Parasitemia due to Sarcocystis neurona‐like infection in a clinically ill domestic cat

An 8‐year‐old, 6‐kg, male neutered Domestic Shorthair cat was presented to The Ohio State University Veterinary Medical Center (OSU‐VMC) for difficulty breathing. Physical examination and thoracic radiographs indicated pneumonia, a soft‐tissue mass in the left caudal lung lobe, and diffuse pleural effusion. The effusion was classified as modified transudate. Rare extracellular elongated (~5–7 μm × 1–2 μm) zoites with a central round to oval‐shaped purple to deep purple vesicular nucleus with coarsely stippled chromatin and light blue cytoplasm were seen on a peripheral blood smear. Serum IgG and IgM were positive for Sarcocystis sp. antibodies and negative for Toxoplasma gondii antibodies, suggesting that the infection was acute rather than a recrudescence of prior infection. This organism was most consistent with either Sarcocystis neurona or Sarcocystis dasypi based on DNA sequence analysis of PCR products using COC ssRNA, ITS‐1, snSAG2, and JNB25/JD396 primer sets. This is the first report to visualize by light microscopy circulating Sarcocystis sp. merozoites in the peripheral blood of a domestic cat. Therefore, Sarcocystis should be considered as a differential diagnosis in cats with suspected systemic protozoal infection.     

Sarcocystosis in a Stillborn Lamb

Confirmed congenital sarcocystosis has been reported extremely rarely in domestic ruminants. Sarcocystosis was diagnosed in a stillborn lamb with microscopic lesions predominantly in the central nervous system and placenta. Encephalitis was characterized by multiple foci of glial nodules some with central necrosis, perivascular cuffing and vascular occlusion, while placental lesions consisted of multifocal necroses, inflammation and mild calcification. Immature and mature schizonts were found in vascular endothelium of several organs. It is suggested that the protozoa were Sarcocystis tenella based on their morphology, location and as this is the most pathogenic Sarcocystis sp. parasitizing sheep.     

Prevalence of agglutinating antibodies to Sarcocystis neurona in raccoons, Procyon lotor, from the United States

Equine protozoal myeloencephalitis (EPM) is the most important protozoal disease of horses in North America and it is caused by Sarcocystis neurona. Natural cases of encephalitis due to S. neurona have been reported in raccoons, Procyon lotor. We examined 99 raccoons for agglutinating antibodies to S. neurona using the S. neurona agglutination test (SAT) employing formalin-fixed merozoites as antigen. Raccoons originated in Florida (N=24, collected in 1996), New Jersey (N=25, collected in 1993), Pennsylvania (N=25, collected in 1999), and Massachusetts (N=25, collected in 1993 and 1994). We found that 58 (58.6%) of the 99 raccoons were positive for antibodies to S. neurona using the SAT; 44 of 99 raccoons (44%) had titers of ≥1:500. This prevalence is similar to the reported seroprevalence of 33–60% for S. neurona antibodies in horses from the United States using the Western blot test.     

Clinical Sarcocystis neurona encephalomyelitis in a domestic cat following routine surgery

Sarcocystis neurona is an important cause of equine protozoal myeloencephalitis (EPM) in horses in the Americas. An EPM-like neurological disease also has been reported from other mammals but it is difficult to induce this disease in the laboratory. A 4-month-old male domestic cat developed neurological signs 3 days following castration. The cat was euthanized 12 days later because of paralysis. Encephalomyelitis was the only lesion and was associated with numerous Sarcocystis schizonts and merozoites in the brain and spinal cord. The protozoa reacted positively with S. neurona-specific polyclonal rabbit antibody. Two unidentified sarcocysts were present in the cerebellum. It may be possible that stress of surgery triggered relapse of S. neurona infection in this cat.     

Equine Protozoal Myeloencephalitis: An Updated Consensus Statement with a Focus on Parasite Biology,Diagnosis, Treatment,and Prevention

Equine protozoal myeloencephalitis (EPM) remains an important neurologic disease of horses. There are no pathognomonic clinical signs for the disease. Affected horses can have focal or multifocal central nervous system (CNS) disease. EPM can be difficult to diagnose antemortem. It is caused by either of 2 parasites, Sarcocystis neurona and Neospora hughesi, with much less known about N. hughesi. Although risk factors such as transport stress and breed and age correlations have been identified, biologic factors such as genetic predispositions of individual animals, and parasite‐specific factors such as strain differences in virulence, remain largely undetermined. This consensus statement update presents current published knowledge of the parasite biology, host immune response, disease pathogenesis, epidemiology, and risk factors. Importantly, the statement provides recommendations for EPM diagnosis, treatment, and prevention.     

