Seroprevalence and associated risk factors for Neospora caninum infection in dairy cattle in South Africa.

Bovine neosporosis is a widespread parasitic disease associated with significant economic losses. Its effects on the reproductive performance of cows have resulted in losses that run into the hundreds of millions of US dollars in dairy industries in various countries (Reichel et al., Int J Parasitol 43:133-142, 2013). Due to outdated and scant information on the occurrence of Neospora caninum infection in South Africa, the study aimed to determine the seroprevalence and risk factors associated with infection in dairy cattle in South Africa. A total of 1401 blood samples were randomly collected from cattle on 48 dairy farms in seven of the nine provinces in South Africa. A close-ended questionnaire was used in a cross-sectional study to obtain farm-level and animal-level data. Serological testing was done using a commercial IDvet Screen® Neospora caninum Indirect ELISA. An overall seroprevalence, adjusted for test sensitivity and specificity, of 2.3% (95% CI, 1.3-4.1) was detected and 48% (23/48) of sampled farms had at least one animal testing positive. The highest seroprevalence of N. caninum was in the KwaZulu-Natal province with 7.5% (95% CI, 3.8-14.3), and the lowest in Western Cape with 0.1% (95% CI, 0-1.2). The highest within-farm seroprevalence of 25% was detected on a farm in the North West Province. In a multivariable logistic regression model, the odds of N. caninum seropositivity were higher in Holstein-Friesian cattle when compared to other breeds. Good hygiene was identified as a protective factor. Cattle left out on pasture had increased odds of testing positive for N. caninum compared to those that were penned. The odds of testing seropositive for N. caninum was higher on farms that practised segregation of cattle into different age groups. The purchase of replacement animals was a significant risk factor, as open herds had increased odds of N. caninum seropositivity. Cattle on farms that did not have a specific calving location were more likely to be seropositive. This is the first such study in South Africa and shows that N. caninum is widely distributed in the country at a low seroprevalence, but it may be a cause of concern on certain farms.

Molecular and serological prevalence rates of Neospora caninum infection in dogs from Jordan.

Neosporosis is a proven disease of farm animals and dogs caused by Neospora caninum. This cross-sectional study investigates N. caninum prevalence and seroprevalence among 268 dogs. Nc5 gene PCR was carried out on dog faeces and confirmed by sequencing. Seroprevalence was detected using an indirect fluorescent antibody test (IFAT). Three age groups, gender, locality (Amman, Irbid, and Zarqa Governorates), dog type (stray, pet, and breeding), place of living (indoor/outdoor), food type (raw/cooked), having diarrhoea, having abortion in the area, and having animals nearby were tested as independent variables for associations with positivity to N. caninum using univariate and multivariable logistic regression analyses. The true prevalence of N. caninum was 34.3% (95% CI 28.4, 40.5) using the Nc5-PCR test. The true seroprevalence rate of N. caninum among dogs in Jordan was 47.9% (95% CI 41.4, 54.5) using IFAT. The sequenced isolates of Nc5-PCR products (n = 85) matched three N. caninum strains, namely, NcHareGre (n = 70, 82.4%, 95% CI 72.6-89), NC MS2 (n = 14, 16.5%, 95% CI 9.3-26.1), and L218 (n = 1, 1.2%, 95% CI 0.03-6.4). The three strains were isolated previously from three different countries and continents. N. caninum shedding is associated with abortion among dogs and animals in the area (odds ratio = 3.6). In Amman and Zarqa, living indoors reduced seroprevalence at 0.45, 0.24, and 0.02 odds ratios, respectively. Jordan shares three molecular N. caninum strains with three different countries and continents.

Herd-level contamination of Neospora caninum, Toxoplasma gondii and Brucella in milk of Iranian dairy farms.

