Exploring associations between the teat apex metagenome and Staphylococcus aureus intramammary infections in primiparous cows under organic directives.

The primary objective of this study was to identify associations between the prepartum teat apex microbiome and the presence of Staphylococcus aureus intramammary infections (IMI) in primiparous cows during the first 5 weeks after calving. We performed a case-control study using shotgun metagenomics of the teat apex and culture-based milk data collected longitudinally from 710 primiparous cows on five organic dairy farms. Cases had higher odds of having S. aureus metagenomic DNA on the teat apex prior to parturition compared to controls (OR = 38.9, 95% CI: 14.84-102.21). Differential abundance analysis confirmed this association, with cases having a 23.8 higher log fold change (LFC) in the abundance of S. aureus in their samples compared to controls. Of the most prevalent microorganisms in controls, those associated with a lower risk of post-calving S. aureus IMI included Microbacterium phage Min 1 (OR = 0.37, 95% CI: 0.25-0.53), Corynebacterium efficiens (OR = 0.53, 95% CI: 0.30-0.94), Kocuria polaris (OR = 0.54, 95% CI: 0.35-0.82), Micrococcus terreus (OR = 0.64, 95% CI: 0.44-0.93), and Dietzia alimentaria (OR = 0.45, 95% CI: 0.26-0.75). Genes encoding for Microcin B17 AMPs were the most prevalent on the teat apex of cases and controls (99.7% in both groups). The predicted abundance of genes encoding for Microcin B17 was also higher in cases compared to controls (LFC 0.26). IMPORTANCE: Intramammary infections (IMI) caused by Staphylococcus aureus remain an important problem for the dairy industry. The microbiome on the external skin of the teat apex may play a role in mitigating S. aureus IMI risk, in particular the production of antimicrobial peptides (AMPs) by commensal microbes. However, current studies of the teat apex microbiome utilize a 16S approach, which precludes the detection of genomic features such as genes that encode for AMPs. Therefore, further research using a shotgun metagenomic approach is needed to understand what role prepartum teat apex microbiome dynamics play in IMI risk.

Calf management and welfare in the Canadian and US dairy industries: Where do we go from here?

The objective of this narrative review was to compare the results of the 2015 Canadian National Dairy Study and the 2014 US National Animal Health Monitoring System (NAHMS) Dairy Studies, with a specific focus on calf management and welfare, and to interpret these findings within more recent calf health research to describe where we need to go next in the North American dairy industry. Situating results of periodic national studies within the context of past and recent research provides an opportunity to gauge adoption of recommendations and best practices and to help identify persistent and new challenges that the industry is wrestling with to help guide research needs. Through this review of the 2 national studies, we identified several strengths of the Canadian and US dairy industries. In each area of calf health management, improvements relative to previous NAHMS studies and the published literature have been found in the level of mortality, amount of colostrum fed, housing, and the number of producers using pain control for disbudding and dehorning. There were, however, some areas that present clear opportunities for improvement. Specifically, although mortality levels have improved, a large number of calves die at birth, within the first 48 h of life, and during the preweaning period. To improve the health of calves in early life, producers could look at feeding high-quality colostrum at 10% of body weight in the first feeding, as well as feeding a larger amount of milk during the preweaning period. The barriers to making these management changes and improving overall calf health need to be identified in future studies. The majority of preweaning calves in Canada and the United States are housed in individual housing. This represents a clear opportunity for improvement because recent research has identified the positive aspects of group housing. Finally, with respect to pain control, improvements are needed (particularly in the United States) to ensure that pain management is provided when disbudding and dehorning calves. Although the science is clear on pain management, discussions with producers are needed to identify reasons for the lack of uptake.

Investigating Ontario dairy farmers motivations and barriers to the adoption of biosecurity and Johne's control practices.

For the control of Johne's disease (JD), management practices to minimize disease transmission must be implemented and maintained. Once infected, animals will enter a latent phase and will typically only manifest clinical symptoms years later. As young calves are the main susceptible group on farm, the observed effects of management practices geared toward minimizing their exposure to infective material may not be realized until years later. This delayed feedback limits the sustained implementation of JD control practices. Although quantitative research methods have demonstrated changes to management practices as well as their association with changes to JD prevalence, dairy farmers can offer insights into the current challenges relating to JD implementation and control. Thus, this study aims to use qualitative methods and in-depth interviews (n = 20) with Ontario dairy farmers who had previously been engaged in a Johne's control program to explore their motivations and barriers to the implementation of JD control practices and general herd biosecurity. A thematic analysis using inductive coding was completed generated the following 4 overarching themes: (1) the hows and whys of Johne's control, (2) barriers to general herd biosecurity, (3) barriers to Johne's control, and (4) overcoming barriers. Farmers no longer believed JD was an issue on their farm. Johne's was low on their list of concerns due to little public discourse, absence of animals displaying clinical signs, and no financial support for diagnostic testing. Producers who were still actively engaged in JD control cited animal and human health as their primary motivations. Financial support, targeted education, and promoting engagement through discourse may help encourage producers to reconsider their participation in JD control. Government and industry collaboration with producers may help to develop more effective biosecurity and disease control programs.

