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.     

A long-term study of Neospora caninum infection in a Swedish dairy herd

A longitudinal study was performed in a Swedish dairy herd where Neospora caninum had been isolated from a stillborn calf. Starting in autumn 1994, blood samples from all female animals in the herd were collected once yearly until 1999. The sera were analysed for presence of IgG1 antibodies to N. caninum by the iscom ELISA, and by an avidity ELISA to establish the timing of infection. In addition, data on reproductive performance were compiled. During the study the percentage of seropositive female animals increased from 63% to 87%. In 1994 a large number of young animals tested seropositive although their dams were seronegative, indicating that a transmission of the parasite other than the vertical had recently occurred. Low avidity values supported this assumption. The annual abortion rate increased from a mean of 2% before the initiation of the study to 9% in 1994-1998. During the same time, as judged by the avidity data, a large proportion of the animals shifted from being recently to being chronically infected. The source of the external infection in the herd could not be identified.     

A longitudinal study of seroprevalence and seroconversion of Neospora caninum infection in dairy cattle in northeast Thailand

A long-term study was carried out in 11 dairy herds in the Khon Kaen province of northeast Thailand between August 2001 and November 2004. The objective was to investigate seroprevalence dynamics of Neospora caninum infection in the herds and to demonstrate patterns of seroconversion in individual cattle. Each herd was visited once a year, in total four times, and sera from cattle > 3 months of age and farm dogs as well as a sample from the bulk milk were collected. All samples were analysed for presence of specific antibodies by an N. caninum iscom ELISA. The overall percentage of antibody-positive cattle was constant and varied only between 10 and 13% over the 4 years, but the variation in within-herd seroprevalence between herds was substantial. Two herds had > or = 20% seropositive animals at all samplings and consistently high bulk milk OD, whereas two herds had no seropositive animal at the last two samplings and low bulk milk OD. Five herds had a decreasing trend of within-herd seroprevalence, whereas the remaining six herds had a higher portion of test-positive individuals at the end of the study. A total of 424 individuals were sampled more than once; 344 (81%) and 32 (8%) were consistently antibody-negative and antibody-positive, respectively. The proportions of animals that changed from being seronegative to seropositive and from being seropositive to seronegative between the years were 3.9-4.6% and 19-39%, respectively. Apparent vertical and horizontal transmission rates were 58% (95% CI; 44-71%) and 5% (95% CI; 3-7%), respectively. In conclusion, the overall percentage of N. caninum antibody-positive cattle was constant over the years, but the within-herd seroprevalence varied substantially between the herds. Seroconversions were likely to occur in individual cattle although most animals had consistent serological status throughout the study.     

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.     

Neospora caninum infections in Australia and New Zealand

OBJECTIVE: To review the current state of knowledge of Neospora caninum infections with particular reference to Australia and New Zealand. PROCEDURE: Several databases were searched electronically including Medline, Current Contents, Vet CD using several key words (Neospora caninum, neosporosis, abortion, cattle, dogs) and authors names. References in original articles were also traced and use made of the author's own original research in the field. CONCLUSION: N caninum is recognised worldwide and is also widespread, in particular in dairy cattle, in Australia and New Zealand. It has been reported in both countries retrospectively (in dogs) from the early 1970s. Abortion storms in dairy herds appear to be the most common feature. Recent reports indicate that the dog is the definitive host, in which the sexual cycle is completed. Further studies are however required to establish important aspects of the epidemiology, such as mode of transmission. In the absence of an effective vaccine, the most effective control strategy is selective culling of infected animals and the prevention of access of dogs to expelled placentas and foetuses and, possibly, to raw beef.     

Chronic Neospora caninum infection and repeat abortion in dairy cows: a 3-year study

The serological status of Neospora was monitored in animals older than 6 months in a dairy herd with a 3-year history of prevalent N. caninum and N. caninum-associated abortions. The numbers of animals in the herd tested each year of the study period were 259, 222 and 231, respectively. A separate analysis was performed on the 122 animals persisting in the herd for the 3 years. The overall seroprevalence of N. caninum in the herd decreased from 31.7% in the first year to 24.8% in the second year and to 19.9% in the third year of the study, while the overall abortion rate decreased from 20.6% in the first year to 5.5% in the second year, and 9.9% in the third. These decreases occurred in response to control measures adopted from the second year onwards, such as culling Neospora-seropositive aborted animals and inseminating Neospora-seropositive dams with beef bull semen. Of the total number of abortions recorded in seropositive animals, 51% were repeat abortions that occurred in 36.8% of the animals with a previous history of abortion. The initial seroprevalence of Neospora in the 122 animals followed for the 3 years was 18%, increasing to 21.3% in the second and third years. Seroconversion only occurred in four animals during the second and third years of the study and abortion occurred only in seropositive individuals. Of the total number of abortions recorded in the 122 animals, 61.5% were repeat abortions that occurred in 26.7% of the animals with a previous history of abortion. These results indicate that Neospora seropositivity can be very stable through time and N. caninum infected cows can show a high rate of repeat abortions. The present data reinforce the idea that annual serological screening for Neospora can be an effective and rapid method of detecting N. caninum infection, such that control measures can be established at the farm level.     

