Mycobacterium Avium Ssp. Paratuberculosis Infection Research Articles

Short communication: Heritability of susceptibility to infection by Mycobacterium avium ssp. paratuberculosis in Holstein cattle

Johne's disease in cattle is the result of infection of the small intestine by Mycobacterium avium ssp. paratuberculosis (MAP), leading to an incurable inflammatory bowel disease (Johne's disease or paratuberculosis). The disease is a concern both for its direct cost to dairy producers and for its zoonotic potential. The objective of this study was to estimate the heritability for susceptibility to infection of cattle by MAP using Johne's testing records (ELISA test for presence of antibodies to MAP in milk or blood) from US Holstein cattle from 2009 to 2016. Data sets were edited to include records from herds with 100 or more total records and sires with 50 or more daughters. Data sets were further edited to include (1) only herds with at least 1 positive test, (2) herds with at least 2.5% positive test results, and (3) herds with at least 5% positive test results to examine the effect of data from herds with higher proportions of positive tests, and presumably higher pathogen exposure, on heritability estimates. Two models were used in this study, a linear sire model and a binary threshold-probit sire model. Both were mixed models considering fixed effects of herd and age at test, the latter as a covariate accounting for linear and quadratic effects; random effects included sire and residual. Analyses were conducted using a restricted maximum likelihood method. Heritability estimates (±standard error) from the linear model were 0.041 ± 0.004, 0.050 ± 0.004, and 0.062 ± 0.007 for data from herds with at least 1 positive test, ≥2.5% positive tests, and ≥5% positive tests, respectively. Heritability estimates from the threshold model were 0.157 ± 0.014, 0.174 ± 0.016, and 0.186 ± 0.021 for data from herds with at least 1 positive test, ≥2.5% positive tests, and ≥5% positive tests, respectively. Heritability estimates from the linear model were affected by population incidence for positive tests, in contrast to estimates from the threshold model, likely accounting for the difference in magnitude of heritability estimates between models and suggesting that the threshold model analysis is the better choice. Heritability estimates increased as data were restricted to herds with presumed higher MAP exposure for both linear model and threshold model analyses. These estimates are similar to previous estimates in other dairy cattle populations and suggest the potential for selection to lessen susceptibility to MAP infection.

The genetic architecture of milk ELISA scores as an indicator of Johne's disease (paratuberculosis) in dairy cattle

Johne's disease (or paratuberculosis), caused by Mycobacterium avium ssp. paratuberculosis (MAP) infection, is a globally prevalent disease with severe economic and welfare implications. With no effective treatment available, understanding the role of genetics influencing host infection status is essential to develop selection strategies to breed for increased resistance to MAP infection. The main objectives of this study were to estimate genetic parameters for the MAP-specific antibody response using milk ELISA scores in Canadian Holstein cattle as an indicator of resistance to Johne's disease, and to unravel genomic regions and candidate genes significantly associated with MAP infection. After data editing, 168,987 milk ELISA records from 2,306 herds, obtained from CanWest Dairy Herd Improvement, were used for further analyses. Variance and heritability estimates for MAP infection status were determined using univariate linear animal models under 3 scenarios: (a) SCEN1: the complete data set (all herds); (b) SCEN2: herds with at least one suspect or test-positive animal (ELISA optical density ≥0.07); and (c) SCEN3: herds with at least one test-positive animal (ELISA optical density ≥0.11). Heritability estimates were calculated as 0.066, 0.064, and 0.063 for SCEN1, SCEN2, and SCEN3, respectively. The correlations between estimated breeding values for resistance to MAP infection and other economically important traits, when significant, were favorable and of low magnitude. Genome-wide association analyses identified important genomic regions on Bos taurus autosome (BTA)1, BTA7, BTA9, BTA14, BTA15, BTA17, BTA19, and BTA25 showing significant association with MAP infection status. These regions included 2 single nucleotide polymorphisms located 2 kb upstream of positional candidate genes CD86 and WNT9B, which play key roles in host immune response and tissue homeostasis. This study revealed the genetic architecture of MAP infection in Canadian Holstein cattle as measured by milk ELISA scores by estimating genetic parameters along with the identification of genomic regions potentially influencing MAP infection status. These findings will be of significant value toward implementing genetic and genomic evaluations for resistance to MAP infection in Holstein cattle.

