Clinicopathologic diagnosis of cutaneous toxoplasmosis in an Angora cat

This report deals with a case of cutaneous toxoplasmosis in a 2 year-old female Angora cat. Cutaneous lesions were characterized by prescapular ulcers and hyperemic nodules in the skin of the inguinal and dorsosacral regions. A skin biopsy sample was collected from the lesioned area and processed for histopathologic examination and immunoperoxidase test using Toxoplasma gondii and Neospora caninum specific antibodies. Toxoplasma gondii immunopositive reactions were detected in keratinocytes and dermal macrophages while no immunoreactivity was detected for N. caninum. The case of cutaneous toxoplasmosis was further confirmed by PCR analysis using T. gondii B1 gene-specific primers. In conclusion, we report the first case of cutaneous toxoplasmosis in Angora cats.     

Cutaneous toxoplasmosis in a female Japanese cat

A 16-year-old female Japanese cat was presented with a single mammary-gland nodule approximately 3 cm in diameter. Histologically, the nodule consisted of necrotizing granulomatous panniculitis, vasculitis, and mastitis, and contained free and clustered protozoal organisms. The organism was present in the cytoplasm of macrophages, fibroblasts, endothelial cells, and mammary-gland epithelia. The organism was positive for anti- Toxoplasma gondii and anti- Neospora caninum antibodies. Electron microscopy showed single and grouped tachyzoites, with morphologic features similar to those of T. gondii. Polymerase chain reaction and deoxyribonucleic acid sequence analysis was consistent with T. gondii infection. This is the first report of cutaneous toxoplasmosis in a Japanese cat.     

A Neospora-like protozoon found in an aborted bovine placenta

Portions of the placenta and kidneys from a 5-month-old bovine fetus were examined histologically. The placental cotyledonary villi were necrotic and protozoa were found in lesions. The organisms were in trophoblasts, periodic acid Schiff-negative and did not react in an immunoperoxidase test using anti-Toxoplasma gondii serum. Individual zoites were approximately 5 x 2 microns and the biggest collection was 45 x 35 microns. The organism in the present case was structurally distinct from Sarcocystis spp. and Toxoplasma gondii, but resembled Neospora caninum.     

Toxoplasma gondii-like organisms in skin aspirates from a cat with disseminated protozoal infection

An 8-year-old, male domestic shorthair cat was referred to the Matthew J. Ryan Veterinary Hospital of the University of Pennsylvania with a 3-day history of lethargy, inappetance, hyperemic skin nodules, coughing, and vomiting. Laboratory results included nonregenerative anemia, lymphopenia, thrombocytopenia, hypoalbuminemia, hyponatremia, and increased alanine aminotransferase and aspartate aminotransferase activities. Cytology of the skin nodules revealed many spindle- to crescent-shaped protozoal organisms, with morphology consistent with Toxoplasma gondii or Neospora caninum. Gross necropsy, histopathologic, immunohistochemical, and transmission electron microscopic findings confirmed a systemic protozoal infection; however, the organism exhibited characteristics of both N caninum and T gondii. Diagnosis of a T gondii-like infection was based on internal structures of the organism and positive reaction to rabbit polyclonal antibodies to T gondii. Reports of toxoplasmic dermatitis are rare in the cat and dog, and this is the first reported diagnosis of T gondii-like protozoa in skin aspirates.     

Studies on serological cross-reaction of Neospora caninum with Toxoplasma gondii and Hammondia heydorni

The enzyme-linked immunosorbent assay (ELISA) was used to examine cross-reactivity of Neospora caninum with Toxoplasma gondii and Hammondia heydorni. Anti-T. gondii mouse and cat sera cross-reacted with N. caninum soluble antigen (NLA), but not with the recombinant surface antigen (NcSRS2). Anti-H. heydorni dog sera showed no cross-reactivity with either the NLA antigen or the NcSRS2. Lack of cross-reactivity between anti-H. heydorni sera and N. caninum antigens, and the cross-reactivity of anti-T. gondii sera with the NLA suggest that N. caninum has common antigens to T. gondii except for NcSRS2 based on serology. In light of several studies suggesting a closer relationship between N. caninum and H. heydorni than with T gondii, examination of serological cross-reactivity with N. caninum may be necessary to further classify the parasites in addition to molecular and morphological studies and clarification of the life cycle.     

