Dogs are definitive hosts of Neospora caninum

Dogs were investigated to determine if they are definitive hosts of Neospora caninum. Four dogs were fed N. caninum tissue cysts in infected mouse tissue, and two negative control dogs were fed uninfected mouse tissue. Dog faeces were examined daily for 30 days using a sucrose flotation technique. Three challenged dogs shed spherical to subspherical unsporulated oocysts, measuring 10 to 11 microns in diameter. Oocysts sporulated within 3 days and contained two sporocysts, each with four sporozoites. Outbred, inbred, and gamma-interferon knockout mice were inoculated with canine faecal extracts and monitored for evidence of neosporosis using a variety of morphologic, immunohistologic, serologic, and genetic analyses. Mice that received faeces from each dog observed to shed oocysts were demonstrated to have neosporosis by two or more techniques. One mouse was demonstrated to be infected with N. caninum by immunohistochemistry, ultrastructural analysis, and a species-specific PCR test. No evidence of neosporosis was observed in control animals. Based on this study, dogs are a definitive host of Neospora caninum.     

Locomotion of neutrophil fragments occurs by graded radial extension

Neospora caninum is a protozoan parasite which causes neurological problems in dogs and abortion in cattle. As N. caninum is difficult to distinguish morphologically from Toxoplasma gondii, we developed a molecular tool capable of discriminating between the two parasites. Genomic DNA was isolated from in vitro cultured N. caninum tachyzoites and cloned into a plasmid vector. Resulting colonies were subsequently screened by differential hybridization using N. caninum and T. gondii DNA. Two clones were characterized in detail: one clone, termed pNc5, was found to be specific for N. caninum whereas the second clone, pNc1, hybridized with DNA from both parasites. The sequence of pNc5 was determined and different oligonucleotide primers were designed for use in the polymerase chain reaction (PCR). A 944 bp fragment was specifically amplified from N. caninum DNA, but not from DNA extracted from T. gondii or different Sarcocystis species. Positive signals in PCR were obtained with as little as 100 pg parasite template DNA. In addition, dual PCR with primer pairs specific for N. caninum and T. gondii allowed the detection of either parasite in mixed samples.     

Quantification of phosphorus metabolites from chemical shift imaging spectra with corrections for point spread effects and B1 inhomogeneity

Neospora caninum is an apicomplexan parasite which is morphologically and ultrastructurally very similar to Toxoplasma gondii. In order to identify molecules involved in host cell entry and subsequent modification of the parasitophorous vacuole, a polyclonal antiserum directed against N. caninum tachyzoites was raised in a rabbit. Subcellular fractionation of tachyzoites was performed using the non-ionic detergent Triton-X-114. Membrane fractions were analysed by immunoblotting using the polyclonal antiserum. One of the immunoreactive protein bands had a mol. wt of 33,000 and was subsequently named Nc-p33. Affinity-purified anti-Nc-p33 antibodies were used to characterise this polypeptide using SDS-PAGE, isoelectric focusing, Western blot analysis and immuno-EM. Nc-p33 was found in two isolates of N. caninum (NC-1 and Liverpool), but could not be detected in T. gondii tachyzoites. Immunogold EM revealed that Nc-p33 constituted a dense granule-associated protein, and Western blotting demonstrated that Nc-p33 was most likely identical to the recently described antigen NCDG1. Shortly after invasion, this dense granule protein was targeted to the parasitophorous vacuole membrane, and, at later timepoints after infection, was also found on the parasitophorous vacuolar network. This suggested that Nc-p33 could play a functional role in the modification of the parasitophorous vacuole and its membrane.     