The bulk milk examination is a reliable screening tool for monitoring the quality of milk in the farms. The infection to Neospora caninum, Toxoplasma gondii and Brucella sp. Was evaluated in bulk milk samples of dairy farms in Hamedan province, West part of Iran. All the dairy farms (n = 149) were examined for N. caninum, T. gondii and Brucella infections using milk ring test (MRT), microbiology, serology (Enzyme-linked Immunosorbent Assay), and molecular techniques. Based on molecular methods, Brucella-infection was negative in all farms; while, 55 %, 5.4 % and 2.7 % of samples were positive for N. caninum, T. gondii and mix infection, respectively. The highest Neospora-infection was detected in the farms with history of abortion in fall and winter. There was significant association between Neospora-infection and the presence of dogs and rodents in the farms, herd size, and age of the animals. Also, a significant association was seen between Toxoplasma-infection and the presence of cats and rodents in the farms, as well as age of the animals. Average total bacterial count (TBC) was calculated 1.14 × 106±1.1 × 106. The highest TBC was in the farms from Central locations of studied area (5.7 × 106±2.24 × 106), farms with more than 120 animals (7.9 × 106±2.8 × 106), and farms with ≥50-months age (1.74 × 106±6.3 × 105) in spring and summer (6.9 × 106±3.7 × 106). The number of somatic cells was estimated between 1 × 104 and 2 × 106 (Average = 4.2 × 105±3.39 × 105). The current study was a comprehensive evaluation of Neospora, Toxoplasma and Brucella infections in milk samples of Iranian dairy farms for the first time. Neospora-infection is responsible for economic losses in the region. Health education and milk pasteurization are so helpful for inhibiting the milk borne diseases. To reduce the risk factors, predict and design the appropriate schemes like redundant of heterogeneous animals are recommended.

Seroprevalence of Toxoplasma gondii and Neospora caninum in dairy cows in Hebei province, China.

Toxoplasma gondii (T. gondii) and Neospora caninum (N. caninum) are protozoan parasites that cause severe disease in animals and significant economic losses for farmers worldwide. Neospora caninum is considered to be a serious parasite for dairy cows, while T. gondii infection is a serious parasitic disease that can infect people and livestock. The aim of the present study was to investigate the seroprevalence of T. gondii and N. caninum in dairy cows in Hebei province, China. A total of 723 dairy cow sera samples from three regions in Hebei were analyzed by enzyme-linked immunosorbent assays (ELISA). The results revealed that 19.92% (144/723) of the individual serum samples were seropositive for T. gondii, 37.34% (270/723) of the individual serum samples were seropositive for N. caninum, and 8.7% (63/723) of the individual serum samples were seropositive for mixed infection. A higher seroprevalence of N. caninum was found in the present study, when compared to previous data. To the best of the author's knowledge, the present study is the first to detect T. gondii and N. caninum antibodies in the vast areas of Hebei province, China.

Urological detection of specific antibodies against Neospora caninum infection in mice: A prospect for novel diagnostic approach of Neospora.

The intracellular protozoan parasite Neospora caninum is incriminated to induce drastic economic losses in both livestock and pet animal industries. Neosporosis is primarily characterized by abortion in cattle and paralytic symptoms in dogs. Because there are no effective treatments or vaccines, diagnosis is critical for Neospora control. Thus, diversification of laboratory tests and specimens used for diagnosis of N. caninum is an essential scientific endeavor to judge and select the most appropriate diagnostic tool. Herein, we provide the first evidence for the utility of urine samples for demonstration of specific antibodies against N. caninum employing an experimentally infected murine model. Specific antibodies to recombinant N. caninum dense granule 7, surface antigen 1, and lysate antigen were assayed using different antibodies-based ELISAs. Urine based IgG ELISA efficiently discriminated between infected mice (acute or chronic infection), and those of non-infected mice. This effect was also noticed for IgG1 and IgG2a suggesting the utility of urine for assessment of T-helper 2- and T-helper 1-mediated immunities, respectively. In addition, reactivity of specific antibody in urine was also confirmed against parasites when indirect fluorescent antibody test was employed. Usefulness of urine as an additional clinical sample for Neospora diagnosis was confirmed via comparison with the relevant control non-infected and infected mouse sera as reference samples. Because of minimum invasiveness and ease of urine collection, this approach might offer new diagnostic opportunities for N. caninum either for the field or research purposes. However, further studies are required to extrapolate this preliminary study and results in the animal species of interest particularly in dogs.

Genetic characterization of Neospora caninum from Northern Italian cattle reveals high diversity in European N. caninum populations.

