Kamil Sedlák

Toxoplasma gondii and Neospora caninum antibodies in sheep in the Czech Republic.

In the Czech Republic, serum from 547 sheep coming from nine farms was examined for antibodies against parasites Toxoplasma gondii and Neospora caninum by screening ELISA. Antibodies against T. gondii were found in 325 sheep (59%) with prevalence ranging from 11% to 96% in different farms. Antibodies against N. caninum were found in 63 sheep (12%) with prevalence ranging from 4% to 21% in different farms. Mixed infections were found in 53 sheep (10%). It was the first evidence of N. caninum antibodies in sheep from the Czech Republic.

Antibodies to Selected Viral Disease Agents in Wild Boars from the Czech Republic

Blood samples were collected from wild boar (Sus scrofa) shot during the hunting season from 1999 to 2005 in the Czech Republic. Sera were tested by enzyme-linked immunosorbent assay for the presence of antibodies against classical swine fever virus (CSFV), swine vesicular disease virus (SVDV), Aujeszkys disease virus (ADV), and bovine viral diarrhea virus (BVDV). Indirect fluorescence antibody test was used for detection of antibodies against porcine circovirus type 2 (PCV-2) and transmissible gastroenteritis virus (TGEV). Antibodies against ADV, BVDV, PCV-2, and TGEV were detected in 30% (101 of 338), 1% (2 of 352), 43% (57 of 134), and 1% (1 of 134) of wild boars, respectively. Sera of 6,471 and 362 tested wild boars were negative for the presence of antibodies against CSFV and SVDV, respectively. This is the first survey of TGEV antibodies in wild boars and the first serologic survey of viral diseases in wild boars in the Czech Republic. Wild boars in the Czech Republic may act as a potential reservoir of ADV and thus have a role in the epidemiology of this disease.

Neospora spp. and Toxoplasma gondii antibodies in horses in the Czech Republic

During January 2007, blood samples were collected from 552 healthy horses from nine different regions of the Czech Republic. Sera were tested for serum antibodies to Neospora caninum by a competitive-inhibition enzyme-linked immunosorbent assay and confirmed by an indirect fluorescent antibody test. The same samples were tested for serum antibodies against Toxoplasma gondii by a latex agglutination test. In total, 131 of 552 (24%) horses reacted positively for Neospora antibodies in competitive-inhibition enzyme-linked immunosorbent assay; seven of them had ≥50% of inhibition. Samples were confirmed in indirect fluorescence test, and only two samples were positive with final titres 50 and 100, while others were negative. Antibodies against T. gondii were found in 125 (23%) horses. This is the first serologic survey for Neospora spp. antibodies performed on horses in the Czech Republic.

High susceptibility of partridges (Perdix perdix) to toxoplasmosis compared with other gallinaceous birds.

Partridges ( Perdix perdix ), chukars ( Alectoris chukar ), wild guineafowl ( Numida meleagris ), wild turkeys ( Meleagris gallopavo ) and chickens ( Gallus domesticus ) were inoculated per os with 103 or 105 Toxoplasma gondii oocysts (K7 strain). Two of five partridges fed 103 oocysts and six of eight partridges fed 105 oocysts died between day 6 and 16 post-inoculation (p.i.); no clinical symptoms were observed in surviving birds. Antibodies to T. gondii were detected in the birds by the indirect fluorescence antibody test (IFAT) first on day 7 p.i. On days 14, 21 and 28 p.i. (end of the experiment), antibodies were found in all partridges, chukars, guineafowl and turkeys. In chickens, IFAT antibodies were first detected on day 14 p.i., and all chickens were serologically positive on days 21 and 28 p.i. Bioassay in mice revealed T. gondii in the brain, liver, spleen, heart and leg muscles of all partridges and chukars. Enteritis was the most striking lesion in partridges that died. Results indicated that partridges are highly susceptible to toxoplasmosis, while chukars, wild guineafowl and turkeys seem to be less susceptible. Chickens are highly resistant to T. gondii infections.

Fatal toxoplasmosis in brown hares (Lepus europaeus): possible reasons of their high susceptibility to the infection.

