Communicating without words: Measuring nonverbal communication between veterinarians and farmers during routine herd health consultations.

Uptake of advice and the ability to facilitate change on-farm are key elements for successful veterinary practice. However, having the necessary clinical skills and knowledge is not enough to achieve this: effective communication skills are essential for veterinarians to realize their advisory role by exploring and understanding the farmer's worldview. Research of verbal aspects of veterinarian communication supports the use of a relationship-centered communication style; we next need to study how veterinarian-farmer nonverbal communication (NVC) can influence interactions and their outcomes, which has been examined in medical and companion animal practice. In this study, we considered which aspects of NVC should be measured, and how, to provide an essential first step toward understanding the significance of NVC for veterinarians working in dairy practice, which should be of interest to researchers, veterinary educators, and practitioners. Eleven video recordings of routine consultations in the UK were analyzed for farmer and veterinarian NVC. The NVC attributes with established links to positive patient and client outcomes from medical and social science studies were chosen, and a methodology developed for their measurement, by adapting measures typically used in NVC research. Each consultation was segmented into intervals defined by the main activity and location on farm: introduction, fertility examination, discussion, and closing. This approach allowed us to analyze the content more consistently, establish which aspects of NVC featured within each interval, and whether the activity and location influenced the observed NVC. We measured 12 NVC attributes, including body orientation, interpersonal distance, head position, and body lean, which have been shown to influence empathy, rapport, and trust: key components of relationship-centered communication. Future research should seek to establish the significance of NVC in effective communication between veterinarian and farmer, building on our findings that show it is possible to measure nonverbal attributes. Veterinarians may benefit from becoming skilled nonverbal communicators and have more effective conversations during routine consultations, motivating farmers to make changes and improve herd health.

Public opinion and perception of rosé veal in the UK.

Rosé veal is understood as meat from male purebred dairy calves. The UK veal market is limited, and calves are often killed at birth due to lack of purpose. A questionnaire, completed by 1002 respondents, investigated the UK public knowledge, perceptions and opinions of rosé veal, and whether raising awareness would sustain the rosé veal market. Most respondents (66%) did not eat rosé veal, mainly due to limited availability or exposure (31%) and animal welfare concerns (17%). A third of respondents knew the differences between rosé and white veal. After defining this, 61% of respondents would eat rosé veal, compared to 23% of respondents who ate it beforehand. Most respondents (91%) were willing to pay more for rosé veal burgers than the proposed average price for beef burgers. Increasing availability is key to encourage the sustainability of rosé veal production in the UK. These findings are useful for dairy farmers to strategically promote the use of purebred dairy bull calves.

Application of multiple behaviour change models to identify determinants of farmers' biosecurity attitudes and behaviours.

It has been recognised that few cattle farmers undertake biosecurity practices on their farms. Approaches that take into consideration individuals' preparedness for change, alongside beliefs thought to motivate the enactment of certain behaviours, may provide a framework for actuating tangible change. The aim of this study was to use a combination of behaviour change models to link beliefs with behaviour and identify possible key interventions to improve the uptake of biosecurity measures by dairy cattle farmers in Great Britain (GB). This is the first study to explore farmers' practices and attitudes in relation to the prevention of direct (animal to animal contact); indirect (via fomites); and other biosecurity measures using a multitheory approach. A cross-sectional study was carried out, with postal questionnaires sent to 2505 dairy cattle farmers. Questions were asked about the extent to which a host of biosecurity measures were used, the influence of various stakeholders (e.g. veterinarians, industry bodies) in informing biosecurity choices, and the perceived control farmers felt they had over biosecurity on their farms. Farmer attitudes towards biosecurity were also explored. Two behaviour change models, the Transtheoretical Model, and the Theory of Planned Behaviour, were utilised. A variety of analysis methods were used to interrogate the data, including multivariable logistic regression. A total of 908/2505 (36.2%) farmers responded, with 757 responses (30.2%) deemed eligible for inclusion. Farmers generally fell into one of two categories: those that reported not applying biosecurity measures with no intention of doing so in the future, and those that reported undertaking biosecurity measures for some time. Farmers felt that biosecurity improved cattle health and welfare, but also felt that disease was inevitable. More farmers agreed with statements relating to their ability to control, rather than prevent disease. Analysis suggested a difference between influencing beliefs and whether specific types of measure were more likely to be undertaken. For example, farmers' beliefs about other stakeholders appeared to play a role in influencing the utilisation of measures preventing direct contact (e.g. nose to nose contact), rather than indirect contact (e.g. fomite transmission). The use of a combination of behaviour change models has identified key variables to use for interventional approaches targeted towards the different type of biosecurity measure (preventing direct or indirect transmission) to improve the uptake of biosecurity on dairy cattle farms in GB. Other industry stakeholders should be aware of these variables when working with farmers to achieve optimal cattle herd health.

