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The mode of action of bismuth subnitrate in teat sealant formulations as a preventative for intramammary infections during the dry period is unknown. Although previous studies proposed an action mechanism—creating a physical barrier in the teat canal to prevent bacterial invasion—it has not been proven experimentally. We hypothesized that bismuth subnitrate has an inhibitory effect on bacterial growth, in addition to its barrier effect. The objective of this study was to assess the effect of bismuth subnitrate on bacterial growth of major mastitis-causing agents. A strain of Streptococcus uberis (SR115), 2 strains of Staphylococcus aureus (SA3971/59 and SA1), and a strain of Escherichia coli (P17.14291) were tested in vitro for their ability to grow in the presence or absence of bismuth subnitrate. Disk diffusion testing, impedance measurement, and evaluation of bacterial growth in shaking conditions were the methods used to test this hypothesis. A reduction of growth in the presence of bismuth subnitrate occurred for all the strains tested. However, we observed strain and species variations in the extent of growth inhibition. These results suggest that an inhibitory effect on bacterial growth by bismuth subnitrate could partially explain the efficacy of bismuth-based formulations for preventing intramammary infections over the dry period. Further research is required to test the effect of teat sealant formulations on bacterial growth.

We conducted a longitudinal study to evaluate the effect of non-aureus staphylococci (NAS) causing subclinical intramammary infections (IMI) on quarter milk somatic cell count (qSCC) and quarter milk yield (qMY). In total, 324 quarters of 82 Holstein Friesian heifers were followed from calving to 130 d in milk (DIM) and were sampled 10 times each at 14-d intervals. The IMI status of each quarter was determined based on bacterial culture results at the current and previous or next sampling day, or both. The qSCC was determined on each sampling day and the average qMY on sampling day was available through stored daily milk weight data in the management program of the automatic milking system. A transient IMI (tIMI) was defined as a case where a specific pathogen was isolated from a quarter on only one sampling day and not on the previous or next sampling day. When the same bacterial strain, as defined by random amplification of polymorphic DNA-PCR, was isolated from the same quarter on multiple sampling days, it was defined as a persistent IMI (pIMI) status on those sampling days; a pIMI episode was defined as the combination of multiple consecutive pIMI statuses with the same bacterial strain on different sampling days. During this study, 142 subclinical IMI with NAS occurred in 116 different quarters from 64 animals, yielding in total 304 NAS isolates belonging to 17 different species. The prevalence of NAS was highest in the first 4 DIM. Overall, the predominant species was Staphylococcus chromogenes (52% of the isolates), followed by S. epidermidis (9.2%), S. xylosus (8.2%), and S. equorum (5.9%). Staphylococcus chromogenes was the only species for which an effect on qSCC and qMY could be analyzed separately; the other NAS species were considered as a group because of their low prevalence. Eighteen out of 40 IMI (45%) caused by S. chromogenes persisted over at least 2 sampling days, whereas only 10 of 102 (9.8%) IMI caused by other NAS species persisted for at least 2 sampling days. The average duration of pIMI episodes was 110.4 d for S. chromogenes and 70 d for the other NAS species. Remarkably, 17 of the 18 pIMI episodes with S. chromogenes started within the first 18 DIM. The qSCC was highest in quarters having a pIMI with a major pathogen, followed by quarters having a pIMI with S. chromogenes, and a pIMI with other NAS. Transient IMI with other NAS or with a major pathogen caused a small but significantly higher qSCC, whereas the qSCC in quarters having a tIMI with S. chromogenes was not statistically different compared with noninfected quarters. No significant differences in qMY were observed between quarters having a pIMI or tIMI with S. chromogenes or with the other NAS species compared with noninfected quarters, despite the higher qSCC. Quarters having a pIMI with major pathogens showed significantly lower daily milk production. Surprisingly, quarters that cured from an IMI with S. chromogenes had a significantly lower qMY than noninfected quarters.

