The mammosphere-derived epithelial cell secretome modulates neutrophil functions in the bovine model.

Background: Innovative therapies against bacterial infections are needed. One approach is to focus on host-directed immunotherapy (HDT), with treatments that exploit natural processes of the host immune system. The goals of this type of therapy are to stimulate protective immunity while minimizing inflammation-induced tissue damage. We use non-traditional large animal models to explore the potential of the mammosphere-derived epithelial cell (MDEC) secretome, consisting of all bioactive factors released by the cells, to modulate host immune functions. MDEC cultures are enriched for mammary stem and progenitor cells and can be generated from virtually any mammal. We previously demonstrated that the bovine MDEC secretome, collected and delivered as conditioned medium (CM), inhibits the growth of bacteria in vitro and stimulates functions related to tissue repair in cultured endothelial and epithelial cells. Methods: The immunomodulatory effects of the bovine MDEC secretome on bovine neutrophils, an innate immune cell type critical for resolving bacterial infections, were determined in vitro using functional assays. The effects of MDEC CM on neutrophil molecular pathways were explored by evaluating the production of specific cytokines by neutrophils and examining global gene expression patterns in MDEC CM-treated neutrophils. Enzyme linked immunosorbent assays were used to determine the concentrations of select proteins in MDEC CM and siRNAs were used to reduce the expression of specific MDEC-secreted proteins, allowing for the identification of bioactive factors modulating neutrophil functions. Results: Neutrophils exposed to MDEC secretome exhibited increased chemotaxis and phagocytosis and decreased intracellular reactive oxygen species and extracellular trap formation, when compared to neutrophils exposed to control medium. C-X-C motif chemokine 6, superoxide dismutase, peroxiredoxin-2, and catalase, each present in the bovine MDEC secretome, were found to modulate neutrophil functions. Conclusion: The MDEC secretome administered to treat bacterial infections may increase neutrophil recruitment to the site of infection, stimulate pathogen phagocytosis by neutrophils, and reduce neutrophil-produced ROS accumulation. As a result, pathogen clearance might be improved and local inflammation and tissue damage reduced.

Greenhouse gas emissions and nutrient use efficiency assessment of six New York organic dairies.

Improving nutrient use efficiency and reducing greenhouse gas (GHG) emissions are important environmental priorities for organic-certified dairy operations. The objectives of this research were to quantify annual nutrient use and GHG emissions in 6 organic New York dairy farms. Farm-gate nutrient mass balances (NMB) were estimated with the Cornell NMB calculator. Whole-farm GHG emissions were estimated using Cool Farm Tool (CFT) and COMET. Farm-gate NMBs were low, ranging from -6.5 to 19 kg N ha-1 for N1 (without legume N fixation), 26 to 71 kg N ha-1 for N2 (including N fixation), -2.4 to 8.2 kg P ha-1 for P, and 1.1 to 19.8 kg K ha-1 for K. Additional nutrient imports, coupled with nutrient management planning, adequate legume stands and diet balancing may help improve P balances, and ensure no N deficiencies in the system. Estimates of annual GHG emission intensity ranged from 0.98 to 2.10 kg CO2-eq per kg of fat and protein corrected milk (FPCM) estimated by CFT, and from 0.69 to 2.48 kg CO2-eq kg FPCM-1 estimated by COMET. Enteric fermentation, feed production and fuel and energy use represented the largest sources of GHGs. For farms with liquid manure storages, manure management was also a significant source. Estimates of soil carbon (C) stock changes from CFT were in agreement or smaller than previous studies, and estimates from COMET were in agreement or greater. Variability and uncertainty in the results for soil C stock change indicate more research and new protocols are needed. Impact of individual management changes on GHG emissions intensity were small, ranging from -8 to +7% in CFT, and -8% to +8% in COMET. The management changes that resulted in the largest reductions in GHG emissions intensity included increasing individual cow productivity and milk to total feed ratio, and implementation of manure treatment systems.

Transition Cow Nutrition and Management Strategies of Dairy Herds in the Northeastern United States: Associations of Nutritional Strategies with Analytes, Health, Milk Yield, and Reproduction.