Recent studies have revealed extensive genetic variations among Neospora caninum, a cyst-forming protozoan parasite that is one of the main causes of bovine abortion in the cattle industry worldwide. Previous genetic studies based on multilocus microsatellite genotyping (MLGs) of different Ibero-American populations showed a high genetic diversity. These studies provided clear clues of a predominant clonal propagation in cattle and population sub-structuring partially associated with geographical origin. Although, these reports were limited to a reduced number of countries. In this study, the N. caninum isolates from aborted bovine fetuses and stillbirths and a goat abortion from Northern Italy were investigated genetically using 9 microsatellite markers. Complete or nearly complete isolate profiles were obtained from 30 fetuses and stillbirths. An extensive genetic diversity was also found in this Italian N. caninum population. The study of genetic relationships among Italian MLGs using network (eBURST) and principal component analyses based on the allele-sharing coefficient (PCoA) showed different clonal subpopulations disseminated throughout Northern Italy without apparent segregation depending on the geographic origin, cattle breed, or time of collection. The presence of linkage disequilibrium supports a predominant clonal propagation of Italian N. caninum. In addition, most of Italian MLGs segregated from other global populations including Spain, Argentina, Mexico, Brazil, Germany, and Scotland, suggesting the existence of specific N. caninum subpopulations in the Northern Italy and different subpopulations of N. caninum circulating in Europe.

Molecular Detection and Genotyping of Toxoplasma gondii and Neospora caninum in Slaughtered Goats in Central China.

Toxoplasma gondii and Neospora caninum infections can cause reproductive failure in animals, including goats, and toxoplasmosis is one of the most important foodborne diseases. However, information on the molecular prevalence and genetic characterization of T. gondii and N. caninum in the tissues of goats in China is limited. In this study, brain samples of 422 slaughtered goats were collected from slaughterhouses in Henan and Anhui provinces, Central China, and examined for the presence of T. gondii and N. caninum by nested polymerase chain reaction (PCR) based on B1 and NC5 genes, respectively. The prevalence of T. gondii and N. caninum DNA was 5.2% and 2.8%, respectively. No significant differences were found between the prevalences of two parasite infections and animal age, sex, and region (p > 0.05). Two of 22 T. gondii-positive samples were completely genotyped at 11 genetic markers (SAG1, [3' + 5'] SAG2, alternative SAG2 [alt. SAG2], SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) using PCR-restriction fragment length polymorphism, and were identified as genotype ToxoDB no. 225, which has not been previously reported in goats in any country worldwide. For N. caninum, two different sequences at the ITS1 region, three genotypes at the MS5 microsatellite locus, and one genotype at the MS8 locus were identified. This study showed that T. gondii and N. caninum are moderately prevalent in goats in Central China; however, it should be emphasized that T. gondii prevalence in goats poses a potential health threat for consumers in the investigated areas. To the best of our knowledge, this is the first report on the genetic characterization of N. caninum isolates from goats in China. Our results have important implications for a better understanding of the genetic diversity of these parasites in China.

Neosporosis among Egyptian camels and its associated risk factors.

Neospora caninum is an obligate intercellular protozoan parasite and considered one of the abortive pathogens in a wide range of ruminants. The present study aimed to investigate the antibodies against N. caninum in camels from three governorates in Egypt to understand the epidemiology of neosporosis and its associated abortion. The study was carried out on 282 camels using commercial enzyme-linked immunosorbent assay (ELISA) kit. The obtained results confirmed the presence of anti-Neospora caninum antibodies in 31 out of 282 camels. Moreover, no significant difference in seroprevalence was observed between different localities under the study and between males and females. On contrary, the number of seropositive camels was increased significantly in adult camels of 8-11 years old (OR = 9.1, 95% CI: 1.1-73.1), during the autumn (OR = 2.3, 95% CI: 0.79-7.1) and mainly in females with history of abortion (OR = 2.8, 95% CI: 1.2-6.5) or in camels raised with dogs (OR = 2.8, 95% CI: 1.1-7.08). The presence of antibodies against N. caninum in camels from Egypt increases the demand for further studies to determine its epidemiological role as a cause of abortion and reduce its economic impacts on camel's industry.