The incidence of milk leakage (ML) after dry-off (DO) and related risk factors was studied in 1,175 dairy cows from 41 commercial herds in 8 European countries: Belgium, Czech Republic, Denmark, France, Germany, Italy, the Netherlands, and Spain. Milk leakage was assessed twice for 30 s each during 3 visits at 20 to 24 h, 30 to 34 h, and 48 to 52 h after DO. Information related to dry-cow management and udder health was collected at herd and cow level, including individual somatic cell count (ISCC) from test-day controls and occurrence of clinical mastitis cases from DO until 30 d in lactation. Mixed-effect logistic regression analyses were used to identify possible risk factors for ML and to study the association between ML and new intramammary infections. Intramammary infections were defined as clinical mastitis cases during the dry period and in the first 30 d in lactation or a rise in ISCC from before to after the dry period (threshold: 200,000 cells/mL) or both. Milk leakage was observed in 24.5% of the cows between 20 and 52 h after DO, where the herd incidence varied between 0.0 and 77.8%. The reduction in number of milkings in the weeks before DO had statistically significant effect on the ML incidence. When the milking frequency was reduced from 3 times/d to 2 or maintained at twice a day, cows had 11 (95% CI = 3.43-35.46) or 9 (95% CI = 1.85-48.22) times higher odds of leaking milk, respectively, compared with cows where the milking frequency was reduced from twice to once a day. Also, the milk production 24 h before DO was associated with ML incidence. Hence, cows with a milk production between 13 and 21 L or above 21 L had 2.3 (95% CI = 1.48-3.53) and 3.1 (95% CI = 1.79-5.3) times higher odds of leaking milk, respectively, compared with cows with a milk production below 13 L. A higher ML incidence was present in the group of cows with an average ISCC in the last 3 mo before DO ≥200,000 cells/mL (odds ratio = 1.7; 95% CI = 1.13-2.41) compared with cows with an average ISCC100,000 cells/mL. Quarters with ML tended to have 2.0 times higher odds of developing clinical mastitis compared with quarters not leaking milk. Cows with ML tended to have 1.5 times higher odds of intramammary infections (i.e., an increase of ISCC or clinical mastitis) compared with cows without ML.

The ability of certain spore-forming bacteria in the order Bacillales (e.g., Bacillus spp., Paenibacillus spp.) to survive pasteurization in spore form and grow at refrigeration temperatures results in product spoilage and limits the shelf life of high temperature, short time (HTST)-pasteurized fluid milk. To facilitate development of strategies to minimize contamination of raw milk with psychrotolerant Bacillales spores, we conducted a longitudinal study of 10 New York State dairy farms, which included yearlong monthly assessments of the frequency and levels of bulk tank raw milk psychrotolerant spore contamination, along with administration of questionnaires to identify farm management practices associated with psychrotolerant spore presence over time. Milk samples were first spore pasteurized (80°C for 12 min) and then analyzed for sporeformer counts on the initial day of spore pasteurization (SP), and after refrigerated storage (6°C) for 7, 14, and 21 d after SP. Overall, 41% of samples showed sporeformer counts of >20,000 cfu/mL at d 21, with Bacillus and Paenibacillus spp. being predominant causes of high sporeformer counts. Statistical analyses identified 3 management factors (more frequent cleaning of the bulk tank area, the use of a skid steer to scrape the housing area, and segregating problem cows during milking) that were all associated with lower probabilities of d-21 Bacillales spore detection in SP-treated bulk tank raw milk. Our data emphasize that appropriate on-farm measures to improve overall cleanliness and cow hygiene will reduce the probability of psychrotolerant Bacillales spore contamination of bulk tank raw milk, allowing for consistent production of raw milk with reduced psychrotolerant spore counts, which will facilitate production of HTST-pasteurized milk with extended refrigerated shelf life.

