RESUMO
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.
Assuntos
Coccidiose , Doenças do Cão , Fezes , Neospora , Animais , Cães , Neospora/genética , Neospora/imunologia , Neospora/isolamento & purificação , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Coccidiose/epidemiologia , Coccidiose/veterinária , Coccidiose/parasitologia , Estudos Soroepidemiológicos , Jordânia/epidemiologia , Estudos Transversais , Feminino , Masculino , Fezes/parasitologia , Prevalência , Anticorpos Antiprotozoários/sangue , Reação em Cadeia da Polimerase/veterinária , Técnica Indireta de Fluorescência para Anticorpo/veterináriaRESUMO
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 Due to outdated and scant information on the occurrence of Neospora caninum infection in South Africa, the study aimed to determine the seroprevalence and risk factors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.
Assuntos
Anticorpos Antiprotozoários , Doenças dos Bovinos , Coccidiose , Neospora , Animais , Bovinos , Coccidiose/epidemiologia , Coccidiose/veterinária , Coccidiose/parasitologia , África do Sul/epidemiologia , Estudos Soroepidemiológicos , Neospora/imunologia , Neospora/isolamento & purificação , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia , Fatores de Risco , Estudos Transversais , Anticorpos Antiprotozoários/sangue , Feminino , Ensaio de Imunoadsorção Enzimática/veterinária , Indústria de Laticínios , Inquéritos e QuestionáriosRESUMO
Neospora caninum is a major cause of reproductive loss in cattle worldwide as it leads to abortion and animal repositioning. Although Toxoplasma gondii does not cause a reproductive problem in cattle, consuming raw or uncooked beef poses the risk of transmission. This study aimed to evaluate the occurrence of anti-N. caninum and anti-T. gondii antibodies in dairy cattle in the West and Northwest regions of São Paulo State, Brazil. A total of 653 serum samples from dairy cows were analyzed using an indirect immunofluorescence assay (IFA). Epidemiological data from the farms were associated with the serological results of the animals by logistic regression based on the presence of antibodies. The frequencies of the antibodies against N. caninum and T. gondii were 41.6% (272/653) and 11.5% (75/653), respectively. A statistically significant association was observed between: the serum anti-N. caninum antibodies and breed, history of food supplementation for calves, introduction of outside animals that later presented reproductive problems, and history of reproductive problems by the trimester of gestation. The present study highlights the importance of neosporosis in dairy cattle in the study regions and that the inclusion of this parasite in the investigation of animals with reproductive disorders is important.
Assuntos
Anticorpos Antiprotozoários , Doenças dos Bovinos , Coccidiose , Neospora , Toxoplasma , Toxoplasmose Animal , Animais , Bovinos , Neospora/imunologia , Brasil/epidemiologia , Coccidiose/veterinária , Coccidiose/epidemiologia , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/diagnóstico , Fatores de Risco , Estudos Soroepidemiológicos , Toxoplasma/imunologia , Feminino , Anticorpos Antiprotozoários/sangue , Indústria de Laticínios , Técnica Indireta de Fluorescência para Anticorpo/veterináriaRESUMO
Livestock industry is a pivotal sector for providing food, work places and monetary support for Egyptian people. Besnoitia besnoiti and Neospora caninum are protozoan parasites that are responsible for significant economic losses in ruminants, particularly in cattle. Besnoitia besnoiti can cause fertility problems and a general loss in productivity, while N. caninum is a major cause of abortion and neonatal abnormalities in infected animals. There is little information on the existence of these protozoa in Egypt, thus we conducted this study to reveal the current situation in cattle (n = 264), sheep (n = 151), and goats (n = 25). Serum samples were collected from governorates of Cairo, Giza, and Beni Suef, representing the most densely populated regions in Egypt. Using commercial ELISAs, an overall estimation among all tested animals (n = 440) revealed 7.7%, 13.2%, and 0.9% as seropositive rates for B. besnoiti, N. caninum, and mixed infection, respectively. Animal species (cattle vs sheep vs goat) and age of cattle (less than vs >1 year old) were analyzed as risk factors for infection. Regarding B. besnoiti, the seroprevalence was significantly higher in cattle than in sheep and goats and in adult cattle than calves. For N. caninum infection, no significant differences were recorded, although the seropositive rates were higher in cattle, and in adult cattle. This study provides the first seroprevalence data for B. besnoiti in all surveyed animals in the regions included, and in sheep and goats from Egypt, and supports the current knowledge for the occurrence of N. caninum in Egypt.