Granulomatous meningoencephalitis due to toxoplasma gondii in a black-headed night monkey (Aotus nigriceps)

This report describes toxoplasmosis infection in a black-headed night monkey (Aotus nigriceps) from the Zoological Park of the Federal University of Mato Grosso, Brazil. The animal was submitted alive to the Veterinary Hospital of the Federal University of Mato Grosso presenting neurologic clinical signs. There were no improvements on the animal's clinical condition after treatment, so it was euthanized. At necropsy, the most significant gross lesions observed were hemorrhages diffusely scattered throughout the cerebral hemispheres. Histologic findings were characterized by nonsuppurative encephalitis and mild multifocal necrotizing nonsuppurative myocarditis. Groups of tachyzoites and cysts were observed surrounding the inflamed areas of cerebral parenchyma. Sections of the brain were submitted for immunohistochemistry examination for Toxoplasma gondii and Neospora caninum and also for polymerase chain reaction (PCR) to determine the presence of Toxoplasma gondii. The samples were positive for Toxoplasma gondii by PCR; however, cross reactivity was observed between Toxoplasma gondii and Neospora caninum antibodies. These findings were consistent with a toxoplasmosis infection in this animal.     

Use of a multiplex polymerase chain reaction to rapidly differentiate Neospora caninum from Toxoplasma gondii in an adult dog with necrotizing myocarditis and myocardial infarct.

This report describes a 3-year-old male castrated Mastiff dog that died unexpectedly with locally extensive, acute, necrotizing myocarditis and myocardial infarction. Intralesional protozoal tachyzoites in the affected myocardium were confirmed to be Neospora caninum by a novel multiplex polymerase chain reaction (PCR) and immunohistochemistry. Protozoal organisms were not identified in other tissues by histology, immunohistochemistry, or PCR. The multiplex PCR assay was used to quickly provide preliminary results on fresh myocardium to differentiate N. caninum and Toxoplasma gondii. Neosporosis is an uncommon cause of myocarditis in adult dogs and differential diagnoses for myocarditis in this population of dogs are reviewed.     

Neospora caninum in sheep: a herd case report

Neospora caninum was detected by means of PCR in the brain of 4 out of 20 aborted fetuses in a flock of 117 sheep exhibiting a persistent abortion problem, and N. caninum tissue cysts were furthermore found in encephalitic lesions in one of the PCR-positive fetuses. Toxoplasma gondii was detected as aborting agent in another 3 out of 20 fetuses. Antibodies to N. caninum (by indirect fluorescence antibody test (IFAT)) were found in 10.3% of 117 ewes and antibodies for T. gondii were found in 97.4% of 117 ewes. Other organisms associated with abortion were Chlamydia psittaci in three fetuses and Pasteurella multocida in one fetus. This is the first report of N. caninum associated abortion in naturally infected sheep.     

Seroprevalence of Neospora caninum in dairy cattle in Bahia, Brazil.

Sera collected from 447 dairy cattle on 14 dairy farms were tested for Neospora caninum antibodies by use of an immunofluorescent antibody technique. Positive reactions with titres > or =1:200 were found in 63 (14.09%) of animals. Neospora positive sera were also tested for Toxoplasma gondii antibodies by using a commercial latex agglutination test. Antibodies to T. gondii were detected in 3 (4.76%) of 63 N. caninum positive sera. These results indicate that N. caninum infection is widespread among dairy cattle in Bahia state.     

Dog shedding oocysts of Neospora caninum: PCR diagnosis and molecular phylogenetic approach

Results of molecular determination of a dog isolate of Neospora caninum in the Czech Republic are presented. Colorless bisporocystic oocysts measuring 10-13 micro m x 10-11 micro m were recovered from feces and used for DNA isolation. A diagnostic PCR procedure using previously described molecular methods was performed to determine the species. The N. caninum species-specific primers based on the Nc 5 region produced a positive result, while primers specific for Hammondia heydorni rDNA internal transcribed spacer 1 (ITS1) was negative. Sequencing and phylogenetic comparison of ITS1 rDNA and the D2 domain of the large subunit rDNA (D2 LSU) determined our isolate to be N. caninum. Phylogenetic analysis of closely related genera Toxoplasma, Neospora and Hammondia based on ITS1 and D2 LSU robustly distinguished three clades: (i). Toxoplasma gondii + Hammondia hammondi, (ii). N. caninum + Neospora hughesi, and (iii). H. heydorni. Based on phylogenetic relationships we propose three acceptable suggestions to solve the problem of taxonomy of these genera.     