Fatal hepatic sarcocystosis in a puppy with eosinophilia and eosinophilic peritoneal effusion

A 3-month-old male Golden Retriever puppy was evaluated for lethargy and fever of 2-days duration. Results of a CBC and biochemical profile revealed marked eosinophilia (6.3 X 10(3)/microL; reference interval 0.1-1.2 X 10(3)/microL), moderate thrombocytopenia, and increased activities of alanine aminotransferase, aspartate aminotransferase, and creatine kinase. Hepatomegaly and peritoneal effusion were found using abdominal ultrasound. Peritoneal fluid analysis revealed eosinophilic inflammation (23,000 nucleated cells/microL with 88% eosinophils). Despite supportive treatment the puppy's condition deteriorated rapidly; euthanasia was requested, and a necropsy performed. Microscopically, there was marked necrosuppurative and eosinophilic hepatitis with vasculitis. Numerous hepatocytes contained protozoal organisms suspected to be Toxoplasma gondii or Neospora caninum. However, serum was negative for both T gondii and N caninum antibodies; polymerase chain reaction assay on hepatic tissue was negative for both organisms; and immunohistochemical evaluation of hepatic tissue using serum raised against T gondii, N caninum, and Sarcocystis neurona also was negative. Schizont morphology suggested that merozoites replicated by endopolygeny, forming rosettes around a central residual body. Transmission electron microscopy revealed that merozoites lacked rhoptries. These findings were consistent with a diagnosis of Sarcocystis canis, an apicomplexan parasite with an unknown life cycle.     

Experimental inoculation of domestic cats (Felis domesticus) with Sarcocystis neurona or S. neurona-like merozoites

Sarcocystis neurona is the parasite most commonly associated with equine protozoal myeloencephalitis (EPM). Recently, cats (Felis domesticus) have been demonstrated to be an experimental intermediate host in the life cycle of S. neurona. This study was performed to determine if cats experimentally inoculated with culture-derived S. neurona merozoites develop tissue sarcocysts infectious to opossums (Didelphis virginiana), the definitive host of S. neurona. Four cats were inoculated with S. neurona or S. neurona-like merozoites and all developed antibodies reacting to S. neurona merozoite antigens, but tissue sarcocysts were detected in only two cats. Muscle tissues from the experimentally inoculated cats with and without detectable sarcocysts were fed to laboratory-reared opossums. Sporocysts were detected in gastrointestinal (GI) scrapings of one opossum fed experimentally infected feline tissues. The study results suggest that cats can develop tissue cysts following inoculation with culture-derived Sarcocystis sp. merozoites in which the particular isolate was originally derived from a naturally infected cat with tissue sarcocysts. This is in contrast to cats which did not develop tissue cysts when inoculated with S. neurona merozoites originally derived from a horse with EPM. These results indicate present biological differences between the culture-derived merozoites of two Sarcocystis isolates, Sn-UCD 1 and Sn-Mucat 2.     

Serologic responses of cats against experimental Sarcocystis neurona infections

Sarcocystis neurona is the most important cause of a neurologic disease of horses, equine protozoal myeloencephalitis (EPM). Cats and other carnivores can act as its intermediate hosts and horses are aberrant hosts. Little is known of the sero-epidemiology of S. neurona infections in cats. In the present study, antibodies to S. neurona were evaluated by the S. neurona agglutination test (SAT). Cats fed sporocysts from the feces of naturally infected opossums or inoculated intramuscularly with S. neurona merozoites developed high levels (> or =1:4000) of SAT antibodies. Antibodies to S. neurona were not found in a cat inoculated with merozoites of the closely related parasite, Sarcocystis falcatula. These results should be useful in studying sero-epidemiology of S. neurona infections in cats.     

