Serological and molecular detection of Theileria equi infection in horses in Hungary.

The prevalence of Theileria equi infection was studied in 324 healthy horses from 27 farms in Hungary with cELISA and IFAT and the blood samples of 101 horses selected randomly were also examined by PCR. The results indicate that there are many stud farms where one or more horses are infected with T. equi. Among 27 farms 17 (67.9%) were found to have seropositive horses. The seroprevalence of theileriosis among the tested stud farms ranged between 0 and 100%. No marked differences were found in seropositivity between geographical areas. The overall prevalence of positive samples was 32.0% with cELISA as well as with IFAT. The results obtained with cELISA and IFAT in this study had the strongest agreement, except for 9 samples in which the two serological tests gave different results. The prevalence of infection among 101 horses was 49% with PCR. All 14 sequenced samples were found by BLAST analysis to be closest to the T. equi 18S rRNA gene sequences in GenBank with a similarity of ≥ 99%. No significant association was found between the seropositivity and the age of horses. Horses below 5 years of age had three times higher chance to be PCR-positive, than older ones. There was no significant association between the gender and the results of diagnostic tests (cELISA: p=0.40; IFAT: p=0.25; PCR: p=0.41). Based on the findings, the prevalence of equine theileriosis is much higher than expected and it occurs in many regions of the country unlike equine babesiosis. To the authors' knowledge, this is the first report of the serological and molecular survey of T. equi infection in horses in Hungary.

Infection with Toxoplasma gondii during pregnancy: seroepidemiological studies in Austria.

Seropositivity among pregnant woman in Austria has decreased from 48% to 50% at the end of the 1970s to 35% in recent years. Despite this decrease, knowledge of possible sources and risk factors for Toxoplasma infection remains important. We reviewed seroepidemiological studies that were undertaken to assess the roles of undercooked meat and oocysts in cat faeces as potential sources of infection in pregnant women. Improved management and hygiene in pig herds raised in confinement have resulted in less contact of pigs with cats and a decrease of infected pigs within one decade from 14% in 1982 to 0.9% in 1992. In Austrian wild boar populations, however, seroprevalences remained essentially unchanged during the same decade (18% in 1983 and 19% in 1990-1993). Austrian sheep and goats are usually kept on small farms where cats abound and are predominantly seropositive (66% in sheep and 69% in goats). The seroprevalence in cats has decreased from approximately 81% in 1987 to 59% in 1996; presumably because of cats' increased consumption of processed food. Despite the decrease of infection in pregnant women via the cat-to-pig pathway, it may be offset by a recent concomitant increase in mutton consumption. Free-ranging chickens are a good indicator of the prevalence of T. gondii oocysts in the soil because chickens are ground feeders. Antibodies to T. gondii, as evaluated by the modified agglutination test, were found in 36% of chickens from 12 Austrian 'biofarms'. Because Austrians rarely consume raw pork, the fraction of human T. gondii infections associated with pig meat consumption is likely small. As meat consumption and lifestyle patterns change in Austria, the risk of human infection with T. gondii via different pathways needs re-evaluation and targeted educational efforts to control transmission.

Seroprevalence of canine babesiosis in Hungary suggesting breed predisposition.

Six hundred fifty-one blood samples were collected from urban and rural dogs in various parts of Hungary to measure antibody levels to Babesia canis with indirect fluorescent antibody test. Thirty-seven (5.7%) of the sera showed positivity with titers between 1:80 and 1:10,240. Seroconverted dogs were found in 13 locations of the country. It is concluded that canine babesiosis is becoming more prevalent in Eastern Hungary. Seropositivity increased then declined with age, reaching a maximum in case of 3.1- to 5-year-old dogs. Prevalence of antibodies to B. canis was significantly higher among german shepherds and komondors. This suggests a genetic predisposition of german shepherd dogs to chronic babesiosis (carrier status) with long-term maintenance of their seropositivity. On the other hand, heavy-coated komondors are phenotypically more suitable for repeated exposure to ticks, potentially infected with B. canis. This is the first report on the seroprevalence of canine babesiosis in Hungary.

Canine neosporosis in Hungary: screening for seroconversion of household, herding and stray dogs.