The objective was to identify relationships between transition cow nutritional strategies and the prevalence of elevated analytes (nonesterified fatty acids (NEFA), ß-hydroxybutyrate (BHB), and haptoglobin (Hp)), disorder incidence (DI), milk yield, and reproductive performance. Multiparous and primiparous cows from 72 farms in the northeastern US were enrolled in a herd-level cohort study. Farms were dichotomized within parity into a nutritional strategy within each period; far-off: controlled energy (CE; <16.5% starch and ≥40% forage neutral detergent fiber (FNDF)) or not CE (NCE; ≥16.5% starch or <40% FNDF or both), close-up: high FNDF (HF; ≥40% FNDF) or low FNDF (LF; <40% FNDF), and fresh: low starch (LS; <25.5% starch) or high starch (HS; ≥25.5% starch). No evidence existed that transition cow nutritional strategies were associated with milk yield outcomes (p ≥ 0.20). In general, our results support feeding multiparous cows HF close-up and HS fresh to minimize excessive BHB and DI; however, multiparous cows fed LF close-up had a higher pregnancy rate, and lower prepartum NEFA and Hp. Similarly, our results support feeding primiparous cows CE far-off, HF close-up, and HS fresh to maximize reproductive performance, and minimize BHB and DI; however, herds fed HF close-up or HS fresh had higher Hp.

Selective dry-cow therapy can be implemented successfully in cows of all milk production levels.

Antibiotic stewardship on dairy farms can be heightened through the implementation of selective dry-cow therapy (SDCT). However, some producers are concerned that this practice may be related to poor udder health outcomes in cows with high milk production at the time of dry-off. The objective of this study was to evaluate if the effect of culture-guided SDCT (Cult-SDCT) and algorithm-guided SDCT (Alg-SDCT) on dry-period intramammary infection (IMI) dynamics and postcalving udder health and performance [when compared with blanket dry-cow therapy (BDCT)] varied according to milk production level before dry-off. Data were compiled from clinical trials conducted in the United States and Canada that compared Cult-SDCT and Alg-SDCT to a positive control, i.e., BDCT. In those trials, cows were enrolled 1-2 d before dry-off, randomized to their dry-cow therapy strategy and followed until 120 d in milk of the subsequent lactation. The number of cows and quarters in the final data set were 1,485 and 5,097, respectively. Measured outcomes included quarter-level antibiotic use at dry-off, quarter-level IMI prevalence after calving, quarter-level dry-period IMI cure risk, quarter-level dry-period new IMI risk, cow-level clinical mastitis and removal from the herd during 1-120 d in milk, and somatic cell count and milk yield during 1-120 DIM. The primary objective of analysis was to investigate if the effect of Cult-SDCT and Alg-SDCT on these outcomes, when compared with BDCT, varied according to milk production level before dry-off. To do this, each cow was classified as having low, mid or high production, based on her milk yield tertile group at the most recent herd test before enrollment (low: <23.7 kg/d, mid: 23.7 to 30.4 kg/d, and high >30.4 kg/d). Multivariable generalized estimating equations were used to estimate risk differences and differences in means, and Cox regression was used to estimate hazard ratios. For Cult-SDCT, the proportion of quarters treated with dry-cow antibiotics within each milk production level were 40.7% (low), 41.7% (mid) and 47.2% (high). For Alg-SDCT, the proportions were 60.6% (low), 38.7% (mid), and 35.1% (high). Measures of udder health were not markedly different when comparing Cult-SDCT to BDCT and Alg-SDCT to BDCT. This was consistently observed in low, mid and high producing cows. In conclusion, the findings from this study indicate that Cult-SDCT and Alg-SDCT can be successfully implemented in cows of all milk production levels.

Contextualized understandings of dairy farmers' perspectives on antimicrobial use and regulation in Alberta, Canada.

Antimicrobial resistance (AMR) has been largely attributed to antimicrobial use (AMU). To achieve judicious AMU, much research and many policies focus on knowledge translation and behavioral change mechanisms. To address knowledge gaps in contextual drivers of decisions made by dairy farmers concerning AMU, we conducted ethnographic fieldwork to investigate one community's understanding of AMU, AMR, and associated regulations in the dairy industry in Alberta, Canada. This included participation in on-farm activities and observations of relevant interactions on dairy farms in central Alberta for 4 mo. Interviews were conducted with 25 dairy farmers. The interviews were analyzed using thematic analysis and yielded several key findings. Many dairy farmers in this sample: (1) value their autonomy and hope to maintain agency regarding AMU; (2) have shared cultural and immigrant identities which may inform their perspectives of future AMU regulation as it relates to their farming autonomy; (3) feel that certain AMU policies implemented in other contexts would be impractical in Alberta and would constrain their freedom to make what they perceive to be the best animal welfare decisions; (4) believe that their knowledge and experience are undervalued by consumers and policy makers; (5) are concerned that the public does not have a complex understanding of dairy farming and, consequently, worry that AMU policy will be based on misguided consumer concerns; and (6) are variably skeptical of a link between AMU in dairy cattle and AMR in humans due to their strict adherence to milk safety protocols that is driven by their genuine care for the integrity of the product. A better understanding of the sociocultural and political-economic infrastructure that supports such perceptions is warranted and should inform efforts to improve AMU stewardship and future policies regarding AMU.