The association of immune response and colostral immunoglobulin G in Canadian and US Holstein-Friesian dairy cows.

In cattle, maternal immunoglobulins are transferred through colostrum to provide passive immunity to the neonatal calf once they are absorbed into circulation. Cows can be assessed for antibody- and cell-mediated immune responses (AMIR and CMIR, respectively), and through estimated breeding values (EBV) and genomic parent averages (GPA), cows can be classified as having high, average, or low immune response (IR). The objective of this study was to identify associations of colostral IgG concentrations with IR in dairy cows. High IR dairy cows identified by GPA or EBV were hypothesized to produce higher colostral IgG concentrations than cows with average or low IR. Colostrum was collected from Holstein dairy cows from 3 large commercial herds (n = 590) in the United States and 1 research herd at the Ontario Dairy Research Centre (n = 275) in Canada. For the US herds, IR GPA were available through genotyping. For the Canadian herd, IR EBV were available through phenotyping and pedigree information. Colostral IgG concentrations were measured by radial immunodiffusion and analyzed using general linear models in SAS. Based on a prediction equation, cows in US herds with a CMIR GPA of 1 would have colostral IgG concentrations 6.3 g/L higher on average than cows with a CMIR GPA of 0. High CMIR cows produced statistically greater colostral IgG concentrations (least squares mean ± standard error of the mean, 107.5 ± 7.7 g/L) than low CMIR cows (91.4 ± 7.1 g/L), with intermediate values for average CMIR cows (105.1 ± 5.6 g/L). No differences were found among AMIR categories in US cows. The Canadian herd showed a trend for cows with high CMIR EBV (continuous variable) to produce greater colostral IgG. No differences were observed among high, average, and low AMIR EBV classifications in Canadian cows. The findings suggest that selective breeding of Holstein cows to enhance CMIR could contribute to higher-quality colostrum in succeeding generations.

Management practices associated with Johne's bulk tank milk ELISA positivity.

Johne's disease (JD) control is often based on the culling of positive animals and the adoption of management practices that minimize exposure of young stock to the pathogen (Mycobacterium avium ssp. paratuberculosis). Throughout 2010 to 2013, the province of Ontario, Canada, instituted a voluntary Johne's control program consisting of whole-herd testing and risk assessment. The JD risk assessment evaluated 5 management areas to characterize herd JD risk. Using a modified milk ELISA technique with an optical density cut-off of 0.089, province-wide bulk tank milk (BTM) testing was used to assess the prevalence of JD high-risk herds at the end of the control program and again 4 yr after its completion. Approximately 71% of Ontario bulk tanks were classified as positive in 2017 compared with roughly 46% in 2013. In 2019, the same JD risk assessment used in the original program was readministered on 180 Ontario dairy farms. Using this cross-sectional approach, logistic regression models were built using data from the original program risk assessment and follow-up risk assessment as well as the BTM ELISA results to determine management factors associated with the control of JD. We demonstrated that management of the maternity area is an important factor in the control of Johne's disease. Although it is believed that the highest risk group for JD infection is calves under 6 mo, the cleanliness scores of older heifers and their exposure to mature cow manure was significantly associated with JD control; farms with highly contaminated weaned and bred heifers and those that had exposure to mature cow manure were more likely to be unsuccessful in their JD control efforts. Careful management of young calves appears to be important for JD control, and this management should continue even after calves have left the maternity area.

Descriptive analysis of the changes in Johne's disease management practices on Ontario dairy farms through repeat risk assessment.