The abrupt cessation of milking at dry-off may induce milk leakage, which may increase the risk of new intramammary infections (IMI). This study assessed the efficacy of 1 i.m. injection of 5.6 mg of cabergoline (Velactis, Ceva Sante Animale, Libourne, France) at drying-off on milk leakage after dry-off and new IMI across the dry period and postcalving compared with a placebo (negative control) and an intramammary antibiotic treatment (positive control) under field conditions. The study was a double-blind, randomized, 3-arm, multicenter, clinical trial performed under Good Clinical Practice conditions. Data from 900 dairy cows of various breeds from 63 farms in France, Germany, and Hungary were analyzed. Only quarters with no bacterial growth at drying-off and a cow somatic cell count ≤200,000 cells/mL were included. Quarters infected with major or minor pathogens or cows with high somatic cell count at time of inclusion were excluded. Cows that qualified for the study were visited 7 times in total before and after drying-off and after calving. Presence (yes/no) of milk leakage was recorded on the day after dry-off. A new infected quarter (new IMI) was defined as one with a major pathogen present in any one of the 2 postcalving samples. Two mixed logistic regression models were fitted to the data to evaluate the efficacy of cabergoline in the reduction of milk leakage and new IMI. One i.m. injection of cabergoline at drying-off significantly reduced the incidence of milk leakage the day after dry-off compared with both placebo and antibiotic treatment. Cabergoline-treated cows significantly reduced the risk of new IMI by major pathogens across the dry period and postcalving by 21% when compared with placebo cows (20.5 vs. 26.0%, respectively). However, when milk leakage was added to the model, the significance of cabergoline was reduced. We interpreted this to show that milk leakage is an intervening variable between treatment with cabergoline and lower risk of new IMI. The antibiotic treatment significantly decreased the odds of new IMI compared with both cabergoline and placebo. However, because several countries are currently disallowing the preventive use of antibiotics at dry-off in noninfected quarters, the dry-off facilitator cabergoline may therefore be of particular value to reduce the risk of new IMI across the dry period.

The objectives of this study were to estimate the direct effects of clinical mastitis (CM) occurring in early productive life (defined as the first 100 d of the first lactation) of Holstein dairy cows on the future rate of CM occurrence and on the length of total productive lifetime. Information on CM cases and other data occurring in 55,144 lactations in 24,831 cows in 5 New York State Holstein herds was collected from January 2004 until February 2014. For the first objective, a generalized linear mixed model with a Poisson distribution was used to study the effects of CM cases occurring in the first 100 d of a cow's first lactation, as well as farm indicator and number of days in the cow's lifetime, on the future lifetime rate of CM. Only cows that had completed their productive life [i.e., all had been culled (or sold) or had died; n = 14,440 cows] were included in this analysis. For the second objective, a Cox proportional hazards model was used to study the effects of CM cases occurring in the first 100 d of a cow's first lactation on the length of total productive lifetime. The model was stratified by farm. All 24,831 cows were included in this analysis with right censoring. Cows experienced between 0 and 4 CM cases in the first 100 d of lactation 1. Over their lifetime, cows experienced between 0 and 25 CM cases. During the study period, 10% of all cows died and nearly half of all cows were culled. The average length of productive life, including censored observations, was 2.0 yr after first calving. Compared with cows having no CM cases in the first 100 d of lactation 1, cows with 1 CM case in that time period had a 1.5 times higher rate of total number of CM cases over their lifetime. Cows with 2 (or 3 or more) CM cases in the first 100 d of lactation 1 had a 1.7 times (or 2.6 times) higher rate of total number of CM cases over their lifetime. For each additional CM case occurring in the first 100 d of lactation 1, the hazard rate of culling increased by 34%. Given economic conditions for preferentially culling mastitic cows, the study findings may help farmers make optimal decisions with regard to culling of such cows.

Johne’s disease (JD) is a chronic enteric disease in cattle caused by Mycobacterium avian subsp. paratuberculosis (MAP). Eradicating JD is a difficult task due to the long incubation period of MAP, inefficient diagnostic tests, and delayed clinical signs. Effective control strategies can help farmers to reduce prevalence, but those most acceptable to farmers combine specific information about lactation performance and testing results, which existing models do not provide. This paper presents an individual-based model of MAP infection dynamics and assesses the relative performance of the applied alternative control strategies. The base dairy herd model included the daily life events of a dairy cow and reflects several current dairy management processes. We then integrated MAP infection dynamics into the model. The model adopted four different test-based control strategies based on risk-based culling decisions and three hygiene scenarios. The model tracked the source of each infection and quantified the efficacy of each control strategy in reducing the risks of different transmission routes. The results suggest that risk-based culling can reduce prevalence compared with no control, but cannot eliminate the infection. Overall, this work provides not only a valuable tool to investigate MAP transmission dynamics but also offers adaptability to model similar infectious diseases.