Seroprevalence of Toxoplasma gondii and Neospora caninum in dairy goats from Romania

Little information is available about the seroprevalence of Toxoplasma gondii and Neospora caninum infections in goats in Romania and even in Europe. During 2007-2010, 735 serum samples were collected from dairy goats located in 4 historical regions (Cri?ana, Maramure?, Transylvania and Muntenia) of Romania. Sera were analyzed for T. gondii and N. caninum antibodies (IgG type) by enzyme-linked immunosorbent assay (ELISA) using two commercial kits (Chekit Toxotest Antibody ELISA and Chekit Neospora caninum Antibody ELISA; Idexx-Bommeli, Switzerland). Three hundred and eighty-eight out of 735 (52.8%) goats presented T. gondii antibodies and 12 out of 512 (2.3%) goats had N. caninum antibodies. The high seroprevalence of T. gondii suggests that infection with this parasite is common in dairy goats in Romania, and less common the infection with N. caninum. This is the first time that infection with N. caninum in goats has been reported in Romania and the first extended study on seroepidemiology of T. gondii.     

Aetiology of bovine abortion in Switzerland from 1986 to 1995--a retrospective study with emphasis on detection of Neospora caninum and Toxoplasma gondii by PCR

In a retrospective study, covering the period from 1986 to 1995, tissues of aborted fetuses were re-examined. A total of 347 cases were tested immunohistochemically, among them samples of 223 brains were examined for Neospora caninum, Toxoplasma gondii and Bovine Virus Diarrhoea Virus (BVDV), and 249 placentae for Chlamydiaceae. Two real-time PCR assays, one for N. caninum, and one for T. gondii, were developed. These potential abortion-inducing agents were detected - and confirmed by PCR, except for BVDV - in 16.1% (N. caninum), 0% (T. gondii), 9.9% (BVDV) and 0.8% (Chlamydiales) of the cases examined. Immunohistochemistry proved to be inadequate for the detection of the protozoal epitopes, whereas it was confirmed as a very useful tool for the detection of BVDV. In abortion material, PCR is considered to be more suitable for the detection of protozoa and Chlamydophila abortus, an adequate sampling presupposed.     

Survey of nine abortifacient infectious agents in aborted bovine fetuses from dairy farms in Beijing, China, by PCR

Abortion in dairy cattle causes considerable economic losses to the dairy industry. Aborted fetuses and samples from the corresponding aborting dams from 12 dairy herds in Beijing were tested for 9 abortifacient infectious pathogens by PCR between 2008 and 2010. From a total of 80 abortion cases collected during this period, infectious agents were detected in 45 (56.3%) cases, 22 (48.9%) of which represented co-infections with two or three infectious agents. The detected pathogens included infectious bovine rhinotracheitis virus (36.3%) and Neospora caninum (31.3%), followed by bovine viral diarrhoea virus (7.5%), Brucella abortus (6.3%), Tritrichomonas foetus (5%) and Toxoplasma gondii (1.3%). Campylobacter fetus, Coxiella burnetii and Chlamydophila psittaci were not detected in any abortion case. Findings from this study indicated that infectious bovine rhinotracheitis virus and Neospora caninum were the main potential causes of abortions in Beijing dairy herds, whereas the bacterial pathogens were not, in contrast to reports from other countries. This is the first study to test nine abortifacient infectious agents by PCR at the same time, and it is also the first time to report the involvement of a variety of infectious agents in bovine abortion cases in China.     