Johne's disease (JD) control is often based on the culling of positive animals and the adoption of management practices that minimize exposure of calves to the pathogen Mycobacterium avium ssp. paratuberculosis (MAP). From 2010 to 2013, Ontario, Canada, instituted a voluntary Johne's control program consisting of whole-herd testing and a Risk Assessment and Management Plan (RAMP). The RAMP consisted of 38 questions that evaluated 5 different management areas to characterize herd risk for MAP introduction and within-herd spread. The RAMP produced a numerical score for each area, with higher scores associated with higher risk. The RAMP focused on animal purchases, calving management, calf management, and heifer and cow cleanliness and management. In the summer of 2019, the RAMP was repeated on 180 farms that had participated in the JD program of 2010 to 2013 and had bulk tank milk ELISA results from 2013 and 2017. This cross-sectional study demonstrated that many producers changed management practices over the 4- to 7-year period. Producers changed their cattle buying practices, with a reduction in purchasing from multiple sources and more herds refraining from buying in animals. However, overall scores were higher in 2019 than in 2013. The 2019 RAMP indicated that fewer farms were utilizing individual calving pens in 2019 than in 2013 (13% vs. 26%), yet more farms had policies in place to deal with sick or suspect JD cows entering the maternity area (92% vs. 74%). Management changes occurred over time, some of which represent increased risk (crowded maternity pens) and others decreased risk (closed herd, protocols in place for JD-positive cows) for MAP introduction and transmission. These results highlight the importance of frequent risk assessments and the documentation of changes to management practices on-farm as a means to assess herd disease risk more accurately.

Factors influencing how Canadian dairy producers respond to a downer cow scenario.

Understanding how downer cattle are managed allows for the evaluation of strengths and weaknesses in these practices, which is an important step toward improving the care these animals receive. The objective of this cross-sectional study was to analyze factors associated with the care and management of downer cattle by Canadian dairy producers. Data were obtained from the 2015 National Dairy Study, and analysis was limited to the 371 respondents completing the downer cow scenario. The scenario described a downer cow that the producer wanted to keep in their herd but must be moved, and was followed by questions addressing the cow's care and management. Using multivariable logistic regression models, associations between respondent demographics and farm characteristics, and the presence of downer cow protocols, we assessed decisions regarding euthanasia and use of behavioral prognostic indicators. Written downer cow protocols were reported by 18.2% of respondents, 67% indicated that they had a nonwritten protocol, and 14.8% reported that they did not have a protocol (either written or nonwritten). Respondents from western provinces were more likely to have a written protocol than those from Ontario. Nineteen percent of the respondents with a written or unwritten protocol reported veterinary involvement in developing their downer cow protocol, which occurred more commonly on farms with more frequent herd health visits and a good producer-veterinarian relationship. An area to move a downer cow to was present on 88% of farms, with respondents who were farm staff being less likely to report having knowledge of a designated area than respondents who were the farm owner. In addition, approximately half (45%) of respondents reported moving downer cattle with hip lifters as their most common method. Behavioral prognostic indicators chosen by respondents were associated with the respondent's geographic region, age, farm size, and education. Most notably, older respondents were more likely to use appetite, and less likely to use attitude, as a prognostic indicator compared with younger respondents. Using perceived pain as a prognostic indicator was more common among respondents from western and Atlantic provinces compared with respondents from Ontario, and more common among respondents with a college or university education. These results highlighted herd and farmer demographics that were associated with how Canadian dairy producers managed downer cattle in 2015 and could be used as a benchmark for evaluating how these management practices compare with those currently implemented.

Communication preferences and social media engagement among Canadian dairy producers.

The objective of this study was to determine communication preferences of dairy producers in Canada. A secondary objective was to evaluate social media engagement of dairy producers. A survey was administered to Canadian dairy producers between March and April, 2015 to collect information on current management practices on their farms. A total of 1,373 Canadian dairy producers responded to the survey, representing a response rate of 12%. The survey consisted of 192 questions; however, only questions regarding producer demographics, importance of information sources, and internet and social media use were evaluated in this study. The primary outcome variables of interest included use of the internet to access dairy information, importance of different sources of information about dairy herd health and management, and use of online search engines and social media platforms. For each outcome, logistic regression analyses were used to investigate associations between the outcome and demographic variables. Veterinarians were viewed as a "very important" source of information by the majority of respondents (79%), whereas milk recording and dairy producer organizations were viewed as a "very important" source of information by 36% of respondents. Other producers (46%) and magazines or newspapers (51%) were commonly viewed as an "important" source of information. Online search engines were commonly used by respondents (94%). Social media was viewed as less important, and had mixed levels of use. YouTube (70%), Facebook (63%), and Twitter (18%) were the most commonly used social media platforms. Eighty percent of Twitter users reported using the platform to interact with and obtain or share information about herd health management online, which was the highest reported interactivy regarding herd health among all social media platforms. This exploratory study offers insight into the communication preferences of Canadian dairy producers and can be used to facilitate future communication strategies aimed at engaging rural farming audiences across Canada.