Association of paratuberculosis sero-status with milk production and somatic cell counts across 5 lactations, using multilevel mixed models, in dairy cows

The aim of this work was to investigate associations between individual cow Mycobacterium avium ssp. paratuberculosis (MAP) seropositivity, 305-d corrected milk production, and somatic cell count during 5 lactations lifespan in Portuguese dairy herds using multilevel mixed models. We used MAP serum ELISA (Idexx MAP Ac, Idexx Laboratories Inc., Westbrook, ME) results (n = 23,960) from all the 20,221 adult cows present in 329 farms and corresponding 47,586 lactation records from the National Dairy Improvement Association. Cows and farms were classified as positive or negative. Multilevel mixed models were used to investigate the association of cow MAP status with variation in milk production and somatic cell count. Cow MAP status, farm status, and lactation number were considered as independent variables. A quadratic function of lactation number was used to mimic the effect of lactation order on milk production. The models considered 3 levels: measurement occasion (level 1) within cow (level 2) and cow within farm (level 3). Four final models were produced, including all herds and cows, to address the effect of farm status (models 1 and 2) or the effect of cow status (models 3 and 4) on the outcome variables. Our results show that MAP status affects milk production. Losses are detectable from third lactation onward. During the first 5 lactations, positive cows accumulated an average loss of 1,284.8 kg of milk when compared with the negative cows. We also observed that somatic cell counts were higher in positive cows and a positive interaction occurs between cow status and lactation number, suggesting a positive association between MAP infection and increased somatic cell counts. Our results are in line with previous studies, suggesting a possible positive relation between cow milk production and susceptibility to MAP infection.

Short communication: Uncovering quantitative trait loci associated with resistance to Mycobacterium avium ssp. paratuberculosis infection in Holstein cattle using a high-density single nucleotide polymorphism panel

Mycobacterium avium ssp. paratuberculosis (MAP) is the etiological agent of Johne's disease in cattle. Johne's disease is a disease of significant economic, animal welfare, and public health concern around the globe. Therefore, understanding the genetic architecture of resistance to MAP infection has great relevance to advance genetic selection methods to breed more resistant animals. The objectives of this study were to perform a genome-wide association study of previously analyzed 50K genotypes now imputed to a high-density single nucleotide polymorphism panel (777K), aiming to validate previously reported associations and potentially identify additional single nucleotide polymorphisms associated with antibody response to MAP infection. A principal component regression-based genome-wide association study revealed 15 putative quantitative trait loci (QTL) associated with the MAP infection phenotype (serum or milk ELISA tests) on 9 different chromosomes (Bos taurus autosomes 5, 6, 7, 10, 14, 15, 16, 20, and 21). These results validated previous findings and identified new QTL on Bos taurus autosomes 15, 16, 20, and 21. The positional candidate genes NLRP3, IFi47, TRIM41, TNFRSF18, and TNFRSF4 lying within these QTL were identified. Further functional validation of these genes is now warranted to investigate their roles in regulating the immune response and, consequently, cattle resistance to MAP infection.

Short communication: Heritability estimates for susceptibility to Mycobacterium avium ssp. paratuberculosis infection in Chinese Holstein cattle

Paratuberculosis in ruminants, which is caused by Mycobacterium avium ssp. paratuberculosis (MAP), is a contagious, chronic enteric disease associated with economic losses, animal welfare, and health implications in dairy cattle production. In this study, we estimated the variance components and heritability of susceptibility to MAP infection in Chinese Holstein cattle. We collected 4,937 serum samples from cows in 7 dairy herds in the Beijing region of China and used the ELISA test to detect antibodies to MAP. Three statistical models were implemented to estimate heritabilities: (1) a linear model (ELISA sample-to-positive ratios as a continuous trait); (2) a binary threshold model (positive/negative from ELISA results); and (3) an ordered threshold model (ELISA results as an ordered categorical model with categories 1 to 5 corresponding to negative, uncertain, mildly positive, intermediate positive, and strongly positive). The heritability estimates ranged from 0.0389 to 0.1069, indicating that genetic factors affect MAP infection susceptibility in Chinese Holstein cattle.