Injection-site lesion prevalence and potential risk factors in UK beef cattle.

Injectable veterinary medicinal products (VMPs) are widely used in cattle in the UK, and in particular vaccines are often used on large numbers of animals in the herd. The formation of injection-site lesions (ISLs) is a risk when using injectable products and has potential consequences for meat quality, animal welfare and beef industry income. This study used carcase observation in four abattoirs in England to determine ISL prevalence in beef cattle. Additionally, a questionnaire survey was used to investigate vaccination technique among UK beef farmers. The ISL prevalence was 4.1 per cent (95 per cent CI 3.4 per cent to 4.9 per cent). A potential difference between sites being used for vaccination and the distribution of ISLs on carcases suggested that factors other than vaccination were contributing to ISL incidence. Questionnaire responses highlighted deficits in good vaccination practices such as using the recommended site of injection and needle hygiene. The role of the veterinarian in knowledge transfer is crucial in providing practical injection advice when prescribing vaccines and other VMPs. This study identified factors to address when aiming to reduce ISL formation in UK beef animals.

Factors influencing veterinary surgeons' decision-making about dairy cattle vaccination.

The use of vaccines in the cattle industry is widespread; however, there is limited published guidance for use by decision-makers such as farmers and vets. To best support vets in advising dairy farmers on the optimisation of vaccination strategies, it is important to understand how and why vets make decisions about recommending the vaccination of cattle. The objective of this study was to explore in-depth farm animal vets' motivators and barriers to the implementation of vaccination strategies on British dairy farms. Semistructured interviews were conducted with 15 vets. Results indicated that vets have a positive attitude towards the use of vaccination and have few barriers to advising its implementation. Vets appear to group farmers into three 'character types'. This characterisation influences the vet-farmer relationship and how the vet communicates with the farmer. Vets required evidence of disease or a risk of disease as a motivator to advise vaccination. However, this seemed to be sometimes overruled by a risk-averse attitude; resulting in vaccination being advised 'just in case'. Crucially, the need for resources to support and build on the vet-farmer relationship is highlighted as an area requiring further exploration in order to optimise vaccination strategies on farm.

Attitudes of farm consultants in the South Island of New Zealand towards the role of veterinarians in farm businesses.

AIMS: To assess the attitude of dairy farm consultants to the role of veterinarians on dairy farms and whether they viewed veterinary advice as impartial, free from commercial bias and central to farm productivity and profitability. METHODS: A telephone survey was commissioned in September 2014 of 36 full-time farm consultants in the Canterbury and Otago region of New Zealand using a pre-prepared questionnaire to explore their attitudes towards the veterinary profession. Of those approached, 25 completed the survey. RESULTS: The consultants surveyed agreed or strongly agreed (21/23) that they were the custodians of animal health expenditure on farms, which was seen as an expense to be controlled (19/25 agreed or strongly agreed). This view was held more strongly (9/9 agreed or strongly agreed) by consultants with >5 years' professional experience, compared to 10/16 consultants with ≤5 years' experience. Most consultants (24/25) disagreed that they did not respect veterinary advice, but agreed (24/25) that veterinarians almost always had a vested interest in the advice that they gave. The role of veterinarians was seen by respondents to be mainly treatment of sick animals (22/25) and provision of animal health products (24/25), but not in maximising farm profitability (selected by 8/25 respondents). Consultants viewed their own roles as providing advice on the influence of animal health on farm profitability (21/25), nutrition (22/25) and reproduction (20/25). Most respondents (21/25) stated that reducing disease and animal health issues was important, but they (21/25) also identified a reduction in farm animal health expenditure as a measure of success. Only 7/25 consultants felt that they should work in partnership with veterinarians. CONCLUSIONS: This was a small scale study from one region of New Zealand but it indicates that consultants are ambivalent about the role and value of working more closely with veterinarians, and about the benefit that veterinary intervention may bring to a farm's profitability. CLINICAL RELEVANCE: More successful veterinary involvement on dairy farms will follow from a better understanding of how other rural professionals are involved in the farm's management and in meeting the individual farmer's goals and motivations.