Herd turnover rate reexamined: a tool for improving profitability, welfare, and sustainability.

Longevity and herd turnover rate are becoming common topics of discussion as the dairy industry strives for continuous improvement in efficiency, profitability, animal welfare, and environmental sustainability. Having the most productive animal fill each slot on a dairy makes strategic replacement and the resulting herd turnover an important tool for producers. Dairy operations can be considered to have slots available to be occupied by cows. The number of slots available is governed by dairy characteristics including parlor size and facility design. With sustainability and profitability goals, producers should aim to fill each slot with the most productive animal. The advantages of a modest surplus of replacement heifers allowing for a higher herd turnover rate are examined and shown to improve herd profitability, enhance welfare, and reduce environmental impact. A model assuming constant demand for dairy foods is presented with increased herd turnover rate leading to more milk production per cow and reduced enteric methane emissions. This analysis demonstrates that all else being equal, raising more replacements (having a relatively higher herd turnover rate and decreased herd-level longevity) improves sustainability compared to management aimed at lower herd turnover rates. Understanding the drivers of herd turnover in dairy production has important implications for the components of one health: animal well-being, food production, and environmental stewardship. The present work examines one tool toward this goal, while the companion Currents in One Health by Nguyen et al, JAVMA, January 2023, takes a broader view of many aspects of dairy sustainability.

Dairy production sustainability through a one-health lens.

Dairy production provides high-quality, healthful nutrients to people on a planet soon to be inhabited by over 9 billion people. In doing so, it is ever more important to continuously improve the care of dairy animals, safeguard the environment we all share, and reliably produce nutritious food while maintaining the economic viability of the people working in dairy agriculture. In this paper, we review some associations between dairy consumption and human health along with the many interconnections between people, dairy animals, plants, and our shared environment. Understanding these relationships is an example of one health at work. In the US, total dairy consumption is at its highest point in the last 50 years, while many objective measures of cow health (eg, subclinical mastitis) have never been better since they have been recorded. Further, indications of food safety such as violative antibiotic residues in milk have never been lower. Dairy foods provide essential nutrients such as protein, vitamin B12, and calcium, while there is also evidence that they are protective against chronic conditions such as cardiovascular disease. Finally, the environmental footprint of dairy production in the US, as measured by metrics such as carbon dioxide-equivalent emissions intensity per unit of dairy nutrient, is the lowest it has ever been. The companion Currents in One Health by Nguyen et al, AJVR, January 2023, discusses some additional animal welfare and environmental impact implications of modern dairy production management in detail.

Effects of dietary branched-chain amino acid supplementation on serum and milk metabolome profiles in dairy cows during early lactation.

The 3 branched-chain AA (BCAA), Val, Leu, and Ile, are essential AA used by tissues as substrates for protein synthesis and energy generation. In addition, BCAA are also involved in modulating cell signaling pathways, such as nutrient sensing and insulin signaling. In our previous study, dietary BCAA supplementation was shown to improve protein synthesis and glucose homeostasis in transition cows. However, a more detailed understanding of the changes in metabolic pathways associated with an increased BCAA availability is desired to fine-tune nutritional supplementation strategies. Multiparous Holstein cows (n = 20) were enrolled 28 d before expected calving and assigned to either the BCAA treatment (n = 10) or the control group (n = 10). Cows assigned to BCAA were fed 550 g/d of rumen-protected BCAA mixed with 200 g/d of dry molasses from calving until 35 DIM, whereas the cows assigned to the control were fed only 200 g/d of dry molasses. Serum samples were collected on d 10 before expected calving, as well as on d 4 and d 21 postpartum. Milk samples were collected on d 14 postpartum. From a larger cohort, we selected 20 BCAA-supplemented cows with the greatest plasma urea nitrogen concentration, as an indicator for greater BCAA availability, for the metabolomics analysis herein. Serum and milk samples were subjected to a liquid chromatography-mass spectrometry-based assay, detecting and measuring the abundance of 241 serum and 211 milk metabolic features, respectively. Multivariable statistical analyses revealed that BCAA supplementation altered the metabolome profiles of both serum and milk samples. Increased abundance of serum phosphocholine and glutathione and of milk Val, Ile, and Leu, and decreased abundance of milk acyl-carnitines were associated with BCAA supplementation. Altered phosphocholine and glutathione abundances point to altered hepatic choline metabolism and antioxidant balance, respectively. Altered milk acyl-carnitine abundances suggest changes in mammary fatty acid metabolism. Dietary BCAA supplementation was associated with a range of alterations in serum and milk metabolome profiles, adding to our understanding of the role of BCAA availability in modulating dairy cow protein, lipid, and energy metabolism on a whole-body level and how it affects milk composition.

