Neospora caninum serostatus and milk production of Holstein cattle

OBJECTIVE: To determine whether Neospora caninum serostatus was associated with milk production among Holstein cattle in Ontario. DESIGN: Case-control study and cross-sectional observational study. ANIMALS: 3,702 Holstein cows in 83 herds (case-control study) and 3,162 Holstein cows in 57 herds. PROCEDURE: Herds in the case-control study were grouped on the basis of N. caninum abortion status. Herds in the observational study were considered representative of Ontario dairy herds. The N. caninum serostatus of individual cows was determined with a kinetic ELISA. Milk production was modeled to compare seropositive with seronegative animals while controlling for parity, days since parturition, and herd clustering. RESULTS: In the case-control study, 305-day milk production of seropositive cows was significantly less than milk production of seronegative cows in herds with abortions attributable to N. caninum infection and in herds with abortions attributable to pathogens other than N. caninum, but not in herds without abortion problems. In the observational study, 305-day milk production for seropositive cows was not significantly different from milk production of seronegative cows. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that the association between N. caninum serostatus and milk production in Ontario Holstein dairy cattle may depend on abortion status of the herd. In herds with abortion problems, regardless of cause, N. caninum-seropositive cattle produced less milk, whereas in herds without abortion problems, N. caninum-seropositive cattle produced the same amount of milk as seronegative cattle.     

Neosporosis in bovine dairy herds from the west of France: detection of Neospora caninum DNA in aborted fetuses, seroepidemiology of N. caninum in cattle and dogs.

Neospora caninum is considered one of the major causes of abortion in cattle in most parts of the world. In this study, the role of N. caninum was investigated in groups of aborted cattle and dairy herds from the west of France. Good correlation was found between parasite DNA detection in fetuses and serologic statuses of dams. In groups with documented abortion status and no antibodies to other pathogens, 17-45% of aborted animals were seropositive for N. caninum, and significant relationship between prevalence of Neospora antibodies and frequency of abortions was found. Neospora-associated abortions were observed all the year round, with a peak in summer. Higher ratios of seropositive abortions were found before the 6th month of gestation. In 12 herds studied in the field, serologic prevalence ranged 6-47%. No difference in age was found between seropositive and seronegative cows. Results indicate that N. caninum is an important and stable cause of abortion in cattle in France.     

The importance of bovine neosporosis for abortion in Switzerland

The protozoan parasite Neospora caninum is one of the most important infectious abortion causes in cattle worldwide. In a Swiss case-control-study we investigated 113 abortion problem farms and 113 control farms. It was possible to detect N. caninum in 21% of 242 investigated aborted fetuses by PCR upon brain samples. Eighty-four % of mothers aborting N. caninum positive fetuses were serologically positive for this parasite. The seroprevalence for N. caninum of all aborting mothers was 44%. Within a period of 3 to 12 months 4,505 cattle from the participating farms were investigated serologically at two different time points. Strong fluctuations of the specific antibody concentration could be observed, resulting in a conversion to seronegativity in the second blood sample in 39% of formerly seropositive animals. Eighteen months after the end of the study, 42 case- and 42 controlfarms were questioned about their present abortion-status. Analysis of the questionnaire revealed that 80% of the former abortion problem farms observed an improvement of the situation. However, in 43% of those farms no preventive measures had been taken and no significant differences in hygiene, forage and animal turnover could be observed between case- and control-farms. Additionally, no significant differences were detected concerning risk factors such as the presence of farm dogs and the disposal procedure for placental and fetal material.     

Neosporosis as a cause of abortion in dairy cattle in Rio Grande do Sul, southern Brazil.