Abortion epidemic in a dairy herd associated with horizontally transmitted Neospora caninum infection

A dairy herd experienced an abortion epidemic during which 43 per cent of the cows at risk aborted. Neospora caninum infection was demonstrated in four of six fetuses suitable for examination and the group of at-risk cows that aborted had significantly higher N. caninum antibody concentrations than the at-risk cows that delivered a live calf at term (P<0.001). The antibody concentrations in the cow herd were significantly higher than in the youngstock (P<0.001), and the concentrations in the youngstock increased significantly (P<0.001) with age. When seven months to a year old, the calves born at term to the at-risk cows had significantly higher (P=0.007) antibody concentrations than age-matched calves born before the epidemic. At the time of the epidemic, there was a significant increase in the antibody levels of the herd that was not consistent with vertical infection alone, indicating that there appeared to have been a sudden large increase in the incidence of horizontal postnatal transmission of N. caninum to the cow herd, or to the surviving offspring of the at-risk cows, or to both of these groups.     

Seroprevalence of Neospora caninum infection following an abortion outbreak in a dairy cattle herd

OBJECTIVE: To investigate the seroprevalence of Neospora caninum infection in a commercial dairy cattle herd, 15 months after detection of an abortion outbreak. PROCEDURE: Sera from the whole herd (n = 266) were examined for N caninum antibodies by indirect fluorescent antibody test (IFAT) and immunoblot analysis. Herd records were reviewed to collate serological results with abortion history, proximity to calving, and pedigree data. RESULTS: The seroprevalence of N caninum infection was 24% (63/266) for IFAT titre > or = 160, 29% (78/266) for immunoblot positive (+ve), and 31% (82/266) for IFAT > or = 160 and/or immunoblot +ve; 94% (59/63) of animals with IFAT > or = 160 were immunoblot +ve. The association between seropositivity (IFAT > or = 160 and/or immunoblot +ve) and history of abortion was highly significant (P < 0.001); the seroprevalence was 86% (18/21) in aborting cows, compared with 30% (50/164) in non-aborting animals. The abortion rate for seropositive cows was 26% (18/68) compared with 3% (3/117) for seronegative animals. IFAT titres of infected cows were higher within 2 months of calving than at other times (P < 0.001). The association between seropositivity in dams and daughters was highly significant (P = 0.009). CONCLUSIONS: The abortions were associated with N caninum infection and there was evidence of reactivation of latent infection close to calving and congenital transmission of infection. Immunodominant antigens identified by immunoblots may prove useful for improved diagnostic tests.     

Seroprevalence of Neospora caninum infection in dairy and beef cattle in Paraguay

Seroprevalence of Neospora caninum in 879 beef and dairy cattle in different locations of Paraguay was determined by an ELISA. In the survey, 262 (29.8%) cattle were positive to N. caninum, and animals with anti-Neospora antibody titre were observed in all the locations tested in the country. Serum samples taken from a herd that exhibited persistent abortion had the highest percentage of animals being positive to the parasite (17/30, 56.7%). In the same herd, abortion was significantly more likely in animals with high anti-Neospora antibody titre. Immunoblot analysis demonstrated that the banding pattern from positive Paraguayan cattle was similar to that seen with the positive control sample. In conclusion, N. caninum infection is present among Paraguayan beef and dairy cattle, and it may be an important cause of bovine abortion in Paraguay.     

Isolation and molecular characterisation of Neospora caninum in cattle in New Zealand

AIM: To isolate Neospora caninum from the brains of naturally infected cattle and use molecular techniques to characterise the isolates. METHODS: Neospora caninum tachyzoites were isolated in Vero cell culture from the brains of a cow and two calves. The isolates were characterised using polymerase chain reaction (PCR) methods, DNA sequencing, an immunofluorescent antibody test (IFAT), transmission electron microscopy (TEM), and immunohistochemistry (IHC). The brains of the three cattle were subjected to histopathological examination. A pathogenicity study was conducted in 120 BALB/c mice. RESULTS: Neospora caninum tachyzoites were isolated from all three cases and first observed in vitro between 14 and 17 days post-inoculation. Parasites were sub-cultured and maintained in Vero cell culture for more than 6 months. PCR products were generated for all three isolates, using two different primers. Sequencing of the PCR products and a subsequent BLAST search identified the isolates as N. caninum. In addition, the isolates tested positive using IFAT and IHC, and ultrastructure revealed by TEM was characteristic of N. caninum. Histopathological examination revealed lesions characteristic of N. caninum in 1/3 brains. In the pathogenicity study using BALB/c mice, the mortality rate was 3-7%. CONCLUSION: This was the first successful isolation of N. caninum in New Zealand confirmed using molecular characterisation tests.     