Calculating clinical mastitis frequency in dairy cows: Incidence risk at cow level, incidence rate at cow level, and incidence rate at quarter level.

The lack of standardization in reporting clinical mastitis incidence limits the ability to compare results across multiple studies without additional calculations. There is both a biological and statistical rationale for evaluating the at-risk period at the quarter level. This study aimed to: (1) to outline an applied method for calculating clinical mastitis (CM) incidence rate at the quarter level using currently available software; and (2) to present the results of three different measurements: incidence risk at cow level, incidence rate at cow level, and incidence rate at quarter level. In an open population prospective cohort of eight commercial dairy farms monitored from May 15, 2016, to May 31, 2017, all CM cases (n = 7513) were identified by trained on-farm personnel, who collected all milk samples from all quarters with visibly abnormal milk. Microbiological identification was determined by culture and MALDI-TOF. All lactating quarters were at risk for CM. A quarter was at risk for a new CM case if there was at least 14 d between a previously diagnosed case and the current case in the same quarter, or if a different pathogen was isolated in the same quarter within 14 d. A total of 17,513,429 quarters days at risk (QDAR) were estimated. A statistical software macro and Structured Query Language (SQL) were used to bring all data together. The monthly incidence rate at the cow level was 16.6 cases per 10,000 cow-days, the monthly incidence rate at the quarter level was 4.4 cases per 10,000 QDAR and the monthly incidence risk at the cow level was 4.8 cases per 100 cows. Although the evaluation of QDAR requires additional computation when compared to other methods, it might allow for a more precise evaluation of the data and a more accurate evaluation of mastitis incidence. Clearly defining the methods used to report mastitis incidence will improve our ability to discuss and learn about the differences and similarities across studies, regions, and countries.

Characterization and comparison of the microbiomes and resistomes of colostrum from selectively treated dry cows.

Professionals in animal agriculture promote prudent use of antimicrobials to address public and animal health concerns, such as reduction of antimicrobial residues and antimicrobial resistance (AMR) in products. Few studies evaluate the effect of selective dry-cow therapy on preservation of the milk microbiome or the profile of AMR genes (the resistome) present at freshening. Our objectives were to characterize and compare the microbiomes and resistomes in the colostrum of cows with low somatic cell count that were treated or not treated with intramammary cephapirin benzathine at dry-off. From a larger parent study, cows on a New York dairy farm eligible for dry-off and with histories of somatic cell counts ≤200,000 cells/mL were enrolled to this study (n = 307). Cows were randomly assigned to receive an intramammary antimicrobial and external teat sealant (ABXTS) or sealant only (TS) at dry-off. Composite colostrum samples taken within 4 h of freshening, and quarter milk samples taken at 1 to 7 d in milk were subjected to aerobic culture. The DNA extraction was performed on colostrum from cows with culture-negative samples (ABXTS = 43; TS = 33). The DNA from cows of the same treatment group and parity were pooled (26 pools; ABXTS = 12; TS = 14) for 16S rRNA metagenomic sequencing. Separately, the resistome was captured using a custom RNA bait library for target-enriched sequencing. Sequencing reads were aligned to taxonomic and AMR databases to characterize the microbiome and resistome, respectively. The R statistical program was used to tabulate abundances and to analyze differences in diversity measures and in composition between treatment groups. In the microbiome, the most abundant phyla were Firmicutes (68%), Proteobacteria (23%), Actinobacteria (4%), and Bacteroidetes (3%). Shannon and richness diversity means were 0.93 and 14.7 for ABXTS and 0.94 and 13.1 for TS, respectively. Using analysis of similarities (ANOSIM), overall microbiome composition was found to be similar between treatment groups at the phylum (ANOSIM R = 0.005), class (ANOSIM R = 0.04), and order (ANOSIM R = -0.04) levels. In the resistome, we identified AMR gene accessions associated with 14 unique mechanisms of resistance across 9 different drug classes in 14 samples (TS = 9, ABXTS = 5). The majority of reads aligned to gene accessions that confer resistance to aminoglycoside (TS = ABXTS each 35% abundance), tetracycline (TS = 22%, ABXTS = 54%), and ß-lactam classes (TS = 15%, ABXTS = 12%). Shannon diversity means for AMR class and mechanism, respectively, were 0.66 and 0.69 for TS and 0.19 and 0.19 for ABXTS. Resistome richness diversity means for class and mechanism were 3.1 and 3.4 for TS and 1.4 and 1.4 for ABXTS. Finally, resistome composition was similar between groups at the class (ANOSIM R = -0.20) and mechanism levels (ANOSIM R = 0.01). Although no critical differences were found between treatment groups regarding their microbiome or resistome composition in this study, a larger sample size, deeper sequencing, and additional methodology is needed to identify more subtle differences, such as between lower-abundance features.

