A direct florescent antibody test for Leptospira sp. was negative.

Contributor's Diagnosis and Comments: Placentitis, diffuse with numerous intralesional encapsulated yeast. Etiology: Cryptococcus neoformans.

Cryptococcal infection in horses generally localizes in the lungs or brain. Infection of the reproductive tract is rare, but has been reported previously in an American Paint mare1 and in an Arabian mare4. The description of the placenta in the latter case was similar to the current submission. Cryptococcal organisms were noted in the lungs of the fetuses in both cases. The foal in the present case developed signs of pneumonia at 5 days of age and died at 9 days of age. Cryptococcal organisms were abundant throughout the lungs and bronchial lymph nodes.

Mycotic placentitis in the horse primarily occurs in late gestation. Filamentous fungi cause locally extensive placentitis at the cervical star and adjacent area which is attributed to an ascending infection. Cases of Histoplasma and Candida infection have involved the entire placenta, resulting in either multifocal granulomatous inflammation or a diffuse necrotizing and proliferative placentitis. Spread of the organisms in these cases may have been through the blood stream to the endometrium. The inflammatory response in the placenta of the present case was minimal to absent. A granulomatous inflammatory response was observed in an endometrial biopsy of the mare, and granulomatous inflammation was also present in the lung of the foal. Although the route of infection of this mare could not be determined, the most likely route was via the urogenital tract.

Clearance of the cryptococcal organisms from the body is dependent upon the pathogenicity of the infecting strain of Cryptococcus as well as an effective cell-mediated immune response in the host. The mare in this case showed no signs of systemic infection. Samples obtained from the uterus and clitoral sinus 6 months post-foaling had no evidence of Cryptococcus by culture or cytologic examination.

AFIP Diagnosis: Chorioallantois: Placentitis, histiocytic, diffuse, mild, with numerous yeasts, Thoroughbred, equine, etiology consistent with Cryptococcus neoformans.

Conference Note: Participants agreed that the morphology of the budding yeast is consistent with Cryptococcus neoformans. In H&E stained sections, C. neoformansappears as 2-20 m diameter, thin walled, slightly basophilic spherical cells with infrequent, single, narrow-based buds; yeast are typically surrounded by a wide non-staining capsule. A mucicarmine stain viewed in conference helped demonstrate the typical outer capsule characteristic of C. neoformans. Histologically, the most striking feature is an almost total lack of inflammation in the face of numerous yeasts. The capsule of Cryptococcus neoformans is composed of mucopolysaccharides which are thought to inhibit macrophage phagocytosis and antigen-antibody interactions, providing an explanation for the lack of inflammation.

Contributor: Livestock Disease Diagnostic Center, University of Kentucky, 1429 Newtown Pike, Lexington, KY 40511-1280.

1. Blanchard PC, Filkins M: Cryptococcal pneumonia and abortion in a equine fetus. J Am Vet Med Assoc. 1992, 201:1591-1592.

2. Hong CB, Donahue JM, Giles RC, et al: Etiology and pathology of equine placentitis. J Vet Diag Invest, 1993, 5:56-63.

3. Petrites-Murphy MB, Robbins LA, Donahue JM, et al: Equine cryptococcal endometritis and placentitis with neonatal cryptococcal pneumonia. J Vet Diag Invest, 1996, (in press).

4. Ryan MJ, Wyand DS: Cryptococcus as a cause of neonatal pneumonia and abortion in two horses. Vet Pathol, 1981, 18:270-272.

5. Jubb KVF, Kennedy PC, Palmer N: Pathology of Domestic Animals, 4th ed., Vol. 2, Academic Press, 1993, p. 668.

International Veterinary Pathology Slide Bank:
Laser disc frame #5081, 20866.


Case III - 96A 3291 (AFIP 2549711)

Signalment: One-year-old male Domestic Shorthair cat.

History: Two day history of pyrexia (105 F), dehydration, anemia and leukocytosis; died despite supportive care.

Gross Pathology: The cat was icteric, and the lungs were edematous. Tracheobronchial lymph nodes were hyperplastic. The spleen was enlarged.

Laboratory Results: FA for FeLV was negative. The hematocrit was 17.8% and the white blood cell count was 23,000 cells/ l.

Contributor's Diagnosis and Comments: Morphologic diagnosis: Diffuse pulmonary alveolitis with cytoplasmic schizonts within intravascular macrophages.