Congenital Neospora caninum infection in dairy cattle and associated calfhood mortality.

A prospective cohort study was undertaken on two central California dairies, A and B, to estimate prevalence of congenital infection with Neospora caninum, to characterize temporal variation in prevalence, to determine if occurrence of congenital infection was associated with specific dam and calf attributes, and to estimate the effect of congenital infection on calfhood mortality. Of the 405 calves enrolled over a period of 2 1/2 y on dairy A and dairy B, 30.6% (85/278) and 53.5% (68/127), respectively, were seropositive precolostrally to N. caninum, as determined by an ELISA test. Adult cow seroprevalence at calving was 36.0% (82/228) for dairy A, and 57.9% (33/57) for dairy B. No evidence was found for a significant increasing or decreasing trend in adult and precolostral seroprevalence through the study period (P > or = 0.26). For both herds combined, 81% of seropositive cows (93/115) and 5% of seronegative cows (8/170) had congenitally infected calves. Seroprevalence did not increase with cow age on either dairy (P > or = 0.47). The probability of a calf being congenitally infected was not associated with dam age, dam lactation number, dam history of abortion, calf gender, or length of gestation (P > or = 0.11). High dam ELISA values at calving were significantly associated (P < or = 0.001) with an increased probability of congenital infection in her calf. Results of survival analyses of female calves available for follow-up indicated a consistently greater survivorship to 90 d in congenitally infected calves than in noninfected calves on both dairies, which was significant for dairy A (P = 0.07, n = 186) but not for dairy B (P = 0.69, n = 72), thus indicating that congenital infection does not necessarily have a detrimental effect on calf health. The findings of a similar magnitude in congenital infection rate and adult cow prevalence, the lack of increasing seroprevalence with cow age, the lack of an effect of dam age on precolostral seropositivity, and the constant seroprevalences during the study period, suggest that, in the two dairies studied, congenital transmission constituted a substantial amount of infection and was likely the major mode of transmission of N. caninum.     

Isolation and molecular characterisation of Neospora caninum in cattle in New Zealand

AIM: To isolate Neospora caninum from the brains of naturally infected cattle and use molecular techniques to characterise the isolates.

METHODS: Neospora caninum tachyzoites were isolated in Vero cell culture from the brains of a cow and two calves. The isolates were characterised using polymerase chain reaction (PCR) methods, DNA sequencing, an immunofluorescent anti-body test (IFAT), transmission electron microscopy (TEM), and immunohistochemistry (IHC). The brains of the three cattle were subjected to histopathological examination. A pathogenicity study was conducted in 120 BALB/c mice.

RESULTS: Neospora caninum tachyzoites were isolated from all three cases and first observed in vitro between 14 and 17 days post-inoculation. Parasites were sub-cultured and maintained in Vero cell culture for more than 6 months. PCR products were generated for all three isolates, using two different primers. Sequencing of the PCR products and a subsequent BLAST search identified the isolates as N. caninum. In addition, the isolates tested positive using IFAT and IHC, and ultrastructure revealed by TEM was characteristic of N. caninum. Histopathological examination revealed lesions characteristic of N. caninum in 1/3 brains. In the pathogenicity study using BALB/c mice, the mortality rate was 3–7%.

CONCLUSION: This was the first successful isolation of N. caninum in New Zealand confirmed using molecular characterisation tests.     

Seroepidemiology of Neospora caninum infection in dairy cattle herds in Mashhad area, Iran

A seroepidemiology study of Neospora caninum was conducted in Mashhad area, Khorasan province, Iran on 337 cattle from 30 dairy using ELISA. All herds had seropositive cattle and antibodies were formed in 46% of the cattle. Abortion was significantly associated with seropositivity of cattle. The highest risk of abortion was observed in the 1-2-year-old cows. Our results indicate that Neospora infection is widespread in Iran.     