Toxoplasma gondii, Neospora caninum and Encephalitozoon cuniculi in Animals from Captivity (Zoo and Circus Animals).

Problems with parasitic infections are common in zoological gardens and circuses. In some animals it can lead to several disorders such as systemic disease, reproductive disorders (abortions and neonatal mortality), and even to death if severe illness is untreated. Thus, the aim of this study was to evaluate the prevalence of three common parasites in 74 animals from three zoos, and four circuses in Southern Italy. Antibodies to Toxoplasma gondii, Neospora caninum, and Encephalitozoon cuniculi were detected in 51%, 12%, and 20% of animals, respectively. Co-infections of T. gondii and N. caninum were reported in seven animals (9%) and co-infection of T. gondii and E. cuniculi in one animal. T. gondii, N. caninum and E. cuniculi seroprevalence differed in type of diet (P ≤ 0.0001; P ≤ 0.037 and P ≤ 0.004, respectively). T. gondii and E. cuniculi seroprevalence also differed in animal families (P ≤ 0.0001) and according to type of housing (P ≤ 0.003), respectively. Statistical differences were not found in other characteristics (gender, age, country of birth, origin, and contact with cats or dogs). This is the first serological study focusing on protozoan and microsporidian parasites in zoo and circus animals from Southern Italy and the first detection of antibodies to E. cuniculi in camels in Europe.

Detection of Neospora caninum DNA in cases of bovine and ovine abortion in the South-West of Scotland.

Neospora caninum is a commonly diagnosed cause of reproductive losses in farmed ruminants worldwide. This study examined 495 and 308 samples (brain, heart and placenta) which were collected from 455 and 119 aborted cattle and sheep fetuses, respectively. DNA was extracted and a nested Neospora ITS1 PCR was performed on all samples. The results showed that for bovine fetuses 79/449 brain [17.6% (14.2-21.4)], 7/25 heart [28.0% (12.1-49.4)] and 5/21 placenta [23.8% (8.2-47.2)] were PCR positive for the presence of Neospora DNA. Overall 82/455 [18.0% (14.6-21.7)] of the bovine fetuses tested positive for the presence of N. caninum DNA in at least one sample. None (0/308) of the ovine fetal samples tested positive for the presence of Neospora DNA in any of the tissues tested. The results show that N. caninum was associated with fetal losses in cattle (distributed across South-West Scotland), compared to sheep in the same geographical areas where no parasite DNA was found. Neospora is well distributed amongst cattle in South-West Scotland and is the potential cause of serious economic losses to the Scottish cattle farming community; however, it does not appear to be a problem amongst the Scottish sheep flocks.

Experimental inoculation of Neospora caninum tachyzoites in eared doves (Zenaida auriculata).

Neospora caninum is an apicomplexan parasite distributed worldwide. Although a positive association between the presence of birds and abortions in cattle associated to N. caninum has been reported, the role of the birds in the epidemiologic cycle of the parasite is unknown. To the best knowledge, no experimental studies have evaluated N. caninum in the eared dove, Zenaida auriculata. Therefore, we aimed to determine whether Z. auriculat can act as intermediate host for N. caninum. Eighteen birds were divided into four groups, G1, G2, G3, and G4 (control); G1, G2 and G3 received 2 × 106 tachyzoites of NC-1 strain via different routes: subcutaneous, intramuscular, and intraperitoneal, respectively. G4 composed of three birds. Serum samples were collected weekly, and one bird each from G1, G2 and G3 was euthanized on the 7th and 14th day post-inoculation (dpi). The remaining birds were euthanized after the 28th dpi. Tissues from the doves were evaluated using histopathological analysis, PCR and dog bioassay to detect the parasite. Dogs were fed with tissues from the birds and monitored for 30 days. Serum samples were collected weekly from the dogs for serological analysis, and feces samples were collected daily until the end of the experiment for coproparasitological examinations. No dove showed clinical signs of the infection; however, all of them seroconverted after the inoculation, with stronger immunological response in the G3 birds. The lung tissue of one G3 bird showed positive PCR results; it was euthanized on the 7th dpi, and an inflammatory infiltrate was observed in the lung and kidney from this dove. The dogs did not shed oocysts or seroconverted. Our results indicate that the intraperitoneal route induced infection in the doves; however, the parasite may have been eliminated by the host, and the doves may be resistant to chronic infection.