Prevalence of antibodies to Neospora caninum in different canid populations

A total of 1,554 dogs from 5 countries on 3 continents were tested for antibodies to Neospora caninum using an indirect fluorescent antibody test. In Australia, overall, 42/451 (9%, 95% confidence interval [CI] 6-12%) dogs were seropositive (Melbourne 11/207 [5%, 95% CI 2-9%]; Sydney 18/150 [12%, 95% CI 7-18%]; Perth 13/94 [14%, 95% CI 8-22%]). Antibodies to N. caninum were also detected in dogs in South America (Uruguay [20%, 95% CI 16-24%, n = 414]) and sub-Saharan Africa (Tanzania [22%, 95% CI 12-36%, n = 49]). In contrast, only 1 of 500 dogs tested from the Falkland Islands and none of 140 dogs from Kenya was seropositive. Of wild canids, 1/54 (2%, 95% CI 0-10%) British foxes and 15/169 (9%, 95% CI 5-14%) Australian dingoes had antibodies to N. caninum.     

Mandibular metastasis of endometrial carcinoma diagnosed via a dental radiograph

OBJECTIVE: To identify risk factors associated with Neospora caninum infection in dairy herds in Québec and to estimate seroprevalence in case and control herds. DESIGN: Herd-based case-control and seroprevalence study. ANIMALS: 3,059 cows from 24 case and 22 control dairy herds in Québec. PROCEDURE: Blood samples were obtained from each cow, and sera were tested, using an ELISA, for antibodies to N caninum. Owners of herds answered questionnaires requesting information on demographics and herd management practices. Seroprevalence was compared between case and control herds, using the Mann-Whitney test. Risk factors were compared between case and control herds, using logistic regression. RESULTS: All case herds and 73% of control herds had at least one seropositive cow. Median seroprevalence was significantly greater among case herds (22.5%) than among control herds (7.5%). Dogs were found most often and in greatest numbers on farms housing case herds compared with control herds during the past 3 years. CLINICAL IMPLICATIONS: Although the exact role that dogs have in transmission of N caninum in dairy herds needs to be elucidated, dogs should have limited access to barns and cattle.     

Male involvement in family planning: a case study spanning five generations of a south Indian family

Neospora caninum is a coccidial protozoan parasite that infects a large range of mammals including dogs, cats, mice, and cattle. Morphologically, N. caninum appears indistinguishable from Toxoplasma gondii, although they are genetically distinct. To date there have been no reported cases of this infection in humans, although nonhuman primates may be susceptible to infection. Inbred A/J mice develop no clinical and little histologic evidence of infection in spite of a high-dose inoculum of N. caninum. Splenocytes obtained from infected mice proliferate in vitro in response to both N. caninum and T. gondii-soluble antigen. A transient state of T cell hyporesponsiveness to parasite antigen and mitogen was observed at Day 7 p.i. This downregulatory response could be partially reversed by the addition of the nitric oxide antagonist LNMMA, but not antibody to IL-10. Mice infected with N. caninum produce significant quantities of IL-12 and IFN gamma, most evident shortly after infection. In vivo, antibody to IL-12 is able to neutralize immune resistance to the parasite. Moreover, in vivo depletion of IFN gamma with antibody renders the mice susceptible to infection. These observations suggest that N. caninum induces a T cell immune response in the infected host that is at least partially mediated by IL-12 and IFN gamma.     