Short communication: Describing mortality and euthanasia practices on Canadian dairy farms.

The objective of this study was to describe mortality rates and euthanasia practices used for cows, heifer calves, and male dairy calves on Canadian dairy farms. An internet survey was administered to Canadian dairy producers between March and April 2015 to collect information on current management practices. Approximately 81% (867/1,076) and 63% (673/1,065) of respondents reported that at least one animal died unassisted and at least one animal was euthanized on the farm in the preceding 12-mo period, respectively. Overall, mean mortality was 8% for cows, and 6 and 2% for preweaning and weaned heifers, respectively. On average, 48, 76, and 89% of all reported mortality events in cows, weaned heifers, and preweaning heifers were recorded as unassisted deaths. Cows that died without assistance were necropsied more often than preweaning heifers dying without assistance; these cows were also necropsied more than cows that were euthanized. Conversely, preweaning heifers that were euthanized were necropsied more frequently than those that died without assistance. Choosing not to perform necropsies on animals that die of unknown causes could represent a missed opportunity to identify cause of death and inform changes to prevent future deaths. The use of a firearm was the most common method for euthanizing cows and heifers (54%) and male dairy calves (51%). Approximately 7% of respondents reported using blunt force to euthanize animals. The use of this method was much higher for euthanasia of male dairy calves (34%) than for heifer calves (7%). Québec respondents euthanized their animals using blunt force more often than farmers from other regions. Further, 53% of respondents using blunt force indicated it was their primary method of calf euthanasia; these results are extremely concerning. Approximately 31% of respondents who reported that they do not use blunt force to euthanize heifers and cows did report using blunt force to euthanize male dairy calves. These results can be used to inform veterinarian-client communication, broader extension tools and programs, and industry policies to improve dairy cattle health and welfare, a goal that is becoming of increasing importance from a consumer assurance perspective.

Canadian National Dairy Study: Describing Canadian dairy producer practices and perceptions surrounding cull cow management.

Farmer decisions surrounding culling have an important effect on the health and welfare of cull cows. The objectives of this study were to describe the self-reported shipment behaviors of Canadian dairy producers and understand farmer perspectives on the factors that were most influential in their decision to cull a cow. A nationwide survey was administered between March and April 2015 that included 192 questions covering producer background information, farm characteristics, biosecurity practices, disease prevalence, calf health, cow welfare, lameness, milking hygiene, reproduction, and Internet and social media use. The survey yielded a 12% response rate; a total of 1,076 respondents (78% of all survey respondents completed the culling section of the survey) were included in this study for analysis. Approximately 80, 51, and 38% of respondents reported shipping at least 1 cow to auction, direct to slaughter, and to another dairy farm in the past 12 mo, respectively. Ability of the cow to remain standing (93% of respondents) and drug withdrawal times (92% of respondents) were identified as the most important factors for consideration when culling cows. The time between culling decision and when the cow was actually transported was longer for lame cows than sick cows; almost 70% of respondents reported that cows culled for illness were typically shipped within 1 wk of culling decision, whereas only 51% of respondents indicated the same was true for lame cows. Last, Canadian dairy producers generally exhibited strong confidence that their culled cows would arrive at slaughter in the same condition as they left, but felt very unsure about knowing the location of their final destination. These results highlight several gaps between producer perceptions and the true situation, and can be used to develop tailored programs and inform policy and regulatory decisions aimed at improving cull cow decisions and cow welfare.

Canadian dairy farmers' perception of the efficacy of biosecurity practices.

The adoption of biosecurity practices on dairy farms is limited worldwide. Multiple aspects, one of which is the perception of the effectiveness of these practices, influence the adoption of preventive behavior. The objectives of the present study were to investigate the general understanding of biosecurity by Canadian dairy farmers and to describe their perception of the effectiveness of specific biosecurity practices. In 2015, 368 Canadian dairy farmers were selected to participate in an on-farm national cross-sectional benchmarking study during which they were asked about their general understanding of biosecurity and their perception of the effectiveness of 17 practices to minimize biosecurity risk. For these same biosecurity practices, respondents were asked whether they discussed that particular practice with a veterinarian and whether they implemented it on their farm. Herd and respondent characteristics were also recorded. Associations between the perception of effectiveness of each biosecurity practice and the discussion with a veterinarian, its implementation on farm, and the herd and respondent characteristics were examined using ordered regression models. Most respondents thought the purpose of on-farm biosecurity was to prevent both entry of a new pathogen and spread of an existing pathogen (73%) and considered general biosecurity to be effective (92%) and important (58%). When asked about specific biosecurity practices, respondents considered most of them to be effective (60-94%). Practices related to direct animal-to-animal contact were perceived as effective by more respondents than biosecurity practices related to fomites and visitors. Less than 20% of the respondents reported discussing the different biosecurity practices with a veterinarian, and less than 60% reported implementing these practices on their farm. Geographical region, involvement of a veterinarian, and implementation of the practice on farm were associated with the perception of effectiveness for multiple practices. Finally, the reasons chosen by most respondents as motivation to implement biosecurity practices were (1) if the practice helped to regionally prevent the introduction or the spread of a disease and (2) if it was proven to be beneficial to the health and welfare of animals. These results show that there is a positive perception of the effectiveness of specific biosecurity practices, that there might be a lack of understanding of the practices involving transmission of diseases via indirect contact, and that the perceived threat associated with not adopting practices is minimal.