Epidemiology and impact of Neospora caninum infection in three Queensland tropical dairy herds

This study investigated the epidemiology of Neospora caninum in three tropical dairy herds in North Queensland, Australia. All animals in the herds were bled, and the sera were tested by ELISA for N. caninum antibodies. Herd records were examined, and the number of calves carried to term and the number of abortions which occurred over the lifetime of each animal were recorded to determine the abortion rate for each animal. Pedigrees were constructed for two of the herds to investigate whether vertical transmission was occurring. The seroprevalence of N. caninum ranged from 23% to 34%. The abortion rate in seropositive animals was significantly (p < 0.001) higher than in seronegative animals in all three herds (12-20.1% cf. 3.6-7%). Overall, the probability of a calf being seropositive was 3.5 times higher when the dam was also seropositive than when the dam was seronegative. Subsequent selective breeding employed by one herd reduced the N. caninum seroprevalence from 23% to 5% over a 9-year period. This study shows that N. caninum infection is prevalent in North Queensland dairy cattle, and both post-natal infection and vertical transmission are common.     

Potential involvement of Neospora caninum in naturally occurring ovine abortions in New Zealand

Neospora caninum is an obligate intracellular parasite and is recognised as the leading cause of bovine abortion worldwide. Natural infection with N. caninum has been described in sheep but it has generally not been regarded as a significant cause of abortion. Recently, there have been several New Zealand cases of foetal abortions where N. caninum was detected which strongly suggested the involvement of Neospora in these abortions. However, there is minimal information about the prevalence of N. caninum infection naturally occurring in New Zealand sheep flocks and particularly its impact on reproduction success. Thus, this present study provides preliminary data on the role that Neospora is playing in ovine reproductive failure by establishing the prevalence of N. caninum antibodies and DNA in ewe blood and foetal material present in 21 New Zealand sheep farms with ongoing unexplained abortion problems and 10 farms with consistently high fertility levels. The results of this study demonstrated an overall seroprevalence of 1.4% which varied between Aborting/non-pregnant (1.8%), age-matched pregnant controls (0.6%) and high fertility (2.1%) ewes. However, despite the variation observed, there was no statistical difference between the three groups. In addition, Neospora DNA was detected by PCR in 13% of submitted foetal brains and in ewe blood from aborting/non-pregnant (6.9%), age-matched pregnant controls (3.6%) and high fertility pregnant (2.1%) ewes. When the PCR results were considered with the IFAT and IDEXX ELISA results, there was no correlation between serology positive and PCR positive blood samples. Taken together, these results reveal that reliance on ELISA-based serology or PCR alone may underestimate the involvement of Neospora. Furthermore, determining the involvement of Neospora appears to require a multi-facetted approach where diagnostic methods and serological cut-off values may need to be adjusted as further information about the effect of natural infections with N. caninum in the ovine host is elucidated.     

Transplacental Neospora caninum infection in dogs

A Beagle bitch was inoculated SC and IM with 1.5 million Neospora caninum tachyzoites on the 35th day of gestation. Eight pups were born alive 28 days after N caninum inoculation of the bitch. Pup 1 did not breathe and died while still enclosed in fetal membranes. Pup 2 died 2 days after birth (DAB). Pups 3, 4, and 5 were euthanatized on 2, 3, and 20 DAB, respectively, because they were hypothermic and not nursing. Pups 6, 7, and 8 remained clinically normal. Indirect fluorescent N caninum serum antibody titers were: less than 50 (pups 1 and 8 at 17 DAB, and the bitch before inoculation), 50 (pups 2 and 3 on 2 DAB), 200 (pups 6 and 7 on 17 DAB), and 800 (bitch on day 17 after parturition). Neospora caninum was recovered in cultured cells inoculated with placenta and tissues from all 5 pups necropsied, and N caninum was seen in histologic sections of the heart of pup 5. Results indicate that N caninum can be transplacentally transmitted in dogs.     

Neospora caninum infection in three dogs

Infection with Neospora caninum in three young dogs is described. The predominant clinical signs were lower motor neuron deficits of the pelvic limbs, bladder and rectum. In two cases there was liver infection and dysfunction. The younger dogs had an acute onset rapidly progressive syndrome. The older dog had a similar but more chronic course. The diagnosis was confirmed by an immunofluorescence antibody test. The parasite is sensitive to clindamycin and trimethoprim/sulphonamide preparations, however the prognosis for return to function is poor especially if muscle contracture has occurred.     

Serological survey of Neospora caninum infection in dairy cattle herds in Aguascalientes,Mexico

A serological survey for antibody activity to Neospora caninum was carried out in Aguascalientes, a state in the central part of Mexico. One-hundred and eighty-seven serum samples from 13 dairy herds were tested by the ELISA test. The herd prevalence was 100% and the overall prevalence was 59% (n=110). Seventy-six of 97 seropositive cows had previous records of abortion. There was a statistically significant difference between the groups (P<0.05). However, the odds ratio was 1.4, suggesting an association between abortion and seropositivity. Neosporosis in dairy cattle appears to be widespread in Mexico, warranting more epidemiological studies to determine the distribution of the causative protozoan.