A survey of epithelial odontogenic tumors and cysts in dogs and cats

A retrospective histologic study of 12 canine and eight feline epithelial odontogenic tumors and cysts was conducted from oral masses (n = 3,917) obtained between 1980 and 1990. No sex or breed predilection was identified. Ameloblastoma was observed in two dogs (case Nos. 1, 2) 6 and 8 months of age. Calcifying epithelial odontogenic tumors were seen in a dog (case No. 3) and in two cats (case Nos. 4, 5) between 8 and 16 years of age. Ameloblastic fibroma (or fibroameloblastoma) was observed in cats (case Nos. 6-10) only. Inductive fibroameloblastoma was observed in four cats (case Nos. 6-9) up to 1 year of age, whereas ameloblastic fibroma was seen in a 14-year-old cat (case No. 10). A single ameloblastic odontoma was identified in a 20-month-old dog (case No. 11). Two complex odontomas occurred in a 6-month-old (case No. 12) and a 4-year-old (case No. 13) dog. Odontogenic cysts were identified in five dogs (case Nos. 14-18) aged 4.5 months to 16 years and in a 1-year-old cat (case No. 19) and have not been previously reported in these species. These cysts were lined by a stratified epithelium reminiscent of the appearance of ameloblastic epithelium. An odontogenic keratocyst with prominent central parakeratotic keratinization was identified in one 9-year-old female dog (case No. 20). Almost all epithelial odontogenic tumors were circumscribed, benign tumors that warranted a good prognosis for survival, although local recurrence may have followed (or may follow) incomplete excision. Calcifying epithelial odontogenic tumors may be locally invasive. Of six odontogenic cysts (case Nos. 14-19), two (case Nos. 15, 18) gave rise to basi-squamous carcinomas. The classification and behavior of epithelial odontogenic tumors and cysts in human beings, dogs, and cats are discussed.     

Distribution of Neospora caninum within the central nervous system and other tissues of six dogs with clinical neosporosis

Six cases of neosporosis in dogs were diagnosed on the basis of clinical signs, serology and immunohistochemistry. The brains and spinal cords, at several levels, and various other tissues were examined using anti-Neospora caninum serum by an immunoperoxidase technique. Parasite density varied markedly from case to case. Although found most consistently in the cerebrum, parasites were distributed throughout the central nervous system (CNS), both within the grey and white matter and within nerve roots. Clinical signs were not related to the position of parasites. Tissue cysts were found infrequently in all areas of the CNS, but not in other tissues. Parasite density was not related to the age of the dog or whether treatment had been given. However, with the exception of the only adult dog examined, more CNS parasites were found in dogs with a longer duration of illness and with higher antibody titres. Tachyzoites were present in skeletal muscles, in the muscularis of the oesophagus, in heart, lung and, less frequently, liver, and rarely in the adrenal gland, thyroid gland and uterus; no clinical signs were seen resulting from damage to these organs. Parasites were not observed in lymphoid tissue. In visceral organs, parasites were most widely distributed in peracute cases. For post mortem diagnosis of neosporosis, the CNS, particularly the cerebrum, is the optimum tissue to examine but parasites may also be found in many other tissues, especially in acute cases. Muscle biopsy of appropriate muscles (as suggested by the clinical signs) provides the possibility of a definitive premortem diagnosis.     

Prevalence of Neospora caninum and Toxoplasma gondii antibodies in sera from camels from Egypt

Sera from camels from Egypt were examined by the direct agglutination tests incorporating mercaptoethanol for antibodies to Neospora caninum and Toxoplasma gondii. Antibodies to N. caninum were found in 6 of 161 camels in titers of 1:40 (2 camels) and 1:80, 1:160, 1:640, and 1:1280 in 1 camel each, using N. caninum formalin preserved whole tachyzoites as antigen. Antibodies to T. gondii were found in 17.4% of 166 camels in titers of 1:25 (3 camels), 1:50 (18 camels). and > 1:500 (8 camels) using T. gondii tachyzoites. All 6 camels with N. caninum antibodies had no T. gondii antibodies in 1:4 dilution of serum, indicating specificity of the reaction. This is the first report of N. caninum prevalence in Egypt.     

Neospora caninum causes abortions in a cattle herd in North Rhine Westphalia]

In a cattle herd in North Rhine-Westphalia a series of eight abortions occurred between 15th August and 14th October, 1996. Four of the aborted fetuses were histopathologically, virologically, bacteriologically and parasitologically examined. A multifocal necrotising encephalitis was observed in two fetuses. In addition, the liver of two fetuses and the placenta of one fetus contained infected foci. In three fetuses protozoan stages were detected which were identified as N. caninum by immunohistochemistry. A polymerase chain reaction conducted with histological material revealed N. caninum-DNA in the placenta of an aborted fetus. When seven cows that had aborted were tested for antibodies directed against N. caninum, six animals were positive by immunofluorescence and all seven by immunoblotting. The serological examination of the entire herd revealed a high herd seroprevalence of antibodies directed against N. caninum. Potential modes of infection with N. caninum in the herd are discussed, in particular the possibility of exposure to a point source (definitive host).     