Farmers' perception of the role of veterinary surgeons in vaccination strategies on British dairy farms.

There is limited research investigating the motivators and barriers to vaccinating dairy cattle. Veterinary surgeons have been identified as important sources of information for farmers making vaccination and disease control decisions, as well as being farmers' preferred vaccine suppliers. Vets' perception of their own role and communication style can be at odds with farmers' reported preferences. The objective of this study was to investigate how dairy farmers perceived the role of vets in implementing vaccination strategies on their farm. Semi-structured interviews were conducted with 24 dairy farmers from across Britain. The data were analysed using thematic analysis. Analysis revealed that farmers perceive vets to have an important role in facilitating decision-making in all aspects of vaccination, including the aspects of vaccine distribution and advice on implementation. This important role is acknowledged by farmers who have regular veterinary contact, but also farmers with solely emergency veterinary contact. Given this finding, future work should investigate the attitudes of vets towards vaccination and how they perceive their role. Combining this knowledge will enable optimisation of vaccination strategies on British dairy farms.

Quantifying veterinarians' beliefs on disease control and exploring the effect of new evidence: a Bayesian approach.

The clinical beliefs (expectations and demands) of veterinarians regarding herd-level strategies to control mastitis, lameness, and Johne's disease were quantified in a numerical format; 94 veterinarians working in England (UK) were randomly selected and, during interviews, a statistical technique called probabilistic elicitation was used to capture their clinical expectations as probability distributions. The results revealed that markedly different clinical expectations existed for all 3 diseases, and many pairs of veterinarians had expectations with nonoverlapping 95% Bayesian credible intervals. For example, for a 3-yr lameness intervention, the most pessimistic veterinarian was centered at an 11% population mean reduction in lameness prevalence (95% credible interval: 0-21%); the most enthusiastic veterinarian was centered at a 58% reduction (95% credible interval: 38-78%). This suggests that a major change in beliefs would be required to achieve clinical agreement. Veterinarians' clinical expectations were used as priors in Bayesian models where they were combined with synthetic data (from randomized clinical trials of different sizes) to explore the effect of new evidence on current clinical opinion. The mathematical models make predictions based on the assumption that veterinarians will update their beliefs logically. For example, for the lameness intervention, a 200-farm clinical trial that estimated a 30% mean reduction in lameness prevalence was predicted to be reasonably convincing to the most pessimist veterinarian; that is, in light of this data, they were predicted to believe there would be a 0.92 probability of exceeding the median clinical demand of this sample of veterinarians, which was a 20% mean reduction in lameness. Currently, controversy exists over the extent to which veterinarians update their beliefs logically, and further research on this is needed. This study has demonstrated that probabilistic elicitation and a Bayesian framework are useful for evaluating the diversity and strength of veterinarians' clinical beliefs. The wide variations observed have implications for designing future projects. Although many factors influence disease control, nonetheless the heterogeneity in beliefs also raises concern over the extent to which a broadly consistent approach is currently being achieved; it supports the argument for more randomized clinical trials and for national programs to control nonstatutory endemic diseases.

Association between somatic cell count during the first lactation and the cumulative milk yield of cows in Irish dairy herds.