An agent-based model evaluation of economic control strategies for paratuberculosis in a dairy herd

This paper uses an agent-based simulation model to estimate the costs associated with Mycobacterium avium ssp. paratuberculosis (MAP), or Johne's disease, in a milking herd, and to determine the net benefits of implementing various control strategies. The net present value (NPV) of a 1,000-cow milking herd is calculated over 20 yr, parametrized to a representative US commercial herd. The revenues of the herd are generated from sales of milk and culled animals. The costs include all variable and fixed costs necessary to operate a representative 1,000-cow milking herd. We estimate the NPV of the herd with no MAP infection, under an expected endemic infection distribution with no controls, and under an expected endemic infection distribution with various controls. The initial number of cows in a herd with an endemic MAP infection is distributed as 75% susceptible, 13% latent, 9% low MAP shedding, and 3% high MAP shedding. Control strategies include testing using ELISA and fecal culture tests and culling of cows that test positive, and culling based on observable milk production decrease. Results show that culling cows based on test results does not increase the herd's NPV and in most cases decreases NPV due to test costs as well as false positives and negatives with their associated costs (e.g., culling healthy cows and keeping infected cows). Culling consistently low producing cows when MAP is believed to be present in the herd produces higher NPV over the strategy of testing and culling MAP infected animals, and over the case of no MAP control.

In vitro bioassessment of the immunomodulatory activity of Saccharomyces cerevisiae components using bovine macrophages and Mycobacterium avium ssp. paratuberculosis

The yeast Saccharomyces cerevisiae and its components are used for the prevention and treatment of enteric disease in different species; therefore, they may also be useful for preventing Johne's disease, a chronic inflammatory bowel disease of ruminants caused by Mycobacterium avium ssp. paratuberculosis (MAP). The objective of this study was to identify potential immunomodulatory S. cerevisiae components using a bovine macrophage cell line (BOMAC). The BOMAC phagocytic activity, reactive oxygen species production, and immune-related gene (IL6, IL10, IL12p40, IL13, IL23), transforming growth factor β, ARG1, CASP1, and inducible nitric oxide synthase expression were investigated when BOMAC were cocultured with cell wall components from 4 different strains (A, B, C, and D) and 2 forms of dead yeast from strain A. The BOMAC phagocytosis of mCherry-labeled MAP was concentration-dependently attenuated when BOMAC were cocultured with yeast components for 6 h. Each yeast derivative also induced a concentration-dependent increase in BOMAC reactive oxygen species production after a 6-h exposure. In addition, BOMAC mRNA expression of the immune-related genes was investigated after 6 and 24 h of exposure to yeast components. All yeast components were found to regulate the immunomodulatory genes of BOMAC; however, the response varied among components and over time. The in vitro bioassessment studies reported here suggest that dead yeast and its cell wall components may be useful for modulating macrophage function before or during MAP infection.

