Associations between pregnancy outcome and serological response to Neospora caninum among a group of dairy heifers

AIM: To monitor pregnancy in a group of rising 2-year-old dairy heifers on a farm on which abortion due to Neospora caninum was known to occur in previous years. METHODS: A prospective cohort study group of 164 rising 2-year-old heifers was pregnancy-tested and blood-sampled at 4-5-week intervals throughout gestation. Sera were tested for antibodies to N. caninum at 3-4-month intervals, using an enzyme-linked immunosorbent assay (ELISA). When loss of pregnancy was detected, an N. caninum indirect fluorescent antibody test (IFAT) was conducted retrospectively on stored sera collected the month before abortion, the month abortion was detected, and for the following 2 months, from heifers that aborted. All fetal and placental material detected following abortion was subjected to gross post-mortem and histopathological examination. RESULTS: Eleven of 18 (61%) heifers that were seropositive and 4/146 (3%) heifers that were seronegative to N. caninum by ELISA, aborted. The relative risk for abortion among ELISA-positive heifers was 23.6. Abortion occurred predominantly between Days 120 and 152 gestation among the ELISA-positive heifers and throughout gestation among the ELISA-negative heifers. IFAT titres rose around the time of abortion in most of the heifers that were previously seropositive by ELISA, but dropped rapidly again in post-abortion samples. IFAT titres among 4/6 ELISA-positive heifers that did not abort increased, but later in gestation than the time other heifers aborted. IFAT titres remained negative in heifers that aborted that were ELISA negative. CONCLUSIONS: Heifers that were seropositive to N. caninum by ELISA had a much greater risk of abortion than seronegative heifers. Most seropositive heifers showed evidence of a reactivation of infection during pregnancy. High (> or =1:2,000) N. caninum IFAT titres also occurred in non-aborting heifers. CLINICAL RELEVANCE: Culling of replacement heifers seropositive to N. caninum may be a cost-effective strategy for minimising risk of abortion. Pregnancy testing heifers before 5 months gestation may overestimate the number that calve in N. caninum-infected herds, but would assist in documenting the occurrence of abortion. Reliance on a high (>1:2,000) IFAT titre to rule-in N. caninum as a cause of abortion is likely to produce false-positive results.     

Some Factors Affecting the Abortion Rate in Dairy Herds with High Incidence of Neospora‐Associated Abortions are Different in Cows and Heifers

The aim of this study was to determine if the factors affecting the abortion rate in dairy herds with high incidence of Neospora‐associated abortions are different in pregnancies of cows and heifers chronically infected with Neospora caninum. In heifers (n = 229), an increase in the cumulative number of days with a mean relative humidity (RH) lower than 60% during the second trimester of gestation increases the risk of abortion. Yet, the likelihood of abortion was 7.6 times lower for pregnant heifers inseminated with Limousin bull semen, compared with those inseminated with Holstein‐Friesian bull semen. In pregnancies of parous cows (n = 521), an increase in rainfall and in the cumulative number of days with a mean RH lower than 60% during the second trimester of gestation increased the abortion rate. However, in contrast, an increase in the lactation number produced a decrease in the abortion rate, with a likelihood of abortion 4.8 times lower for pregnant cows inseminated with Limousin bull semen, and three times lower for those inseminated with Belgian Blue bull semen, compared with dairy cows inseminated with Holstein‐Friesian bull semen. Finally, the likelihood of abortion was 3.2 times lower for pregnancies of parous cows with low antibody titres against N. caninum (6–30 units) as compared to those with high antibody titres (≥30 units), whereas in heifers this variable had no effect. The practical recommendations of the present study include the control of the cow environment during the second trimester of gestation, the priority of culling for parous cows with higher antibody titres against N. caninum and the insemination of Neospora‐seropositive cows with semen from the Limousin breed.     

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.     

Immune response to Neospora caninum in naturally infected heifers and heifers vaccinated with inactivated antigen during the second trimester of gestation