Assuntos
Anticorpos Antiprotozoários , Doenças dos Bovinos , Coccidiose , Doenças das Cabras , Cabras , Neospora , Sarcocystidae , Doenças dos Ovinos , Animais , Egito/epidemiologia , Neospora/imunologia , Coccidiose/veterinária , Coccidiose/epidemiologia , Coccidiose/parasitologia , Bovinos , Doenças dos Ovinos/epidemiologia , Doenças dos Ovinos/parasitologia , Ovinos , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia , Doenças das Cabras/epidemiologia , Doenças das Cabras/parasitologia , Anticorpos Antiprotozoários/sangue , Estudos Soroepidemiológicos , Sarcocystidae/imunologia , Sarcocystidae/isolamento & purificação , Cabras/parasitologia , Feminino , Ensaio de Imunoadsorção Enzimática/veterináriaRESUMO
As for many other organisms, CRISPR-Cas9 mediated genetic modification has gained increasing importance for the identification of vaccine candidates and drug targets in Neospora caninum, an apicomplexan parasite causing abortion in cattle and neuromuscular disease in dogs. A widely used approach for generating knock-out (KO) strains devoid of virulence factors is the integration of a drug selectable marker such as mutated dihydrofolate reductase-thymidylate synthase (mdhfr-ts) into the target gene, thus preventing the synthesis of respective protein and mediating resistance to pyrimethamine. However, CRISPR-Cas9 mutagenesis is not free of off-target effects, which can lead to integration of multiple mdhfr-ts copies into other sites of the genome. To determine the number of integrated mdhfr-ts in N. caninum, a duplex quantitative TaqMan PCR was developed. For this purpose, primers were designed that amplifies a 106 bp fragment from wild-type (WT) parasites corresponding to the single copy wtdhfrs-ts gene, as well as the mutated mdhfrs-ts present in KO parasites that confers resistance and were used simultaneously with primers amplifying the diagnostic NC5 gene. Thus, the dhfr-ts to NC5 ratio should be approximately 1 in WT parasites, while in KO parasites with a single integrated mdhrf-ts gene this ratio is doubled, and in case of multiple integration events even higher. This approach was applied to the Neospora KO strains NcΔGRA7 and NcΔROP40. For NcΔGRA7, the number of tachyzoites determined by dhfr-ts quantification was twice the number of tachyzoites determined by NC5 quantification, thus indicating that only one mdhfr-ts copy was integrated. The results obtained with the NcΔROP40 strain, however, showed that the number of dhfr-ts copies per genome was substantially higher, indicating that at least three copies of the selectable mdhfr-ts marker were integrated into the genomic DNA during gene editing by CRISPR-Cas9. This duplex TaqMan-qPCR provides a reliable and easy-to-use tool for assessing CRISPR-Cas9 mediated mutagenesis in WT N. caninum strains.
Assuntos
Sistemas CRISPR-Cas , Técnicas de Inativação de Genes , Neospora , Tetra-Hidrofolato Desidrogenase , Timidilato Sintase , Tetra-Hidrofolato Desidrogenase/genética , Neospora/genética , Timidilato Sintase/genética , Animais , Reação em Cadeia da Polimerase em Tempo Real/métodos , Resistência a Medicamentos/genética , Edição de Genes/métodos , Coccidiose/parasitologia , Complexos MultienzimáticosRESUMO
Neospora spp. and Toxoplasma gondii are two closely related protozoan parasites that are widely distributed throughout the world. Horses can act as intermediate hosts for both parasites and can acquire disease. Blood samples were taken from 487 clinically healthy horses from 17 different mechanized stables in Tehran, the capital of Iran, during September and November of 2022. IFAT and ELISA were employed to detect antibodies directed against Neospora spp. and T. gondii. The anti-N. caninum antibodies were detected in 52 of the horses (10.67%) based on IFAT and in 86 of the 487 horses (17.65%) based on the ELISA test. Also, antibodies against T. gondii were detected in 41 horses (8.42%) based on IFAT and in 63 of 487 horses (12.94%) based on the ELISA test. Also, in 6 of the horses (1.23%) based on IFAT and in 13 of the 487 horses (2.67%) based on the ELISA test, double positivity suggested co-infection with both parasites. Gender, age groups, and the presence of dogs for neosporosis, and age groups and the presence of cats for toxoplasmosis, could be considered factors having an influence on the seroprevalences (P < 0.05). The results proved the importance of the urgent implementation of stringent regulatory measures to prevent and control the spread of these parasites.