P067 INVESTIGATION OF THE PRESENCE OF MYCOBACTERIUM AVIUM SSP. PARATUBERCULOSIS IN CHILDREN WITH CROHN DISEASE USING QUANTITATIVE DNA SEQUENCE-BASED APPROACHES

Mycobacterium avium ssp. paratuberculosis (MAP) has been suspected to play a role in Crohn disease (CD) pathogenesis. Prior studies of MAP detection in dairy cows have been more extensively investigated, successfully detecting the organisms by culture and PCR. These studies have shown the organism to be the causative agent in Johne’s disease, an enteritis in ruminants that is histopathologically similar to CD. We aim to use techniques which successfully detect MAP in cow feces and apply them to human feces and intestinal biopsy samples of patients with CD and healthy controls to see if MAP infection is correlated with active disease in children with CD. Using feces and intestinal biopsy samples from known MAP-positive and negative calves, we performed several Mycobacterium-specific DNA extraction methods. We used quantitative PCR and DNA-sequencing methods. We applied these methods to samples from pediatric patients with CD and non-CD controls. Two samples were taken from each patient: one from the terminal ileum, and the other from either the cecum, left colon, or rectum. Five out of the eight patients were treatment naïve at the time of sample collection. The other three received antibiotics leading up to sample collection. We correctly identified the presence of MAP in intestinal biopsies from infected dairy calves using qPCR, which aligned with culturing data from these biopsies. In our small subset of samples, we were unable to detect the presence of MAP in CD patients by any of these techniques. The detection of MAP from intestinal biopsies in calves demonstrates that we can use these techniques to quantitatively identify the number of organisms in intestinal biopsies, without the use of culture-based methods. We have confirmed the difficulty in detecting this organism in humans. Work is ongoing to improve the sensitivity of our techniques and increase the number of patient samples tested.

Evaluation of the association of SLC11A1 gene polymorphism with incidence of paratuberculosis in goats.

Paratuberculosis is one of the chronic granulomatous enteritis that predominantly affects ruminants world wide, caused by Mycobacterium avium ssp. paratuberculosis (MAP). In ruminants, microsatellite polymorphisms of the 3' untranslated region (3'UTR) of the solute carrier family 11 member A1 (SLC11A1) gene were associated with resistance to intracellular pathogen infections. This research was carried out to detect the polymorphisms in A and B regions of the 3'UTR of SLC11A1 gene and to evaluate the potential association between these polymorphisms and MAP infection in goats. MAP-specific antibodies were detected by ELISA and MAP infection was confirmed by IS900 PCR in 150 adult goats from different regions of Kerala, India. The polymorphism of microsatellite regions A and B at 3'UTR of the SLC11A1 gene was analysed in goats by an automated technique, fragment analysis, using fluorescent-tagged forward primers. Eight alleles with sizes ranging from 221 to 239 bp were found in region A. Region B revealed two alleles, 117 bp (B₇) and 119 bp (B₈). Animals with B₈ alleles were found to have higher incidence of paratuberculosis than animals with B₇ alleles (P < 0.01). There was no statistically significant association found between region A genotypes and paratuberculosis incidence. These results suggest that caprine SLC11A1 gene has significant role in paratuberculosis resistance in goats and further studies might help in development of a PCR-based genotyping test for paratuberculosis resistance and selection of superior animals for future goat breeding programmes.

Seroprevalence of Mycobacterium avium ssp. paratuberculosis in small ruminants in a flock in Antioquia, Colombia

Paratuberculosis is a chronic infectious disease caused by Mycobacterium avium ssp. paratuberculosis (MAP). This study aimed at determining the presence of antibodies against MAP, and the association between the serological status of MAP and individual factors in goats and sheep in a flock in Antioquia, Colombia. We studied fifty-three goats and 6 sheep over two years old, and took blood samples to obtain serum and information on individual factors. The presence of antibodies against MAP was determined by ELISA, whereas the information was descriptively analyzed. ELISA did not detect any positive or suspect animal among the 59 tested animals, so an association between serological status to MAP and individual factors could not be established. This is the first and the fourth study reporting the search for antibodies against MAP in a goat and in a sheep population in Colombia, respectively. Several factors may explain the serological status of the sampled animals. It is necessary to continue with similar studies in other flocks of the region and the country to determine the current MAP infection status in small ruminants in Colombia.

