Serological responses to Neospora caninum in experimentally and naturally infected water buffaloes (Bubalus bubalis)

The water buffalo (Bubalus bubalis) is an important natural host for Neospora caninum. Serologic responses to N. caninum were studied in experimentally and naturally infected water buffaloes in Brazil. Antibodies were assayed by the indirect fluorescent antibody test (IFAT) using a cut off value of 1:25. Six buffaloes were each inoculated subcutaneously with 5 x 106 live culture-derived tachyzoites of the cattle Illinois strain of N. caninum, and two calves were kept as uninoculated controls. Post-inoculation (p.i.) blood samples were collected weekly for 8 weeks and then monthly until 1 year p.i. All inoculated buffaloes developed IFAT titers of 1:100 or more between 7 and 11 days p.i. and the titers remained elevated until 7 weeks p.i. Antibody titers peaked to 1:1600 in 1, 1:800 in 3 and 1:400 in 2, usually by 3 weeks p.i. Antibody titers declined to 1:25 or 1:50 in all the six buffaloes by 12 months p.i. IFAT titers to N. caninum remained at an undetectable level (< 1:25) in both control uninoculated buffaloes. To follow the dynamics of N. caninum antibodies, sera from 29 buffaloes and their calves were collected for 1 year and assayed for N. caninum antibodies; 23 of 29 calves were seropositive (IFAT of 1:100 or more) at 1-2 day of age. of these 23 calves, 17 remained seropositive during the study, while six became seronegative at four (two calves), six (one calf) seven (two calves) and eight (one calf) months of age. These findings suggest a high rate of neonatal transmission of N. caninum in buffaloes. Published by Elsevier B.V.

Antibodies to Neospora caninum and Toxoplasma gondii in domestic cats from Brazil

The occurrence of antibodies to Neospora caninum and Toxoplasmagondii was determined in 400 domestic cats (Siamese, Persian, and undetermined breeds) from the Municipality of Aracatuba, São Paulo, Brazil, through the indirect fluorescence antibody test (IFAT). of the 400 cats, 100 were seropositive to T. gondii (25%, titer >= 64) and 98 to N. caninum (24.5%, titer >= 16). The rate of seropositive cats for T. gondii was correlated with age (chi(2)=35.7; p < 0.001), with a higher number of infected animals at older ages. of the 219 cats younger than 1-year-old, 13.2% were seropositive for T. gondii, while 39.2% were positive in the 181 older animals. The presence of N. caninum was also correlated with age (divided by(2)=8.8; p < 0.01), with 18.7% (41/219) and 31.5% (57/181) of positive animals at ages below and above 12-month, respectively. An association between the occurrences of both protozoa in the felines was also observed (chi(2)=19.6; p < 0.001).

Evaluation of serological tests for the diagnosis of Neospora caninum infection in dogs: Optimization of cut off titers and inhibition studies of cross-reactivity with Toxoplasma gondii

Diagnosis of Neospora caninum infection in dogs is based on serological assays such as the indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assays (ELISA). This study evaluated two serological tests (IFAT and ELISA) for the detection of IgG antibodies to N. caninum in 300 serum samples of dogs through the optimization of cut off titers by using the two-graph receiveroperating characteristic (TG-ROC) curve. In addition, the identification of major cross-reactive antigens with Toxoplasma gondii was investigated by inhibition ELISA and immunoblotting (IB) assays. IFAT and ELISA results showed 74% agreement, with a good negative concordance (P-neg=0.83), but a poor positive concordance (P-pos=0.42). The great majority (86%) of sera with positive concordant results (IFAT+/ELISA+) recognized at least two out of three N. caninum immunodominant antigens, particularly the 29-32 and 35-37 kDa bands. Optimization of cut off titers in IFAT and ELISA was performed considering the reactivity to at least two out of three N. caninum immunodominant antigens as infection markers, obtaining a titer of 50 for IFAT and 200 for ELISA. Seropositivity to N. caninuin was significantly associated with T gondii-seropositive samples, particularly in ELISA (55.4%). Inhibition ELISA curves for N. caninum showed a partial heterologous inhibition, indicating some degree of cross-reactivity between N. caninum and T gondii antigens. Inhibition IB assays showed a moderate heterologous inhibition for N. caninum antigens above 45-50 kDa. These results indicate that ELISA should be used critically when crude tachyzoite antigen preparations are employed, due to possible cross-reactivity with other related parasites as T gondii. Also, the cut off dilution of 1:50 in IFAT showed to be the most appropriated for N. caninum serology in dogs. Therefore, we suggest that N. caninum immunodominant antigens, specially the 17 and 29-32 kDa proteins, should be selected markers in serological assays for canine neosporosis. (c) 2006 Elsevier B.V. All rights reserved.