Disease diagnosis: Cytauxzoonosis.

Cause: Cytauxzoon felis.

Histopathology: In pulmonary venules, large (40 m diameter) intravascular macrophages with abundant foamy cytoplasm contain schizonts of Cytauxzoon felis. These intracellular schizonts are filled with numerous 1-5 m basophilic merozoites. Some pulmonary venules are characterized by fibrinoid necrosis with degenerate leukocytes within the walls. The lungs are edematous as evidenced by a patchy distribution of proteinic fluid and fibrin in alveoli, numerous alveolar macrophages and scattered hemorrhage.

Cytauxzoonosis is an uncommon fatal disease of domestic cats in the southeastern and midwestern United States. Cytauxzoon is a protozoan classified in the order Piroplasmida and family Theileriidae. The family Theileriidae also includes the genera Theileria and Gonderia. In the family Theileriidae there is an erythrocytic stage of infection and a leukocytic or tissue phase. The schizonts of Cytauxzoon develop within macrophages while Theileria and Gonderia have their leukocytic stage in lymphocytes. The bobcat is thought to be a reservoir host for Cytauxzoon felis and the brown dog tick (Dermacentor variabilis) is known to transmit the protozoa. Domestic cats with C.felisinfection usually present with a hemolytic crisis. An antemortem diagnosis was made in this case by demonstrating piroplasms in erythrocytes of Giemsa and Wright's stained blood smears.

AFIP Diagnosis: Lung: Pneumonia, interstitial, subacute, diffuse, moderate, with intramonocytic, intravascular schizonts, Domestic Shorthair, feline, etiology consistent with Cytauxzoon felis.

The domestic cat is considered to be a dead-end host due to the rapidly fatal course of the disease. Infected cats become febrile and have signs typical of an acute hemolytic anemia (depression, generalized pallor and icterus). Clinically, cats are depressed, anoretic, pyrexic, dehydrated, pale and jaundiced. Death commonly occurs within a few days of onset of clinical signs. Gross lesions include splenomegaly, enlarged edematous mesenteric lymph nodes often with petechia, ecchymotic hemorrhages in the lung and other viscera, and distended abdominal vessels. Microscopically, the presence of prominent, large schizont-containing phagocytic cells within and often occluding the lumens of vessels in the lung and spleen is considered pathognomonic.

The erythrocytic forms of C. felis are generally single, 1-1.5 m diameter organisms within the erythrocyte having a signet-ring like appearance. Differential diagnosis for this phase should include Haemobartonella felis, Babesia felis, Howell-Jolly bodies, or stain precipitate. The leukocytic or tissue phase must be distinguished from Toxoplasma, Leishmania, and Histoplasma. Organisms are best seen in Wright's or Giemsa-stained thin blood smears or impression smears of the spleen.

In domestic cats, cytauxzoonosis is almost universally fatal, although some cats survive for unknown reasons. Immunocompetence appears to influence the outcome of cytauxzoonosis in bobcats. In these animals, the disease is generally benign, but may be fatal if the animal is stressed by capture or transport. Protective immunity against apicomplexan hemoparasites requires specific T-cell cytotoxicity with resultant killing of infected leukocytes and erythrocytes.

Contributor: College of Veterinary Medicine, Department of Pathology, University of Georgia, Athens, GA 30602.

1. Garner MM, Lung NP, et al. Fatal Cytauxzoonosis in a captive-reared white tiger (Panthera tigris). Vet Pathol, 1996, 33:82-86.

2. Kier AB. Cytauxzoon. In : Infectious Diseases of the Dog and Cat. Green CE (ed). W.B. Saunders Company. pp 792-795, 1990.

3. Kier AB. The pathology of experimental Cytauxzoonosis. J Comp Path. 1987, 97:415-432.

4. Hoover JP, Walker DB, Hedges JD. Cytauxzoonosis in cats: Eight cases (1985-1992). J Am Vet Med Assoc, 1994, 205(3): 455-460.

5. Walker DB, Cowell RL. Survival of a domestic cat with naturally acquired cytauxzoonosis. J Am Vet Med Assoc, 1995, 206(9): 13631365.

International Veterinary Pathology Slide Bank:
Laser disc frame #5298, 5829, 5830, 5831, 7986, 9642, 13102, 13673, 14300, 14534.