The pathogenicity of two Neospora caninum goat strains in a BALB/c mouse model.

Goats are frequently described as an intermediate host for the protozoan Neospora caninum, manifesting the disease mainly by recurrent abortions with placentitis and encephalitis in fetuses. Several reports of natural and experimental infections in cattle and mice show differences in the immune response, and the outcome of the infection can be variable depending on the species affected and by the behavior of the infective strain. This study describes for the first time two Neospora caninum strains isolated from naturally infected goats from the state of Minas Gerais, Brazil. One placenta and one brain from different goats were processed for a first bioassay in gerbils (Meriones unguiculatus). Subsequently, a second bioassay was performed by inoculating the processed brain samples from gerbils into Interferon gamma (IFN-γ) knockout mice (KO mice). Tachyzoites collected from the peritoneal fluid of the KO mice were inoculated into VERO cell monolayers, where they presented a very slow growth rate. The tachyzoites were also inoculated into BALB/c mice with a dose of 106 tachyzoites per animal. After a 5-week follow up, the animals infected with both of the strains developed a strong polarized Th1 response with increased serum and spleen gene expression levels of pro-inflammatory cytokines (mainly IFN-γ and TNF-α) in the first week. Tissue lesions were mild in the animals infected with both strains. Despite the strong immune response preventing an infection in the visceral organs, the parasite was able to reach the brain, causing progressive brain lesions from the second to fifth week post infection. The NC-goat1-infected mice presented with severe meningoencephalitis, but the NC-goat2-infected animals had considerable histological brain lesions only at week 5. Immunohistochemical analysis of the mouse brains revealed a different pattern of inflammatory cells compared to the naturally infected goats. A severe inflammatory infiltrate of CD3+ T lymphocytes was found in the NC-goat1-infected mice. A more discrete infiltrate of CD3+ T cells was found in the NC-goat2-infected animals. Additionally, IBA1 IHC revealed an intense microglial reaction and monocyte perivascular cuffs in the NC-goat1-infected animals and lower microglia/monocyte infiltrates in the NC-goat2-infected mice. This work contributes knowledge on the pathogenicity of new Neospora caninum strains in mice, comparable with other well-established mouse models of the disease, and demonstrates the importance of studying goats as an intermediate host of this parasite.

Evidence of Neospora caninum infection in buffaloes (Bubalus bubalis) from Northwestern Romania.

Neospora caninum is a heteroxenous parasitic Apicomplexan protozoan, able to infect a variety of domestic and wild animals, mainly associated with reproductive disorders and abortions in susceptible species. Considering the importance of Romania in the European buffalo industry and the severe economic losses caused by N. caninum infection in livestock, the aim of the present study was to assess the occurrence of neosporosis in buffaloes raised in household and farming systems in northwestern Romania. Overall, 197 serum samples were tested for the presence of N. caninum-specific antibodies by ELISA. Additionally, from 74 slaughtered buffaloes, diaphragm digest, heart, and lymph node samples were tested for the presence of N. caninum DNA. The overall seroprevalence was 68.5%. The seroprevalence was significantly higher in adult animals compared to calves and young ones and in the household system compared to farm. By nPCR, six samples were found to be positive for N. caninum DNA, of which three were serologically negative. The present study reveals a high prevalence of N. caninum-specific antibodies in buffaloes in northwestern Romania and to our best knowledge, it is the first one performed in Romania.

Comparison of PCR assays to detect Toxoplasma gondii oocysts in green-lipped mussels (Perna canaliculus).