Forty-six aborted bovine fetuses submitted to the Faculty of Veterinary, Department of Clinical Pathology, Universidade Federal do Rio Grande do Sul, were examined histopathologically. Non-suppurative inflammation was observed mainly in the brain and heart of 22 fetuses. Brain lesions consisted primarily of mononuclear inflammatory cell infiltrates with occasional foci of necrosis. Protozoa that reacted with Neospora caninum antisera were seen in 18 of the 22 (81.8%) brain specimens from fetuses with encephalitis. Blood samples collected from 223 Holstein cows on five dairy herds were tested for N. caninum antibodies by an immunofluorescent antibody technique. These samples were obtained from aborting cattle and normally calving cattle (control group). Overall, 11.2% of cows sampled had N. caninum antibodies at a dilution of 1:200. Seroprevalence was higher (P = 0.0053) in aborting (23.3%) than in non-aborting cows (8.3%). Association between seropositivity to N. caninum and abortion was found, with seropositive cows being 3.3 times more likely to abort than seronegative cows (OR = 3.33; 95% CI: 1.38, 8.062). Additionally, N. caninum antibodies were detected in sera from seven cows that had aborted fetuses with lesions suggestive of protozoal infection. These results suggest that N. caninum is an important cause of abortion in dairy cattle in Rio Grande do Sul State, Brazil.     

Proportion of abortions due to neosporosis among dairy cattle in Japan

A retrospective cohort study was conducted to clarify the association between seropositive reactions to Neospora caninum and subsequent reproductive disorders among dairy cattle in Japan. A statistically significant association between Neospora seropositive reactions and abortions was observed (P=0.016), and seropositive cattle were 6.1 times more likely to abort compared to seronegative cows. No significant differences were observed between seropositive reactions and other reproductive disorders such as conception failure, perinatal death and calf mortality. As indicated by estimation of the attributable fraction, 83.6% of abortions in Neospora seropositive animals may be attributed to N. caninum. Considering seroprevalence of N. caninum in the cattle which aborted in Japan, 21.8% of abortions were estimated to be caused by neosporosis in Japan.     

Outbreak of abortion associated with Neospora caninum infection in a beef herd

Abortion outbreaks caused by infection with Neospora caninum in beef cattle have not been well documented. Neospora caninum infection was confirmed in 4 fetuses that were aborted by cattle in a 350-head beef herd; an additional 58 cattle aborted during the next 2.5 months. Overall, 44.4% (157/354) of the cattle in the herd did not become pregnant or experienced fetal loss during this period. Initially, 81.3% (282/347) of the herd was seropositive for antibodies to N caninum, and seropositive cows were 6.2 times as likely to not be pregnant as were seronegative cows. Other potential causes of abortion were not identified in this herd. Following the outbreak, few cattle in the herd or in a second exposed herd seroconverted, but high antibody titers persisted in most seropositive cattle through the end of the calving season. Evidence of cow-to-fetus transmission of the organism was detected in > 82% of seropositive cows.     

Cytokine gene expression profiles in peripheral blood mononuclear cells from Neospora caninum naturally infected dams throughout gestation

Neospora caninum is a major cause of abortion in cattle but it is not known why some infected animals suffer abortion while others do not. An essential role in protective immunity against N. caninum has been proposed for Th1 cytokines such as IFN-γ and IL-12 although cytokine patterns in N. caninum infected pregnant cattle have been scarcely addressed. In this study, gene expression of the cytokines IFN-γ, IL-12, IL-10, IL-4 and TNF-α was analyzed by real time RT-PCR in peripheral blood mononuclear cells in N. caninum naturally infected dams throughout pregnancy. Blood samples were drawn from 18 cows (13 N. caninum seropositive and 5 N. caninum seronegative) on Days 45, 90, 120, 150, 180 and 210 of pregnancy or until abortion. Four seropositive animals aborted. Compared to the seronegative animals, N. caninum infected dams showed up-regulated mRNA levels of the Th1 cytokines, IFN-γ, TNF-α and IL-12p40, along with up-regulation of the T regulatory (Treg) cytokine IL-10. In contrast, expression levels of IL-4 (Th2 cytokine) did not differ significantly among the different groups throughout the study period. Our findings indicate clear differences in peripheral blood cytokine gene expression levels during pregnancy between animals naturally infected with N. caninum and seronegative control animals. To the best of our knowledge, this is the first study to examine the gene expression of Th1, Th2 and regulatory cytokines in the peripheral blood of pregnant cows naturally infected with N. caninum.     