Description of the Characteristics of Five Bedding Materials and Association With Bulk Tank Milk Quality on Five New York Dairy Herds.

Environmental mastitis represents a major challenge on dairy farms where contagious pathogens are controlled by improved milking procedures. Therefore, research focused on the environment is important to improve udder health programs. The objectives of this prospective and descriptive study were to (1) describe bedding bacterial counts, pH, and dry matter (DM) of five different bedding types (organic: manure solids, straw, paper fiber; inorganic: sand, recycled sand) and (2) explore the association between bedding bacterial counts with bulk tank milk quality. This study took place within five conveniently selected commercial dairy herds, each with a predominant bedding material in lactating pens. Bedding samples (used n = 237; fresh n = 53) were collected monthly from July 2018 to July 2019 following a standard operating procedure (SOP) to minimize sampling variability. Additionally, a bulk tank (BT) milk sample (n = 40) was collected on the same day unless milk had been picked up prior to arrival. Both BT and bedding samples were submitted to the laboratory for culture and bacterial identification and quantification of Streptococcus spp, coliforms, and non-coliforms as well as detection of several pathogens of mastitis importance. Somatic cell count was evaluated in BT samples. Within bedding type, the correlation between bedding characteristics and bacterial counts in bedding was evaluated using Pearson correlation. Within bedding type, the correlation between bacterial counts in bedding samples and bacterial counts in BT were determined. The Kruskal-Wallis test was used to evaluate the bacterial count by bedding type and to evaluate BT somatic cell count differences based on bedding type. In fresh bedding, bacterial counts were generally higher for manure solids for all bacterial groups compared with other materials. In used samples, organic materials had the highest levels of all bacterial groups. The proportion of samples with detectable organisms of mastitis importance varied within and among herds in both bedding and BT samples throughout the study period. In bedding samples, a higher DM content had the lowest levels of bacterial growth compared with those with lower DM content. Most bedding samples were on the alkaline side within a pH range of 8-11. No relationship between bacterial counts and pH was observed. No associations between BT bacteria counts and bedding bacterial counts were observed. No association between bulk tank somatic cell counts based on bedding type were observed. Despite using an SOP for bedding sampling in an effort to consistently collect samples, we still observed a large amount of variability both within and among bedding samples. This variability may have obscured any potential association between BT milk quality and bedding type.

Evaluation of rapid culture, a predictive algorithm, esterase somatic cell count and lactate dehydrogenase to detect intramammary infection in quarters of dairy cows at dry-off.

Our objective was to compare four tests to standard milk culture followed by MALDI-ToF in quarters of cows at dry-off. Cows (n = 432) were randomly selected from seven U.S. dairy herds already participating in a multi-site clinical trial in summer 2018. Aseptic foremilk samples were collected from quarters (n = 1728) two days prior to dry-off, and subjected to index and reference tests. The four index tests included rapid culture, a predictive algorithm, an esterase strip test measuring somatic cell count (SCC) and a cow-side lactate dehydrogenase (LDH) test. Rapid culture was performed by inoculating quarter milk samples onto a commercial rapid culture plate. Plates were evaluated by technicians after 30-40 h of incubation at 37 ± 2 °C. Quarters were classified as infected if any bacterial growth was observed. For the algorithm test method, all quarters were classified as infected if the cow met any of the following criteria: 1) any Dairy Herd Improvement Association (DHIA) test with a SCC > 200,000 cells / ml during the current lactation or 2) two or more clinical mastitis cases during the current lactation. Esterase-SCC and cow-side LDH tests involved adding milk to the test strip and reading for color changes. For esterase-SCC and cow-side LDH tests, low (≥250 cells / ml and ≥100 U / L) and high (≥500 cells / ml and ≥200 U / L) thresholds were used to classify quarters as infected or not. Composite samples (4 × 2 mL quarter-milk samples commingled) were also tested for rapid culture, esterase-SCC and cow-side LDH tests, such that if a composite sample was positive, then all quarters contributing to that sample were classified as infected. The reference test was traditional aerobic culture conducted in an accredited laboratory using MALDI-ToF for identification of isolates. Traditional culture was only conducted on quarter-milk samples, and consequently, IMI was always considered at the quarter-level. Unconditional logistic regression was used to estimate sensitivity (SE), specificity (SP), apparent prevalence, positive predictive values (PPV) and negative predictive values (NPV) for each index test. Cohen's Kappa (κ) was used to measure agreement between tests. Algorithm, esterase-SCC and cow-side LDH tests had poor agreement with the reference test (κ ranging from 0.01 to 0.12), while rapid culture had fair agreement (κ = 0.28). No test had concurrently high SE and SP. Negative predictive values were moderate to high for all tests.