The objective of this study was to compare the immune response to Neospora caninum in naturally infected heifers and heifers inoculated with a killed whole N. caninum tachyzoite preparation during the second trimester of gestation. Nine Holstein heifers were used in this study; three naturally infected heifers were born from seropositive dams, and six seronegative heifers were born from seronegative dams. Four seronegative heifers were subcutaneously vaccinated with a killed whole N. caninum tachyzoite preparation at weeks 13, 15 and 17 of gestation. A killed whole N. caninum tachyzoite preparation containing 45 mg of protein/5 ml dose was formulated with 70% of mineral oil adjuvant (13% consisting of Arlacel C, 85% Marcol 52 and 2% Tween-80). Similarly, two seronegative heifers (negative controls) were inoculated with mock-infected bovine monocytes in oil adjuvant. Humoral immune responses were tested by using an indirect fluorescent antibody test (IFAT) and an indirect enzyme-linked immunosorbent assay (ELISA) for detecting isotype specific antibodies. Cellular immune responses were assessed by lymphocyte proliferation test (LPT) and IFN-gamma production. N. caninum-specific antibody responses increased in immunized cattle by week 15 of gestation (mean reciprocal antibody titers 450+/-252), peaked at week 23 (mean 16,000+/-6400). Maximum antibody response in naturally infected heifers was observed at week 19 of gestation (mean: 3467+/-2810). Mean serum IFAT titers were significantly higher in immunized heifers compared with those in naturally infected heifers from weeks 17 to 25 (P < 0.05). Analysis of isotype specific antibodies in naturally infected heifers revealed a predominant IgG1 response in one heifer and a predominant IgG2 response in the other two. Similar titers of IgG1 and IgG2 occurred in immunized heifers. Control heifers remained seronegative throughout the study by IFAT and ELISA. Significant antigen-specific proliferation responses were only detected in naturally infected heifers in week 19 of gestation. Peripheral mononuclear blood cells (PMBC) from immunized animals produced IFN-gamma in similar concentrations to those of infected animals (P > 0.05). No abortion was seen in any experimental group; however, one calf from a vaccinated heifer died due to dystocia. All calves from vaccinated and control heifers were seronegative by IFAT at 6 months of age; in contrast, calves born from naturally infected heifers remained seropositive with titers > or = 200. Killed vaccine induced similar immune responses to those found in chronically, naturally infected cattle which did not abort; however, different immune pathways may be followed in vaccinated and natural infected heifers with differences in degree of protective immunity.     

Effects of crossbreed pregnancies on the abortion risk of Neospora caninum-infected dairy cows

Previous studies have shown that the use of beef bull semen significantly reduces the rate of abortions due to Neospora caninum in artificially inseminated (AI) seropositive dairy cows. In addition, certain beef breeds could be more resistant to N. caninum infection and abortion than others. The aim of the present study was to determine whether different crossbreed pregnancies, those derived from Limousin, Charolais, Piedmontese or Belgian Blue semen, carry different risks of abortion in Neospora-infected dairy cows. The effects of possible interactions between maternal levels of N. caninum antibodies and the different breed crosses were also evaluated. The study was performed on five commercial Holstein–Friesian dairy herds in Northeast Spain with previously confirmed diagnoses of N. caninum infection in aborted foetuses. The study population was comprised of 1115 pregnancies: 633 pregnancies recorded after AI using Holstein–Friesian semen from 18 bulls and 482 after AI using beef semen from 27 bulls (304 inseminations using semen from Limousin bulls, 191 from Belgian Blue bulls, 89 from Piedmontese bulls and 49 from Charolais bulls). Abortion rates were 32.2% (155/482) and 15.2% (96/633) for seropositive cows inseminated with Holstein–Friesian and beef breed semen, respectively. Logistic regression analysis revealed the herd and the interaction between maternal N. caninum antibody titre and the different crossbreeds as significant factors affecting the abortion rate. Lowest abortion rates, similar to that shown by seronegative animals in the analysed herds (3.2%, 239/7432), were observed in dams AI using Limousin semen that had low (<30 relative index (RI) units) N. caninum antibody titres (2.1% abortion, 3/145) and these cows were used as reference. Compared to the cows used as reference, cows with low N. caninum antibody titres (<30 RI units) showed a similar risk of abortion when inseminated with Piedmontese or Charolais bull semen, but higher risk of abortion when inseminated with Holstein (17.9 times) or Belgian Blue (7.2 times) bull semen. All cows with high N. caninum antibody titres (≥30 RI units) had a higher risk of abortion, ranging from 8.9 times (cows inseminated with Limousine semen) to 37.8 times (cows inseminated with Piedmontese semen), compared to the cows used as reference. In conclusion, different crossbreed pregnancies carried different abortion risks in Neospora-infected dairy cows. The use of beef bull semen dramatically reduced the risk of abortion in dairy cows, especially if Limousin breed semen was used. Moreover, this reduction was found to be dependent on the N. caninum antibody titre such that the lowest incidence of abortions was recorded in Limousin semen inseminated cows with low antibody titres. Insemination of Neospora-seropositive cows with beef bull semen could both reduce the risk of abortion and avoid breeding replacements for infected cattle.     