Nucleocytoplasmic shuttling by protein nuclear import factors

Anti-Neospora caninum antibody was detected in anti-Toxoplasma gondii positive and negative human sera by ELISA, western blot and immunofluorescence assay (IFA). Twelve cases out of 172 (6.7%) Toxoplasma-positive sera cross-reacted with both T. gondii and N. caninum antigens, and one out of 110 Toxoplasma-negative sera reacted with N. caninum antigen by ELISA. By western blot, all 12 sera reacted with T. gondii antigens with various banding patterns but specifically at 30 kDa (SAG1) and 22 kDa (SAG2) bands. With N. caninum antigen, the number of reactive bands was reduced, however a 43 kDa band reacted in three cases in Toxoplasma-positive sera in addition to one in Toxoplasma-negative control sera. All sera of the Toxoplasma-positive group labeled surface membrane of T. gondii, but reacted differently with N. caninum. Fluorescence was detected in surface membrane, subcellular organelles, or both in N. caninum. And one case in the Toxoplasma-negative group also reacted with N. caninum strongly in subcellular organelles. This suggested that the antibody against N. caninum may be present in human sera although the positive rate was very low in this study. The possibility of human infection with N. caninum remains to be evaluated further.     

Identification of a major surface protein on Neospora caninum tachyzoites

Neospora caninum is a recently identified coccidian parasite that is closely related to Toxoplasma gondii. Molecules associated with the surface of N. caninum tachyzoites are likely to be involved in the process of adhesion and invasion of host cells. They probably also participate in the interaction of the parasite with the immune system, and they could play an important role in the pathogenesis of the parasite. To identify such surface molecules, we performed subcellular fractionation studies of isolated N. caninum tachyzoites. Employing the nonionic detergent Triton-X-114, we prepared a membrane fraction. Immunoblot analysis of this fraction using polyclonal antisera directed against tachyzoites of N. caninum and T. gondii resulted in the identification of a protein of approximately 43 kDa (Nc-p43). This molecule was present in two isolates of Neospora (Nc-1 and Liverpool) but was absent in Toxoplasma (RH-strain) tachyzoites. Further immunofluorescence and immunogold transmission electron microscopy (TEM) studies using affinity-purified anti-Nc-p43 antibodies demonstrated the presence of this molecule on the surface of N. caninum tachyzoites.     

Changes of biochemical markers during fracture healing

Clinical diagnosis of canine nasal mite (Pneumonyssoides caninum) infection is difficult due to the mite's location in the caudal nasal cavity and frontal sinuses. The current study was performed to evaluate the efficacy of milbemycin oxime in treating dogs with nasal mite infection. A prospective open uncontrolled study included 20 dogs with case histories indicating possible nasal mite infection. Inclusion criteria consisted of either nasal mites being demonstrated (group 1, four dogs), or suspicious clinical signs with no other apparent causes, combined with eosinophilia (group 2, 16 dogs). Milbemycin oxime 1 mg/kg was given orally three times at 10-day intervals. In 17 (85 per cent) dogs, clinical signs resolved completely following milbemycin therapy; within 10 days of the first treatment in 13 cases (group 1, four dogs; group 2, nine dogs) and within 14 days in four cases. In the remaining three dogs clinical signs persisted but were diminished.     