Detection of immunoglobulin G antibodies to Neospora caninum in humans: High seropositivity rates in patients who are infected by human immunodeficiency virus or have neurological disorders

Considering that little is known about the epidemiology of Neospora caninum infection in humans, particularly in populations with high Toxoplasma gondii infection rates, the present study aimed to investigate the presence of antibodies to N. caninum in T. gondii-seropositive and -seronegative individuals. A total of 256 serum samples divided into four groups (61 samples from human immunodeficiency virus [HIV]-positive patients, 50 samples from patients with neurological disorders, 91 samples from newborns, and 54 samples from healthy subjects) were assessed for N. caninum and T. gondii serologies by indirect fluorescent-antibody test, enzyme-linked immunosorbent assay, and immunoblotting (IB). Immunoglobulin G antibodies to N. caninum were predominantly detected in HIV-infected patients (38%) and patients with neurological disorders (18%), while newborns and healthy subjects showed lower seropositivity rates (5% and 6%, respectively). Seropositivity to N. caninum was significantly associated with seropositivity to T. gondii in both HIV-infected patients and patients with neurological disorders. Seroreactivity to N. caninum was confirmed by IB, with positive sera predominantly recognizing the 29-kDa antigen of N. caninum. The results of this study indicate the presence of N. caninum infection or exposure in humans, particularly in HIV-infected patients or patients with neurological disorders, who could have opportunistic and concurrent infections with T. gondii. These findings may bring a new concern for the unstable clinical health of HIV-infected patients and the actual role of N. caninum infection in immunocompromised patients.

Anticorpos para Leptospira spp., Toxoplasma gondii e Neospora caninum em cães errantes albergados em canil privado

The serological profile of 300 mongrel dogs of various ages and gender were investigated. Animals were captured in the streets and afterwards directed to a private kennel in Avare city (SP) to search for leptospirosis, toxoplasmosis, and neosporosis. Blood samples were obtained from jugular or cephalic vein for the obtention of sera. The microscopic agglutination test (MAT) was used to leptospirosis. MAT detect the prevalence of 9.3%. The most frequent reactant serovars were Bratislava (35.7%), Cynopteri (17.9%), Autumnalis (14.3%), and Copenhageni (10.7%), besides 7.1% to others serovars: Icterohaemorrhagiae, Canicola, and Hardjo. The modified agglutination test used for the diagnosis of toxoplasmosis showed 26% of positive animals, with titers varying from 16 to 256, with 16 in 3.3%, 64 in 13.7%, and 256 in 9% of the samples. To canine neosporosis, it was used the indirect fluorescent antibody test, and two animals (0.7%) demonstrated antibodies with titers 25 and 100. The results show the participation of the animals in the epidemiological chain of the researched diseases.

Antibodies to Toxoplasma gondii and Neospora caninum in Captive Neotropical and Exotic Wild Canids and Felids

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação clínica e hematológica em bezerros Nelore infectados experimentalmente com isolados de Babesia bigemina das regiões Sudeste, Nordeste e Norte do Brasil

A comparative study was made regarding the clinical and hematological alterations caused by isolates of Babesia bigemina from southeastern, northeastern and northern Brazil in experimentally infected Nelore calves. Eighteen calves between 7 and 9 months of age, without antibodies against Babesia sp and raised free from ticks, were used. Three animals were previously inoculated with 2.0×109 parasitic erythrocytes (PE) for each stabilate. The other 15 calves were subdivided into three groups, with five animals each, that were subinoculated with 1.0×1010 PE of the respective isolates. The clinical and hematological alterations were evaluated by the determination of parasitaemia, haemogram, plasmatic fibrinogen, reticulocyte count, descriptive analysis of the bone marrow and erythrocytic osmotic fragility, for 30 days, totalizing seven moments of observation. The follow-up of the immunological response by the indirect fluorescent antibody test was carried out daily until the 10th day after inoculation (DAI) and after that, on the 15th, 20th, 25th and 30th DAI. A mild clinical manifestation of the disease was observed. The laboratory findings revealed low levels of parasitaemia; decrease of the erythrogram values; absence of reticulocytes, initial decrease in the total count of leukocytes, neutrophils and lymphocytes with a posterior elevation of the number of these cells; hypercellularity of the erythrocytic series and decrease of the myeloid: erythroid relation which was more accentuated between the 8th and 12th DAI, and an increase of the erythrocytic osmotic fragility in the groups inoculated with the Southeast and Northeast isolates. None of the three isolates of B. bigemina gave rise to the clinical characteristic form of the disease, although they induced an humoral immune response.