Longitudinal Phenotypes Improve Genotype Association for Hyperketonemia in Dairy Cattle.

The objective of our study was to identify genomic regions associated with varying concentrations of non-esterified fatty acid (NEFA), ß-hydroxybutyrate (BHB), and the development of hyperketonemia (HYK) in longitudinally sampled Holstein dairy cows. Our study population consisted of 147 multiparous cows intensively characterized by serial NEFA and BHB concentrations. To identify individuals with contrasting combinations in longitudinal BHB and NEFA concentrations, phenotypes were established using incremental area under the curve (AUC) and categorized as follows: Group (1) high NEFA and high BHB, group (2) low NEFA and high BHB), group (3) low NEFA and low BHB, and group (4) high NEFA and low BHB. Cows were genotyped on the Illumina Bovine High-density (777K) beadchip. Genome-wide association studies using mixed linear models with the least-related animals were performed to establish a genetic association with HYK, BHB-AUC, NEFA-AUC, and the comparisons of the 4 AUC phenotypic groups using Golden Helix software. Nine single-nucleotide polymorphisms were associated with high longitudinal concentrations of BHB and further investigated. Five candidate genes related to energy metabolism and homeostasis were identified. These results provide biological insight and help identify susceptible animals thus improving genetic selection criteria thereby decreasing the incidence of HYK.

Understanding the effect of producers' attitudes, perceived norms, and perceived behavioral control on intentions to use antimicrobials prudently on New York dairy farms.

Understanding farmers' behavior, motivations, and perceptions toward antimicrobial use can influence how veterinarians translate research into practice and guide effective ways of implementing protocols. A multidisciplinary team investigated behavioral tendencies of New York dairy farmers toward antimicrobial use by administering a survey modeled with the reasoned action approach. This approach is a framework from social psychology containing the constructs attitude, perceived norms, and perceived behavioral control, and is used in combination with structural equation modeling to determine what drives intentions. Multiple indicators and multiple causes (MIMIC) models were then used to determine the effects of beliefs on their underlying constructs. The objective of the study was to provide direct and indirect measures of the constructs using survey data to determine importance of and associations with intention to use antimicrobials prudently. The structural equation model indicated that perceived behavioral control explained intention. Thus, farmers who feel capable of prudent use expressed positive intentions. Attitude and perception of others also had influence to a lesser extent. MIMIC models showed that the most important attributes of instrumental attitude were increasing profitability, decreasing risk of residues, and increasing herd health. Contributing attributes of affective attitude were job satisfaction, decreasing resistance, and increasing milk production. For perceived norms, the attributes were opinions/approval of family and peers, veterinarians, and milk processors. Finally, for perceived behavioral control, attributes focused on saving money on labor and treatment, ability to fit into the daily routine, and effectiveness with veterinary guidance. In conclusion, the best approach for adoption of practices might be presentation of examples of successful strategies by other producers, particularly in peer groups. In addition, veterinarians should provide the tools and guidance needed to produce economic gain, reduction of risks associated with residues and resistance, and positive experiences when using the tactics.

Short communication: Oral and intranasal administration of a modified-live Salmonella Dublin vaccine in dairy calves: Clinical efficacy and serologic response.

Our objectives were to evaluate the clinical efficacy of oral and intranasal administration of a commercial modified-live Salmonella Dublin vaccine in dairy calves and to determine the serologic response associated with these extralabel routes of administration. We conducted a randomized field trial with calves from a New York dairy farm following an outbreak of Salmonella Dublin. A total of 399 Holstein calves were allocated by pen to 3 treatment groups: oral vaccination, intranasal vaccination, and an unvaccinated control group. Administration of the vaccine through oral and intranasal routes did not have a significant effect on pneumonia incidence risk or weight gain; however, calves vaccinated orally and intranasally had lower mortality risk as compared with control calves. Among calves tested using a Salmonella Dublin ELISA, vaccination did not induce an increase in antibody production relative to control calves, indicating that oral and intranasal administration will not hinder diagnosis based on this assay.

A longitudinal prospective cohort study investigating the association of premilking stimulation and teat-end shape on milking characteristics and teat tissue condition in dairy cows.