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.

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.

Some strains of sporeforming bacteria (e.g., Bacillus spp. and Paenibacillus spp.) can survive pasteurization and subsequently grow at refrigeration temperatures, causing pasteurized fluid milk spoilage. To identify farm management practices associated with different levels of sporeformers in raw milk, a bulk tank sample was obtained from and a management and herd health questionnaire was administered to 99 New York State dairy farms. Milk samples were spore pasteurized [80°C (176°F) for 12 min] and subsequently analyzed for most-probable number and for sporeformer counts on the initial day of spore pasteurization (SP), and after refrigerated storage (6°C) at 7, 14, and 21 d after SP. Management practices were analyzed for association with sporeformer counts and bulk tank somatic cell counts. Sixty-two farms had high sporeformer growth (≥3 log cfu/mL at any day after SP), with an average sporeformer count of 5.20±1.41 mean log10 cfu/mL at 21 d after SP. Thirty-seven farms had low sporeformer numbers (25% of cows with dirty udders in the milking parlor were 3.15 times more likely to be in the high category than farms with ≤10% of milking cows with dirty udders. Farms with 25% of cows with dirty udders observed in the milking parlor. Changes in management practices associated with cow cleanliness may directly ensure longer shelf life and higher quality of pasteurized fluid milk.

Clinical Staphylococcus aureus mastitis is difficult to cure. Extended antimicrobial treatment is often advocated as a practical approach to improve cure rates; however, scientific evidence of this hypothesis is lacking. A multi-centered, nonblinded, randomized, positive-controlled clinical trial was conducted in 5 European countries-France, Hungary, Italy, the Netherlands, and the United Kingdom-to study the efficacy of an extended intramammary cefquinome treatment (5 d) compared with a standard intramammary cefquinome treatment (1.5 d) of Staph. aureus clinical mastitis. Least squares means estimates of bacteriological cure during lactation were 34% [standard error (SE). = 9.9%] for the standard treatment group and 27% (SE. = 8.4%) for the extended treatment group. In the final model, extended therapy was not significantly better. The only factor predicting bacteriological cure was pretreatment cow somatic cell count (SCC). Cows with >250,000 cells/mL in milk before treatment were less likely to cure. Least squares means of clinical cure during lactation was 60% (SE. = 19%) for the standard treatment group and 82% (SE. = 12%) for the extended treatment group. In the final model, clinical cure after extended treatment was significantly better. Pretreatment cow udder firmness predicted clinical cure. Firm udders were less likely to cure clinically. Irrespective of treatment regimen, new infection rates with pathogens other than Staph. aureus were higher (42%) after bacteriological cure than after nonbacteriological cure (22%) and cured cows had a significantly lower SCC. In conclusion, independent of the treatment protocol, cows with an SCC

We examined consistency of the relationship between intramammary infection (IMI) and somatic cell score (SCS) across several classes of cow, herd, and sampling time variables. Microbial cultures of composite milk samples were performed by New York Quality Milk Production Services from 1992 to 2004. SCS was from the most recent Dairy Herd Improvement test before IMI sampling. Records were analyzed from 79,308 cows in 1,124 commercial dairy herds representing a broad range of production systems. Three binary dependent variables were presence or absence of contagious IMI, environmental IMI, and all IMI. Independent variables in the initial models were SCS, SCS2, lactation number, days in milk, sample day milk yield, use of coliform mastitis vaccine, participant type (required by regulation or voluntary), production system (type of housing, milking system, and herd size), season of sampling, year of sampling, and herd; also the initial models included interactions of SCS and SCS2 with other independent variables, except herd and milk yield. Interaction terms characterize differences in the IMI-SCS relationship across classes of the independent variables. Models were derived using the Glimmix macro in SAS (SAS Institute Inc., Cary, NC) with a logistic link function and employing backward elimination. The final model for each dependent variable included all significant independent variables and interactions. Simplified models omitted SCS2 and all interactions with SCS. Interactions of SCS with days in milk, use of coliform mastitis vaccine, participant type, season, and year were not significant in any of the models. Interaction of SCS with production system was significant for the all IMI model, whereas interaction of SCS with lactation number was significant for the environmental and all IMI models. Each 1 point increase in SCS (or doubling of somatic cell count) was associated with a 2.3, 5.5%, and 9.1% increase in prevalence of contagious, environmental, and all IMI, respectively. Empirical receiver operator characteristic curves and areas under the curve were derived for final and simplified models. The areas under the curve for simplified and final models within each type of IMI differed by 0.009 or less. We concluded that the relationship of IMI with SCS was generally stable over time and consistent across seasons, production systems, and cow factors.