Efficacy of ivermectin in a beef-based chewable formulation against Ancylostoma caninum and Uncinaria stenocephala in dogs

The effective dosage of a chewable formulation of ivermectin was determined in 35 young dogs with induced infections of Ancylostoma caninum and Uncinaria stenocephala. Dogs were inoculated with these parasites and held until the infections were patent. Within each of 7 replicates, dogs were allocated randomly to 1 of 5 treatment groups: vehicle control, or ivermectin at 6, 12, 18, or 24 micrograms/kg. Chewable treatments were tailored to body weight. Seven or 8 days after treatment, parasites were recovered using standard techniques. All 7 controls had adult A. caninum (geometric mean = 35.5) and U. stenocephala (geometric mean = 82.6). Against A. caninum, the efficacy of ivermectin was 52%, 98%, 95%, and 97% at 6, 12, 18, and 24 micrograms/kg, respectively. The statistical model that best described the dose response was linear to 12 micrograms/kg with a plateau thereafter. Using this model, the estimated reduction from the predicted control mean was 97.2%; the estimated dose to eliminate 90% of the worms (ED90) was 8.4 micrograms/kg, and the ED95 was 10.5 micrograms/kg. Against U. stenocephala, the dose response was linear in the range studied, with an ED90 of 20.8 micrograms/kg; it was estimated that 93.2% of the worms would be eliminated.     

Identification of Propionibacterium acnes and related organisms by precipitin tests with trichloroacetic acid extracts

During the months May-July, 1974, a follow-up investigation was carried out on hookworm infections in dogs and cats in order to study changes in the hookworm distribution in Surinam since 1967. Fifty-two cats and 152 dogs were examined for the presence of helminths in the oesophagus, small intestine, coecum and heart. In total 15,942 helminths were isolated; most of these (14,006) were hookworms belonging to the species Ancylostoma caninum, A. tubaeforme, A. ceylanicum and A. braziliense. The average size of the hookworn load in dogs and cats appeared to have decreased since 1967, especially as regards A. ceylanicum. The distribution area of A. braziliense had extended largely by 1974 to the east and south in Surinam. In west Surinam A. braziliense infection was not found, just as in 1967.. The remaining parasites were Dipylidium caninum, Toxocara sp., Dirofilaria immitis, Spirocerca sanguinolenta, Spirometra mansonoides, Taenia taeniaeformis and Trichurisvulpis.     

Culling associated with Neospora caninum infection in dairy cows

OBJECTIVES: To estimate the extent to which cows infected with Neospora caninum were culled, compared with noninfected cows, and to identify differences in reasons for culling between infected and noninfected cows. ANIMALS: 442 Holstein cows on a commercial dairy with 36% seroprevalence for N caninum. PROCEDURE: Culling of cows was done after first calving without knowledge of N caninum serologic status. RESULTS: Risk of a seropositive cow dying was not different from that of a seronegative cow (P = 0.50). Seropositive cows were culled 6.3 months earlier than seronegative cows, and had a 1.6 times greater risk of being culled, compared with seronegative cows (P = 0.004), after adjusting for culling risk associated with abortion. For cows culled for low milk production, culling risk for a seropositive cow was twice that for a seronegative cow (P = 0.007). CONCLUSIONS: The economic impact of N caninum infection in dairy cattle can be expected to extend beyond that for abortion alone. Costs of the disease also may include premature culling and diminished milk production. CLINICAL RELEVANCE: Plans to control N caninum infection on dairies should include consideration that benefits may include reduction in premature culling and increase in milk production.     

Serological survey of antibody to Neospora caninum in Japanese dogs

Prevalence of antibody to Neospora caninum (NC) in Japanese dogs were examined. The antibody was positive in 15 of 48 dogs (31.3%) reared in the dairy farms that had case of the abortions due to NC infection or had the cattle seropositive to NC, whereas the prevalence was 7.1% (14 of 198 dogs) among the dogs kept in urban areas. In one dog breeder, all 17 Shetland sheepdogs older than 7 months were seropositive, and one pup was diagnosed as neosporosis 2 months before the first serological examination. The antibody titers of the dogs kept at this breeder were almost unchanged for 1.5 years. Serological evidence of the dogs in the dairy farms and urban areas indicates the transmission of NC between dogs and cattle. Also serological results of the dogs in one breeder may suggest the potential horizontal transmission among dogs.     