Reduced potential milk yield is an important component of mastitis costs in dairy cows. The first aim of this study was to assess associations between somatic cell count (SCC) during the first lactation, and cumulative milk yield over the first lactation and subsequent lifetime of cows in Irish dairy herds. The second aim was to assess the association between SCC at 5 to 30d in milk during parity 1 (SCC1), and SCC over the entire first lactation for cows in Irish dairy herds. The data set studied included records from 51,483 cows in 5,900 herds. Somatic cell count throughout the first lactation was summarized using the geometric mean and variance of SCC. Data were analyzed using linear models that included random effects to account for the lack of independence between observations, and herd-level variation in coefficients. Models were developed in a Bayesian framework and parameters were estimated from 10,000 Markov chain Monte Carlo simulations. The final models were a good fit to the data. A 1-unit increase in mean natural logarithm SCC over the first lactation was associated with a median decrease in first lactation and lifetime milk yield of 135 and 1,663kg, respectively. A 1-unit increase in the variance of natural logarithm SCC over the first lactation was associated with a median decrease in lifetime milk yield of 719kg. To demonstrate the context of lifetime milk yield results, microsimulation was used to model the trajectory of individual cows and evaluate the expected outcomes for particular changes in herd-level geometric mean SCC over the first lactation. A 75% certainty of savings of at least €199/heifer in the herd was detected if herd-level geometric mean SCC over the first lactation was reduced from ≥120,000 to ≤72,000cells/mL. The association between SCC1 and SCC over the remainder of the first lactation was highly herd dependent, indicating that control measures for heifer mastitis should be preferentially targeted on an individual-herd basis toward either the pre- and peripartum period, or the lactating period, to optimize the lifetime milk yield of dairy cows.

A questionnaire-based survey on the uptake and use of cattle vaccines in the UK.

BACKGROUND: Vaccination is a widely used strategy for disease control in cattle in the UK and abroad. However, there has been limited research describing the uptake and use of cattle vaccines on UK farms. AIM: To describe the current uptake and usage of cattle vaccines in the UK. DESIGN: A questionnaire, available in paper and online format, was distributed to cattle farmers by convenience sampling. PARTICIPANTS: All UK cattle farmers were eligible to participate in the study. RESULTS: Eighty-six per cent of respondents (n=229/266) had vaccinated their cattle in the past year. Diseases most commonly vaccinated against were Bovine Viral Diarrhoea, Leptospirosis and Infectious Bovine Rhinotracheitis. Vaccination compliance was limited in certain areas, for example only 48 per cent of respondents stated that they administered the second dose in the primary course within the recommended timeframe, and 14 per cent of respondents stated that they vaccinated earlier than the youngest recommended age. Although outside the scope of this study, further work is needed to establish the extent of inadequate compliance and the effect this has on vaccine efficacy. The role of the veterinarian was highlighted as the main supplier of vaccines and preferred source of vaccination information. Respondents preferred to receive recommendations regarding vaccination by face-to-face communication with the veterinarian. CONCLUSIONS: The results provide a description of the current uptake and usage of cattle vaccines in the UK. Uptake is generally high but there are areas of usage of vaccines which could be improved upon. The veterinarian plays a key role as supplier of vaccines and a source of information for the majority of farmers. Although outside the scope of this study, further work is needed to establish the extent of inadequate compliance and the effect this has on vaccine efficacy. Although the respondents in this study represent a biased population of farmers, the findings indicate areas for future investigation in order to improve vaccination strategies in cattle in the UK.

Bayesian evaluation of budgets for endemic disease control: An example using management changes to reduce milk somatic cell count early in the first lactation of Irish dairy cows.

The aim of this research was to determine budgets for specific management interventions to control heifer mastitis in Irish dairy herds as an example of evidence synthesis and 1-step Bayesian micro-simulation in a veterinary context. Budgets were determined for different decision makers based on their willingness to pay. Reducing the prevalence of heifers with a high milk somatic cell count (SCC) early in the first lactation could be achieved through herd level management interventions for pre- and peri-partum heifers, however the cost effectiveness of these interventions is unknown. A synthesis of multiple sources of evidence, accounting for variability and uncertainty in the available data is invaluable to inform decision makers around likely economic outcomes of investing in disease control measures. One analytical approach to this is Bayesian micro-simulation, where the trajectory of different individuals undergoing specific interventions is simulated. The classic micro-simulation framework was extended to encompass synthesis of evidence from 2 separate statistical models and previous research, with the outcome for an individual cow or herd assessed in terms of changes in lifetime milk yield, disposal risk, and likely financial returns conditional on the interventions being simultaneously applied. The 3 interventions tested were storage of bedding inside, decreasing transition yard stocking density, and spreading of bedding evenly in the calving area. Budgets for the interventions were determined based on the minimum expected return on investment, and the probability of the desired outcome. Budgets for interventions to control heifer mastitis were highly dependent on the decision maker's willingness to pay, and hence minimum expected return on investment. Understanding the requirements of decision makers and their rational spending limits would be useful for the development of specific interventions for particular farms to control heifer mastitis, and other endemic diseases.