The effect of Mycobacterium avium ssp. paratuberculosis infection on clinical mastitis occurrence in dairy cows

Endemic diseases can be counted among the most serious sources of losses for livestock production. In dairy farms in particular, one of the most common diseases is Johne's disease, caused by Mycobacterium avium ssp. paratuberculosis (MAP). Infection with MAP causes direct costs because it affects milk production, but it has also been suspected to increase the risk of clinical mastitis (CM) among infected animals. This might contribute to further costs for farmers. We asked whether MAP infection represents a risk factor for CM and, in particular, whether CM occurrences were more common in MAP-infected animals. Our results, obtained by survival analysis, suggest that MAP-infected cows had an increased probability of experiencing CM during lactation. These results highlight the need to account for the interplay of infectious diseases and other health conditions in economic and epidemiological modeling. In this case, accounting for MAP-infected cows having an increased CM occurrence might have nonnegligible effects on the estimated benefit of MAP control.

Longitudinal relationship between fecal culture, fecal quantitative PCR, and milk ELISA in Mycobacterium avium ssp. paratuberculosis-infected cows from low-prevalence dairy herds.

Mycobacterium avium ssp. paratuberculosis (MAP), the causative agent of ruminant Johne's disease, presents a particular challenge with regard to infection mitigation on dairy farms. Diagnostic testing strategies to identify and quantify MAP and associated antibodies are imperfect, and certain facets of the relationship between diagnostic tests remain to be explored. Additional repeated-measures data from known infected animals are needed to complement the body of cross-sectional research on Johne's disease-testing methods. Statistical models that accurately account for multiple diagnostic results while adjusting for the effects of individual animals and herds over time can provide a more detailed understanding of the interplay between diagnostic outcomes. Further, test results may be considered as continuous wherever possible so as to avoid the information loss associated with dichotomization. To achieve a broader understanding of the relationship between diagnostic tests, we collected a large number of repeated fecal and milk samples from 14 infected cows, in addition to bulk milk samples, from 2 low-prevalence dairy herds in the northeast United States. Predominately through the use of mixed linear modeling, we identified strong associations between milk ELISA optical density, fecal quantitative PCR, and fecal culture in individual animals while concurrently adjusting for variables that could alter these relationships. Notably, we uncovered subtleties in the predictive abilities of fecal shedding level on milk ELISA results, with animals categorized as disease progressors reaching higher ELISA optical density levels. Moreover, we observed that spikes in fecal shedding could predict subsequent high ELISA values up to 2 mo later. We also investigated the presence of MAP in individual milk samples via PCR and noted an association between poor udder hygiene and MAP positivity in milk, suggesting some level of environmental contamination. The paucity of positive milk samples and the complete absence of detectable MAP in the bulk tank throughout the study period indicate that contamination of milk with MAP may not be of chief concern in low-prevalence herds. An enhanced understanding of the interrelationships between diagnostic tests can only benefit the development of testing strategies and objectives, which in turn may lessen MAP infection prevalence in dairy herds.

Systems Analysis of Early Host Gene Expression Provides Clues for Transient Mycobacterium avium ssp avium vs. Persistent Mycobacterium avium ssp paratuberculosis Intestinal Infections.

