Effect of Holstein genotype on immune response to an intramammary Escherichia coli challenge.

Selective breeding of US dairy cows since the mid-1960s has contributed to remarkable gains in milk yield per cow. This increased milk yield has been associated with an increase in health issues. Since 1964, the University of Minnesota has selectively bred a Holstein herd to maintain genetically static, unselected Holsteins (UH). Comparison of these UH cows with contemporary Holsteins (CH) has demonstrated that the UH cows not only produce less milk but also have fewer health concerns than their CH herdmates. The objective of this study was to determine the effects of Holstein genotype on innate immune response in an experimental intramammary Escherichia coli challenge model. Primiparous UH (n = 5) and CH (n = 7) cows received 430 cfu of E. coli strain P4 in 1 quarter. Blood and affected quarter milk samples were collected at 0, 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 7, 9, and 11 d relative to E. coli infusion. Rectal temperatures were recorded at each milking through d 4 of the experiment. Milk bacterial counts, somatic cell count and BSA concentrations, complete blood cell counts, rectal temperature, and serum and milk whey cytokine (IL-1ß and IL-6) concentrations were used as metrics to determine infection severity. Longitudinal (repeated) data were analyzed using general linear models with PROC MIXED with day of study as the repeated effect. Whole blood transcriptomes were generated by RNA sequencing. Transcripts with a false discovery rate of P < 0.05 and a delta log2 expression value greater than 0.7 or less than -0.7 were used for functional enrichment analysis. Bacterial counts were consistently greater in milk from CH than UH cows from d 0.25 through d 2.5. Milk somatic cell count increased within 6 h (d 0.25) after E. coli administration in CH and UH cows but did not differ between genotypes after d 1. Rectal body temperature peaked at d 1 in CH and UH cows but was greater in CH cows. Milk BSA, IL-1ß, and IL-6 concentrations were greater in CH than UH cows after E. coli administration. Blood lymphocyte and neutrophil counts were decreased at 0.5 and 1 d in CH but not in UH cows. The number of differentially expressed transcripts at each of the postinfusion sampling times was consistently greater (4- to 90-fold) in CH than in UH cows. A key difference between the immune reaction of the 2 genotypes was that the immune response to E. coli was largely contained within the mammary gland of the UH cows but became more systemic in the CH cows. These data demonstrate that UH cows exerted more effective control of E. coli infused into the mammary gland and thus support the hypothesis that selection practices since the mid-1960s have resulted in CH cows with an immune system that is less effective in fighting intramammary infections. Identification of genetic factors associated with enhanced immune functions that differ between the UH and CH cows could contribute to efforts to reintroduce or enhance beneficial components that have been lost or reduced in the CH cows since the mid-1960s.

Bovine NK-lysin-derived peptides have bactericidal effects against Mycobacterium avium subspecies paratuberculosis.

Infection with Mycobacterium avium subspecies paratuberculosis (MAP) is complex, but little is known about the role that natural killer (NK) cells play. In the present study, four bovine NK-lysin peptides were synthesized to evaluate their bactericidal activity against MAP. The results demonstrated that bNK-lysin peptides were directly bactericidal against MAP, with bNK1 and bNK2A being more potent than bNK2B and bNK2C. Mechanistically, transmission electron microscopy revealed that the incubation of MAP with bNK2A resulted in extensive damage to cell membranes and cytosolic content leakage. Furthermore, the addition of bNK2A linked with a cell-penetrating peptide resulted in increased MAP killing in a macrophage model.

Milk biosynthesis requires the Golgi cation exchanger TMEM165.

Milk is a hallmark of mammals that is critical for normal growth and development of offspring. During biosynthesis of lactose in the Golgi complex, H+ is produced as a by-product, and there is no known mechanism for maintaining luminal pH within the physiological range. Here, using conditional, tissue-specific knockout mice, immunostaining, and biochemical assays, we test whether the putative H+/Ca2+/Mn2+ exchanger known as TMEM165 (transmembrane protein 165) participates in normal milk production. We find TMEM165 is crucial in the lactating mammary gland for normal biosynthesis of lactose and for normal growth rates of nursing pups. The milk of TMEM165-deficient mice contained elevated concentrations of fat, protein, iron, and zinc, which are likely caused by decreased osmosis-mediated dilution of the milk caused by the decreased biosynthesis of lactose. When normalized to total protein levels, only calcium and manganese levels were significantly lower in the milk from TMEM165-deficient dams than control dams. These findings suggest that TMEM165 supplies Ca2+ and Mn2+ to the Golgi complex in exchange for H+ to sustain the functions of lactose synthase and potentially other glycosyl-transferases. Our findings highlight the importance of cation and pH homeostasis in the Golgi complex of professional secretory cells and the critical role of TMEM165 in this process.

Case report: characterization of a persistent, treatment-resistant, novel Staphylococcus aureus infection causing chronic mastitis in a Holstein dairy cow.