P‐33 Poxvirus infection in two cats

A 3‐year‐old Burmese cat was presented with a history of nonresolving crusted and papular lesions of the face and prior treatment with prednisolone. Skin biopsies revealed typical pox lesions with hyperplasia and ulceration of the epidermis and eosinophilic cytoplasmic inclusion bodies in the epidermal cells. In the upper dermis there was prominent diffuse mast cell infiltration and mild neutrophilic and eosinophilic inflammation. Rare cytoplasmic inclusion bodies were also present in swollen endothelial cells of dermal venules, which showed no other degenerative changes. Histological diagnosis was confirmed by electron microscopic evidence of pox virus particles in inclusion bodies of epidermal cells. The lesions resolved within 6 weeks with systemic antibiotic therapy and supportive care. A 2‐year‐old domestic short‐haired cat was presented with multiple disseminated papular and ulcerative pox lesions with central eschar over the entire body. Histologically, large epidermal inclusion bodies (up to 6 μm in diameter) were present. Widespread haemorrhage and vascular wall necrosis was visible in the dermis and subcutis. Some subcutaneous vessels showed neutrophilic vasculitis. In addition to diffuse dermal neutrophilic and eosinophilic inflammation, a lymphohistiocytic panniculitis was also present. The cat died as a result of massive haemorrhage and lymphedema, despite supportive care. Funding: Self‐funded.     

Cerebral phaeohyphomycosis caused by Cladosporium spp. in two domestic shorthair cats

Two domestic shorthair cats presented for clinical signs related to multifocal central nervous system dysfunction. Both cats had signs of vestibular system involvement and anisocoria, and one had generalized seizure activity. Cerebrospinal fluid analysis revealed a neutrophilic pleocytosis with protein elevation in one cat and pyogranulomatous inflammation in the second. Electroencephalography and brain-stem auditory-evoked potentials in the first cat confirmed cerebral cortical and brain-stem involvement. Euthanasia was performed in both cats, and postmortem diagnoses of phaeohyphomycosis secondary to Cladosporium spp. were made based on histopathology and fungal culture in both cats.     

Detection of Zoonotic Protozoa Toxoplasma gondii and Sarcocystis suihominis in Wild Boars from Spain

Food safety regulations require the control of the presence of protozoa in meats destined for human consumption. Wild boar (Sus scrofa) meat may constitute a source of zoonoses. A 23.8% (688/2881) seroprevalence of anti‐Toxoplasma gondii antibodies and 72.2% (662/910) Sarcocystis sarcocysts prevalence were detected among wild boars hunted in Southwestern areas of Spain. Identity of Sarcocystis spp. was performed by RFLP‐PCR and sequencing, detecting S. miescheriana (7/8) and the zoonotic S. suihominis (1/8). Risk assessment studies of these coccidian in meats destined to human consumption are needed.     

Penetration of equine leukocytes by merozoites of Sarcocystis neurona

Horses are considered accidental hosts for Sarcocystis neurona and they often develop severe neurological disease when infected with this parasite. Schizont stages develop in the central nervous system (CNS) and cause the neurological lesions associated with equine protozoal myeloencephalitis. The present study was done to examine the ability of S. neurona merozoites to penetrate and develop in equine peripheral blood leukocytes. These infected host cells might serve as a possible transport mechanism into the CNS. S. neurona merozoites penetrated equine leukocytes within 5 min of co-culture. Infected leukocytes were usually monocytes. Infected leukocytes were present up to the final day of examination at 3 days. Up to three merozoites were present in an infected monocyte. No development to schizont stages was observed. All stages observed were in the host cell cytoplasm. We postulate that S. neurona merozoites may cross the blood brain barrier hidden inside leukocytes. Once inside the CNS these merozoites can egress and invade additional cells and cause encephalitis.     

What is your diagnosis? Rectal scraping from a dog with diarrhea

Abstract: A 3‐year‐old, spayed female, mixed‐breed dog was evaluated for chronic diarrhea, vomiting, and weight loss. A marked inflammatory leukogram, mild regenerative anemia, and marked hypoalbuminemia were noted. Cytologic evaluation of a rectal scraping revealed numerous round to ovoid protozoal cysts, 5–25 μm in diameter, with small to moderate amounts of pale blue cytoplasm and round eosinophilic nuclei. A distinct, variably sized, round to oval vacuole was often seen within the cytoplasm and frequently displaced the nucleus. The cysts were morphologically similar to Blastocystis sp., an amoeba‐like protozoal parasite found in both diseased and asymptomatic humans and animals. Histologic findings in endoscopic biopsies from the stomach, duodenum, ileum, and colon were unremarkable and protozoal organisms were not observed. The dog was diagnosed with exocrine pancreatic insufficiency based on markedly decreased serum levels of trypsin‐like immunoreactivity. Alteration of gastrointestinal flora secondary to the underlying pancreatic disease likely allowed overgrowth of the protozoa, which were considered an incidental finding. Their identification was important in avoiding an incorrect diagnosis and unwarranted treatment.     