Assuntos
Anticorpos Antiprotozoários , Coccidiose , Doenças dos Cavalos , Neospora , Toxoplasma , Toxoplasmose Animal , Animais , Cavalos/parasitologia , Toxoplasma/imunologia , Neospora/imunologia , Irã (Geográfico)/epidemiologia , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/parasitologia , Toxoplasmose Animal/sangue , Toxoplasmose Animal/imunologia , Estudos Soroepidemiológicos , Doenças dos Cavalos/epidemiologia , Doenças dos Cavalos/parasitologia , Doenças dos Cavalos/sangue , Coccidiose/epidemiologia , Coccidiose/veterinária , Coccidiose/sangue , Masculino , Feminino , Anticorpos Antiprotozoários/sangue , Ensaio de Imunoadsorção Enzimática/veterináriaRESUMO
This study documents the presence of anti-Neospora caninum antibodies and their association with certain risk factors in 2 deer species from the central region of Veracruz State, Mexico. A total of 90 blood samples, 20 from temazate deer (Mazama temama) and 70 from white-tailed deer (Odocoileus virginianus), were taken from 3 farms, and serum samples were subjected to ELISA indirect test to detect N. caninum antibodies; the association between the serological status and the possible risk factors was then estimated. The overall presence of anti-N. caninum antibodies was 57.7% (52/90; 95% CI 46.9-67.9), with positive animals identified on all farms; in white-tailed deer it was 57% and in temazate deer 60%. Prevalence was higher in females than males. Adult animals had a higher prevalence than young ones. The risk analysis identified the age in the adult animal category (odds ratio 5.8) as being associated with the presence of anti-N. caninum antibodies. These results provide evidence of the significant contamination of oocysts in the environment and allow us to estimate the contribution of deer to the sylvatic cycle.
Assuntos
Anticorpos Antiprotozoários , Coccidiose , Cervos , Ensaio de Imunoadsorção Enzimática , Neospora , Animais , Anticorpos Antiprotozoários/sangue , Coccidiose/veterinária , Coccidiose/epidemiologia , Coccidiose/parasitologia , Cervos/parasitologia , México/epidemiologia , Feminino , Masculino , Neospora/imunologia , Ensaio de Imunoadsorção Enzimática/veterinária , Estudos Soroepidemiológicos , Fatores de Risco , Fatores Etários , Fatores SexuaisRESUMO
This study evaluated the reproductive, productive and financial consequences of chronic Trypanosoma vivax infection in a dairy cattle herd located in a region without the cyclic vector during two years. Animals were categorized as either positive (chronically infected) or negative for T. vivax antibodies using a commercial rapid test. Additionally, serum samples from cows were analyzed for the presence of anti-Neospora caninum antibodies. Pregnancy diagnoses were performed through rectal palpation and ultrasonography after 30, 60 and every 21 days until the 144th day of pregnancy. If an abortion occurred in the final trimester, serology and cPCR were performed on calves for T. vivax and N. caninum. The breeding period, calving interval and pregnancy losses were recorded. The milk production of each animal during the 305 days of lactation was measured, and the annual financial impact of milk production was calculated using a revenue minus feed cost (RMFC) indicator. Out of 177 cows, 71.75â¯% were chronically infected, and 13.50â¯% were T. vivax-negative. No correlation (p = 0.8854) of co-infection between T. vivax and N. caninum was observed. Negative cows required fewer (p≤0.05) artificial inseminations than chronically infected ones. T. vivax was not significantly associated (p = 0.7893) with pregnancy loss up to 81 days of pregnancy. Cows chronically infected by T. vivax had 4-fold greater chance (p = 0.0280) of experiencing pregnancy loss between 82 and 144 days of gestation. Eighteen cows aborted, two were positive for T. vivax antibodies, and one for N. caninum antibodies. The calves were negative for T. vivax and N. caninum. Chronically infected cows and negative cows for T. vivax that experienced pregnancy loss (82-144 days of pregnancy) had a longer (p≤0.05) breeding period to become pregnant, and consequently a longer calving interval compared to cows that maintained pregnancy. The difference (p≤0.05) in milk production was evident when pregnancy loss occurred between 82 and 144 days of gestation in cows chronically infected by T. vivax. The RMFC indicated a negative impact of 38.2â¯% on the farm's annual milk revenue due to the presence of chronically infected cows.
Assuntos
Doenças dos Bovinos , Indústria de Laticínios , Reprodução , Trypanosoma vivax , Animais , Bovinos , Feminino , Gravidez , Doenças dos Bovinos/parasitologia , Doenças dos Bovinos/economia , Indústria de Laticínios/economia , Doença Crônica/veterinária , Tripanossomíase Africana/veterinária , Tripanossomíase Africana/parasitologia , Tripanossomíase Africana/epidemiologia , Anticorpos Antiprotozoários/sangue , Coccidiose/veterinária , Coccidiose/parasitologia , Coccidiose/economia , Aborto Animal/parasitologia , Lactação , Leite , Neospora/imunologiaRESUMO
Toxoplasma gondii and Neospora caninum infections may be associated with neuromuscular disorders in dogs. The aim of this study was to assess the seroprevalence to these protozoan parasites in dogs with neuromuscular disease from urban areas of Buenos Aires province, Argentina, over a period of 20 years, and to evaluate the association of seropositivity and antibody titres with different variables such as sex, breed and age. For this, a total of 7238 serum samples from urban owned dogs were analysed by the indirect fluorescence antibody test (IFAT) for specific IgG antibodies. The observed seropositivity rates were 35.7â¯% for T. gondii and 25.7â¯% for N. caninum. Crossbred dogs had a significantly higher seroprevalence for T. gondii than purebred dogs (41â¯% vs. 29.3â¯%), while a trend towards significance was observed for N. caninum, which was slightly higher in purebred dogs (26â¯% vs. 23.6â¯%). Seroprevalence for both parasites increased with age and was higher in older animals. Regarding the distribution of specific antibody titres, the most frequent IFAT T. gondii titre found was 100 and for N. caninum it was ≥800. For toxoplasmosis, there was no association with age group, and low titres (50, 100 and 200) predominated in all groups. However, for neosporosis, age and titres were significantly associated for one age group, with dogs under 12 months of age having a higher proportion of high titres (400 and 800). The trend in the seroprevalence for T. gondii is increasing over the years and lower antibody titres predominate in the dogs studied, which may be more related to the presence of chronic infections and not necessarily to the clinical signs of the animals. Despite the generally low titres observed for toxoplasmosis in this study, it is important to highlight the high seroprevalence found in our region, as dogs can act as sentinels of environmental contamination and as indicators of possible human infection. In the case of neosporosis, although the trend in seroprevalence in dogs with signs appears to be decreasing over the years, our work shows that higher antibody titres predominate, and are probably related to the clinical signs presented by the dogs. This study provides the most recent epidemiological data and serological profiles of T. gondii and N. caninum infections in a large number of canine sera from urban areas in Argentina, providing relevant information for clinical veterinarians and epidemiologists in order to understand the circulation of the parasites.