Association between somatic cell count after first parturition and cumulative milk yield in dairy cows.

The aim was to assess the association between the somatic cell count of parity 1 cows between 5 and 30 days in milk (SCC1), and subsequent cumulative milk yield over approximately two years for cows in English and Welsh dairy herds. The dataset included records from 43,461 cows in 2111 herds, from 2004 to 2006. Cumulative milk yield was the model outcome, and a random effect was included to account for variation between herds. The model fitted the data well and was used to make predictions of cumulative milk yield, based on SCC1. A unit increase in the natural logarithm of SCC1/1000 was associated with a median decrease in cumulative milk yield of 482 kg, over a median study period of 868 days.

Proactive dairy cattle disease control in the UK: veterinary surgeons' involvement and associated characteristics.

Characteristics of 94 veterinary surgeons associated with delivering preventive herd-level strategies to control mastitis, lameness and Johne's disease were investigated using two multinomial models. The response variables were 'Gold Standard Monitoring' (including on-going data analysis, risk assessments and laboratory testing), and a lower level of involvement called 'Regular Control Advice'. Although the sample was biased towards those who spend the majority of their time with dairy cows, 69 per cent currently had no involvement in Gold Standard Monitoring for lameness, 60 per cent no involvement with Johne's, and 52 per cent no involvement with mastitis. The final model predicted that an assistant without a postgraduate cattle qualification, who had spent no time on dairy cattle continuous professional development (CPD) in the last year, had an 88 per cent chance of having no involvement with Gold Standard Monitoring for any disease, versus <5 per cent chance for a CPD 'enriched' partner with a postgraduate cattle qualification; there was <1 per cent chance this assistant would be involved with Gold Standard Monitoring of all three diseases on one or more farms, versus a 58 per cent chance for this partner. CPD and employment status were also associated with markedly different probabilities for delivering Regular Control Advice. Increased postgraduate education may further veterinary involvement of this nature.

Association between somatic cell count early in the first lactation and the lifetime milk yield of cows in Irish dairy herds.

Change in lifetime milk yield is an important component of the cost of diseases in dairy cows. Knowledge of the likelihood and scale of potential savings through disease prevention measures is important to evaluate how much expenditure on control measures is rational. The aim of this study was to assess the association between somatic cell count (SCC) at 5 to 30 d in milk during parity 1 (SCC1), and lifetime milk yield for cows in Irish dairy herds. The data set studied included records from 53,652 cows in 5,922 Irish herds. This was split into 2 samples of 2,500 and 3,422 herds at random. Linear models with lifetime milk yield and first-lactation milk yield as the outcomes and random effects to account for variation between herds were fitted to the data for the first sample of herds; data for the second sample were used for cross-validation. The models were developed in a Bayesian framework to include all uncertainty in posterior predictions and parameters were estimated from 10,000 Markov chain Monte Carlo simulations. The final model was a good fit to the data and appeared generalizable to other Irish herds. A unit increase in the natural logarithm of SCC1 was associated with a median decrease in lifetime milk yield of 864 kg, and a median decrease in first-lactation milk yield of 105 kg. To clarify the meaning of the results in context, microsimulation was used to model the trajectory of individual cows, and evaluate the expected outcomes for particular changes in the herd-level prevalence of cows with SCC1 ≥ 400,000 cells/mL. Differences in mean lifetime milk yield associated with these changes were multiplied by an estimated gross margin for each cow to give the potential difference in milk revenue. Results were presented as probabilities of savings; for example, a 75% probability of savings of at least€97 or€115/heifer calved into the herd existed if the prevalence of cows with SCC1 ≥ 400,000 cells/mL was reduced from ≥ 20 to <10 or <5%, respectively, and at least€71/heifer calved into the herd if the prevalence of cows with SCC1 ≥ 400,000 cells/mL was reduced from ≥ 10 to <5%. The results indicate large differences in lifetime milk yield, depending on SCC early in the first lactation and the findings can be used to assess where specific interventions to control heifer mastitis prepartum are likely to be cost effective.