Antibody coefficients for the diagnosis of equine protozoal myeloencephalitis

Background: Diagnosis of equine protozoal myeloencephalitis (EPM) remains a challenge for equine practitioners. Current utilized methods have inadequate sensitivity and specificity, because of a high number of false positive results. Hypothesis/Objective: Evaluation of antibody indices to Sarcocystis neurona should provide high sensitivity and specificity for diagnosis of EPM. Animals: Archived samples from 29 clinical patients. Methods: Archived serum and cerebrospinal fluid (CSF) samples from clinical patients with either EPM (14) or cervical vertebral compressive myelopathy (CVM) (15) were examined and tested for anti‐S. neurona antibodies by the SnSAG2 ELISA. The results were used to calculate the antibody index (AI) and C‐value. Sensitivity and specificity were calculated, and the AI, C‐value, immunoglobulin G (IgG) concentrations, and anti‐S. neurona titers compared. In addition, negative CSF was spiked in varying concentrations with blood from a horse with a high anti‐S. neurona titer, and the tests repeated. Results: Results demonstrated that the IgG concentration, anti‐S. neurona titer, AI, and C‐value were significantly higher (P < .05) in horses with EPM than in those with CVM. Sensitivity and specificity of the AI was 71 and 100%, respectively, and that of the C‐value was 86 and 100%, respectively. In addition, the AI and C‐value from the samples spiked with S. neurona positive blood remained below 1 (eg, negative) in CSF with a red blood cell (RBC) count up to 10⁵ RBC/μL. Conclusions/Clinical Importance: Results of the study demonstrate the value of calculating the AI and C‐value in the diagnosis of EPM in horses. In addition, the test is robust in the presence of blood contamination.     

Epidemiology of Sarcocystis neurona infections in domestic cats (Felis domesticus) and its association with equine protozoal myeloencephalitis (EPM) case farms and feral cats from a mobile spay and neuter clinic

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease in the horse most commonly caused by Sarcocystis neurona. The domestic cat (Felis domesticus) is an intermediate host for S. neurona. In the present study, nine farms, known to have prior clinically diagnosed cases of EPM and a resident cat population were identified and sampled accordingly. In addition to the farm cats sampled, samples were also collected from a mobile spay and neuter clinic. Overall, serum samples were collected in 2001 from 310 cats, with samples including barn, feral and inside/outside cats. Of these 310 samples, 35 were from nine horse farms. Horse serum samples were also collected and traps were set for opossums at each of the farms. The S. neurona direct agglutination test (SAT) was used for both the horse and cat serum samples (1:25 dilution). Fourteen of 35 (40%) cats sampled from horse farms had circulating S. neurona agglutinating antibodies. Twenty-seven of the 275 (10%) cats from the spay/neuter clinic also had detectable S. neurona antibodies. Overall, 115 of 123 (93%) horses tested positive for anti-S. neurona antibodies, with each farm having greater than a 75% exposure rate among sampled horses. Twenty-one opossums were trapped on seven of the nine farms. Eleven opossums had Sarcocystis sp. sporocysts, six of them were identified as S. neurona sporocysts based on bioassays in gamma-interferon gene knockout mice with each opossum representing a different farm. Demonstration of S. neurona agglutinating antibodies in domestic and feral cats corroborates previous research demonstrating feral cats to be naturally infected, and also suggests that cats can be frequently infected with S. neurona and serve as one of several natural intermediate hosts for S. neurona.     

High prevalence of Sarcocystis calchasi sporocysts in European Accipiter hawks

The emerging Sarcocystis calchasi induces a severe and lethal central nervous disease in its intermediate host, the domestic pigeon (Columba livia f. domestica). Experimental studies have identified the Northern goshawk (Accipiter g. gentilis) as final host. Phylogenetically closely related European sparrowhawks (Accipiter n. nisus) and wood pigeons (Columba palumbus) have been found to harbor genetically closely related Sarcocystis spp. However, data on the prevalence and potential interspecies occurrence of these parasites are lacking. Here, we report that European Accipiter hawks (Accipitrinae) are highly infected with S. calchasi, S. columbae and Sarcocystis sp. ex A. nisus in their small intestine. Thirty-one of 50 (62%) Northern goshawks necropsied during 1997-2008 were positive for S. calchasi in a newly established species-specific semi-nested PCR assay based on the first internal transcribed spacer region. Unexpectedly, 14 of 20 (71.4%) European sparrowhawks tested also positive. In addition, birds of both species were found to be infested with S. columbae and an, as yet, unnamed Sarcocystis sp. recently isolated from European sparrowhawks. These findings raise new questions about the host specificity of S. calchasi and its high virulence in domestic pigeons, since sparrowhawks only rarely prey on pigeons. Notably, isolated sporocysts from both infected Accipiter spp. measured 8 μm × 11.9 μm, precluding a preliminary identification of S. calchasi in feces of Accipiter hawks based on morphology alone. Importantly, three of four Northern goshawks used in falconry tested positive for S. calchasi. In conclusion, the results indicate that both European Accipter spp. in Germany serve as natural final hosts of S. calchasi and suggest that falconry and pigeon sport may serve as risk factors for the spread of this pathogen in domestic pigeons.     