BACKGROUND: Mastitis is the most common health concern plaguing the modern dairy cow and costs dairy producers estimates of two billion dollars annually. Staphylococcus aureus infections are prevalent, displaying varied disease presentation and markedly low cure rates. Neutrophils are considered the first line of defense against mastitis causing bacteria and are frequently targeted in the development of treatment and prevention technologies. We describe a case of naturally occurring, chronic mastitis in a Holstein cow (1428), caused by a novel strain of S. aureus that was not able to be cleared by antibiotic treatment. CASE PRESENTATION: The infection was identified in a single quarter, 2 months into the cow's first lactation. The infection persisted for the following 20 months, including through dry off, and a second calving and lactation. This case of mastitis was associated with a consistently high somatic cell count, however presented with no other clinical signs. This cow was unsuccessfully treated with antibiotics commonly used to treat mastitis, consisting of two rounds of treatment during lactation and an additional round at the beginning of dry off. The chronic infection was also unchanged through an experimental mid-lactation treatment with pegylated granulocyte-colony stimulating factor (PEG-gCSF) and an additional periparturient treatment with PEG-gCSF. We isolated milk neutrophils from 1428 and compared them to two cows challenged with experimental S. aureus, strain Newbould 305. Neutrophils from 1428's milk had higher surface expression of myeloperoxidase compared to experimental Newbould challenged animals, as well as increased presence of Neutrophil Extracellular Traps. This suggests a heightened activation state of neutrophils sourced from 1428's naturally occurring infection. Upon postmortem examination, the affected quarter revealed multifocal abscesses separated by fibrous connective tissues. Abscesses were most common in the gland cistern and collecting duct region. Microscopically, the inflammatory reaction was pyogranulomatous to granulomatous and consistent with botryomycosis. Colonies of Gram-positive cocci were found within the eosinophilic matrix of the Splendore-Hoeppli reaction within granulomas and intracellularly within the acinar epithelium. CONCLUSIONS: Collectively, we describe a unique case of chronic mastitis, the characterization of which provides valuable insight into the mechanics of S. aureus treatment resistance and immune escape.

Wildland firefighter smoke exposure and risk of lung cancer and cardiovascular disease mortality.

Wildland firefighters are exposed to wood smoke, which contains hazardous air pollutants, by suppressing thousands of wildfires across the U. S. each year. We estimated the relative risk of lung cancer and cardiovascular disease mortality from existing PM2.5 exposure-response relationships using measured PM4 concentrations from smoke and breathing rates from wildland firefighter field studies across different exposure scenarios. To estimate the relative risk of lung cancer (LC) and cardiovascular disease (CVD) mortality from exposure to PM2.5 from smoke, we used an existing exposure-response (ER) relationship. We estimated the daily dose of wildfire smoke PM2.5 from measured concentrations of PM4, estimated wildland firefighter breathing rates, daily shift duration (hours per day) and frequency of exposure (fire days per year and career duration). Firefighters who worked 49 days per year were exposed to a daily dose of PM4 that ranged from 0.15 mg to 0.74 mg for a 5- and 25-year career, respectively. The daily dose for firefighters working 98 days per year of PM4 ranged from 0.30 mg to 1.49 mg. Across all exposure scenarios (49 and 98 fire days per year) and career durations (5-25 years), we estimated that wildland firefighters were at an increased risk of LC (8 percent to 43 percent) and CVD (16 percent to 30 percent) mortality. This unique approach assessed long term health risks for wildland firefighters and demonstrated that wildland firefighters have an increased risk of lung cancer and cardiovascular disease mortality.

Factors affecting smoke and crystalline silica exposure among wildland firefighters.

Smoke exposure data among U.S. wildland firefighters for carbon monoxide, respirable particulate and respirable crystalline silica are presented from a field surveillance program between 2009 and 2012. Models to predict fireline-average exposure to each inhalation hazard were developed and fit to the available data. The models identify factors to consider when defining similar exposure groups and designing future data collection. Task-based rather than shift-average data collection is important because the work activity representing the majority of fireline time, the position up- or downwind of the fire, and the proportion of time this combination represented were significant factors in the model for carbon monoxide, and all but wind position were significant for respirable particulate matter. The wind position versus the fire was not important for respirable quartz exposure. The crew type was an important factor in each model.

Carbon monoxide exposures among U.S. wildland firefighters by work, fire, and environmental characteristics and conditions.