Association between somatic cell count early in the first lactation and the longevity of Irish dairy cows.

Reduced longevity of cows is an important component of mastitis costs, and increased somatic cell count (SCC) early in the first lactation has been reported to increase culling risk throughout the first lactation. Generally, cows must survive beyond the first lactation to break even on their rearing costs. The aim of this research was to assess the association between SCC of primiparous cows at 5 to 30 days in milk (SCC1), and survival over a 5-y period for cows in Irish dairy herds. The data set used for model development was based on 147,458 test day records from 7,537 cows in 812 herds. Cows were censored at their last recording if identified at a later date in other herds or if recorded at the last available test date for their herd, otherwise, date of disposal was taken to be at the last test date for each cow. Survival time was calculated as the number of days between the dates of first calving and the last recording, which was split into 50-d intervals. Data were analyzed in discrete time logistic survival models that accounted for clustering of 50-d intervals within cows, and cows within herds. Models were fitted in a Bayesian framework using Markov chain Monte Carlo simulations. Model fit was assessed by comparison of posterior predictions to the observed disposal risk for cows aggregated by parameters in the model. Model usefulness was assessed by cross validation in a separate data set, which contained 144,113 records from 7,353 cows in 808 herds, and posterior predictions were compared with the observed disposal risk for cows aggregated by parameters of biological importance. Disposal odds increased by a factor of 5% per unit increase in ln SCC1. Despite this, posterior predictive distributions revealed that the probability of reducing replacement costs by >€10 per heifer calved, through decreasing the herd level prevalence of cows with SCC1 ≥ 400,000 cells/mL (from an initial prevalence of ≥ 20 to <10%) only exceeded 50% for less than 1 in 5 Irish herds. These results indicate that the effect of a reduction in the prevalence of cows with SCC1 ≥ 400,000 cells/mL on replacement costs alone for most Irish dairy herds is small, and future research should investigate other potential losses, such as the effect of SCC1 on lifetime milk yield.

Opinions and practices of veterinarians and dairy farmers towards herd health management in the UK.

The objective was to compare farm veterinary surgeons' and dairy farmers' opinions on herd health plans and herd health and production management with the aim of discovering and better understanding the differences. Two comparable questionnaires, one for farm veterinarians and one for dairy farmers, were distributed throughout the UK. While listing the 'major roles' of the veterinarian on the farm, veterinarians considered 'optimising milk production', 'decreasing overall cost' and 'being an independent adviser' as important roles, but these were not seem to be perceived as such by the farmers. In addition, when presenting themselves to clients, veterinarians seemed to favour the 'friend of the farmer' approach; a much smaller proportion of farmers seemed to prefer this approach. The majority of farm respondents (98 of 121; 81 per cent) valued the discussions with their veterinarian, and it was apparent from the relatively small proportion of veterinarians instigating a discussion on farm (33 of 125; 26 per cent) that there is the opportunity for a more proactive approach from veterinarians. The study underlines that 'demonstrating cost-effectiveness' is still a main concern for veterinarians and farmers and identifies areas that can be improved by more training and effective communication.

Evaluation of four serological techniques to determine the seroprevalence of Neospora caninum in foxes (Vulpes vulpes) and coyotes (Canis latrans) on Prince Edward Island, Canada.

The objectives of this study were (1) to evaluate the performance and agreement of serological assays (ELISA, IFAT, Neospora caninum agglutination test and immunoblot) using reference sera and field sera from foxes and coyotes and (2) to estimate the N. caninum seroprevalence in foxes and coyotes on Prince Edward Island, Canada. With fox and coyote reference sera the test performance of the ELISA, IFAT and IB was excellent (100% sensitivity and specificity). NAT showed a low sensitivity (50%). Serum was collected from 201 coyotes and 271 foxes. The seroprevalence observed in the different assays ranged from 0.5 to 14.0% in coyotes and 1.1 to 34.8% in foxes. The seroprevalence, when taking more than one test positive as cut-off value was 3.3 and 1.1% for coyotes and foxes, respectively. From the N. caninum-positive group, all coyotes were older than 3 years. Agreement among assays (measured as prevalence-adjusted bias-adjusted kappa) using the field sera ranged from 0.17 to 0.97. Best agreement was observed between ELISA and IFAT, poor agreement was observed between NAT and the other assays. Positive agreement was moderate to poor among all assays utilized in this study. Although the seroprevalence observed was low, N. caninum antibodies are present in foxes and coyotes on Prince Edward Island (PEI) and their role in the N. caninum epidemiology needs further study.