Case IV - N95 1782 (AFIP 2549850)

Signalment: 5-day-old female Landrace pig.

History: Signs of illness seen from day 1 to day 3. Initially slight dehydration and increased respiratory rate. Advanced clinical signs developed within 24 hours; severe dehydration and fluid accumulation in abdomen, dyspnea, pyrexia, scours, high mortality. No response to antibiotics.

Gross Pathology: Five piglets received - 4 live, 1 dead. Similar gross lesions in all five. Severe mesocolonic edema and ascites. Congestion of colonic serosa and mucosa. Colonic lumen contains variable amounts of green semifluid to solid or dry fecal material. Large volume (20 ml) of clear fluid in thoracic cavity.

Laboratory Results: Clostridium difficile toxins were demonstrated in colonic contents by mouse inoculations. Mouse protection achieved with specific antiserum.

Contributor's Diagnosis and Comments:
1. Diffuse acute colitis and edema.
2. Multifocal erosive fibrinonecrotizing colitis.
3. Severe edema and inflammation of mesocolon.

Disease: Clostridium difficile - Colitis and enterotoxemia.

There is severe diffuse edema of the mesocolon, colonic serosa, submucosa and lamina propria. Edema is accompanied by light to focally heavy infiltration of neutrophils, with focal heavy infiltration of walls of venules in the mesocolonic tissue. The colonic lamina propria is focally extremely dilated and the surface epithelium is focally necrotic or lost. These foci are characterized by heavy neutrophil infiltrations of the lamina propria and/or nodular attached masses of fibrinopurulent exudate projecting into the lumen from the eroded surface. In other areas, the surface epithelium is cuboidal, and there is blebbing and shedding of epithelium. Crypt epithelium is hyperplastic. Numerous large bacilli are present in the lumen.

AFIP Diagnosis: Colon: Colitis, acute, diffuse, moderate, with erosions, vasculitis, marked transmural edema, and marked venular neutrophilic margination and emigration, Landrace, porcine.

Conference Note: Clostridium difficile is the most frequently identified enteric pathogen in humans with antibiotic-associated diarrhea and colitis. It is infrequently reported in animals but has been implicated in antibiotic-associated typhlitis/colitis in hamsters and guinea pigs, acute hemorrhagic necrotizing enteritis in neonatal foals and severe ulcerative colitis in dogs. Clostridium spiroforme is another causative agent in antibiotic-associated colitis in laboratory animals.

C. difficile is not a significant component of the microflora of healthy adult humans or animals; however, it can establish large populations in antibiotic treated or gnotobiotic animals and in infants before they acquire a complete flora.

C. difficile is a gram-positive anaerobic bacillus that produces two protein exotoxins: toxin A (Tx-A) and toxin B, which have different physicochemical and biological properties. Both are lethal toxins when administered parentally to experimental animals, and the minimum lethal dose is about the same for each (~100 fold less than that for C. botulinum toxin). The role of toxin A and toxin B as lethal toxins remain unclear, but what is known is that toxin A is responsible for nearly all changes within the gastrointestinal tract. Toxin B is known to be cytopathic in tissue culture.

Toxin A causes a marked increase in leukocyte adherence and emigration accompanied by albumen leakage, mast cell degranulation, release of prostaglandin E2, leukotriene B4 (LTB4) and platelet activating factor (PAF).

Contributor: Western College of Veterinary Medicine, 52 Campus Drive, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5B4.

1. Wilson KH. The Microecology of Clostridium difficile. Clinical Infectious Diseases. 1993;16(Suppl 4):5214-218.

2. Bartlett JG. Clostridium difficile: History of its Role as a Enteric Pathogen and the Current State of Knowledge About the Organism. Clin Inf Dis. 1994;18(Suppl 4):S265-272.

3. Kurose I. et al. Clostridium difficile Toxin A-induced Microvascular Dysfunction. J Clin Invest. 1994;94:1919-1926.

4. Jubb KVF, Kennedy PC, Palmer N. Pathology of Domestic Animals, 4th ed., Vol. 2, 1993, pg. 244.

5. Jones TC, Mohr U, Hunt RD (eds). Monographs on Pathology of Laboratory Animal Sponsored by The International Life Sciences Institute: Digestive System, Springer-Verlag, 1985, pp. 343-346.

International Veterinary Pathology Slide Bank: None

Lance Batey
Captain, VC, USA
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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