BACKGROUND: Premilking udder preparation is essential for harvesting high-quality milk as gently, completely, and quickly as possible. The associations between characteristics such as teat-end shape and premilking stimulation on milking characteristics and machine milking-induced changes to the teat tissue condition have not been rigorously investigated. The primary objective was to investigate the interactive effects of manual premilking stimulation (i.e., preparation lag time) and teat-end shape on total milk yield, two-minute milk yield, milking unit-on time, and time in low milk flow rate. Our secondary objective was to study the association of manual premilking stimulation and changes to the teat tissue condition after machine milking (i.e., short-term changes). In a longitudinal prospective cohort study, 384 milking observations from 129 cows were analysed. Holstein cows were housed in sand-bedded free-stall pens, fed a total mixed ration, and milked 3 times a day. Cows were classified by teat-end shape into 1 of 3 categories: pointed, flat, or round. Individual cow milking characteristics were recorded with electronic on-farm milk meters. The duration of manual stimulation, preparation lag time, and presence of short-term changes were documented for each milking observation. General linear mixed models were used to study the interactive effects of preparation lag time and teat-end shape on milking characteristics. RESULTS: There was an interaction between preparation lag time and teat-end shape for two-minute milk yield and time in low milk flow rate. The preparation lag time effect was modified by teat-end shape, while no interaction was observed for total milk yield or milking unit-on time. A generalized linear mixed model revealed that preparation lag time was associated with short-term changes in teat tissue condition, where the odds of short-term changes decreased as preparation lag time increased. CONCLUSIONS: In summary, cows with different teat-end shapes may require different premilking stimulation regimens. Increasing preparation lag time benefits teat tissue condition during machine milking. Further research is warranted to optimize individual premilking stimulation in dairy cows.

The effect of the transition period and postpartum body weight loss on macrophage infiltrates in bovine subcutaneous adipose tissue.

The transition period in dairy cattle is characterized by a rapid increase in metabolic demands due to the onset of lactation in parallel with a voluntary drop in dry matter intake, which slowly increases compared with the rapidly increasing milk production. The resulting deficit in energy intake is largely met by lipolysis of adipose tissue stores liberating nonesterified fatty acids (NEFA) as energy substrates to support milk production and for the synthesis of milk fat. Previous work in cattle and other species has documented an increase in adipose tissue macrophages (ATM) during periods of feed restriction and lipolysis; however, alterations in ATM during the transition period have only recently received interest because of the role that adipose tissue remodeling and inflammation may play during this time. The primary objective of this study was to evaluate the number of macrophages in subcutaneous adipose tissue from dairy cattle at multiple time points throughout the transition period. Secondary objectives included the evaluation for potential associations of ATM with changes in body condition score (BCS) and concentrations of circulating NEFA and ß-hydroxybutyrate. Holstein cows (n = 25) were enrolled in the study 28 d before expected parturition and fed the same prepartum and postpartum rations for ad libitum consumption. On d 10 before expected parturition, and on d 4 and 21 after calving, subcutaneous adipose tissue biopsies were taken from the paralumbar fossa and preserved in formalin. Paraffin-embedded sections of tissue from each of the 25 cows at the 3 time points (total of 75 samples) were labeled with an immunohistochemical marker of macrophages, ionized calcium-binding adapter molecule 1 (Iba1, also known as allograft inflammatory factor 1). The number of Iba1-positive cells, the number of crown-like structures, and the number of immunopositive aggregates were enumerated in each sample. The number of Iba1-positive macrophages increased by 60 and 93% on d 4 and 21, respectively, when compared with the average prepartum number. Additionally, cows that lost ≥0.50 points of BCS had a 57 and 52% higher number of ATM on d 4 and 21 postpartum, respectively, when compared with cows that lost ≤0.25 points of BCS. No association was found between ATM numbers and NEFA and ß-hydroxybutyrate concentrations or milk production in early lactation. Cows experiencing a more pronounced decrease in BCS postpartum had the largest increase in adipose tissue macrophage counts, suggesting that a greater degree of adipose tissue remodeling during the period of greatest nutrient deficit may be associated with higher numbers of ATM.

Association of herd-level risk factors and incidence rate of clinical mastitis in 20 Brazilian dairy herds.

