Bovine Neosporosis: The Development of Evidence-Based Control Systems

Final report summary: Neospora caninum is a newly recognised protozoan parasite of dogs and cattle, first described in 1988. Now recognised as one of the major infectious cause of abortion in dairy cattle, it is known that cattle can be infected transplacentally or by ingestion of oocysts excreted by dogs. However, very little is known about the frequency and intensity of oocyst shedding by dogs or the infectivity and clinical consequences of oocyst infection in cattle. The importance of environmental infection from dogs to cattle, is a critical question since it has profound implications for control. Environmental contamination with N. caninum oocysts also has potential implications for humans, Given its relationship to the ubiquitous and common zoonotic protozoan Toxoplasma gondii, and the fact that N. caninum will infect primates and grow in human cells in vitro, there is concern to determine if N. caninum is infecting humans with, as yet, unidentified consequences. Ingestion of oocysts, as well as with parasite stages in undercooked beef, are the two major potential routes of human infection.
This project has addressed these key questions about N. caninum and has successfully achieved its major objectives to determine the zoonotic potential of N. caninum and to elucidate the significance of oocyst infection for cattle. <P>

In collaboration with the Health Protection Agency, the most comprehensive serological survey of a human population yet undertaken was conducted using over 3200 samples randomly drawn from anonymised specimens accumulated by the Public Health Laboratories services (PHLS) and representing the general population in England, together with over 500 samples from a targeted high-risk group (farm workers, participants in the PHLS Farm Cohort). Using a quantitative, high-throughput inhibition ELISA, and an Indirect Immunofluorescence Antibody test (IFAT) as a confirmatory assay, none of the 3750 individual human samples tested gave evidence of exposure to N. caninum. It can be concluded that in the UK there is no evidence of human infection from this study. Given the sample size and the population size from which it was drawn, the maximum possible number of exposed persons in England which could exist is fewer than one per thousand. <P>
A major and logistically challenging experiment was conducted to determine the effect of N. caninum oocysts infection on pregnant cattle. Whilst it has been presumed that oocyst infection might cause the abortion ‘storms’ (epidemic outbreaks) characteristic of N. caninum, until this project began almost no oocyst infections of pregnant cattle had been conducted and abortion had not been provoked. This is due to the substantial logistical problems in conducting such experiments because of difficulties of conducting dog infection (to get oocysts) and the extremely low numbers of oocysts produced. Accordingly, we collaborated with Professor Milt McAllister of the University of Illinois who had first demonstrated that dogs are a definitive host of N. caninum. Using 18 cows in three groups with synchronised oestrus and artificial insemination, transplacental infection was confirmed consistently (4/5 calves infected) when cows were infected at 210 days gestation, but not at 70 days (0/6 calves infected) and in only 1/6 pregnancies at 120 days, in which case the foetus was aborted. This experiment demonstrated that oocyst infection could cause transplacental infection in mid to late pregnancy; that it could cause abortion – albeit infrequently; and that those effects could be produced by relatively low numbers of oocysts (estimated oocyst dose between 127 – 40,000 oocysts; because oocyst concentrations were too low to count viable dose was estimated by bioassay). Because it is now realised that chronic maternal infections recrudescing in pregnancy are an important feature of N. caninum in cows, we exploited the unique opportunity of having oocyst-infected cows, to investigate whether such post-natal infection led to chronic infection which could infect subsequent pregnancies. In a second experiment, seven cows shown by immunological responses to have been infected with N. caninum oocysts in their first pregnancy, were rebred. There was no evidence of spontaneous infection in their calves. <P>
The cattle experiments revealed the possible effects of oocyst consumption by cows. A third part of the project investigated the natural production of oocysts by dogs. In a cross-sectional survey of faecal samples from foxhounds, farm dogs and pet dogs, N. caninum oocysts were found in only 1/261 foxhounds, and 0/69 farm dogs and 0/105 pet dogs. Morphologically similar oocysts of Hammondia heynorni were found in 13 foxhounds, one farm dog, and one pet dog and were identified by species-specific PCR. In the N. caninum positive foxhound sample there were <100 oocysts /g faeces. <P>
In the fourth major part of the project the possible relationship of infections in dogs and cattle was investigated on 96 Cheshire dairy farms using a questionnaire for risk factors, estimates of herd prevalence from bulk-milk tank assays for N. caninum-specific antibody and determination of farm dog antibody status by N. caninum-specific IFAT, analysed by logistic regression and multivariate analysis. No significant (at p<0.05) risk factors were identified that would suggest that high herd prevalence (>6%) was associated with transmission from dogs to cattle, but a number of identified risk factors suggested dogs acquire infections from cattle. <P>
In conclusion
<UL> <LI> in the largest study yet conducted, there was no evidence of exposure to N. caninum in humans
<LI> this is consistent with very low levels of environmental contamination likely by N. caninum based on the cross-sectional faecal survey. It also indicates that there is negligible risk from consumption of meat.
<LI> low numbers of oocysts were experimentally shown to cause abortion in 1/6 cows when infection was in mid-pregnancy, and to congenitally infect 4/5 calves when infection was in late-pregnancy – but chronic infections which infected a subsequent pregnancy were not established.
<LI> in the first study of risk factors on British dairy farms, no factors were identified suggesting higher herd prevalence than normal was associated with transmission from dogs. However, there was an association between seropositivity in dogs and herd seroprevalence. Without indicating whether dogs are infected cattle or cattle infected from dogs, this result nonetheless indicates that farmers should be advised to minimise contact between dogs and cattle, to prevent access of dogs to calving membranes, and to prevent dog fouling of cattle feed. </ul>
Overall whilst this project confirmed that oocysts can infect pregnant cows and cause abortion or congenital infection the likelihood of this occurring appears low based on epidemiological risk factors and the prevalence and intensity of oocyst shedding. Control of bovine neosporosis in dairy herds should focus on identifying positive cows and removing them from the replacement breeding programme. The study suggests that oocyst infection may be most significant in causing congenital infection in heifer calves. If congenitally infected female calves are retained they may go on to endogenously infect all their progeny with an associated incidence of sporadic or endemic abortion. This possibility requires to be proven.
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