Risk of abortion associated with Neospora caninum during different lactations and evidence of congenital transmission in dairy cows

OBJECTIVE: To examine the relationship between exposure to Neospora caninum and abortion in dairy cows during their first, second, third, and fourth or later lactations and to establish the main mode of transmission in female calves from birth until their first pregnancy was terminated by abortion or parturition. DESIGN: Prospective observational study. ANIMALS: 460 Holstein cows and 79 female calves. PROCEDURE: Cows were classified as seropositive or seronegative to N caninum within 7 days after calving; incidence of abortion was compared between groups during different lactations. Blood samples were collected from female calves before ingestion of colostrum and every 6 months until their first pregnancy was terminated by abortion or parturition; number of seropositive calves was compared between seropositive and seronegative dams. RESULTS: During the first pregnancy of their second lactation, risk of abortion for seropositive cows was 2.8 times that of seronegative cows. Among 10 calves born to seropositive cows, 4 were classified as seropositive at birth and thereafter. Among 69 calves born to seronegative cows, all were classified as seronegative at birth; 67 calves remained seronegative thereafter. CONCLUSIONS AND CLINICAL RELEVANCE: Exposure to N caninum alone was not significantly associated with abortion in cows during the first, third, and fourth or later lactations. Seropositive cows that have aborted previously may have subsequent abortions attributable to N caninum. Congenital infection was the main mode of N caninum transmission in a cohort of female calves.     

Seroepidemiology of neosporosis in dairy cattle in Uruguay

A seroepidemiological survey of Neospora caninum was done in a dairy farm in Uruguay employing indirect fluorescent antibody test (IFAT). Sera from 217 cattle (155 cows, 31 heifers and 31 calves) were tested for N. caninum antibodies in July 2000 and it was found that abortions were significantly associated with the seropositivity of the cows by chi(2)-test (P = 0.025). The cows with an IFAT titre of 1:3200 were at a significantly higher risk of abortion than seronegative cows (P = 0.0013). Neospora organisms were detected in the brains of two aborted foetuses by immunohistochemical examination. The infection of N. caninum was considered postnatal because of the even distribution of seropositive cows (60%), low seroprevalence (20%) in calves and no significant correlation in the serostatus between calves and their dams (P = 0.43). A fitted binomial generalised linear model revealed that there was an increasing risk of abortion with increasing IFAT titre and increasing age between 2 and 6 years.     

Neospora caninum in sheep: a herd case report

Neospora caninum was detected by means of PCR in the brain of 4 out of 20 aborted fetuses in a flock of 117 sheep exhibiting a persistent abortion problem, and N. caninum tissue cysts were furthermore found in encephalitic lesions in one of the PCR-positive fetuses. Toxoplasma gondii was detected as aborting agent in another 3 out of 20 fetuses. Antibodies to N. caninum (by indirect fluorescence antibody test (IFAT)) were found in 10.3% of 117 ewes and antibodies for T. gondii were found in 97.4% of 117 ewes. Other organisms associated with abortion were Chlamydia psittaci in three fetuses and Pasteurella multocida in one fetus. This is the first report of N. caninum associated abortion in naturally infected sheep.     

First Confirmed Case of Neospora caninum-associated Abortion Outbreak in Portugal

In a dairy herd experiencing an abortion outbreak, 49% of the 119 cows were positive for Neospora caninum antibodies and the remaining 51% were seronegative. Using immunohistochemical analysis of foetal brain tissue, Neospora cysts and bradyzoite clusters were identified in two of the 15 aborted foetuses submitted for testing. Other agents that commonly cause abortions were not found. These results, coupled with neurological clinical symptoms in new-born calves, implicated N. caninum as the major cause of abortions in this dairy herd. This is the first report that associates N. caninum infection with bovine abortions in Portugal.     

Serological investigation of an outbreak of Neospora caninum-associated abortion in a dairy herd in southeastern United States