In order to assess the seroprevalence of canine neosporosis 651 blood samples were collected from 586 household, 41 herding and 24 stray dogs, at small animal clinics in four large cities and other places of Hungary. Nineteen (2.9%) showed positivity in the IFAT with titres between 1:80 and 1:10240. Two dogs with high titres of antibodies to Neospora caninum had neuromuscular signs (imbalance, tremor) and a further one developed papulomatous, ulcerative and necrotizing dermatitis. There was no correlation between titers and age, sex, breed or keeping place. Although more male dogs had antibodies to N. caninum than females in case of both household and herding dogs, this association was not significant. No breed predisposition was observed. However, dogs with seroconversion were significantly more prevalent among rural (6%) than among urban dogs (1%), indicating that dogs in the countryside may have contact with or access to potentially infected offal from cattle and other intermediate hosts more frequently than those in large cities. Furthermore, significantly more herding dogs (29.3%) had antibodies to N. caninum than household dogs (1.2%), confirming the association between the occurrence of neosporosis and dog keeping on farms. The 12 dogs found seropositive among herding ones lived on 6 farms, on 5 of which seropositive cattle were also identified. This is the first report on the prevalence of N. caninum infection in dogs in Hungary.

Genetic and biologic characteristics of Toxoplasma gondii infections in free-range chickens from Austria.

The prevalence of Toxoplasma gondii in free-ranging chickens is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. The prevalence of T. gondii in free-range chickens (Gallus domesticus) from 11 Bio-farms in Austria was determined. Antibodies to T. gondii assayed by the modified agglutination test (MAT) were found in 302 of 830 (36.3%) chickens with titers of 1:10 in 50, 1:20 in 69, 1:40 in 53, 1:80 in 40, 1:160 or higher in 90. Hearts of 218 chickens with MAT titers of 10 or higher were bioassayed individually in mice. Tissues from 1183 chickens were pooled and fed to 15, T. gondii-free cats. Feces of the cats were examined for oocysts; 11 cats shed T. gondii oocysts. T. gondii was isolated from 56 chickens by bioassay in mice. Thus, there were 67 isolates of T. gondii from these chickens. Genotyping of these 67 isolates using the SAG2 locus indicated that all 33 were Type II. Phenotypically and genetically these isolates were different from T. gondii isolates from Brazil. None of the isolates was virulent for mice. This is the first report of isolation of T. gondii from chickens from Austria.

Toxoplasmosis in Pallas' cats (Otocolobus manul) raised in captivity.

Manuls or Pallas' cats (Felis manul, syn. Otocolobus manul) are endangered wild cats from Central Asia kept and bred in many zoos. Despite good breeding success young cats frequently die from acute toxoplasmosis. From 1998 to 2002, a breeding pair in the Schönbrunn Zoo in Vienna, Austria, gave birth to 24 kittens; 58 % of kittens died between the 2nd and the 14th week of life, mostly due to acute toxoplasmosis. The epidemiology of toxoplasmosis in Pallas' cats was examined and a control strategy to protect the kittens from fatal toxoplasmosis was developed. One 12-week-old kitten from a litter of 6 born in 2001 died of generalized toxoplasmosis. This kitten had shed T. gondii oocysts that were bioassayed in mice. Toxoplasma gondii was isolated in tissue culture inoculated with tissues of these mice. The surviving animals were immediately treated with clindamycin for 16 weeks; they acquired a natural infection and seroconverted by the end of this time without clinical signs.

Prevalence of antibodies against Neospora caninum and Toxoplasma gondii in dogs and foxes in Austria.

Sera from 1770 dogs and 94 red foxes from Austria were examined for antibodies against Neospora caninum using the indirect immunofluorescent antibody test (IFAT). 3.6% of the dogs were seropositive with titres ranging from 1:50 to 1:6400. Dogs from rural areas were significantly more often seropositive for N. caninum than those from the urban area of Vienna (5.3% versus 2.1%). There were no significant differences in sex or breed, but a slight increase in seropositivity with age was apparent, indicating postnatal infection. None of the foxes had antibodies against N. caninum. Additionally, sera from 242 dogs and 94 foxes were examined for antibodies against Toxoplasma gondii using the IFAT. Thirty-five percent foxes and 26% of the dogs were positive; 1.7% of the dogs were positive for both parasites. This is the first report of the prevalence of N. caninum infections in dogs and foxes in Austria.

Cross-sectional survey in pig breeding farms in Hesse, Germany: seroprevalence and risk factors of infections with Toxoplasma gondii, Sarcocystis spp. and Neospora caninum in sows.