Use of a multiplex polymerase chain reaction assay in the antemortem diagnosis of toxoplasmosis and neosporosis in the central nervous system of cats and dogs

OBJECTIVE: To develop a multiplex polymerase chain reaction (PCR) assay for the detection of Toxoplasma gondii and Neospora caninum DNA in canine and feline biological samples. SAMPLE POPULATION; Biological samples from 7 cats with systemic (n = 4) or CNS (3) toxoplasmosis, 6 dogs with neospora- or toxoplasma-associated encephalitis, and 11 animals with nonprotozoal disease. PROCEDURE: Primers for T gondii, N caninum, and the canine ferritin gene (dogs) or feline histone 3.3 gene (cats) were combined in a single PCR assay. The DNA was extracted from paraffin-embedded brain tissue, CSF, or skeletal muscle. The PCR products with positive results were cloned, and sequence identity was confirmed. RESULTS: Of 7 cats and 4 dogs with immunohistochemical or serologic evidence of toxoplasmosis, PCR results were positive for all cats and 3 dogs for T gondii, and positive for T gondii and N caninum for 1 dog. Another dog had negative PCR results for both parasites. Of 2 dogs with immunohistochemical or serologic evidence of neosporosis, PCR results were positive for 1 for N caninum and positive for the other for T gondii. All negative-control samples yielded negative results for T gondii and N caninum on the PCR assay. CONCLUSIONS AND CLINICAL RELEVANCE: Standard tests for toxoplasmosis or neosporosis associated with the CNS rely on serologic, histologic, or immunohistochemical analysis and can be difficult to interpret. The multiplex PCR assay with built-in control reactions could be a complementary clinical tool for the antemortem diagnosis of toxoplasmosis or neosporosis associated with the CNS.     

Occurrence of Neospora caninum and Toxoplasma gondii infections in ovine and caprine abortions

Neospora caninum and Toxoplasma gondii are closely related cyst-forming apicomplexan parasites identified as important causes of reproductive failure in cattle and small ruminants, respectively. Protozoan abortion in small ruminants is traditionally associated with T. gondii, but the importance of N. caninum remains uncertain. The aim of this study was to investigate the presence of N. caninum and T. gondii infections in abortion cases in small ruminants submitted for diagnosis. For this purpose, 74 ovine and 26 caprine aborted foetuses were recovered from different areas in Spain. Foetal histopathology was used to detect the presence of protozoal-associated lesions in brain. The presence of N. caninum and T. gondii was confirmed by PCR. Protozoal infection was detected in 17 out of 100 (17%) foetuses examined by at least one of the diagnostic techniques used. Lesions suggestive of protozoal infection were observed in 10.8% (8/74) and 15.4% (4/26) of the ovine and caprine abortions respectively. N. caninum and T. gondii infection was detected by PCR in 6.8% (5/74) and 5.4% (4/74) of sheep foetuses, respectively, of which five showed protozoal-associated lesions. N. caninum DNA was detected in 11.5% (3/26) of goat foetuses, of which two showed protozoal-associated lesions, whereas T. gondii DNA was detected in one goat foetus with no lesions. The simultaneous presence of N. caninum and T. gondii DNA was detected in one sheep foetus with severe lesions. This study demonstrates that N. caninum plays a significant role in abortion in small ruminants in the studied population. In addition, our results highlight the importance of differentiating between protozoa whenever characteristic lesions are observed.     

Seroepidemiology of Neospora caninum and Toxoplasma gondii in cattle and water buffaloes (Bubalus bubalis) in the People's Republic of China

A seroepidemiological survey of Neospora caninum and Toxoplasma gondii in cattle and water buffaloes was carried out in the People's Republic of China. Serum samples were obtained from dairy (n=262, 9 herds in 9 provinces) and beef cattle (n=10, 1 herd) and water buffaloes (n=40) in China. All sera were tested for antibodies to N. caninum and T. gondii by an enzyme-linked immunosorbent assay (ELISA) and an indirect agglutination test (IAT), respectively. The overall seroprevalence of N. caninum in dairy cattle was 17.2% (45/262), and the herds seroprevalence of N. caninum was 88.9% (8/9), and antibodies to T. gondii were present in 6 cows (2.3%). None of the cows had antibodies against both T. gondii and N. caninum. Antibodies to T. gondii or N. caninum were not found in beef cattle or water buffaloes. The seroprevalence of N. caninum in aborting cows (20.2%) was higher than that in non-aborting cows (16.6%) with an odds ratio of 1.26 (95% CI, 0.54-2.95), but the difference was not statistically significant (P>0.05). There was no apparent association of N. caninum seropositivity with age or number of pregnancies. This is the first report on the seroprevalence of N. caninum in cattle and water buffaloes in China.     