An outbreak of Neospora caninum-associated abortion occurred in a South Carolina dairy wherein greater than 10% of the herd aborted over a 4-month period. Of the total number of cows at mid-late gestation, nearly 40% (28/71) aborted while the remaining 60% (43/71) gave birth to normal calves. Immunohistochemical examination of brain tissue from a subset of aborted fetuses confirmed N. caninum as the causative agent of abortion in these animals. A variety of serological assays, including indirect fluorescence antibody test (IFAT), recombinant enzyme-linked immunosorbent assay (rELISA), ISCOM-ELISA, avidity ELISA, and Neospora agglutination test (NAT), were used to evaluate sera collected during the outbreak from 240 cows for antibodies to N. caninum. IFAT and ISCOM-ELISA testing showed that nearly 80% of the dairy cows had antibodies to N. caninum. NAT and rELISA had similar levels of seropositivity relative to IFAT and ISCOM-ELISA, but the percentage of positive sera was dependent on the cut-off value chosen. As indicated by kappa coefficient statistical analysis, ISCOM-ELISA and IFAT exhibited the highest level of agreement in identifying N. caninum-positive and -negative cows. A decrease in the percentage of seropositive cows as determined by ISCOM-ELISA and IFAT with increasing cow age was noted. However, no significant difference was observed between cow age and abortion status. In addition to these tests, an avidity ELISA was performed on all sera with high (> or =0.4) ISCOM-ELISA readings. Avidity index (AI) increased with time post-abortion suggesting that most abortions were due to recent N. caninum infection. Of the cows at risk for abortion, the mean serological AI of aborting cows was significantly lower (P<0.05) than mean serological AI of non-aborting cows.     

Diagnostic survey of bovine abortion with special reference to Neospora caninum infection: importance, repeated abortion and concurrent infection in aborted fetuses in Southern Brazil

The protozoa Neospora caninum is an important cause of bovine abortion world-wide. The objective of this survey was to determine the distribution pattern of infectious abortion in Southern Brazil with special reference to N. caninum infection. A total of 161 bovine aborted fetuses from 149 farms were analysed during a 1.5 year period. The cause of abortion was identified in 51.5% of cases. Overall, 23% (37/161) of the fetuses were considered to be infected with N. caninum. Bacterial infection accounted for 17.4% (28/161) of cases, fungal infection for 3.1% (5/161) of cases and viral aetiology for 1.8% (3/161). Six fetuses had concurrent infection with N. caninum and Leptospira spp. Data from 111 fetuses and the respective aborted cows were analysed to investigate the association between previous abortion and current N. caninum infection. The prevalence of N. caninum-infected fetuses from cows with and without a history of previous abortion was 44% (11/25) and 24.4% (21/86), respectively. Cows aborting a N. caninum-infected fetuses were 2.4 times more likely to have aborted previously than cows aborting for other reasons (95% CI of odds ratio=0.9-6.8, P=0.06).     

Neospora caninum serostatus and culling of Holstein cattle

OBJECTIVE: To determine whether time until culling or risk of culling was associated with Neospora caninum serostatus among Holstein cattle in dairy herds in Ontario. DESIGN: Retrospective cohort study. ANIMALS: 3,412 cows in 56 herds. PROCEDURE: Blood samples were collected, and serum was tested for antibodies against N. caninum. Information on cows that were culled was collected during the 1- to 2-year period that producers were unaware of serostatus of individual cows in their herds. RESULTS: Herd prevalence of N. caninum-seropositive cows ranged from 0 to 68.3% (median, 7.0%). During the time of the study, 184 of 359 (51.3%) N. caninum-seropositive cows were culled, compared with 1,388 of 3,053 (45.5%) seronegative cows. Mean time from blood sample collection to culling for seronegative cows (289 days; 95% confidence interval, 280 to 299 days) was not significantly different from mean time for seropositive cows (296 days; 95% confidence interval, 269 to 323 days). Survival analysis indicated that N. caninum serostatus was not associated with time to culling or risk of culling. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that N. caninum serostatus of Holstein cows in Ontario was not significantly associated with either time to culling or risk of culling. Thus, N. caninum serostatus alone should not be used to determine whether cows should be culled.     