Assuntos
Anticorpos Antiprotozoários , Coccidiose , Doenças do Cão , Neospora , Toxoplasma , Toxoplasmose Animal , Animais , Cães , Neospora/imunologia , Argentina/epidemiologia , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Coccidiose/veterinária , Coccidiose/epidemiologia , Coccidiose/parasitologia , Toxoplasma/imunologia , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/parasitologia , Estudos Soroepidemiológicos , Feminino , Masculino , Anticorpos Antiprotozoários/sangueRESUMO
Toxoplasma gondii and Neospora caninum are major worldwide morbidity-causing pathogens. Bumped kinase inhibitors (BKIs) are a compound class that has been optimized to target the apicomplexan calcium-dependent protein kinase 1 (CDPK1) - and several members of this class have proven to be safe and highly active in vitro and in vivo. BKI-1708 is based on a 5-aminopyrazole-4-carboxamide scaffold, and exhibited in vitro IC50 values of 120 nM for T. gondii and 480 nM for N. caninum ß-galactosidase expressing strains, and did not affect human foreskin fibroblast (HFF) viability at concentrations up to 25 µM. Electron microscopy established that exposure of tachyzoite-infected fibroblasts to 2.5 µM BKI-1708 in vitro induced the formation of multinucleated schizont-like complexes (MNCs), characterized by continued nuclear division and harboring newly formed intracellular zoites that lack the outer plasma membrane. These zoites were unable to finalize cytokinesis to form infective tachyzoites. BKI-1708 did not affect zebrafish (Danio rerio) embryo development during the first 96 h following egg hatching at concentrations up to 2 µM. Treatments of mice with BKI-1708 at 20 mg/kg/day during five consecutive days resulted in drug plasma levels ranging from 0.14 to 4.95 µM. In vivo efficacy of BKI-1708 was evaluated by oral application of 20 mg/kg/day from day 9-13 of pregnancy in mice experimentally infected with N. caninum (NcSpain-7) tachyzoites or T. gondii (TgShSp1) oocysts. This resulted in significantly decreased cerebral parasite loads and reduced vertical transmission in both models without drug-induced pregnancy interference.
Assuntos
Coccidiose , Fibroblastos , Neospora , Pirazóis , Toxoplasma , Animais , Neospora/efeitos dos fármacos , Toxoplasma/efeitos dos fármacos , Camundongos , Coccidiose/tratamento farmacológico , Coccidiose/parasitologia , Pirazóis/farmacologia , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/parasitologia , Humanos , Antiprotozoários/farmacologia , Concentração Inibidora 50 , Toxoplasmose/tratamento farmacológico , Toxoplasmose/parasitologia , Modelos Animais de Doenças , Inibidores de Proteínas Quinases/farmacologia , Proteínas QuinasesRESUMO
Neosporosis is the major infectious cause of abortion and reproductive losses in cattle worldwide; however, there are no available vaccines or drugs to control this disease. Recently, a dual (positive and negative) DIVA-like (Differentiation of Infected from Vaccinated Animals) vaccine was evaluated in a pregnant mouse model of neosporosis, showing promising immunogenic and protective results. The current report aimed to study the safety, the dose-dependent immunogenicity and the dual DIVA-like character of a recombinant subunit vaccine composed of the major surface antigen from Neospora caninum (rNcSAG1) and the carrier/adjuvant Heat shock protein 81.2 from Arabidopsis thaliana (rAtHsp81.2) in cattle. Healthy heifers were separated and assigned to experimental groups A-F and subcutaneously immunized with 2 doses of vaccine formulations 30 days apart as follows: A (n = 4): 50 µg rNcSAG1 + 150 µg rAtHsp81.2; B (n = 4): 200 µg rNcSAG1 + 600 µg rAtHsp81.2; C (n = 4): 500 µg rNcSAG1 + 1,500 µg rAtHsp81.2; D (n = 3): 150 µg rAtHsp81.2; E (n = 3):1,500 µg rAtHsp81.2, and F (n = 3) 2 ml of sterile PBS. The immunization of heifers with the different vaccine or adjuvant doses (groups A-E) was demonstrated to be safe and did not modify the mean value of the evaluated serum biomarkers of metabolic function (GOT/ASP, GPT/ALT, UREA, Glucose and total proteins). The kinetics and magnitude of the immune responses were dose-dependent. The higher dose of the vaccine formulation (group C) stimulated a broad and potent humoral and cellular immune response, characterized by an IgG1/IgG2 isotype profile and IFN-γ secretion. In addition, this was the first time that dual DIVA-like character of a vaccine against neosporosis was demonstrated, allowing us to differentiate vaccinated from infected heifers by two different DIVA compliant test approaches. These results encourage us to evaluate its protective efficacy in infected pregnant cattle in the future.