It has long been a quest in ruminants to understand how two very similar mycobacterial species, Mycobacterium avium ssp. paratuberculosis (MAP) and Mycobacterium avium ssp. avium (MAA) lead to either a chronic persistent infection or a rapid-transient infection, respectively. Here, we hypothesized that when the host immune response is activated by MAP or MAA, the outcome of the infection depends on the early activation of signaling molecules and host temporal gene expression. To test our hypothesis, ligated jejuno-ileal loops including Peyer’s patches in neonatal calves were inoculated with PBS, MAP, or MAA. A temporal analysis of the host transcriptome profile was conducted at several times post-infection (0.5, 1, 2, 4, 8 and 12 hours). When comparing the transcriptional responses of calves infected with the MAA versus MAP, discordant patterns of mucosal expression were clearly evident, and the numbers of unique transcripts altered were moderately less for MAA-infected tissue than were mucosal tissues infected with the MAP. To interpret these complex data, changes in the gene expression were further analyzed by dynamic Bayesian analysis. Bayesian network modeling identified mechanistic genes, gene-to-gene relationships, pathways and Gene Ontologies (GO) biological processes that are involved in specific cell activation during infection. MAP and MAA had significant different pathway perturbation at 0.5 and 12 hours post inoculation. Inverse processes were observed between MAP and MAA response for epithelial cell proliferation, negative regulation of chemotaxis, cell-cell adhesion mediated by integrin and regulation of cytokine-mediated signaling. MAP inoculated tissue had significantly lower expression of phagocytosis receptors such as mannose receptor and complement receptors. This study reveals that perturbation of genes and cellular pathways during MAP infection resulted in host evasion by mucosal membrane barrier weakening to access entry in the ileum, inhibition of Ca signaling associated with decreased phagosome-lysosome fusion as well as phagocytosis inhibition, bias toward Th2 cell immune response accompanied by cell recruitment, cell proliferation and cell differentiation; leading to persistent infection. Contrarily, MAA infection was related to cellular responses associated with activation of molecular pathways that release chemicals and cytokines involved with containment of infection and a strong bias toward Th1 immune response, resulting in a transient infection.

Short communication: Herd-level prevalence of Mycobacterium avium ssp. paratuberculosis is not associated with participation in a voluntary Alberta Johne’s disease control program

Johne’s disease (JD) control programs for dairy farms have the general objective of reducing both cow- and herd-level prevalence of Mycobacterium avium ssp. paratuberculosis (MAP). An important aspect of many programs is herd testing for MAP to determine the infection status of participating farms. However, it is uncertain whether MAP herd-level prevalence on farms voluntarily participating in a JD control program is different from that on nonparticipating farms. Therefore, the aim was to compare MAP infection status of participants and nonparticipants in the Alberta Johne’s Disease Initiative (AJDI), a voluntary JD control program initiated in 2010 in Alberta, Canada. Between September 2012 and August 2013, environmental fecal samples were collected from 93 randomly selected farms not enrolled in the AJDI. Additionally, 81 farms that initially enrolled in the AJDI during the same time interval were also sampled. Samples were collected from 6 defined locations on each farm and cultured for MAP. Results were confirmed using conventional IS900 PCR and F 0285 quantitative PCR. Overall, 51% of participating and 51% of nonparticipating farms were identified as being MAP-infected. Furthermore, based on multivariable logistic regression, the number of MAP-positive samples was not associated with AJDI participation (taking herd size into account as a potentially modifying or confounding variable). In conclusion, there was no indication that voluntary participation in the AJDI was associated with herd-level MAP prevalence.

The effects of progressing and nonprogressing Mycobacterium avium ssp. paratuberculosis infection on milk production in dairy cows

Longitudinal data from 3 commercial dairy herds in the northeast United States, collected from 2004 to 2011, were analyzed to determine the effect of Mycobacterium avium ssp. paratuberculosis (MAP) infection status and progression path on milk production. Disease status, as indicated by MAP test results, was determined through quarterly ELISA serum testing, biannual fecal culture, and culture of tissues and feces at slaughter. Milk production data were collected from the Dairy Herd Information Association. Animals with positive MAP test results were categorized, based on test results over the full course of the study, as high path (at least one high-positive culture) or low path (at least one positive culture or ELISA). The cumulative numbers of positive ELISA and culture results were recorded. The effects of both MAP infection path, status, and number of positive tests on milk production were analyzed using a mixed linear model with an autocorrelation random effect structure. Low- and high-path animals produced more milk before their first positive test than always-negative animals, especially high-path animals. Although mean production decreased after a first positive test, low-path animals were shown to recover some productivity. High-path animals continued to exhibit a decrease in milk production, especially after their first high-positive fecal culture. These results show that not all animals that test positive for MAP will have long-term production losses. Milk production decreased significantly with each additional positive test. Ultimately, production loss appeared to be a function of MAP infection progression.