Diagnostic significance of Neospora caninum DNA detected by PCR in cattle serum

A nested PCR that successfully detected Neospora caninum DNA in serum of cattle was used for investigation of selected abortion cases and in a study of healthy pregnant cows at an abattoir. N. caninum DNA was not detected in serum from antibody positive dams that aborted due to N. caninum, but was present in serum of some antibody negative dams that aborted due to other causes. N. caninum DNA was also found in the serum of about half of the animals that aborted of undetermined cause, but was not detected in cow sera from two beef cattle herds in Western Australia with no recent history of abortion. In the abattoir study of 79 dams and their foetuses N. caninum DNA was found in serum of 3 dams and in material from 11 foetuses. The majority of the cows and all foetuses were antibody negative. Our findings suggest that there is no obvious relationship between the presence or absence of N. caninum DNA in serum and the presence of antibodies to N. caninum in dams, the presence of N. caninum DNA in foetuses or abortion due to N. caninum. This is the first report of the detection of N. caninum DNA in serum of cattle rather than the white blood cell fraction. It indicates the presence of free tachyzoites and/or parasite DNA in circulation. The results suggest that persistent infection in the absence of antibodies is a possible outcome of N. caninum infection. Infection of foetuses in the absence of antibodies supports the possibility of persistent infection due to immunotolerance to an early in utero infection. It is therefore important to test for N. caninum DNA as well as antibodies for the detection of exposed and/or infected animals. However, the presence or absence of N. caninum antibodies or DNA did not support nor exclude N. caninum as the cause of abortion. Additional criteria are required for a positive diagnosis of abortion caused by N. caninum.     

Neospora caninum infection does not affect the fertility of dairy cows in herds with high incidence of Neospora-associated abortions

This present study, was designed to establish whether Neospora caninum infection affects the fertility of high-producing dairy cows, and was based on yearly serological screening for neosporosis and on the confirmation of N. caninum infection in aborted foetuses. Assessment was made of 7518 artificial inseminations (AI) (applied to parous cows) performed in three herds. Of these inseminations, 2540 (33.8%) resulted in pregnancy; 34% of which corresponded to seronegative cows (2226 of 6556 AI performed in seronegative cows) and 32.6% to seropositive animals (314 of 962). Abortion occurred in 97 (30.1%) of the 314 pregnancies recorded in dams seropositive for N. caninum, while only 93 (4.2%) of the 2226 pregnancies in seronegative animals ended in abortion. Logistic regression analysis indicated no significant effects of N. caninum antibody titre, days in milk and milk production at insemination on fertility. Factors found to affect fertility were herd, season of insemination, lactation and insemination number, semen-providing bull and AI technician. Our results indicate that N. caninum infection does not affect the fertility of high-producing dairy cows.     

Application of the Neospora caninum IgG avidity ELISA in assessment of chronic reproductive losses after an outbreak of neosporosis in a herd of beef cattle.

Point-source infections are most likely the cause for Neospora caninum-induced abortion outbreaks in cattle, whereas an increased annual abortion rate may be a consequence of vertical transmission. The aims of the present study were to examine the reproductive effects of neosporosis in a beef herd for 3 years, after a point-source outbreak and to use IgG avidity serology to examine the chronicity of infections and patterns of transmission. During the study, 76-78% of animals were seropositive for N. caninum. The pregnancy rate varied from 88% to 94%, without any reduction in the pregnancy rate of seropositive cows compared with seronegative cows. The annual abortion rate was 2.5-5.5%, and all but 1 abortion occurred in seropositive dams. The efficiency of vertical transmission was estimated to be 85%. Several calves, born to seronegative dams, were seropositive at 6-13 months of age, indicating a 22% mean annual rate of horizontal transmission. The mean avidity in seropositive cows increased from 30 during the initial outbreak to 74 after 3 years. The mode of IgG avidity was 21-40 during the initial abortion outbreak, 41-60 after 1 year, and 61-80 after 2 and 3 years. The results reveal high annual rates of both vertical and horizontal transmission of N. caninum in a herd of beef cows and provide further validation on the ability of the N. caninum IgG avidity ELISA to accurately assess the chronicity of infection.     

Seroprevalence of Neospora caninum in aborting dairy cattle in the Czech Republic