Assuntos
Doenças dos Bovinos , Coccidiose , Neospora , Vacinas Protozoárias , Vacinas Sintéticas , Animais , Bovinos , Coccidiose/prevenção & controle , Coccidiose/veterinária , Coccidiose/imunologia , Doenças dos Bovinos/prevenção & controle , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/parasitologia , Neospora/imunologia , Feminino , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Protozoárias/imunologia , Vacinas Protozoárias/administração & dosagem , Anticorpos Antiprotozoários/sangue , Antígenos de Protozoários/imunologia , Proteínas de Protozoários/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Imunogenicidade da Vacina , GravidezRESUMO
Neospora caninum is an apicomplexan and obligatory intracellular parasite, which is the leading cause of reproductive failure in cattle and affects other farm and domestic animals, but also induces neuromuscular disease in dogs of all ages. In cattle, neosporosis is an important health problem, and has a considerable economic impact. To date there is no protective vaccine or chemotherapeutic treatment on the market. Immuno-prophylaxis has long been considered as the best control measure. Proteins involved in host cell interaction and invasion, as well as antigens mediating inflammatory responses have been the most frequently assessed vaccine targets. However, despite considerable efforts no effective vaccine has been introduced to the market to date. The development of effective compounds to limit the effects of vertical transmission of N. caninum tachyzoites has emerged as an alternative or addition to vaccination, provided suitable targets and safe and efficacious drugs can be identified. Additionally, the combination of both treatment strategies might be interesting to further increase protectivity against N. caninum infections and to decrease the duration of treatment and the risk of potential drug resistance. Well-established and standardized animal infection models are key factors for the evaluation of promising vaccine and compound candidates. The vast majority of experimental animal experiments concerning neosporosis have been performed in mice, although in recent years the numbers of experimental studies in cattle and sheep have increased. In this review, we discuss the recent findings concerning the progress in drug and vaccine development against N. caninum infections in mice and ruminants.
Assuntos
Coccidiose , Neospora , Vacinas Protozoárias , Animais , Coccidiose/prevenção & controle , Coccidiose/veterinária , Coccidiose/parasitologia , Coccidiose/tratamento farmacológico , Coccidiose/imunologia , Neospora/imunologia , Vacinas Protozoárias/imunologia , Bovinos , Doenças dos Bovinos/prevenção & controle , Doenças dos Bovinos/parasitologia , Desenvolvimento de VacinasRESUMO
Neosporosis is a worldwide parasitic disease caused by the protozoan Neospora caninum. It can cause economic losses to farmers due to its relationship with abortions and neonatal mortality in cows. Dogs play a key role in its spread as they are considered definitive hosts. In this study, we determined for the first time the seropositivity of N. caninum infection in dogs from Ecuador and evaluated potential risk factors. A total number of 339 free-roaming dogs from the three main regions of Ecuador (Coastal, Andean, and Amazonian regions) were included in the study and classified either as urban or rural dogs. Serum samples were collected from November 2018 to May 2019, and analyzed with a commercial ELISA test. An overall seropositivity of 6.8 % (CI: 95 %, 2.8 % - 11.7 %) was found in N. caninum infection with no statistical differences among regions or urban/rural dogs. This is the first surveillance of N. caninum in Ecuador, confirming a country-wide distribution of this pathogen. Considering the large populations of free-roaming dogs in Ecuador, a One Health approach for surveillance and managing N. caninum in dogs is needed to protect either livestock or wildlife.