Prevalence, fecal egg counts, and species identification of gastrointestinal nematodes in replacement dairy heifers in Canada.

Information is scarce regarding the epidemiology of gastrointestinal nematodes in Canadian dairy heifers. The objectives of this study were to estimate the prevalence and fecal egg counts of gastrointestinal nematodes in dairy heifers, and using a novel deep-amplicon sequencing approach, to identify the predominant gastrointestinal nematode species in Canadian dairy replacement heifers. Fresh environmental fecal samples (n = 2,369) were collected from replacement heifers on 306 dairy farms across western Canada, Ontario, Québec, and Atlantic Canada. Eggs per gram of feces (EPG) were determined using a modified Wisconsin double-centrifugation sugar flotation technique. Predominant nematode species at the farm level were identified by deep-amplicon nemabiome sequencing of the internal transcribed spacer-2 rDNA locus of nematode third-stage larvae. Generalized estimating equations were used to estimate predicted parasite prevalence and mean EPG in all heifers and by province, allowing for clustering within herds. Individual heifer egg counts ranged from 0 to 141 EPG (median: 0 EPG; interquartile range: 0 to 71 EPG). Gastrointestinal nematodes were detected in 20.9% (95% confidence interval: 17.2 to 24.6%) of heifers, and the predicted mean strongylid EPG accounting for clustering on farms was 1.1 (95% confidence interval: 0.6 to 1.6). The predominant parasite species were Cooperia oncophora and Ostertagia ostertagi. This is the first study in Canada to use a combination of deep-amplicon nemabiome sequencing and a traditional egg count method to describe the epidemiology of gastrointestinal nematodes in dairy heifers.

Biosecurity and herd health management practices on Canadian dairy farms.

The Canadian dairy industry has created national standards to support the adoption of biosecurity practices and to reduce disease risks across the country. There is, however, very little information on the adoption of these practices. The present study aimed to describe the adoption of biosecurity practices on Canadian dairy farms shortly after the creation of the national standards and to identify demographic factors associated with practice adoption. In 2015, 2 questionnaires (phase 1 and 2) were administered to Canadian dairy farmers during an extensive cross-sectional study. Associations between adoption of biosecurity practices as well as associations between adoption of these practices and demographic variables were tested using multiple correspondence analysis. A total of 1,157 questionnaires were completed in phase 1, and a subsample of 368 respondents was selected using stratified random sampling to complete phase 2 during visits to the farms. There was a lack of investigation into general disease syndromes such as a high prevalence of abortion or an unexplained death (38 and 22% of respondents, respectively). Biosecurity measures within herds and between herds to minimize the spread of infection were not widely adopted (e.g., 27% of the respondents never housed sick or lame animals in their calving pen, 41% had closed herds, and 25 and 48% of the open herds had no strategy for introducing new additions and reintroducing returning animals, respectively). Cleanliness of the cows before calving was always ensured by 29% of the respondents, and 27% of the respondents reported always sanitizing the calving pen after each calving. Less than 15% of the respondents had measures in place to limit or control visitors coming on their farm. Moreover, less than half of the respondents reported requiring family members, employees, and visitors to wear farm-designated or clean boots and coveralls. From the multiple correspondence analysis, 2 dimensions were retained and were summarized as "animal movement, calving area, and visitor biosecurity" and "employee biosecurity." Geographical region, type of housing, and milk production were associated with the "employee biosecurity" dimension. The present study demonstrates that many important biosecurity practices are not implemented on Canadian dairy farms; therefore, efforts to promote the adoption of these practices will be necessary to improve biosecurity in dairy herds.