The objectives of this study were to estimate the direct effects of clinical mastitis (CM) occurring in early productive life (defined as the first 100 d of the first lactation) of Holstein dairy cows on the future rate of CM occurrence and on the length of total productive lifetime. Information on CM cases and other data occurring in 55,144 lactations in 24,831 cows in 5 New York State Holstein herds was collected from January 2004 until February 2014. For the first objective, a generalized linear mixed model with a Poisson distribution was used to study the effects of CM cases occurring in the first 100 d of a cow's first lactation, as well as farm indicator and number of days in the cow's lifetime, on the future lifetime rate of CM. Only cows that had completed their productive life [i.e., all had been culled (or sold) or had died; n = 14,440 cows] were included in this analysis. For the second objective, a Cox proportional hazards model was used to study the effects of CM cases occurring in the first 100 d of a cow's first lactation on the length of total productive lifetime. The model was stratified by farm. All 24,831 cows were included in this analysis with right censoring. Cows experienced between 0 and 4 CM cases in the first 100 d of lactation 1. Over their lifetime, cows experienced between 0 and 25 CM cases. During the study period, 10% of all cows died and nearly half of all cows were culled. The average length of productive life, including censored observations, was 2.0 yr after first calving. Compared with cows having no CM cases in the first 100 d of lactation 1, cows with 1 CM case in that time period had a 1.5 times higher rate of total number of CM cases over their lifetime. Cows with 2 (or 3 or more) CM cases in the first 100 d of lactation 1 had a 1.7 times (or 2.6 times) higher rate of total number of CM cases over their lifetime. For each additional CM case occurring in the first 100 d of lactation 1, the hazard rate of culling increased by 34%. Given economic conditions for preferentially culling mastitic cows, the study findings may help farmers make optimal decisions with regard to culling of such cows.

Johne's disease, a granulomatous enteritis of ruminant animals, is a hidden threat on dairy farms, adversely affecting animal welfare as well as herd productivity. Control programs in the United States advocate for specific management practices to temper the spread of the causal organism (Mycobacterium avium ssp. paratuberculosis, MAP), such as improving calving area hygiene and limiting introduction of replacement stock with unknown infection status. A need remains for direct exploration of Johne's disease prevention strategies in the United States with respect to production type. Alongside the growing demand for organic products, the safety of organic dairy practices with respect to MAP control is warranted. Further, conventional herds for which organic practices such as pasture grazing are used should be situated within the risk spectrum. We developed a risk assessment model using the US Voluntary Bovine Johne's Disease Control Program as a framework, with the goal of evaluating the risk of new cow-level MAP infections. A total of 292 organic and conventional farms in 3 states were surveyed on management practices, and an overall analysis was conducted in which each farm was first scored on individual practices using a range of “no risk” to “high risk,” according to the literature. The sum of all risk factors was then analyzed to quantify and compare the risk burden for each production type. Organic herds received higher overall risk scores compared with both conventional grazing and nongrazing subtypes. To identify which factors contributed to the overall increased risk for organic herds, the management practices were categorized and evaluated by logistic regression. We determined that the increased risk incurred by organic herds was predominantly due to decisions made in the calving area and preweaned calf group. However, although certain individual risk factors related to calf management are commonly involved in prevention strategies (e.g., cow/calf separation) and were thus included in the overall risk assessment, empirical evidence linking them to the spread of MAP is lacking. Instead, these factors are problematic when executed with other management decisions, leading to a hypothesized synergism of transmission risk. To this end, we developed a set of compound risk factors, which were also evaluated as outcomes in logistic regression models, with production type serving as the predictor of interest. Organic farms in our study were more susceptible to risks associated with the synergism of study variables. Notably, organic producers were most likely to allow calves to spend extended time with the dam, while also lacking a dedicated calving area. Additionally, calves in organic herds were more often permitted to nurse even with poor udder hygiene on farm. A heightened vigilance toward calving area hygiene is therefore indicated for these herds.