Brown hares (Lepus europaeus) trapped in the countryside and domestic rabbits were experimentally infected with Toxoplasma gondii (K7 strain) oocysts. Hares (n=12) were divided into groups of 4 and infected with 10, 10(3) and 10(5) oocysts. Rabbits (n=12) were infected in the same way. The experimentally infected animals were monitored for 33 days after infection (p.i.). Most of the infected hares demonstrated behavioural changes, and all of them died between 8 and 19 days p.i. Three of the rabbits demonstrated only clinical changes related to the concurrent pasteurellosis. The typical pathological finding in the hares were haemorrhagic enteritis, enlargement and hyperaemia of mesenteric lymph nodes, splenomegaly and multiple miliary necrotic lesions in the parenchyma of the liver and other organs. Pathological changes in the rabbits were less pronounced than in the hares. In rabbit brains, tissue cysts of the T. gondii were found. The incidence of T. gondii antibodies both in the hares and the rabbits was first ascertained on day 7 p.i. On day 12 p.i., antibodies were already found in all the animals infected. Antibody titres in indirect fluorescence antibody test (IFAT) using the anti-rabbit conjugate were markedly higher in rabbits than in hares. In all hares, T. gondii was isolated post mortem from the liver, brain, spleen, kidney, lung, heart and skeletal muscles. Although T. gondii was also isolated in all rabbits, it was not always isolated in all their organs. In all hares, parasitemia was demonstrated on days 7 and 12 p.i. The percentage of rabbits with detected parasitemia was lower. In hares, a decrease in the numbers of leukocytes during the infection was observed. No such decrease was observed in the rabbits. The lymphocyte activity after the stimulation with non-specific mitogens showed significant differences between the hares and the rabbits even before the infection. After the infection, the hares infected with 10(3) and 10(5) doses and in rabbits infected with a 10(5) dose showed a decrease of lymphocyte activity. Rabbits infected with a 10(3) dose showed an increase of the lymphocyte activity. While in hares toxoplasmosis was an acute and fatal disease, the infection in rabbits had subclinical manifestations only and easily passed to a latent stage. The different courses of toxoplasmosis in the hare and the rabbit may be due to the differences in the natural sensitivity of the two species to the T. gondii infection or a negative impact of stress to the immune status of hares.

Serologic survey for toxoplasmosis in domestic birds from the Czech Republic

The present study was carried out on domestic fowls in the Czech Republic in the years 2003 and 2004. A total of 1108 samples obtained from 538 water birds (geese and ducks) and 570 gallinaceous birds (chickens and turkeys) were tested for specific antibodies against Toxoplasma gondii using the indirect fluorescent antibody test. In geese, 43% prevalence (77 positive out of 178 examined) was found with titres of 40, 320 and 2560 in 40, 29 and eight geese, respectively. In ducks, 14% prevalence (52 out of 360) was found with titres of 40 and 320 in 46 and six ducks, respectively. In gallinaceous birds, only one of 293 broiler chickens was positive with a titre of 40; sera of 217 hens and 60 turkeys were negative. We found extremely high antibody prevalence in geese and ducks compared with gallinaceous birds. This is the first seroprevalence study in domestic birds by indirect fluorescent antibody test in Europe.

Seroprevalence and risk factors of Neospora spp. in donkeys from Southern Italy

In some European countries there is an increasing interest on donkey. Despite there are few data regarding the donkeys parasitic diseases especially those with a protozoal etiology as neosporosis. Samples used in the study were collected from 238 domestic donkeys during year 2010 in Southern Italy from 207 females and 31 males of five breeds (Martina-Franca, Amiata, Sicilian-Grey, Ragusano, Sardinian) and crossbreeds with the average age 9 years (1 month - 24 year). Sera were tested by a competitive-inhibition enzyme-linked immunosorbent assay for antibodies against Neospora caninum; the sera were marked positive, if more than 30% inhibition was found. Out of a total 238 donkeys, 28 (11.8%) were found positive for Neospora antibodies with 12% in females and 6% in males. Different seroprevalence 15.4%, 16%, 12% and 8.8% were found in age categories <1 year, 1-4 years, 5-9 years and ≥10 years, respectively. The seroprevalence ranged in different breeds from 36% (Sicilian-Grey) to 0% (Sardinian) and in different use from 17% (for breeding) to 0% (for meat production). Logistic regression analysis demonstrated evidence of a significant (P<0.05) association between crossbreed origin of samples and risk of protozoan infection; age of donkeys was also significant risk factor for protozoan infection. No statistical significant difference (P>0.05) was found among genders and use of donkeys and risk of N. caninum infection. This is the first serological survey for Neospora spp. performed in donkeys.