Encephalitozoon cuniculi–Associated Equine Encephalitis: A Case Report

A case of encephalitis of unknown origin in the horse was investigated. Postmortem examination findings revealed a nonsuppurative granulomatous meningoencephalitis in the right hemisphere of the cerebral cortex. Testing for West Nile virus, equine herpes virus, equine infectious anemia, Toxoplasma gondii, Neospora caninum, and Sarcocystis neurona were negative. The horse had a titer for Encephalitozoon cuniculi, and sections from the affected area of the brain tested positive for the organism using both polymerase chain reaction (PCR) and immunohistochemistry. Amplicons generated using PCR were sequenced, and E. cuniculi genotype II was identified. This is the first case of E. cuniculi genotype II associated with encephalitis in the horse.     

Study on the prevalence of Toxoplasma gondii and Neospora caninum and molecular evidence of Encephalitozoon cuniculi and Encephalitozoon (Septata) intestinalis infections in red foxes (Vulpes vulpes) in rural Ireland

Thoracic fluid (pleural fluid and clotted blood) from 206 foxes were examined for antibodies to Toxoplasma gondii and 220 thoracic fluid samples were tested for Neospora caninum antibodies using indirect immunofluorescent antibody tests (IFAT). A total of 115 (56%) and six (3%) foxes had antibodies to T. gondii and N. caninum, respectively. The brains from 148 foxes were examined for histological lesions and pathological changes suggestive of parasitic encephalitis were observed in 33 (22%). Two thirds of these foxes had antibodies to T. gondii and one fox had antibodies to both T. gondii and N. caninum. PCR assays carried out on DNA extracted from the 33 brains with histological lesions were negative for N. caninum but one of the brains was positive for T. gondii. Microsporidian DNA was also amplified from the brains of two of these foxes. Sequencing these amplicons revealed 100% homology with Encephalitozoon (Septata) intestinalis in one fox and Encephalitozoon cuniculi in the second fox. This is the first report of Encephalitozoon infections in wildlife in Ireland.     

Cutaneous manifestations of disseminated toxoplasmosis in an immunosuppressed dog

A 3.5-year-old, castrated male, giant schnauzer was presented with alopecic pustular dermatitis. Immune-mediated hemolytic anemia had been diagnosed 45 days previously. At the time of presentation, the dog was receiving prednisone, azathioprine, and cyclosporine. Cutaneous protozoosis was diagnosed, and postmortem examination revealed protozoa within cutaneous, cardiac, pancreatic, and pulmonary tissues. The protozoa divided by endodyogeny, had the morphology of Toxoplasma gondii (T. gondii) tachyzoites, and stained positively with T. gondii polyclonal antibodies but not with antibodies to Neospora caninum or Sarcocystis neurona. Immunosuppression may have predisposed this dog to disseminated toxoplasmosis.     

Toxoplasma gondii and Neospora caninum in wildlife: common parasites in Belgian foxes and Cervidae?

Sera from Cervidae were tested for the presence of antibodies against Neospora caninum using ELISA; and against Toxoplasma gondii using SAG1-ELISA and a commercially available agglutination test. The T. gondii seroprevalence was 52% (38/73) in roe deer (Capreolus capreolus), 0% in bred fallow deer (0/4) (Dama dama) and red deer (0/7) (Cervus elaphus). We found 2.7% of the roe deer samples and none of the bred deer samples positive for N. caninum. Brain samples from wild roe deer, red deer and red foxes (Vulpes vulpes) were tested for the presence of T. gondii and N. caninum DNA using multiplex real-time PCR. We detected T. gondii in 18.8% (57/304) of the red foxes and in 1 of the 33 deer samples. N. caninum was found in 6.6% of the red foxes and in 2 roe deer samples. Twenty-six of the T. gondii positive DNA extracts from the red fox samples were genotyped. Twenty-five were type II and only one was found to be type III.