The objectives of this study were to characterize the pathogen frequency and severity of clinical mastitis (CM) in 20 dairy herds of southeastern Brazil; and to determine the incidence rate of clinical mastitis (IRCM; overall and based on specific-pathogen groups) based on quarter time at risk and its association with risk factors at the herd-level. Data were recorded in each herd for a period of 8 to 15 months. The association between herd-level risk factors and IRCM were determined by two groups of mixed regression models: one based on the overall IRCM, and five based on the following specific-pathogen groups: contagious, other Gram-positive, Gram-negative, other, and negative culture. The following herd-level risk factors were evaluated: herd size, housing system, average daily milk yield per cow, bulk milk somatic cell count (BMSCC), and bulk milk total bacterial count (BMTBC). A total of 5957 quarter-cases of CM were recorded from 2637 cows, but only 4212 cases had milk samples collected for culture. The most frequently isolated pathogens were Escherichia coli (6.6% of total cultures), Streptococcus uberis (6.1%), and Streptococcus agalactiae (5.9%). The majority of CM cases were mild (60.3%), while 34.1% were moderate and 5.6% severe. The frequency of severe CM cases was lower for those with a Gram-positive result (4.6%) compared to a Gram-negative result (11.4%). Overall, monthly mean IRCM was 9.7 cases per 10,000 quarter-days at risk (QDAR). Herds with a geometric mean BMSCC ≥ 601 × 103 cell/mL had higher overall IRCM (16/10,000 QDAR) than those with BMSCC ≤ 600 × 103 cell/mL (≤7.7/10,000 QDAR). When the specific-pathogen groups were evaluated, for contagious pathogens, variables housing (free-stalls or compost-bedded pack barns), BMSCC (≥601 × 103 cells/mL), and average daily milk yield per cow (21 and 25 Kg/d) presented the highest IRCM. Furthermore, in Gram-negative group, herds with BMTBC ≥ 31 × 103 cfu/mL had higher IRCM compared with herds with BMTBC ≤ 30 × 103 cfu/mL. Although environmental pathogens were the most common cause of CM in this study, contagious pathogens (e.g., Strep. agalactiae and Staph. aureus) are still a concern in dairy herds of Brazil. Additionally, as there were some herd-level risk factors associated with the IRCM, there may be opportunity for management strategies aiming to improve the control of CM in dairy herds.

The secretome from bovine mammosphere-derived cells (MDC) promotes angiogenesis, epithelial cell migration, and contains factors associated with defense and immunity.

Treatment of bovine mastitis with intramammary antibiotics is common, yet several concerns exist including failed efficacy for individual hosts or pathogens and the inability of approved drugs to revert mastitis-induced tissue damage to healthy tissue capable of returning to full milk production. These issues, in addition to aspects of public health such as accidental antibiotic residues in saleable milk and the potential for antimicrobial resistance, support the need to find alternative therapies for this costly disease. This study shows that the secretome, or collective factors, produced by mammosphere-derived cells (MDC) promotes angiogenesis, epithelial cell migration, and contains proteins associated with immunity and defense; all of which are necessary for healing damaged mammary gland tissue. Furthermore, we found that the MDC secretome remains effective after freezing and thawing, enhancing its therapeutic potential. Our results provide a foundation for further characterization of the individual secreted factors and the rationale for using the MDC secretome as a complementary treatment for bovine mastitis.

SURVEILLANCE FOR HEMORRHAGIC SEPTICEMIA IN BUFFALO ( BUBALUS BUBALIS) AS AN AID TO RANGE EXPANSION OF THE JAVAN RHINOCEROS ( RHINOCEROS SONDAICUS) IN UJUNG KULON NATIONAL PARK, INDONESIA.

The Javan rhinoceros ( Rhinoceros sondaicus) of Ujung Kulon National Park (UKNP) is the crown jewel of Indonesia's rich natural history. The park lies on a peninsula surrounded by coastline and agriculture-dominated landscapes. The invasion of water buffalo ( Bubalus bubalis) into the park carries a substantial health risk to the Javan rhinoceros and threatens plans to establish a new population outside of its only current range in UKNP. Hemorrhagic septicemia (HS), known locally as septicemia epizootica and caused by Pasteurella multocida B:2, could thwart Indonesia's efforts to expand the range of the Javan rhinoceros. Because HS was considered eradicated from Banten Province, few preventative programs have been available to farmers. During June 2012-July 2013, biologic samples were collected from 770 water buffalo in 19 villages. Deep nasal swabs ( n=85) were taken for bacterial culture and blood samples ( n=770) were collected for serologic testing. No animals were positive on culture. The prevalence of antibody to P. multocida in this population was 1.8% (14 of 770 animals). A structured questionnaire was used to gather information about possible risk factors. Husbandry practices associated with presence of antibody in water buffalo included lack of a permanent area to house buffalo at night, low body condition score (=2), high body temperature (≥40 C), a history of clinical signs or sudden death in the previous year, and a grazing system that utilized significant forage inside the park. Antibody was not associated with sex, age, vaccination status, or season. Understanding HS disease dynamics in the buffalo adjacent to UKNP may improve the livelihoods of people and health of endangered rhinoceroses in this ecosystem.