A serological survey for antibodies against Neospora caninum in aborting cattle was carried out in the Czech Republic. Serum samples from 463 aborting dairy cows originated from 137 farms from different parts of the Czech Republic were tested for presence of N. caninum antibodies by use of an enzyme-linked immunosorbent assay (ELISA) and an indirect fluorescent antibody test (IFAT). Antibodies (> or = 1:640) to N. caninum were found in 18 (3.9%) of 463 aborting cows. Farm prevalence in aborting cows was 12.4% (17/137). The antibody titres of cows were 1:200 (9 cows), 1:640 (7 cows), 1:1280 (3 cows), 1:2560 (3 cows), 1:5120 (3 cows), 1:10,240 (2 cows) and 1:20,480 (0 cow). A case-control study was conducted to estimate the association of N. caninum infection and abortion. For this 407 serum samples were collected from cows on five dairy farms with repeated occurrence of endemic and sporadic abortion of unidentified etiology. These samples were obtained from aborting cattle (n=44) and normally calving cattle (control group; n=363) and tested for N. caninum antibodies by an immunofluorescent antibody test (IFAT). Overall, 3.19% (13/407) of cows sampled had positive N. caninum fluorescence with a cut-off titre of 1:200. The prevalence of N. caninum was significantly higher (P<0.05) in the aborting group (13.64%; 95% confidence interval (CI): 5.2, 27.4) than in the control group (1.93%; 95% CI: 0.8, 3.9). A strong association between seropositivity and abortion was found, with seropositive cows being eight times more likely to abort than seronegative cows (odds ratio=8; 95% CI: 2.6, 25.1). This first report on the serological prevalence of N. caninum in cows in the Czech Republic verified a strong association between N. caninum infection and abortions in five dairy farms. Thus, the neosporosis should be considered in differential diagnosis of bovine abortion.     

Neospora-associated abortion episode over a 1-year period in a dairy herd in north-east Spain

This report describes a retrospective study based on serological screening, performed on a Neospora-associated abortion episode over a 12-month period in a dairy herd in north-east Spain. During the 1-year study period, the overall abortion rate for the herd was 23.2% (38 abortions of 164 diagnosed pregnancies). The data analysed were those derived from blood samples collected from the whole herd (n = 237) and from diagnosed pregnancies. Antibodies to Neospora caninum were found in 35.4% of the cattle with 44% of seropositive pregnant animals aborting over 1-year period. Based on the odds ratio, the risk of abortion was 12.2 times higher (P < 0.0001) in the Neospora-seropositive animals than in seronegative animals and significantly higher during the second term of gestation (P < 0.01) than during the first and third terms. Abortions were not found to be associated with parity status or season of pregnancy, and the common risk factors associated with pregnancy loss in the geographical area of the study. Age-related differences in N. caninum seroprevalence were not statistically significant, indicating vertical transmission as the main route of infection. Indeed, a high percentage of congenitally infected offspring was observed (90.6%) and the farm had been free of dogs for the last 7 years. Our results suggest that, when a dairy herd shows an increased incidence of abortions due to Neosporosis, maternal serology can be a good indicator of the abortion risk in individual cows, and that the effects of factors normally related to abortion, such as parity and pregnancy season, may be masked.     

A longitudinal study of Neospora caninum infection on three dairy farms in Brazil

Neospora caninum is an apicomplexan protozoan parasite that is one of the most important infectious causes of abortion in both dairy and beef cattle in many countries. The objectives of this longitudinal study were to determine the prevalence, rates of vertical and horizontal transmission of N. caninum and hazard for culling of N. caninum-seropositive animals in three Brazilian dairy herds. Blood samples from all animals were collected nine times at each of the three farms over a two-year period. Serum was tested for antibodies against N. caninum using the indirect fluorescent antibody test with a cutoff value of 1:100. The percentage of N. caninum-positive samples at each sampling time ranged at Farm I from 3.32% to 11.71%, at Farm II from 3.90% to 22.06% and at Farm III from 3.90% to 22.06%. The number of positive serological reactions varied in relation to the number of repeated samples taken from individual animals at each farm. In all herds, there was a high degree (P<0.05) of association between the N. caninum serological status of dams and daughters. The seropositive conversion rate was estimated as 0.37%, 3.00% and 6.94% per 100 cow-years at Farms I, II and III, respectively. The seronegative conversion rate was estimated as 31.58% and 11.11% per 100 cow-years at Farms I and III, respectively. In all herds, there was no difference (P>0.05) in the culling rate between the cattle that were seropositive cattle and seronegative for N. caninum infection. The results from this study confirm the importance of vertical transmission in the epidemiology of N. caninum. Although a few positive seroconversions indicated horizontal transmission, it does not appear to be the major route of infection for N. caninum.