Gamma Interferon Production and Plasma Concentrations of Pregnancy‐Associated Glycoproteins 1 and 2 in Gestating Dairy Cows Naturally Infected With Neospora caninum

Gamma interferon (IFN‐γ) production and cross‐breed pregnancy have been attributed a role in protecting dairy cows infected with Neospora caninum against abortion. Plasma levels of pregnancy‐associated glycoproteins‐1 (PAG‐1) are a marker of placental/foetal well‐being and of PAG‐2 is an abortion risk indicator in chronically N. caninum‐infected animals. The present study examines, in cross‐breed pregnancies, interactions between IFN‐γ production and levels of PAG‐1 and PAG‐2 in non‐aborting naturally Neospora‐infected dairy cows. Data were obtained from 60 pregnant Holstein‐Friesian cows: 44 Neospora‐seropositive and 16 Neospora‐seronegative; 12 became pregnant using Holstein‐Friesian semen and 48 using Limousin semen. Blood samples were collected on Days 40, 90, 120, 150, 180 and 210 of gestation. Gamma interferon was only detected in the plasma of nine of the 44 Neospora‐seropositive cows, all of them became pregnant using Limousin semen. Through GLM procedures, in cows inseminated with Limousin semen and Neospora‐seropositive cows showing no IFN‐γ production, PAG‐1 concentrations were high and increased throughout gestation compared to the levels detected in cows inseminated with Holstein‐Friesian semen and Neospora‐seropositive cows producing IFN‐γ, respectively. In Neospora‐seronegative cows and in Neospora‐seropositive cows showing no IFN‐γ production, significantly increased PAG‐2 concentrations were observed on gestation Day 120. Our findings indicate that IFN‐γ production correlates negatively and the production of antibodies against N. caninum is uncorrelated with plasma PAG concentrations during gestation in Neospora‐infected dairy cows. Accordingly, IFN‐γ production could be linked to the transplacental migration of tachyzoites, which may cause a reduction in PAG levels.     

Relationship between Rainfall and Neospora caninum‐associated Abortion in Two Dairy Herds in a Dry Environment

The possible direct relationship between climate variations and abortion in Neospora caninum‐infected cows has not been studied. The objective of this study was to determine whether climate changes could be a risk factor for abortion in N. caninum‐infected cows, and was based on yearly serological screening for neosporosis and on the confirmation of N. caninum infection on aborted fetuses in two high‐producing dairy herds with a mean 27% seroprevalence of N. caninum antibodies. The final population study was comprised of 357 pregnancies in seropositive animals. Logistic regression analysis indicated no significant effects of herd, N. caninum antibody titre, climate variables during the first and third trimesters of gestation, mean and maximum temperature–humidity index values during the second trimester of gestation, and previous abortion on the abortion rate. Based on the odds ratio, a 1‐unit increase in lactation number yielded a 0.85‐fold decrease in the abortion rate. The likelihood of abortion was 1.9 times (1/0.54) lower for pregnant cows inseminated with beef bull semen compared with Holstein‐Friesian bull semen. The likelihood of abortion decreased significant and progressively by factors of 0.5, 0.41 and 0.3 for the respective classes 40–49, 30–39 and <30 rainfall mm during the second trimester of gestation (using the class ≥60 rainfall mm as reference). As a general conclusion, it seems that increased rainfall in a dry environment can compromise the success of gestation in N. caninum‐infected cows. Attempts should therefore be made to reduce environment effects during the second trimester of gestation, a period in which the immune response of cows is diminished.     

Serological investigation of aborted sheep and pigs for infection by Neospora caninum

Serology for Neospora caninum was undertaken using direct ELISAs on sera from 660 aborted sheep and 454 breeding sows, which had aborted or were considered infertile. All ovine sera were further tested by indirect fluorescent antibody test (IFAT) for N. caninum, and a latex agglutination test (LAT) for Toxoplasma gondii was performed on 423 of the samples, including all those positive by ELISA. ELISA-positive porcine sera were tested by IFAT and an inhibition ELISA for antibodies to N. caninum and by LAT for T. gondii. Only 3 (0.45%) of the ovine sera were seropositive for N. caninum by both ELISA and IFAT whereas although 40 porcine sera were seropositive by ELISA all were negative by IFAT. The results suggest that environmental exposure to N. caninum occurs rarely in sheep and pigs.     