Toxoplasma gondii is recognised as an important pathogen in the marine environment, with oocysts carried to coastal waters in overland runoff. Currently, there are no standardised methods to detect T. gondii directly in seawater to assess the extent of marine ecosystem contamination, but filter-feeding shellfish may serve as biosentinels. A variety of PCR-based methods have been used to confirm presence of T. gondii DNA in marine shellfish; however, systematic investigations comparing molecular methods are scarce. The primary objective of this study was to evaluate analytical sensitivity and specificity of two nested-PCR (nPCR) assays targeting dhps and B1 genes and two real-time (qPCR) assays targeting the B1 gene and a 529-bp repetitive element (rep529), for detection of T. gondii. These assays were subsequently validated for T. gondii detection in green-lipped mussel (Perna canaliculus) haemolymph using oocyst spiking experiments. All assays could reliably detect 50 oocysts spiked into mussel haemolymph. The lowest limit of detection was 5 oocysts using qPCR assays, with the rep529 primers performing best, with good correlation between oocyst concentrations and Cq values, and acceptable efficiency. Assay specificity was evaluated by testing DNA from closely related protozoans, Hammondia hammondi, Neospora caninum, and Sarcocystis spp. Both nPCR assays were specific to T. gondii. Both qPCR assays cross-reacted with Sarcocystis spp. DNA, and the rep529 primers also cross-reacted with N. caninum DNA. These studies suggest that the rep529 qPCR assay may be preferable for future mussel studies, but direct sequencing is required for definitive confirmation of T. gondii DNA detection.

Isolation and molecular characterization of four novel Neospora caninum strains.

Neospora caninum causes neosporosis, a leading cause of bovine abortion worldwide. Uruguay is a developing economy in South America that produces milk to feed seven times its population annually. Naturally, dairy production is paramount to the country's economy, and bovine reproductive failure impacts it profoundly. Recent studies demonstrated that the vast majority of infectious abortions in dairy cows are caused by N. caninum. To delve into the local situation and contextualize it within the international standing, we set out to characterize the Uruguayan N. caninum strains. For this, we isolated four distinct strains and determined by microsatellite typing that these represent three unique genetic lineages, distinct from those reported previously in the region or elsewhere. An unbiased analysis of the current worldwide genetic diversity of N. caninum strains known, whereby six typing clusters can be resolved, revealed that three of the four Uruguayan strains group closely with regional strains from Argentina and Brazil. The remaining strain groups in an unrelated genetic cluster, suggesting multiple origins of the local strains. Microsatellite typing of N. caninum DNA from fetuses opportunistically collected from local dairy farms correlated more often with one of the isolates. Overall, our results contribute to further understanding of genetic diversity among strains of N. caninum both regionally and worldwide.

Seroprevalence of Neospora caninum infection and associated risk factors in dairy cattle in Serbia.

Neosporosis in cattle is a globally important disease, causing abortions and significant economic losses if epidemic abortions occur. In Serbia, however, studies of Neospora caninum infection in cattle are few and are based on limited samples and/or from limited areas. We thus performed a nationwide study to examine the seroprevalence of N. caninum infection in a sample of cows randomly selected from dairy farms in 12 epidemiological units from throughout Serbia, as well as the possible transmission risk factors. Sera from a total of 1496 cattle were tested by competitive ELISA, and N. caninum-specific antibodies were shown in 7.2% animals (95% confidence interval CI, 6.6-7.9%), ranging from 2.2 to 12% across the epidemiological units. At least one seropositive animal was detected on 10.7% (95% CI, 9.7-11.8%) of farms, with a range of 5.9-25.9%. Logistic regression analysis showed that the single risk factor for infection in individual animals was keeping cows in loose-stalls (OR = 3.31, 95% CI = 1.95-5.60, P < 0.001). Risk factors for the presence of infection on farms also included housing in loose-stalls (OR = 18.49, 95% CI = 5.40-63.36, P < 0.001), and herd size > 100 animals (OR = 24.08, 95% CI = 3.85-150.50, P = 0.001). In view of the relatively low prevalence of infection showed at both the individual and farm level, this is the perfect time to undertake appropriate preventive measures to improve animal health and reduce economic losses.

Vertical transmission of Neospora caninum in bovine fetuses from a slaughterhouse in Brazil.