Stress-Responsive Alternative Sigma Factor SigB Plays a Positive Role in the Antifungal Proficiency of Bacillus subtilis.

Different Bacillus species with PGPR (plant growth-promoting rhizobacterium) activity produce potent biofungicides and stimulate plant defense responses against phytopathogenic fungi. However, very little is known about how these PGPRs recognize phytopathogens and exhibit the antifungal response. Here, we report the antagonistic interaction between Bacillus subtilis and the phytopathogenic fungus Fusarium verticillioides We demonstrate that this bacterial-fungal interaction triggers the induction of the SigB transcription factor, the master regulator of B. subtilis stress adaptation. Dual-growth experiments performed with live or dead mycelia or culture supernatants of F. verticillioides showed that SigB was activated and required for the biocontrol of fungal growth. Mutations in the different regulatory pathways of SigB activation in the isogenic background revealed that only the energy-related RsbP-dependent arm of SigB activation was responsible for specific fungal detection and triggering the antagonistic response. The activation of SigB increased the expression of the operon responsible for the production of the antimicrobial cyclic lipopeptide surfactin (the srfA operon). SigB-deficient B. subtilis cultures produced decreased amounts of surfactin, and B. subtilis cultures defective in surfactin production (ΔsrfA) were unable to control the growth of F. verticillioidesIn vivo experiments of seed germination efficiency and early plant growth inhibition in the presence of F. verticillioides confirmed the physiological importance of SigB activity for plant bioprotection.IMPORTANCE Biological control using beneficial bacteria (PGPRs) represents an attractive and environment-friendly alternative to pesticides for controlling plant diseases. Different PGPR Bacillus species produce potent biofungicides and stimulate plant defense responses against phytopathogenic fungi. However, very little is known about how PGPRs recognize phytopathogens and process the antifungal response. Here, we report how B. subtilis triggers the induction of the stress-responsive sigma B transcription factor and the synthesis of the lipopeptide surfactin to fight the phytopathogen. Our findings show the participation of the stress-responsive regulon of PGPR Bacillus in the detection and biocontrol of a phytopathogenic fungus of agronomic impact.

Regulation of Biofilm Aging and Dispersal in Bacillus subtilis by the Alternative Sigma Factor SigB.

Bacterial biofilms are important in natural settings, biotechnology, and medicine. However, regulation of biofilm development and its persistence in different niches is complex and only partially understood. One key step during the biofilm life cycle is dispersal, when motile cells abandon the mature biofilm to spread out and colonize new niches. Here, we show that in the model bacterium Bacillus subtilis the general stress transcription factor SigB is essential for halting detrimental overgrowth of mature biofilm and for triggering dispersal when nutrients become limited. Specifically, SigB-deficient biofilms were larger than wild-type biofilms but exhibited accelerated cell death, significantly greater sensitivity to different stresses, and reduced dispersal. Interestingly, the signal detected by SigB to limit biofilm growth was transduced through the RsbP-dependent metabolic arm of the SigB regulatory cascade, which in turn positively controlled expression of SinR, the master regulator of biofilm formation and cell motility. This novel SigB-SinR regulatory circuit might be important in controlling the fitness of biofilms (either beneficial or harmful) in diverse environments.IMPORTANCE Biofilms are crucial for bacterial survival, adaptation, and dissemination in natural, industrial, and medical systems. Sessile cells embedded in the self-produced extracellular matrix of the biofilm benefit from a division of labor and are protected from environmental insults. However, as the biofilm ages, cells become stressed because of overcrowding, starvation, and accumulation of waste products. How does the sessile biofilm community sense and respond to stressful conditions? Here, we show that in Bacillus subtilis, the transcription factors SigB and SinR control whether cells remain in or leave a biofilm when metabolic conditions become unfavorable. This novel SigB-SinR regulatory circuit might be important for controlling the fitness of biofilms (either beneficial or harmful) in diverse environments.

Evaluation of bulk tank milk PCR and bulk tank milk modified ELISA tests for the detection of paratuberculosis at the herd level in goat and sheep dairies in Ontario, Canada.