Serology of a Neospora abortion outbreak on a dairy farm in New Zealand: a case study

AIM: To describe the kinetics of serological titres after an abortion outbreak in April-May 1995 due to Neospora caninum affected 17 dairy cows in a herd of 320. METHODS: Thirty-five cows, that had either aborted, carried mummified calves, were not pregnant or calved normally were: bled several times at regular intervals and the sera tested for Neospora antibodies in the indirect fluorescence antibody test (IFAT). RESULTS: Maximal IFAT titres of up to 1:4000 occurred within 6 weeks of the abortion outbreak, decreased over the next 2 months to < or = 1:200 and remained at this level until the next scheduled bleed a further 2 months later. A rise in titres was subsequently observed in the cows that had aborted or were not pregnant (at the time of the abortions) or had carried mummified foetuses. Seroconversion was also observed in some of the control cows, which had, up until then, remained seronegative. A dog and cat in contact with the cows in the herd investigated were, however, negative in the IFAT. CONCLUSIONS: Maximal serological titres in Neospora abortions are observed within weeks of the abortion event and then quickly return to very low levels. Subsequently, a recrudescence of titres can be observed in infected cows during the next pregnancy, without it being associated with repeat abortions.     

Plasma Concentrations of Pregnancy‐Associated Glycoproteins Measured Using Anti‐Bovine PAG‐2 Antibodies on Day 120 of Gestation Predict Abortion in Dairy Cows Naturally Infected with Neospora caninum

The present study sought to determine: (i) the effects of Neospora caninum infection and twin pregnancy on plasma pregnancy‐associated glycoprotein‐2 (PAG‐2) concentrations throughout pregnancy and (ii) whether plasma PAG‐2 concentrations could predict abortion in N. caninum‐infected cows. The study was performed on a commercial Holstein‐Friesian dairy herd in northeastern Spain and the final data included those recorded in 53 non‐aborting and 19 aborting animals. Blood samples were collected immediately before pregnancy diagnosis (on Days 40, 90, 120, 150, 180 and 210 post‐insemination) in non‐aborting cows or until the time of abortion detection in aborting cows. General lineal models (GLM) repeated measures anova revealed the different behaviour of PAG‐1 and PAG‐2, and significant effects of Neospora seropositivity, cool season and twin pregnancy on plasma PAG‐2 concentrations throughout gestation (between‐subject effects). In addition, based on the odds ratios, the likelihood of abortion increased in Neospora‐seropositive cows (by a factor of 7.0) compared to seronegative animals and decreased in cows with a high plasma PAG‐2 concentration (>4.5 ng/ml) on Day 120 of pregnancy (by a factor of 0.24), compared to the remaining cows. In conclusion, there is a relationship between plasma PAG‐2 concentrations and the risk of abortion in Neospora‐infected dairy cows. Thus, plasma PAG concentrations measured using anti‐boPAG‐2 antiserum on Day 120 of gestation could serve as an indicator of the abortion risk in N. caninum infected animals; values <4.5 ng/ml indicating a high risk of abortion in chronically infected animals.     

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‐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.     

Pattern of recognition of Neospora caninum tachyzoite antigens by naturally infected pregnant cattle and aborted foetuses

Different aspects of Neospora tachyzoite antigen recognition by Neospora-infected heifers and cows and aborted foetuses were studied. The pattern of antigen recognition and the relationship between IFAT titres and number of Neospora antigens detected, were evaluated. In addition, the tachyzoite antigens involved in the humoral immune response developed against infection in normal cows and cows that aborted were also characterised throughout pregnancy. Comparison of tachyzoite antigen recognition was carried out in 13 thoracic and/or abdominal fluids from Neospora aborted foetuses and 33 sera from Neospora infected cows that had aborted. The kinetics of Neospora-antigen recognition was studied in Neospora-infected heifers and cows that had aborted foetuses (7) or not (14) during pregnancy.Based on the frequency and intensity of recognition, four IDAs-17-18, 34-35, 37 and 60-62kDa antigens-have been described. Moreover, a correlation was found between Western blot results and IFAT titres in both age groups. In relation to antigen recognition throughout pregnancy by seropositive cows that had aborted or not, the antibody fluctuations throughout pregnancy described in the literature could be due to differences in the intensity and frequency of recognition of particular antigens, especially the 17-18kDa antigen. We emphasize the important role that the 17-18kDa antigen could play in the serological diagnosis of Neospora infection in cattle as this was intensely detected in 100% of the animals.     