The objective of this study was to evaluate the efficacy of intramammary immunization with UV-killed Escherichia coli ECC-Z on prevention of intramammary colonization after a challenge with a dose of the homologous E. coli ECC-Z live bacteria. A total of 10 cows were included in a study to evaluate the efficacy of intramammary immunization. All 10 cows received an intramammary immunization of 100 cfu of UV-killed E. coli ECC-Z bacteria into one hind quarter at the time of dry off. Approximately 2 wk before the anticipated calving date, both hind quarters of all cows were challenged with 100 cfu of live E. coli ECC-Z bacteria. Five of the cows were vaccinated parenterally with a commercial J5 bacterin, and 5 cows served as controls with no parenteral vaccination. The cows were then followed over time and infection risk, clinical scores, somatic cell count, and milk production were observed over time. The results of these 10 cows showed partial protection of intramammary immunization on the outcome of a subsequent homologous intramammary challenge. Immunization resulted in a lower probability of infection, a lower bacteria count, lower somatic cell counts and milk conductivity, a lower clinical mastitis score, and increased milk production compared with unimmunized control quarters. Once the analysis was corrected for immunization, parenteral J5 vaccination had no significant effect on any of the measured parameters. These results provide the first evidence that intramammary immunization may improve the outcome of an intramammary E. coli infection in late gestation and onset of mastitis immediately following parturition. Unlike systemic vaccination, which generally does not reduce the intramammary infection risk, the intramammary immunization did show a 5-times reduced odds of an established intramammary infection after challenge. Cytokine profiles indicated a local return of proinflammatory response after challenge as the data showed a more pronounced increase in in IFN-γ with a subsequent negative feedback due to a spike in the level of IL-10 in immunized quarters relative to nonimmunized quarters. Although these results are preliminary and obtained on only 10 cows, the results provide insight into the biological benefits of triggering mucosal immunity in the mammary gland.

Mycobacterium avium ssp. paratuberculosis (MAP), the etiologic agent of Johne's disease in dairy cattle, may enter the bulk tank via environmental contamination or direct excretion into milk. Traditionally, diagnostics to identify MAP in milk target either MAP antibodies (by ELISA) or the organism itself (by culture or PCR). High ELISA titers may be directly associated with excretion of MAP into milk but only indirectly linked to environmental contamination of the bulk tank. Patterns of bulk-milk ELISA and bulk-milk PCR results could therefore provide insight into the routes of contamination and level of infection or environmental burden. Coupled with questionnaire responses pertaining to management, the results of these diagnostic tests could reveal correlations with herd characteristics or on-farm practices that distinguish herds with high and low environmental bulk-tank MAP contamination. A questionnaire on hygiene, management, and Johne's specific parameters was administered to 292 dairy farms in New York, Oregon, and Wisconsin. Bulk-tank samples were collected from each farm for evaluation by real-time PCR and ELISA. Before DNA extraction and testing of the unknown samples, bulk-milk template preparation was optimized with respect to parameters such as MAP fractionation patterns and lysis. Two regression models were developed to explore the relationships among bulk-tank PCR, ELISA, environmental predictors, and herd characteristics. First, ELISA optical density (OD) was designated as the outcome in a linear regression model. Second, the log odds of being PCR positive in the bulk tank were modeled using binary logistic regression with penalized maximum likelihood. The proportion of PCR-positive bulk tanks was highest for New York and for organic farms, providing a clue as to the geographical patterns of MAP-positive bulk-tank samples and relationship to production type. Bulk-milk PCR positivity was also higher for large relative to small herds. The models revealed that bulk-milk PCR result could predict ELISA OD, with PCR-positive results corresponding to high bulk-milk ELISA titers. Similarly, ELISA was a predictor of PCR result, although the association was stronger for organic farms. Despite agreement between high bulk-milk ELISA titers and positive PCR results, a large proportion of high ELISA farms had PCR-negative bulk tanks, suggesting that farms are able to maintain satisfactory hygiene and management despite a presence of MAP in these herds.

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.