Conflicting results of serological, PCR and microscopic methods clarify the various risk levels of canine babesiosis in Slovakia: a complex approach to Babesia canis diagnostics.

We have performed a survey of Babesia canis prevalence within group of dogs living in Southern and Western Slovakia. Blood samples and sera from 217 dogs, including individuals suspected of having babesiosis, were examined by nested PCR-RFLP, light microscopy and indirect fluorescence antibody test (IFAT). The detection of B. canis DNA revealed the highest number of infected dogs in the region of Nové Zámky, with 23 B. canis-positive blood samples (35.4%, n=65), followed by an area close to Komárno (both areas of Southern Slovakia), where 1 dog out of 52 collected (1.9%) had detectible B. canis DNA in the blood stream. The serological method revealed an opposing pattern, with only 3 dogs (4.8%, n=63) sampled at Nové Zámky presenting IgG antibodies against B. canis, while in Komárno region such antibodies were detected in 15 dogs (28.8%, n=52). This discrepancy may be because the majority of samples from Nové Zámky were dogs suspected of an acute phase of canine babesiosis, whereas dogs at Komárno were sampled during a vaccination campaign, and thus were without any clinical signs of the disease. The latter group contains evidently recovered carriers of IgG against B. canis. Hence, the combination of PCR-based and serological methods enabled us to discover both recently infected as well as recovered dogs, thus obtaining a more realistic view on the epidemiological situation. Remarkably, we did not find any positive samples in the vicinity of Stupava (district Malacky, Western Slovakia), either by PCR-RFLP, microscopy or IFAT (n=100). Considering the numerous falsely diagnosed cases of canine babesiosis, we suggest that light microscopy as the simplest and most accessible diagnostic test. Southern Slovakia was confirmed as an area of high risk of canine babesiosis, whereas conclusions about B. canis spreading over Western Slovakia should be considered with wariness.

Seroprevalence of Toxoplasma gondii in Donkeys (Equus asinus) in Italy

ABSTRACT Toxoplasmosis, an important zoonosis, can be transmitted by eating meat or drinking milk of animals infected with Toxoplasma gondii. Samples were collected from 238 donkeys in the year 2010 in Italy, which included 207 females and 31 males of five breeds and crossbreeds with the average age 9 years (1 month−24 years). Sera were tested for T. gondii antibodies using a latex agglutination test and the indirect fluorescent antibody test; 5 and 8% seropositivity were recorded, respectively. We found significant correlation between the presence of T. gondii antibodies and sex, age, grazing and presence of cats on the farms and their access to donkey feed. This is the first detection of T. gondii antibodies in donkeys in Italy.

Seroprevalence of antibodies of Neospora spp. and Toxoplasma gondii in horses from southern Italy

The consumption of horse meat has been epidemiologically linked to clinical toxoplasmosis in humans and neosporosis that may cause clinical illness in horses. Here we determined seroprevalence of antibodies against Toxoplasma gondii Nicolle et Manceaux, 1908 and species of Neospora Dubey, Carpenter, Speer, Topper et Uggla, 1988 in horses from Italy. Blood samples were collected from 643 apparently healthy horses from 60 farms of 51 municipalities in southern Italy. The presence of antibodies against T. gondii and Neospora spp. were detected by indirect fluorescence antibody test (IFAT); a titre ≥ 50 was considered positive. The same sera were also tested for antibodies against Neospora spp. by a competitive-inhibition enzyme-linked immunosorbent assay (cELISA); samples with ≥ 30% inhibition were considered positive. Antibodies against T. gondii and Neospora spp. were detected in 19 (3.0%) and 15 (2.3%) horses by IFAT, respectively, without statistical difference between gender, age and breeds (p-value ≥ 0.05). Antibodies against species of Neospora were detected in 70 (10.9%) horses by cELISA with statistical difference in gender (6.0-18.5%, p-value ≥ 0.05) and breeds (0-19.4%, p-value ≥ 0.05). Although T. gondii infection rates were low, the risk of human infection should not be dismissed, particularly in Italy where consumption of raw or undercooked horse meat has a long tradition.