Association of TLR4 polymorphisms with Mycobacterium avium subspecies paratuberculosis infection status in Canadian Holsteins.

Mycobacterium avium ssp. paratuberculosis (MAP) causes chronic enteritis in cattle that results in substantial financial losses to the cattle industry worldwide. Given that susceptibility to MAP infection is determined in part by genetics, marker-assisted selection may help in the breeding of animals that are more resistant to MAP infection. The toll-like receptor 4 gene (TLR4) was selected as a potential candidate gene because of its role in innate immunity and its involvement in MAP recognition and infection. The objective of this study, therefore, was to identify associations between TLR4 polymorphisms and susceptibility to MAP infection in Canadian Holstein cows. Two biologically relevant SNPs, including c.-226G>C in the 5'-untranslated region and the non-synonymous SNP c.2021C>T in the potential TIR domain, were selected for an association analysis with MAP infection status in 409 Canadian Holsteins. The haplotype C-T from these combined SNPs yielded significant association with susceptibility to MAP infection, supporting the involvement of TLR4 in susceptibility to MAP infection.

Factors affecting management changes on farms participating in a Johne’s disease control program

Modern Johne’s disease programs aim to control Mycobacterium avium ssp. paratuberculosis (MAP) infection through implementation of management practices that reduce the probability of MAP introduction and within-herd transmission on dairy farms. Success of these programs depends on whether weaknesses in management are corrected through implementation of management improvements. The objectives of this study were, therefore, to (1) assess whether scores in risk-assessment (RA) questions predicted suggestions for management changes for the upcoming year; and (2) determine factors as assessed in an RA that motivated producers to make management improvements and assess whether management improvements were influenced by previously received test results. The RA determining on-farm management related to MAP introduction and transmission were conducted annually by herd veterinarians on 370 dairy farms participating in the Alberta Johne’s Disease Initiative. A maximum of 3 management changes that the farmer and the veterinarian agreed upon were recorded in a management plan. The MAP infection status of the herds was assessed through culture of 6 environmental samples. Whereas a management change was proposed for only 4% of questions with scores 1 or 2 (low risk), a change was proposed for 19% of questions with scores >2 [high risk; odds ratio (OR)=11.4]. Improvement in RA question scores was more likely between the first and second annual RA than between the second and third RA (OR=1.6). Farms with >3 culture-positive environmental samples collected in the previous year were more likely to improve their management than environmental sample culture-negative farms (OR=1.3). In conclusion, proposed management changes were oriented toward previously identified weaknesses in management practices, suggesting that the RA was properly used to design targeted management suggestions. Furthermore, improvements in management were not randomly distributed among farms participating in the control program. Instead, knowledge of MAP infection status of a herd, suggestions for management improvements, and duration of participation all influenced implementation of management improvements.

The long subclinical phase of Mycobacterium avium ssp. paratuberculosis infections explained without adaptive immunity.

Mycobacterium avium ssp. paratuberculosis (MAP) is an infection of the ruminant intestine. In cows, a long subclinical phase with no or low intermittent shedding precedes the clinical phase with high shedding. It is generally considered that an adaptive cell-mediated immune response controls the infection during the subclinical phase, followed by unprotective antibodies later in life. Based on recent observations, we challenge the importance of adaptive immunity and instead suggest a role of the structural organization of infected macrophages in localized granulomatous lesions. We investigated this hypothesis by mathematical modelling. Our first model describes infection in a villus, assuming a constant lesion volume. This model shows the existence of two threshold parameters, the MAP reproduction ratio RMAP determining if a lesion can develop, and the macrophage replacement ratio RMF determining if recruitment of macrophages is sufficient for unlimited growth. We show that changes in RMF during a cow’s life – i.e. changes in the innate immune response – can cause intermittent shedding. Our second model describes infection in a granuloma, assuming a growing lesion volume. This model confirms the results of the villus model, and can explain early slow granuloma development: small granulomas grow slower because bacteria leave the granuloma quickly through the relatively large surface area. In conclusion, our models show that the long subclinical period of MAP infection can result from the structural organization of the infection in granulomatous lesions with an important role for innate rather than adaptive immunity. It thus provides a reasonable hypothesis that needs further investigation.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-015-0202-3) contains supplementary material, which is available to authorized users.