Neospora caninum is considered as one of the main causes of reproductive failure in cattle. Vertical transmission is the main route of infection in the bovine host and plays an important role in maintaining the parasite in the herd. Molecular detection of N. caninum is important to determine the occurrence of the disease and to evaluate the genetic diversity of the parasite. The present study aimed at assessing the vertical transmission of N. caninum using molecular techniques to detect the parasite in tissue samples from bovine fetuses collected in a slaughterhouse in the state of São Paulo, Brazil. Seventy fetuses and 70 blood samples from pregnant cows were collected in a slaughtering line. Fresh samples of heart and brain tissue from fetuses were analyzed using molecular assays. Serum samples from fetuses and cows were subjected to an indirect fluorescent antibody test (IFAT) to detect antibodies against N. caninum. Nested PCR targeting the internal transcriber 1 (ITS1) region of the protozoan organism was used in the molecular testing. From the total of fetuses examined, 71.42% were positive for N. caninum by PCR. A higher number of heart samples (47.1%) were positive for the parasite using this technique. Antibodies against the protozoa were detected in 12.9% of serum samples of cows; 2.8% of fetuses were seropositive for this pathogen. Our results show that vertical transmission of N. caninum occurs in cattle from this region of Brazil, and that the use of different diagnostic techniques contributes to successful diagnosis of congenital transmission of the parasite in cattle.

Diagnostic accuracy of enzyme-linked immunosorbent assay (ELISA) and immunoblot (IB) for the detection of antibodies against Neospora caninum in milk from dairy cows.

The aim of the study was to estimate the sensitivity and specificity of enzyme-linked immunosorbent assay (ELISA) and immunoblot (IB) for detecting antibodies of Neospora caninum in dairy cows, in the absence of a gold standard. The study complies with STRADAS-paratuberculosis guidelines for reporting the accuracy of the test. We tried to apply Bayesian models that do not require conditional independence of the tests under evaluation, but as convergence problems appeared, we used Bayesian methodology, that does not assume conditional dependence of the tests. Informative prior probability distributions were constructed, based on scientific inputs regarding sensitivity and specificity of the IB test and the prevalence of disease in the studied populations. IB sensitivity and specificity were estimated to be 98.8% and 91.3%, respectively, while the respective estimates for ELISA were 60% and 96.7%. A sensitivity analysis, where modified prior probability distributions concerning IB diagnostic accuracy applied, showed a limited effect in posterior assessments. We concluded that ELISA can be used to screen the bulk milk and secondly, IB can be used whenever needed.

Isolation of Neospora caninum from kidney and brain of a bovine foetus and molecular characterization in Brazil.

Bovine neosporosis has become a disease of international concern as it is among the main causes of abortion in cattle. Viable N. caninum has been isolated from brains of fetuses and neonatal calves, and there is no report of isolation of tachyzoites from kidney. Also, detailed information about the genetic diversity of N. caninum is scarce. N. caninum tachyzoites were isolated from the kidney and the brain of an aborted 4-month-old bovine foetus. The parasite was confirmed to be N. caninum by PCR. The tachyzoites of the new isolate, named BNC-PR4, were propagated in Vero cell cultures. Pathogenicity of the parasite was examined in BALB/c mice. Mice inoculated intraperitoneally with BNC-PR4 failed to yield clinical signs of disease and did not induce severe brain lesions, suggesting a bovine isolate with low virulence. The N. caninum-positive DNA sample was further analyzed by multilocus microsatellite (MS) genotyping for MS4, MS5, MS6A, MS6B, MS7, MS8, MS10, MS12, and MS21. Multilocus-microsatellite genotyping revealed a unique genetic profile that differed from previously reported isolates.

Detection of Neospora caninum (Toxoplasmatidae) in wild small mammals from Thailand.

The presence of Neospora caninum Dubey, Carpenter, Speer, Topper et Uggla, 1988 in small mammals (i.e. murid rodents, Erinaceomorpha, Eulipotyphla and Scadentia) was explored for first time in South-East Asia. A total of 192 individuals from six localities across Thailand were analysed. A general prevalence of N. caninum of 22% was observed, with some variation among localities (5-36%). Four main types of habitat were included and rodents trapped in dry-land habitat (17 positive among 41 individuals) were more likely to be infected with N. caninum than those from other habitats (forest, rain-fed land and settlement). Rodent species identity and individual rodent weight had no influence on individual infection. Our results provided the first data on the presence of N. caninum in rodents in South-East Asia and first report of N. caninum in the order Scadentia.