Serological Status and Abortion Rate in Progeny Obtained by Natural Service or Embryo Transfer from Neospora caninum‐Seropositive Cows

Bovine neosporosis is an important disease in dairy herds in Argentina. Embryo transfer (ET) technology offers the opportunity to minimize vertical transmission of neosporosis. The objective of this work was to compare the serological status and the incidence of abortion between heifers obtained via natural service and heifers obtained by ET from Neospora‐infected cows. Progeny from eight Neospora‐infected cows belonging to a commercial dairy farm where ET was routinely performed were studied. Between 1996 and 2001, 16 and 15 heifers were obtained by natural service and ET, respectively. Serostatus and reproductive performance were recorded in both progeny. Serostatus was determined by indirect fluorescent antibody test (IFAT). All heifers obtained by natural service were seropositive and six of 16 pregnancies ended in abortion. All heifers obtained by ET were seronegative and only two abortions of 15 pregnancies were recorded.     

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.     

Risk factors of Neospora caninum infection in dogs and cats in dairy farms in Western Thailand

The objectives of this study were to investigate the risk factors of Neospora caninum infection in dogs and cats found in dairy herds in western Thailand. A case-control study was conducted in dairy herds in three western provinces including Nakhon Pathom, Ratchaburi and Kanchanaburi. Blood samples of pets from 14 positive dairy herds and 26 herds from negative neighbouring farms which were randomly selected, in total blood samples from dogs and cats from 40 herds were collected and examined for antibodies against N. caninum infections using competitive ELISA and indirect fluorescent antibody test (IFAT). No seropositive cats were found. Of the 38 positive dogs, four (10.5%) were seropositive to N. caninum, which was higher than the proportion of seropositive in the negative herd population (one of 76, 1.3%). The higher proportion of seropositive farm dogs as compared with neighbouring dogs tended to be significant (P = 0.055).     

Effect of vaccination of cattle with the low virulence Nc-Spain 1H isolate of Neospora caninum against a heterologous challenge in early and mid-gestation

Live vaccines have emerged as one of the most potentially cost-effective measures for the control of bovine neosporosis. Previous studies have shown that Nc-Spain 1H is a naturally attenuated isolate of Neospora caninum and that immunisation with live Nc-Spain 1H tachyzoites generated a protective immune response in mice. The aim of this study was to evaluate the safety and efficacy of immunisation in cattle. N. caninum-seronegative heifers were immunised subcutaneously twice with 10⁷ live Nc-Spain 1H tachyzoites prior to artificial insemination. No adverse reactions or negative effects on reproductive parameters were recorded following immunisation. In immunised and non-challenged heifers, no foetal deaths were observed, and none of the calves was congenitally infected. The efficacy against N. caninum-associated foetal death and vertical transmission was determined after challenge with high doses of the Nc-1 isolate at 70 and 135 days of gestation, respectively. After the challenge in early gestation, the immunisation induced a protection of 50% against foetal death. In addition, the microsatellite analysis performed in PCR-positive tissue samples from foetuses that died after challenge infection showed that the profiles corresponded to the challenge isolate Nc-1. A degree of protection against vertical transmission was observed after challenge at mid-gestation; calves from immunised heifers showed significantly lower pre-colostral Neospora-specific antibody titres than calves from the non-immunised/challenge group (P < 0.05). Strong antibody and interferon gamma responses were induced in the immunised heifers. This study indicates that the immunisation before pregnancy with the Nc-Spain 1H vaccine isolate appeared to be safe and reduced the occurrence of N. caninum-associated abortion and vertical transmission in experimentally infected cattle. In light of these encouraging results, the next step for testing this live attenuated candidate should be the assessment of its efficacy and safety in naturally infected cattle.     

Neospora caninum Infection in a Clinically Healthy Calf: Parasitological Study and Serological Follow‐up

In this study a case of congenital infection in a clinically healthy calf is reported. The mother showed high antibody levels (IFAT) at 230 days of gestation (IgG titres 1:1600, IgM titres 1:320) and the parasite was isolated from placental cotyledonary villi at calving. The IgM values are indicative of a recent infection in the third trimester of gestation. The calf was monitored serologically for IgM and IgG from birth until slaughtering, at 8 months of age. IgM titre showed a peak at birth, while IgG peak was observed at 40–60 days of age. Parasitic isolation was obtained by biological tests using Swiss mice or VERO cell cultures inoculated with brain and spinal cord tissues. The parasitic presence in the calf was also evidenced in the myocardium with immunohistochemical method. The results are very important because they demonstrate that the period of gestation in which the cow becomes infected is an important factor in the pathogenesis of N. caninum induced abortion: in fact, the acquisition of infection in the third trimester of gestation allowed the foetus to develop a sufficient grade of immunocompetency to limit parasite multiplication with the result of a calf born clinically healthy.