Evaluation of testing strategies to identify infected animals at a single round of testing within dairy herds known to be infected with Mycobacterium avium ssp. paratuberculosis

As part of a broader control strategy within herds known to be infected with Mycobacterium avium ssp. paratuberculosis (MAP), individual animal testing is generally conducted to identify infected animals for action, usually culling. Opportunities are now available to quantitatively compare different testing strategies (combinations of tests) in known infected herds. This study evaluates the effectiveness, cost, and cost-effectiveness of different testing strategies to identify infected animals at a single round of testing within dairy herds known to be MAP infected. A model was developed, taking account of both within-herd infection dynamics and test performance, to simulate the use of different tests at a single round of testing in a known infected herd. Model inputs included the number of animals at different stages of infection, the sensitivity and specificity of each test, and the costs of testing and culling. Testing strategies included either milk or serum ELISA alone or with fecal culture in series. Model outputs included effectiveness (detection fraction, the proportion of truly infected animals in the herd that are successfully detected by the testing strategy), cost, and cost-effectiveness (testing cost per true positive detected, total cost per true positive detected). Several assumptions were made: MAP was introduced with a single animal and no management interventions were implemented to limit within-herd transmission of MAP before this test. In medium herds, between 7 and 26% of infected animals are detected at a single round of testing, the former using the milk ELISA and fecal culture in series 5 yr after MAP introduction and the latter using fecal culture alone 15 yr after MAP introduction. The combined costs of testing and culling at a single round of testing increases with time since introduction of MAP infection, with culling costs being much greater than testing costs. The cost-effectiveness of testing varied by testing strategy. It was also greater at 5 yr, compared with 10 or 15 yr, since MAP introduction, highlighting the importance of early detection. Future work is needed to evaluate these testing strategies in subsequent rounds of testing as well as accounting for different herd dynamics and different levels of herd biocontainment.

Off-site rearing of heifers reduces the risk of Mycobacterium avium ssp. paratuberculosis ELISA seroconversion and fecal shedding in a California dairy herd

The objective of this cohort study was to evaluate whether rearing dairy heifers at different premises than the dairy of origin (off-site) reduced the risk of Mycobacterium avium ssp. paratuberculosis (MAP) infection more effectively than rearing on the dairy of origin (on-site). From 2003 to 2005, 3 cohorts of Jersey heifers were born on a single California dairy, with heifers in the first cohort raised on-site until first calving (n=797); heifers in the second cohort raised on-site until approximately 5mo of age and off-site until about 1 to 2mo precalving (n=791); and heifers in the third cohort raised off-site from d 1 until about 1 to 2mo before first calving (n=797). Cohorts were sequentially enrolled, and heifers were followed until death, culling, or up to 6 yr of age. Heifers were tested annually for MAP infection by serum ELISA and bacterial culture of feces, from lactation 1 until they were 6 yr old, and all mortality and culling events were recorded. Compared with cohort 1, cohort 3 had lower hazards of seroconverting and shedding of MAP in feces, approximately 70 and 38%, respectively. Cohort 2 was not significantly different from cohort 1 for the same outcomes. Mortality hazards were only significantly different between cohorts before first calving, with calves raised completely off-site at lower risk than the remaining 2 cohorts. Additionally, the hazards for culling in cohorts 2 and 3 were only significantly different from cohort 1 after the first calving. To our knowledge, the current study is the first cohort study to evaluate the association between off-site heifer rearing and risk of MAP infection, mortality, and culling. Rearing heifer calves off-site, away from infected adult dairy cows, may have allowed for reduced exposure to MAP in the environment of the calves and, hence, served as a control strategy for Johne’s disease.