Assuntos
Anticorpos Antiprotozoários , Coccidiose , Doenças do Cão , Neospora , Animais , Cães , Equador/epidemiologia , Neospora/imunologia , Coccidiose/veterinária , Coccidiose/epidemiologia , Coccidiose/parasitologia , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Fatores de Risco , Estudos Soroepidemiológicos , Anticorpos Antiprotozoários/sangue , Feminino , Masculino , Ensaio de Imunoadsorção EnzimáticaRESUMO
Neospora caninum is an obligate intracellular parasite that infects a wide range of mammalian species, and particularly causes abortions in cattle and nervous system dysfunction in dogs. Dense granule proteins (GRAs) are thought to play an important role in the mediation of host-parasite interactions and facilitating parasitism. However, a large number of potential GRAs remain uncharacterized, and the functions of most of the identified GRAs have not been elucidated. Previously, we screened a large number GRAs including NcGRA27 and NcGRA61 using the proximity-dependent biotin identification (BioID) technique. Here, we identified a novel GRA protein NcGRA85 and used C-terminal endogenous gene tagging to determine its localization at the parasitophorous vacuole (PV) in the tachyzoite. We successfully disrupted three gra genes (NcGRA27, NcGRA61 and NcGRA85) of N. caninum NC1 strain using CRISPR-Cas9-mediated homologous recombination and phenotyped the single knockout strain. The NcGRA61 and NcGRA85 genes were not essential for parasite replication and growth in vitro and for virulence during infection of mice, as observed by replication assays, plaque assays and in vitro virulence assays in mice. Deletion of the NcGRA27 gene in the NC1 strain reduced the in vitro replication and growth of the parasite, as well as the pathogenicity of the NC1 strain in mice. In summary, our findings provide a basis for in-depth studies of N. caninum pathogenesis and demonstrate the importance of NcGRA27 in parasite growth and virulence, most likely a new virulence factor of N. caninum.
Assuntos
Sistemas CRISPR-Cas , Coccidiose , Neospora , Proteínas de Protozoários , Animais , Neospora/genética , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Camundongos , Coccidiose/parasitologia , Coccidiose/veterinária , Feminino , Camundongos Endogâmicos BALB C , Virulência/genética , Técnicas de Inativação de Genes , CãesRESUMO
The family Sarcocystidae includes several intracellular coccidial parasites such as Toxoplasma gondii, Neospora caninum, Sarcocystis spp. and Hammondia spp. with heteroxenous life cycles involving different parasitic stages (oocysts/sporocysts, tachyzoites and bradyzoites in tissue cysts). The aim of this work was to evaluate monoclonal antibodies (MAb) (anti NcSAG1, anti NcSAG4 and anti TgCC2) and/or polyclonal antibodies (PAb) (anti NcSAG4 and anti TgBAG1) to label specific immunodominant antigens in different parasitic stages of N. caninum (oocyst, bradyzoite and tachyzoite), T. gondii (oocyst, cyst and tachyzoite), H. heydorni (oocyst), S. cruzi (cyst and bradyzoite) and S. falcatula (sporocyst). It was observed that the MAb directed against NcSAG1 reacted exclusively with N. caninum tachyzoites. In contrast, the MAb directed against NcSAG4 did not react with any of the parasites tested at any stage. The MAb directed against NcSAG4 reacted with both N. caninum and T. gondii tachyzoites, T. gondii tissue cysts and S. cruzi tissue cysts and bradyzoites. As expected, the MAb directed against the T. gondii tissue cyst wall antigen TgCC2 reacted with T. gondii tissue cysts, N. caninum bradyzoites, but also with T. gondii and H. heydorni oocysts and S. falcatula sporocysts. Finally, the PAb directed against the T. gondii bradyzoite proteinTgBAG1 reacted with T. gondii tissue cysts, N. caninum bradyzoites, and also with S. cruzi tissue cysts and bradyzoites. These data reveal a wide range of cross-reactions between different species of protozoa and between different developmental stages, which should be taken into account in the design and evaluation of diagnostic tests, as well as in the assessment of vaccination and challenge studies.
Assuntos
Anticorpos Monoclonais , Antígenos de Protozoários , Neospora , Sarcocystis , Toxoplasma , Sarcocystis/imunologia , Neospora/imunologia , Animais , Antígenos de Protozoários/imunologia , Toxoplasma/imunologia , Anticorpos Monoclonais/imunologia , Camundongos , Sarcocystidae/imunologia , Sarcocystidae/crescimento & desenvolvimento , Epitopos Imunodominantes/imunologia , Ensaio de Imunoadsorção Enzimática/veterinária , Anticorpos Antiprotozoários/sangue , Anticorpos Antiprotozoários/imunologia , Camundongos Endogâmicos BALB C , CoelhosRESUMO
OBJECTIVES: Studies in humans have demonstrated the role of Toxoplasma gondii, a protozoan parasite, in epileptic seizures. This study aimed to investigate the serological correlation between T. gondii and N. caninum and epilepsy in dogs. MATERIALS AND METHODS: The medical record database of the Veterinary Teaching Hospital, University of Perugia, was searched for dogs serologically tested by IFAT for T. gondii and N. caninum and following specific inclusion criteria. Dogs were stratified by having a clinical diagnosis of epilepsy or suffering different conditions. RESULTS: One-hundred and twenty-eight dogs were included, 64 with epilepsy and 64 without clinical signs of epilepsy. Seventeen of the 64 epileptic dogs (26.6%; 95% CI: 15.7% to 37.4%) and twenty-one of the 64 non-epileptic dogs (32.8%; 95% CI: 21.3% to 44.3%) tested positive for T. gondii. Eight of the epileptic dogs (12.5%; 95% CI: 4.4% to 20.6%) and three of the non-epileptic dogs (4.7%; 95% CI: 0% to 9.9%) tested positive for N. caninum. There was no statistically significant difference in the rate of T. gondii or N. caninum seroreactivity between epileptic and non-epileptic dogs. CLINICAL SIGNIFICANCE: The results obtained do not seem to support the role of T. gondii and N. caninum as causative agents of dog epilepsy.