Serological profile of anti-Toxoplasma gondii antibodies in apparently healthy dogs of the city of Botucatu, São Paulo state, Brazil

Toxoplasmosis is a zoonosis caused by Toxoplasma gondii, a coccidian protozoan of worldwide distribution. The seroprevalence in canine population can be an alternative for measuring T. gondii urban spreading. A total of 780 blood samples from dogs were collected, during the yearly anti-rabies campaign, carried out by the Department of Veterinary Hygiene and Public Health, School of Veterinary Medicine and Animal Husbandry (FMVZ), São Paulo State University, UNESP, together with the county health authorities, in August 1999. Using Indirect Fluorescent Antibody Test (IFAT) for detecting antibodies anti-T. gondii in the sera samples, we observed that 258 dogs (33.1%) were positive. The associations between the serological results and the epidemiological variables were studied. Statistically significant differences were not found regarding sex (32.2% male and 34.3% female reactors). Dogs without a defined breed showed seropositivity statistically higher than the pedigreed group. The occurrence of infection was considered higher with age.

Presença de anticorpos anti-Neospora caninum e Toxoplasma gondii em cães com leishmaniose visceral da região de Araçatuba, São Paulo, Brasil

Dogs that had positive and negative sera to Leishmania chagasi from the region of Araçatuba, São Paulo, Brazil, were evaluated for the presence of anti-Neospora caninum and anti-Toxoplasma gondii antibodies as potential co-infecting agents. Blood samples were collected from 204 dogs and out of them 98 were carriers of leishmaniosis. Sera were tested for the presence of anti-L. chagasi antibodies by ELISA, and anti-T. gondii and anti-N. caninum by an indirect fluorescent antibody test (IFAT). Age, gender, and association between the presences of anti-L. chagasi antibodies and seroprevalence to N. caninum and T. gondii were analyzed by chi-square test. Out of the 204 sera investigated, 36 (17.6%) were positive for N. caninum (IFAT=50) and 75 (36.8%) to T. gondii (IFAT=16) with titers that varied from 50 to 6400 for N. caninum, and from 16 to 16384 for T. gondii. The copresence of anti-L. chagasi, N. caninum and T. gondii antibodies was observed in 17 (8.3%) dogs. Antibodies to N. caninum were observed in four (3.8%) out of 106 dogs that were negative for L. chagasi, and in 32 (32.6%) out of the 98 dogs that were positive for L. chagasi. Anti-T. gondii antibodies were found in 40 (41.0%) and in 35 (33.0%) of the 98 positive dogs and in 106 negative dogs for L. chagasi, respectively. An association between the presence of antibodies against L. chagasi and a positive response to N. caninum (p<0.001) was observed. The gender and age of the dogs did not show an association between the presence of antibodies and any of the agents studied (p>0.05), with the exception of age and presence of anti-L. chagasi antibodies, in which only a slight association was observed (p=0.038). Within this interaction, a higher number of dogs, older than four years, were positive for this agent when compared to other age groups.

Ações de vigilância continuada, papel do cão como animal sentinela para toxoplasmose.

Toxoplasmosis is a worldwide zoonosis caused by Toxoplasma gondii and its definitive host is the domestic and wild felids infecting human beings and other warmblooded animals. Dogs are considered a potential risk on the transmission due they can mechanically transmit oocysts to man. In this study, a retrospective analysis of toxoplasmic infection in dog serum samples sent to Serviço de Diagnóstico de Zoonoses/FMVZ-UNESP/Botucatu, SP, in the period of 1998 to 2007 was performed. During this period 1097 serum samples were analyzed by the indirect fluorescent antibody test (IFAT), with 299 (27.25%) positive. The most frequent titer was 16 (42.80%), followed by 64 (37.79%). The results indicate that T.gondii is distributed in the environment showing the role of the dog as sentinel animal to toxoplasmosis to monitor public health actions to the control of this zoonosis.