A cross-sectional survey was performed to estimate the prevalences of antibodies to Toxoplasma gondii (ELISA, IFAT), Sarcocystis spp. (ELISA, using S. miescheriana as antigen) and Neospora caninum (ELISA, immunoblotting) in sows from breeding farms in southern Hesse, Germany. A total of 2041 plasma samples of sows from 94 randomly selected farms was examined. Data on farm profiles, husbandry management and sows were collected by a questionnaire and exploratively analysed. For T. gondii the ELISA results agreed well with the results obtained by IFAT (kappa=0.71). Antibodies to T. gondii were detected by ELISA in 19% of the sows. Sixty-nine percent of the farms had at least one seropositive sow, and a within-farm seroprevalence of >or=50% was observed in 14% of all farms. The prevalence of anti-T. gondii antibodies was positively correlated with the age of sows. The within-herd seroprevalence was significantly higher in farms with reproductive disorders than in those without such problems. On the farm level, the farm type 'piglet production' (versus 'pedigree breeding' or 'farrow-to-finish') was the only risk factor associated with the presence of T. gondii-seropositive sows. Antibodies to Sarcocystis spp. were found in 29% of the sows. Seventy-two percent of the farms harboured at least one seropositive sow, and a within-farm seroprevalence of >or=50% was detected in 23% of all farms. The seroprevalence increased significantly with the age of sows. On the farm level, only the farm type 'piglet production' (versus 'pedigree breeding') and the replacement of sows by purchasing (versus raising on the own farm) were identified as risk factors for seropositivity. Antibodies to N. caninum were detected in one sow using both the screening ELISA and the confirmatory immunoblotting technique. This may indicate the first natural N. caninum infection in pigs.

Use of purified tachyzoite surface antigen p38 in an ELISA to diagnose bovine neosporosis.

Affinity-purified 38 kDa surface antigen of Neospora caninum tachyzoites was used to develop an enzyme-linked immunosorbent assay (ELISA) to diagnose N. caninum-associated abortion in cattle. The specificity of this antigen was demonstrated by using sera from cattle experimentally infected with other apicomplexan parasites. In a panel of field sera, serological differences could be demonstrated between herds with epidemic and endemic abortions. Optimal ELISA cut-offs were determined for the detection of association between seropositivity and abortion in herds with N. caninum-associated abortions. These optimal cut-offs differed markedly when only sera from either endemic or epidemic cases were used for cut-off determination. It may thus be appropriate to apply different cut-offs in serological tests depending on the abortion pattern to be analysed.

Serological differences in Neospora caninum-associated epidemic and endemic abortions.

A sandwich enzyme-linked immunosorbent assay (ELISA) for the sensitive and specific detection of bovine antibodies to Neospora caninum was developed and evaluated using sera from cattle experimentally infected with N. caninum, Toxoplasma gondii, Sarcocystis cruzi, Sarcocystis hominis, Sarcocystis hirsuta, Eimeria bovis, Cryptosporidium parvum, Babesia divergens, and field sera from naturally exposed animals. Field sera were classified using a gold standard that included the results from an indirect fluorescent antibody test (IFAT) and an immunoblot (IB). Based on these gold standard results, i.e., IFAT-IB results, an equal relative sensitivity and specificity of 94.2%(theta0) was reached when a cutoff of 0.034 (d0) was employed. The analysis of IFAT-IB-positive field sera showed that within groups of aborting and nonaborting dams, the animals from herds with endemic N. caninum-associated abortions had significantly higher ELISA indices than animals from herds with N. caninum-associated epidemic abortions. By contrast, IFAT-IB-positive aborting dams from herds with endemic N. caninum-associated abortions had significantly lower IFAT titers than IFAT-IB-positive aborting dams from herds with epidemic N. caninum-associated abortions. This is the first time that statistically significant serological differences between herds exhibiting epidemic and endemic N. caninum-associated abortions are described.

Improved disease resistance after Babesia divergens vaccination.

The efficacy of a new inactivated vaccine against Babesia divergens was evaluated by means of inoculation tests. The infection was initiated by i.v. injection of blood containing 2 x 10(9) living parasites into splenectomized and non-splenectomized calves. Clinical status and hematological parameters were determined. Serology examinations for antibodies against B. divergens were carried out by indirect fluorescent antibody test (IFAT). Non-vaccinated and splenectomized animals exhibited experimental infections. In vaccinated and splenectomized animals, clinical symptoms and prolonged incubation periods were observed.

Prevalence of antibodies against Toxoplasma gondii in pigs in Austria--an evaluation of data from 1982 and 1992.

To obtain information on the prevalence of antibodies against Toxoplasma gondii in pigs in Austria, 4697 blood samples from pigs were collected in the years 1982 and 1992 and tested by an indirect fluorescence antibody test. The results of the studies show that due to modern fattening systems the prevalence of infections with T. gondii in slaughtered pigs has obviously decreased in Austria during the last 10 years. The investigations indicate a significant difference in the rate of infection of pigs between 1982 (13.7%) and 1992 (0.9%). Additionally, the prevalence of T. gondii antibodies in breeding sows (1982, 43.4%; 1992, 4.3%) was higher than that in fattening pigs (1982, 12.2%; 1992, 0.8%).