Neosporosis-associated bovine abortion in Pennsylvania

Neospora caninum was found in fetal tissues of 34 of 688 cases of bovine abortion submitted to the Pennsylvania Animal Diagnostic Laboratory System during the period from May 1994 to November 1996. The aborted fetuses ranged in gestational age from 3 to 8 months. Microscopic lesions consisted primarily of encephalitis and myocarditis. A labeled (strept) avidin-biotin staining procedure using anti-N. caninum polyclonal rabbit serum revealed N. caninum organisms within the fetal brain (27 of 27), heart (10 of 13), placenta (5 of 6), kidney (2 of 2), liver (1 of 4) and skeletal muscle (1 of 1).     

Neosporosis and bovine abortion in Scotland

Serum samples were collected over a two-year period from aborting cows and their fetuses from throughout Scotland; 465 maternal sera were examined using an immunofluorescent antibody test for IgG antibody to Neospora caninum and 547 fetal sera were examined for IgM and IgG antibody to the parasite; 355 of the maternal sera were paired with their respective fetal samples. The maternal titres ranged from 1/8 to 1/16,384, with 59 per cent (275) having titres of 1/128 or 1/256. Of the 547 fetal sera, 9.9 per cent had IgM titres equal to or greater than 1/64, 11.2 per cent had IgG titres equal to or greater than 1/64 and 15.9 per cent had IgM and/or IgG titres equal to or greater than 1/64. It is concluded that a fetal IgM or IgG titre of 1/64 or more is evidence of fetal exposure to N caninum but that the examination of maternal sera is less reliable on an individual basis, although valuable for indicating the degree of infection in a herd. Although fetal seropositivity does not necessarily provide proof that N caninum was the cause of death in a given case, the observation that 15.9 per cent of aborted fetuses had antibody to the parasite shows that vertical transmission from dam to fetus is relatively common and may be an important cause of fetal loss in cattle in Scotland.     

Histology and cytology of laparoscopically operated "simple ovarian cysts"

In a retrospective study the histopathological findings of 127 laparoscopically operated unilocular anechoic smooth-walled ovarian cysts have been correlated with clinical characteristics (age, duration of observation, complaints, hormonal treatment), size by ultrasound, kind and colour of cysts content as well as cytological findings. The age of patients differed from 16-61 years (mean +/- s: 36 +/- 16). The histopathologic findings yielded 15 (11.8%) functional cysts, 30 (23.6%) persistent corpus luteum cysts, 9 (7.1%) endometriomas, 7 (5.5%) cystic teratomas, 9 (7.1%) undifferenciated cysts and 57 (44.9%) cystadenomas. There were no differences between histopathologic diagnosis groups according to age and cysts size by ultrasound. Functional cysts with complaints (n = 6) may explain that the observation time in 60% of all functional cysts was smaller than 6 weeks, whereas persistent corpus luteum cysts, endometriomas, cystic teratoma and cystadenomas had been observed for longer than 6 weeks in more than two thirds. Intraoperative evaluation of cysts content as "chocolate"-like was suspicious of endometriomas, but was also present in cysts of other histopathological findings. By means of cytology, endometrioma (siderophages) was suspected in 44.4% and a cystadenoma in 42.1% of all histopathologically verified cases. In all, the cytologic findings were useful for correct histopathological diagnosis in only 33.9% of all 127 cases. It is concluded that differential diagnosis of simple ovarian cysts is not possible by clinical characteristics, neither by ultrasound nor by cytological evaluation. Ovarian cysts should be observed for at least two hormonal cycles. A hormonal treatment by combination preparations containing high doses of oestrogen is also recommended. In cases of persisting ovarian cysts laparoscopic removal is necessary.