Soroprevalência de anticorpos contra Leishmania infantum e outras espécies de protozoários e rickettsia em cães

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Detecção de anticorpos anti-Toxoplasma gondii em cavalos carroceiros da região metropolitana de Curitiba, Paraná, Brasil

Toxoplasma gondii, the agent for toxoplasmosis, has worldwide distribution. Horses normally play a secondary role in its life cycle, but movement around urban areas, feeding on grass and the increasing use of carthorses for gathering recyclable material in some urban areas of Brazil may increase their exposure to T. gondii infection. The aim of the present study was to investigate the frequency of anti-T. gondii antibodies in carthorses in the metropolitan region of Curitiba, PR. IgG antibodies against T. gondii were detected using the indirect fluorescence antibody test (IFAT) (titers ≥ 64). Seventeen (17.0%) of the 100 horses sampled were seropositive. There were no statistical differences in relation to sex (p = 0.28) or age (p = 0.15). Our findings suggest that carthorses are exposed to T. gondii infections and that no associations with age or sex exist.

Falhas reprodutivas associadas com a presença de do vírus da artrite-encefalite caprina, toxoplasma gondii e neospora caninum em caprinos no estado de São Paulo, Brasil

This study aimed at assessing the occurrence of antibodies against the caprine arthritis-encephalitis virus (CAEV), Toxoplasma gondii and Neospora caninum, as well as the associations between the presence of antibodies and the occurrence of reproductive failures in goats. Serum samples were collected from 923 goats of both sexes, over 3 months of age, from 17 dairy farms located in different municipalities of São Paulo State, Brazil. Infections by T. gondii, N. caninum and CAEV were evaluated by indirect methods of diagnosis based on indirect fluorescence antibody test (IFAT), Neospora agglutination test (NAT), and agar gel immunodiffusion (AGID), respectively. A survey was conducted on the farms to obtain information about reproduction dates (abortions, stillbirths and births of weak and premature kids) and zoosanitary management. Antibodies against CAEV, T. gondii and N. caninum was found in 37.81%, 23.62% and 17.23% respectively. There was no significant association between the presence of anti-CAEV antibodies and CAEV/T. gondii or CAEV/N. caninum co-infection, suggesting that CAEV does not predispose goats to infection by these agents. However, when CAEV/T. gondii (p<0.01) or CAEV/N. caninum (p<0.001) co-infection was present, the occurrence of reproductive failures was significantly higher what could indicate that CAEV-induced immunosuppression may predispose goats to develop the clinical symptoms of toxoplasmosis and neosporosis increasing the risks of the reproductive failures.

Prevalência e fatores de risco associados à toxoplasmose ovina no Nordeste do Brasil

In this study, we aimed to determine the prevalence and risk factors associated to Toxoplasma gondii infection in sheep from northeastern Brazil. A total of 932 ovine serum samples from 54 properties in 19 municipalities of the state of Sergipe were collected and assayed using indirect fluorescent antibody test. The assay used antibodies against Toxoplasma gondii (IFAT-IgG) with a cutoff point of 1:64. We observed that 28.22% (263/932) of the ovine samples were serum-reactive. In a logistic regression, factors such as consumption of water directly from the source, consumption of water from a deep well, and age below 12 months were associated with protection; whereas factors such as presence of cats on the property, presence of slatted floor, and use of exchanged or borrowed breeding males were associated with infection. The studied area can be considered endemic for toxoplasmosis, so it is necessary to adopt preventive and control measures because this zoonotic infection poses risks to public health.

Epidemiology of human immunodeficiency virus-visceral leishmaniasis-co-infection

In Brazil, the rates of mother-to-child-transmission (MTCT) of human immunodeficiency virus (HIV) decreased from 20% to 1-2% in some regions. However, the country contains 90% of individuals infected with visceral leishmaniasis (VL) in Latin America, and the west region of São Paulo state faces an alarming expansion of the disease. We describe the epidemiological aspects of the expanding infection of VL and a case report of an HIV-VL-co-infected child from the west region of São Paulo state. The patient was an AIDS-C3 with low levels of CD4, high viral load, severe diarrhea, oral and perineal candidiasis, severe thrombocytopenia, and protein-caloric malnourishment. She evolved with sepsis, renal and cardiac failure. An rK rapid diagnosis test, indirect fluorescent antibody test (IFAT), and bone marrow aspirate were performed for VL. Her symptoms improved significantly after liposomal amphotericin B administration. From the 45 municipalities that compose the Regional Health Department of Presidente Prudente, Lutzomyia longipalpis vectors were found in 58% of them. VL infected dogs were found in 33% of those municipalities, infected dogs and humans were found in 29%, 20% are starting and 33% of the municipalities are preparing VL investigation. It is likely, in this patient, that VL advanced the clinical progression of the HIV disease and the development of AIDS severity. Supported by favorable conditions, the region becomes a new frontier of VL in Brazil. © 2013.