Assuntos
Coccidiose , Doenças do Cão , Epilepsia , Neospora , Toxoplasma , Toxoplasmose Animal , Cães , Animais , Doenças do Cão/parasitologia , Toxoplasmose Animal/complicações , Toxoplasmose Animal/epidemiologia , Toxoplasma/imunologia , Epilepsia/veterinária , Epilepsia/parasitologia , Coccidiose/veterinária , Coccidiose/complicações , Masculino , Feminino , Anticorpos Antiprotozoários/sangue , Estudos RetrospectivosRESUMO
In bovines few studies addressed the contribution of adipose tissue to the host immune response to infection. Here we evaluated the in vitro response of bovine adipose tissue stromal vascular fraction (SVF) cells to the protozoan parasite Neospora caninum, using live and freeze-killed tachyzoites. Live N. caninum induced the production of IL-6, IL-1ß and IL-10 by SVF cells isolated from subcutaneous adipose tissue (SAT), while in mesenteric adipose tissue (MAT) SVF cell cultures only IL-1ß and IL-10 production was increased, showing slight distinct responses between adipose tissue depots. Whereas a clear IL-8 increase was detected in peripheral blood leucocytes (PBL) culture supernatants in response to live N. caninum, no such increase was observed in SAT or MAT SVF cell cultures. Nevertheless, in response to LPS, increased IL-8 levels were detected in all cell cultures. IL-10 levels were always increased in response to stimulation (live, freeze-killed N. caninum and LPS). Overall, our results show that bovine adipose tissue SVF cells produce cytokines in response to N. caninum and can therefore be putative contributors to the host immune response against this parasite.
Assuntos
Coccidiose , Neospora , Animais , Bovinos , Interleucina-10 , Interleucina-8 , Lipopolissacarídeos/farmacologia , Fração Vascular Estromal , Citocinas , Tecido Adiposo , Coccidiose/parasitologiaRESUMO
BACKGROUND: Toxoplasma gondii and Neospora caninum are closely related protozoan parasites that are considered important causes of abortion in livestock, causing huge economic losses. Hunan Province ranks 12th in the production of beef and mutton in China. However, limited data are available on the seroprevalence, risk factors and molecular characterization of T. gondii and N. caninum in beef cattle and goats in Hunan province, China. METHODS: Sera of 985 beef cattle and 1147 goats were examined for the presence of specific antibodies against T. gondii using indirect hemagglutination test (IHAT) and anti-N. caninum IgG using competitive-inhibition enzyme-linked immunoassay assay (cELISA). Statistical analysis of possible risk factors was performed using PASW Statistics. Muscle samples of 160 beef cattle and 160 goats were examined for the presence of T. gondii DNA (B1 gene) and N. caninum DNA (Nc-5 gene) by nested PCR. The B1 gene-positive samples were genotyped at 10 genetic markers using the multilocus nested PCR-RFLP (Mn-PCR-RFLP). RESULTS: Specific IgG against T. gondii were detected in 8.3% (82/985) and 13.3% (153/1147) and against N. caninum in 2.1% (21/985) and 2.0% (23/1147) of the beef cattle and goats, respectively. Based on statistical analysis, the presence of cats, semi-intensive management mode and gender were identified as significant risk factors for T. gondii infection in beef cattle. Age was a significant risk factor for T. gondii infection in goats (P < 0.05), and age > 3 years was a significant risk factor for N. caninum infection in beef cattle (P < 0.05). PCR positivity for T. gondii was observed in three beef samples (1.9%; 3/160) and seven chevon samples (4.4%; 7/160). Genotyping of PCR positive samples identified one to be ToxoDB#10. The N. caninum DNA was observed in one beef sample (0.6%; 1/160) but was negative in all chevon samples. CONCLUSIONS: To our knowledge, this is the first large-scale serological and molecular investigation of T. gondii and N. caninum and assessment of related risk factors in beef cattle and goats in Hunan Province, China. The findings provide baseline data for executing prevention and control of these two important parasites in beef cattle and goats in China.