Comparison of serological methods for the diagnosis of Neospora caninum infection in cattle.

The aims of this study were to evaluate the performance and agreement of various commercial and in-house Neospora caninum antibody assays used in dairy cattle in North America, and to investigate reproducibility of two assays performed in different laboratories. From 1998 to 2005, three enzyme linked immunosorbent assays (ELISAs, a competitive ELISA-VMRD Inc., an indirect ELISA-Biovet Inc., and another indirect ELISA-Herdchek IDEXX Corp.), two indirect fluorescent antibody tests (IFATs, VMRD Inc., and in-house USDA) and one N. caninum agglutination test (NAT, in-house USDA) were utilized to test 397 randomly selected dairy cattle serum samples from 34 herds in eastern Canada for antibodies to N. caninum. The manufacturers' recommended cut-off values were used to evaluate test performance and agreement between tests. One IFAT (VMRD Inc.) performed well (sensitivity and specificity: 0.97 and 0.97, respectively) using reference sera (n = 452), therefore, results from this IFAT on the 397 samples could subsequently be used as the reference standard to calculate test characteristics for the other assays. Only 11% of the 397 sera were found to be N. caninum-positive with the IFAT. Prevalence-adjusted bias-adjusted kappa (PABAK) ranged from 0.06 to 0.99. Positive agreement was moderate to very good (P(pos) = 0.25-0.96). Negative agreement was very good for all assays (P(neg) > 0.94) except NAT (P(neg) = 0.66). Sensitivity was > or =0.89 for all assays except the NAT, which had a significantly lower sensitivity (0.66). Specificity was high (>0.94) for all assays except for one indirect ELISA (specificity = 0.52). This indirect ELISA did not perform satisfactorily when used in 1998, but an improved version of the ELISA performed as one of the best assays in 2004. Reproducibility of the competitive ELISA was excellent, but the reproducibility of the indirect ELISA that was improved was low (concordance correlation coefficient = 0.90 and 0.36, respectively). The performance characteristics observed for most assays in this study make them useful for screening antibodies to N. caninum in cattle.

HPLC purification of recombinant NcGRA6 antigen improves enzyme-linked immunosorbent assay for serodiagnosis of bovine neosporosis.

The gene for a dense granule protein (NcGRA6) of Neospora caninum was expressed in Escherichia coli as a His-tag fusion protein and purified by NiNTA affinity chromatography. In a preliminary study, high binding of antibodies from N. caninum-negative cows was observed in enzyme-linked immunosorbent assay (ELISA) using NiNTA-purified NcGRA6. Analysis of NiNTA eluates revealed a significant number of E. coli proteins that co-purified with recombinant NcGRA6. In an attempt to improve the relative sensitivity and specificity of the NcGRA6-based ELISA, the rNcGRA6 eluates were subjected to a secondary purification using reverse phase-high performance liquid chromatography (RP-HPLC). Analysis of RP-HPLC eluates by SDS-PAGE/silver staining revealed the purification of recombinant NcGRA6 from contaminating E. coli proteins. ELISAs using the RP-HPLC purified NcGRA6 (dELISA) or singly purified NcGRA6 (sELISA) for identifying seropositive and seronegative cows in a beef herd experiencing an epidemic outbreak of neosporosis were compared to standard assays based on native tachyzoite protein-immunofluorescence antibody test, immunoblot assay, and ISCOM-ELISA. The relative sensitivity, specificity, and kappa value of the NcGRA6d-ELISA were greatly improved over the NcGRA6s-ELISA when compared to the three native antigen immunoassays. These results indicate that removal of contaminating E. coli proteins improves the performance of recombinant NcGRA6 ELISA in diagnosing bovine neosporosis, and may have applicability to the use of recombinant proteins in diagnosing other infectious agents.