Molecular characterization of Sarcocystis spp. as a cause of protozoal encephalitis in a free-ranging black bear

A free-ranging juvenile male black bear (Ursus americanus), found dead in Alberta, Canada, had severe nonsuppurative encephalitis. Lesions in the brain were most severe in the gray matter of the cerebral cortex, and included perivascular cuffs of lymphocytes and plasma cells, areas of gliosis that disrupted the neuropil, and intralesional protozoan schizonts. The left hindlimb had suppurative myositis associated with Streptococcus halichoeri. Immunohistochemistry and molecular analyses (PCR and sequencing of 4 discriminatory loci: 18S rDNA, ITS-1 rDNA, cox1, rpoB) identified Sarcocystis canis or a very closely related Sarcocystis sp. in the affected muscle and brain tissues. The main lesion described in previously reported cases of fatal sarcocystosis in bears was necrotizing hepatitis. Fatal encephalitis associated with this parasite represents a novel presentation of sarcocystosis in bears. Sarcocystosis should be considered a differential diagnosis for nonsuppurative encephalitis in bears.     

Seroprevalence of Sarcocystis neurona and Its Association With Neurologic Disorders in Argentinean Horses

Equine protozoal myeloencephalitis (EPM) is generally caused by Sarcocystis neurona and can produce substantial economic losses on equine production in America. The aims of the present study were to evaluate the seroprevalence of S. neurona in the main horse-production area of Argentina and associate it with the occurrence of neurologic disorders. Serum samples were collected from 640 horses in nine Argentinean provinces. Most of the samples correspond to animals ≥1.5-year-old from different breeds (n = 628); 12 samples were from younger horses. Further seroprevalence comparison was conducted from the older animals grouped with (n = 148) or without neurologic signs (n = 480). Immunoblot: proteins from 2 × 10⁷S. neurona merozoites were used as antigen on each membrane. Reactivity to antigens with relative mobility of 7, 10, and 16 kDa was considered specific for antibodies against S. neurona; reactivity at 30 kDa was recorded separately. The overall seroprevalence for S. neurona was 26.1% (167/640), and all the provinces had positive horses. Seroprevalence of animals with neurologic signs was greater (P < .001) than what was observed in normal horses (39.2% vs. 22.1%), with an odds ratio of 2.27. Reactivity at 30 kDa was detected in 71% of all samples. This study identified a wide distribution of S. neurona–positive animals in Argentina and horses with neurologic signs having a greater seroprevalence than normal horses. Sarcocystis neurona infection should be considered for early differential diagnosis and treatment of animals with neurologic disorders to decrease the economic impact of EPM in Argentina.     

Sarcocystis encephalitis in a cockatiel

A sporozoan organism was considered to be the causative agent of central nervous system disease in a cockatiel. The ultrastructural characteristics were typical of the coccidian group Apicomplexa, and the fact that organisms were free within the cytoplasm of infected cells and not within a vacuole, indicated they were Sarcocystis. Light and electron microscopic evaluation of brain tissue demonstrated protozoal organisms associated with areas of necrosis. Differential diagnosis of central nervous system disease in pet birds should include protozoal encephalitis.     

Coccidia (Protozoa : Sporozoasida) of cats and dogs. III. The occurrence of a species of Besnoitia in cats

Macroscopically visible spherical cysts measuring up to 260/μm in diameter and showing characteristics typical of those of the genus Besnoitia were observed in rats dosed with isosporan oocysts recovered from the faeces of a feral cat. Kittens fed these rats shed similar oocysts measuring 16.9 ± 0.1 × 14.6 ± 0.1 μm in their faeces 11–12days later. The feeding of sporulated oocysts of this coccidian to mice, rats, rabbits and guinea pigs resulted in the formation of Besnoitia cystsin all hosts except the last.

Besnoitia is the fourth sporozoan genus, together with Isospora, Toxoplasma, and Sarcocystis to be recovered from New Zealand cats.