Assuntos
Anticorpos Antiprotozoários , Doenças dos Bovinos , Coccidiose , Doenças das Cabras , Cabras , Neospora , Toxoplasma , Toxoplasmose Animal , Animais , Cabras/parasitologia , Neospora/genética , Neospora/imunologia , Neospora/isolamento & purificação , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasma/isolamento & purificação , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/parasitologia , China/epidemiologia , Bovinos , Estudos Soroepidemiológicos , Coccidiose/veterinária , Coccidiose/epidemiologia , Coccidiose/parasitologia , Doenças das Cabras/epidemiologia , Doenças das Cabras/parasitologia , Anticorpos Antiprotozoários/sangue , Feminino , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia , Masculino , Fatores de Risco , Imunoglobulina G/sangue , DNA de Protozoário/genética , Ensaio de Imunoadsorção Enzimática/veterinária , Genótipo , Reação em Cadeia da Polimerase/veterináriaRESUMO
Abortion caused by the parasite Neospora caninum is an important threat to the livestock industry worldwide. Trophoblasts and caruncular cells play major roles in initiating innate immune responses and controlling parasite infection at the fetal-maternal interface. In the present study, bovine uterine epithelial cells (BUECs) and bovine trophoblastic (BT) cells treated with bovine interferon-gamma (IFN-γ), IFN-alpha (IFN-α) and IFN-tau (IFN-τ) followed by infection with N. caninum were examined by measuring the mRNA expression levels of numerous pregnancy-associated proteins and observing parasite growth to elucidate the host-parasite interaction at the uteroplacental region. N. caninum infection increased the expression of prolactin-related protein 1 (PRP1), pregnancy-associated glycoprotein 1 (PAG1), and cytokines (TNF-α, IL-8 and IL-10) in BUECs and of IL-8 in BT cells. Bovine IFN-γ inhibited IL-8 and TNF-α expression in BUECs and IL-8 in BT cells. In contrast, the expression of the interferon-stimulated gene OAS1 was significantly increased by treatment of the infected BT cells with IFN-γ. However, treatment with bovine IFNs did not inhibit N. caninum growth in either cell line. In conclusion, our results suggest that bovine IFN-γ plays a crucial role in control of pathogenesis in uterus and induction of inflammatory response in the placental region following N. caninum infection, rather than growth inhibition of the parasites.
Assuntos
Coccidiose , Citocinas , Endométrio , Células Epiteliais , Neospora , Proteínas da Gravidez , Trofoblastos , Animais , Bovinos , Neospora/fisiologia , Trofoblastos/parasitologia , Trofoblastos/metabolismo , Feminino , Citocinas/metabolismo , Citocinas/genética , Células Epiteliais/parasitologia , Endométrio/parasitologia , Endométrio/metabolismo , Endométrio/citologia , Coccidiose/parasitologia , Coccidiose/veterinária , Proteínas da Gravidez/genética , Proteínas da Gravidez/farmacologia , Gravidez , Doenças dos Bovinos/parasitologia , Regulação da Expressão Gênica , Interações Hospedeiro-ParasitaRESUMO
Bovine Trypanosomiasis and other infectious diseases cause relevant loss for the livestock industry impacting productive/reproductive indices. This study intended to better understand the frequency, seasonality, and profile of infections associated with Bovine Trypanosomiasis. A total of 1443 serum samples were screened for T. vivax infection and other infectious diseases: Neosporosis, Leptospirosis, Bovine Leukosis Virus infection/(BLV), Infectious Bovine Rhinotracheitis/(IBR) or Bovine Viral Diarrhea/(BVD). Distinct methods were used for screening and diagnosis: immunofluorescence assay (Trypanosomiasis), ELISA (Neosporosis,BLV,IBR,BVD) and microscopic agglutination test (Leptospirosis). Our findings demonstrated that the seropositivity for Trypanosomiasis=57% was similar to Neosporosis=55%, higher than Leptospirosis=39% and BVL=34%, but lower than IBR=88% and BVD=71%. The seropositivity for Trypanosomiasis was higher in the autumn and lower in the winter. Regardless the season, the IBR seropositivity (min=73%;max=95%) was higher than Trypanosomiasis (min=48%;max=68%). Moreover, Neosporosis (min=71%;max=100%) and BVD (min=65%;max=76%) were more frequent than Trypanosomiasis in the summer, winter and spring. The diagnosis outcome revealed that Trypanosomiasis&IBR=43% and Trypanosomiasis&Neosporosis=35% were the most frequent co-infections with higher seropositivity in the autumn (58%) and summer (80%), respectively. Noteworthy, high seropositivity to Trypanosomiasis&BVD was registered in the autumn (46%). Together, our data re-enforce the relevance of differential diagnosis between Trypanosomiasis with other bovine infectious diseases and that differences in the seasonality profile is a relevant aspect to be considered while selecting the differential diagnosis to be applied.