Early identification of dairy goat herds and dairy sheep flocks infected with Mycobacterium avium ssp. paratuberculosis is important for controlling this infection and minimizing economic losses. The objective of this study was to evaluate 2 bulk tank milk (BTM) paratuberculosis tests (PCR and modified ELISA) as potential herd-level tests. These tests were compared with the results obtained from testing 20 randomly selected lactating animals per farm (>2 yr) with an individual animal test (fecal culture, fecal PCR, serum ELISA, and milk ELISA). The study was conducted using 29 dairy goat herds and 21 dairy sheep flocks in Ontario, Canada, visited between October 2010 and August 2011. The sensitivity of the BTM PCR was poor in both the dairy goat herds (0.0%) and dairy sheep flocks (25.0%), but exhibited 100% specificity in both species. In comparison, the BTM modified ELISA demonstrated higher sensitivity. In goats, sensitivity ranged from 33.3 to 34.8% when fecal culture and PCR were the reference tests, respectively (specificities were both 100%), and 71.4 to 87.5% when the milk and serum ELISA, respectively, were the reference tests (specificities were 86.4 and 95.2%). The BTM modified ELISA in dairy sheep demonstrated comparable sensitivities, but lower specificities. When fecal culture and PCR were the reference test, sensitivities were 50.0 and 46.7%, respectively (specificities were 77.8 and 83.3%). The sensitivities when the milk and serum ELISA were the reference tests were 87.5 and 72.7%, respectively (specificities were 92.3 and 100%). Fecal PCR was the only individual animal test to identify significantly more farms as positive than the BTM PCR and modified ELISA test in both species. Therefore, whereas the BTM modified ELISA may provide an organization or control program with a high level of confidence that a BTM-positive farm is actually positive (high positive predictive value), if a producer wishes to increase the odds that a positive farm will test positive, so as not to miss an infection, then sampling and testing 20 animals with fecal PCR will better meet that objective.

Training method for animal-based measures in dairy cattle welfare assessments.

Quantitative assessments of animal welfare are increasingly being used in the dairy industry. It is important to have good precision and accuracy within and among assessors. This study explored the effectiveness of a 3-d training workshop for animal-based measures (ABM) of welfare in dairy cows, in which 14 people were trained to evaluate 6 ABM, specifically hock injuries (HI), lameness (LM), body condition score (BCS), and udder, flank, and leg cleanliness (collectively CLN). All scoring systems were modified to a dichotomous outcome, acceptable or unacceptable. Acceptable HI included no swelling or hair loss, unacceptable HI included swelling or scab; acceptable BCS was >2 on a 5-point scale, unacceptable BCS was ≤2. Acceptable CLN was up to minor splashing, unacceptable CLN was distinct plaques to a solid manure plaque. Lameness was evaluated using locomotion score or in-stall lameness score (SLS) in tiestalls; unacceptable LM was ≥3 (lameness score) on a 5-point scale, where 3 equals mild lameness or ≥2 of 4 behavioral in-stall lameness score indicators were detected. Classroom instruction took place on d 1 of training. Day 2 consisted of group assessment of LM (n = 25 cows), and HI, CLN, and BCS (n = 30 cows), and individual assessment of HI, CLN, and BCS (n = 20 cows) were performed. Day 3 included individual assessments of HI, CLN, and BCS (n = 33 cows), and individual video assessment of LM (n = 27 cows). An additional training video for LM was sent to trainees 3 wk after the workshop, and another follow-up assessment of LM took place via video (n = 37 cows). Repeatability and accuracy of the trainees was assessed using Fleiss's κ (FK) and Byrt's κ (BK) to examine group-level inter-rater agreement and expert-trainee agreement, respectively. The kappa systems use a scale of poor (<0), slight (0.01 to 0.20), fair (0.21 to 0.40), moderate (0.41 to 0.61), substantial (0.61 to 0.80), or almost perfect (0.81 to 1.00). At the conclusion of the workshop, FK was 0.66 for HI and 0.43 for LM, and BK mean (and range) was 0.85 (0.63 to 1.00) for HI and 0.66 (0.56 to 0.85) for LM. Each trainee achieved perfect agreement for BCS [BK mean = 1.00 (1.00 to 1.00)] and each trainee achieved almost perfect agreement for CLN [BK mean 0.90 (0.82 to 0.94)]. After the follow-up video and 3 wk of experience, trainees achieved a FK of 0.66 and a BK mean of 0.74 (0.62 to 0.89) for LM. In conclusion, multiple assessors can achieve substantial agreement for ABM with adequate training.

Canadian National Dairy Study: Herd-level milk quality.

The objective of this study was to estimate Canadian national milk quality parameters and estimate the bulk tank milk (BTM) prevalence of 4 mastitis pathogens, Staphylococcus aureus, Streptococcus agalactiae, Mycoplasma bovis, and Prototheca spp., on Canadian dairy farms. A questionnaire was sent to all Canadian dairy producers. Of the 1,062 producers who completed the questionnaire, 374 producers from across the country were visited and milking hygiene was assessed. Farm-level milk quality data for all Canadian dairy producers was collected from the provincial marketing boards and combined with the questionnaire and farm visit data. In addition, a BTM sample was collected either during the farm visit or by the marketing board in November of 2015 and was tested for 4 major mastitis pathogens using the PathoProof Mastitis Major 4 PCR Assay (Thermo Fisher Scientific Inc., Waltham, MA). Apparent herd-level prevalence was 46% for S. aureus, 6% for Prototheca spp., 0% for M. bovis, and 0% for Strep. agalactiae. Due to the low prevalence of M. bovis and Strep. agalactiae and a lack of significant factors associated with farms testing positive for Prototheca spp., an association analysis could only be carried out for Staph. aureus-positive farms. Factors associated with Staph. aureus-positive farms were not fore-stripping cows before milking (odds ratio = 1.87), milking with a pipeline system (odds ratio = 2.21), and stall bases made of a rubberized surface (mats and mattresses), whereas protective factors were using blanket dry cow therapy (odds ratio = 0.49) and applying a tag or visible mark on cows known to have chronic mastitis infections (odds ratio = 0.45). The Canadian national production-weighted geometric mean somatic cell count was determined to be 208,000 cells/mL. This is the first national dairy study conducted in Canada. Participating farms had higher milk yield; were more likely to have a loose housing system, parlor, or automated milking system; and had lower weighted mean BTM somatic cell count than the national level. Sampling larger farms with better milk quality means the apparent prevalence of the 4 mastitis pathogens likely underestimates the true levels.

The Canadian National Dairy Study 2015-Adoption of milking practices in Canadian dairy herds.

Several studies have investigated which management practices have the greatest effect on udder health, but little information is available on how broadly the recommended milking practices are adopted across Canada. The National Dairy Study 2015 was designed to gather dairy cattle health and management data on dairy farms across Canada. The objectives of the present study were to describe the current proportions of adoption of milking practices on Canadian dairy farms, and identify factors associated with their use on farms. A bilingual questionnaire measuring use of various practices, including an udder health-specific section, was developed and sent to all Canadian dairy farms. The questions in the udder health section of the questionnaire were adapted from a bilingual questionnaire previously validated and containing questions regarding general milking hygiene and routine, and on-farm mastitis management. Chi-squared tests were used to investigate simple associations between adoption of practices and various explanatory variables including region, milking system, herd size, and bulk tank somatic cell count. In total, 1,373 dairy producers completed the survey. The regional distribution of the participants was representative of the Canadian dairy farm population, and milk quality was, on average, similar to nonparticipants. Overall, Canadian dairy producers followed the recommendations for milking procedures, but some were more extensively used than others. Fore-stripping, cleaning teats, wiping teats dry, using single-cow towels, and use of postmilking teat disinfectant were widely adopted. Use of gloves and glove hygiene, use of a premilking teat disinfectant, and use of automatic takeoffs were not as extensively implemented. Adoption percentages for several practices, including use of gloves, use of a premilking teat disinfectant, teat drying methods, and use of automatic takeoffs were significantly associated with milking system, herd size, and region. It would be helpful to explore the reasons why producers choose to adopt or ignore recommended milking practices as most are easy to include in the routine and are fairly inexpensive.

Identifying management and disease priorities of Canadian dairy industry stakeholders.

The objective of this study was to identify the key management and disease issues affecting the Canadian dairy industry. An online questionnaire (FluidSurveys, http://fluidsurveys.com/) was conducted between March 1 and May 31, 2014. A total of 1,025 responses were received from across Canada of which 68% (n=698) of respondents were dairy producers, and the remaining respondents represented veterinarians, university researchers, government personnel, and other allied industries. Participants were asked to identify their top 3 management and disease priorities from 2 lists offered. Topics were subsequently ranked from highest to lowest using 3 different ranking methods based on points: 5-3-1 (5 points for first priority, 3 for second, and 1 for first), 3-2-1, and 1-1-1 (equal ranking). The 5-3-1 point system was selected because it minimized the number of duplicate point scores. Stakeholder groups showed general agreement with the top management issue identified as animal welfare and the number one health concern as lameness. Other areas identified as priorities were reproductive health, antibiotic use, bovine viral diarrhea, and Staphylococcus aureus mastitis with these rankings influenced by region, herd size, and stakeholder group. This is the first national comprehensive assessment of priorities undertaken in the Canadian dairy industry and will assist researchers, policymakers, program developers, and funding agencies make future decisions based on direct industry feedback.