Carbon monoxide (CO) exposure levels encountered by wildland firefighters (WLFs) throughout their work shift can change considerably within a few minutes due to the varied tasks that are performed and the changing environmental and fire conditions encountered throughout the day. In a U.S. Forest Service study during the 2009-2012 fire seasons, WLFs from 57 different fires across the U.S. were monitored for CO using CO data-logging detectors while an observer recorded worker tasks, fire characteristics, and environmental conditions at scheduled intervals. Exposures to CO for 735 WLF's work shifts were analyzed to assess the effect of variations among work tasks, fire characteristics, and environmental conditions. Geometric mean full shift time-weighted averages were low at 2.4 parts per million (ppm) and average length of work shift was 11 hr and 15 min. The task with the highest mean CO exposure was sawyer/swamper at 6.8 ppm; workers performing that task had an estimated 9 times higher odds of a having a 1-min CO measurement exceeding 25 ppm than the referent pump task (OR = 8.89, 95% CI = 1.97, 40.24). After adjusting CO exposure limits for shift length, elevation, and work level, 2% and 4% of the WLF's work shifts exceeded the National Institute for Occupational Safety and Health's recommended exposure level and the American Conference of Governmental Industrial Hygienist's threshold limit value, respectively. In regression modeling, variables that were significantly associated with elevated levels of CO exposure included: task, fuel model, wind orientation, crew type, relative humidity, type of attack, and wind speed. In the absence of instruments such as CO detectors that can determine and alert WLFs to elevated CO levels, recognition of the conditions that lead to elevated levels of CO exposure can assist WLFs to effectively use administrative controls, such as work rotations, to minimize exposures.

Genomic and Transcriptomic Analysis of Escherichia coli Strains Associated with Persistent and Transient Bovine Mastitis and the Role of Colanic Acid.

Escherichia coli is a leading cause of bacterial mastitis in dairy cattle. It is most often transient in nature, causing an infection that lasts 2 to 3 days. However, E. coli has been shown to cause a persistent infection in a minority of cases. Mechanisms that allow for a persistent E. coli infection are not fully understood. The goal of this work was to determine differences between E. coli strains originally isolated from dairy cattle with transient and persistent mastitis. Using RNA sequencing, we show gene expression differences in nearly 200 genes when bacteria from the two clinical phenotypes are compared. We sequenced the genomes of the E. coli strains and report genes unique to the two phenotypes. Differences in the wca operon, which encodes colanic acid, were identified by DNA as well as RNA sequencing and differentiated the two phenotypes. Previous work demonstrated that E. coli strains that cause persistent infections were more motile than those that cause transient infections. Deletion of genes in the wca operon from a persistent-infection strain resulted in a reduction of motility as measured in swimming and swarming assays. Furthermore, colanic acid has been shown to protect bacteria from complement-mediated killing. We show that transient-infection E. coli strains were more sensitive to complement-mediated killing. The deletion of genes from the wca operon caused a persistent-infection E. coli strain to become sensitive to complement-mediated killing. This work identifies important differences between E. coli strains that cause persistent and transient mammary infections in dairy cattle.

Parainfluenza Virus 3 Blocks Antiviral Mediators Downstream of the Interferon Lambda Receptor by Modulating Stat1 Phosphorylation.

UNLABELLED: Parainfluenza viruses are known to inhibit type I interferon (IFN) production; however, there is a lack of information regarding the type III IFN response during infection. Type III IFNs signal through a unique heterodimeric receptor, IFN-λR1/interleukin-10R2 (IL-10R2), which is primarily expressed by epithelial cells. Parainfluenza virus 3 (PIV-3) infection is highly restricted to the airway epithelium. We therefore sought to examine type III IFN signaling pathways during PIV-3 infection of epithelial cells. We used three strains of PIV-3: human PIV-3 (HPIV-3), bovine PIV-3 (BPIV-3), and dolphin PIV-1 (Tursiops truncatus PIV-1, or TtPIV-1). Here, we show that message levels of IL-29 are significantly increased during PIV-3 infection, yet downstream antiviral signaling molecules are not upregulated to levels similar to those of the positive control. Furthermore, in Vero cells infected with PIV-3, stimulation with recombinant IL-29/-28A/-28B does not cause upregulation of downstream antiviral molecules, suggesting that PIV-3 interferes with the JAK/STAT pathway downstream of the IFN-λR1/IL-10R2 receptor. We used Western blotting to examine the phosphorylation of Stat1 and Stat2 in Vero cells and the bronchial epithelial cell line BEAS-2B. In Vero cells, we observed reduced phosphorylation of the serine 727 (S727) site on Stat1, while in BEAS-2B cells Stat1 phosphorylation was decreased at the tyrosine 701 (Y701) site during PIV-3 infection. PIV-3 therefore interferes with the phosphorylation of Stat1 downstream of the type III IFN receptor. These data provide new evidence regarding strategies employed by parainfluenza viruses to effectively circumvent respiratory epithelial cell-specific antiviral immunity. IMPORTANCE: Parainfluenza virus (PIV) in humans is associated with bronchiolitis and pneumonia and can be especially problematic in infants and the elderly. Also seen in cattle, bovine PIV-3 causes respiratory infections in young calves. In addition, PIV-3 is one of a number of pathogens that contribute to the bovine respiratory disease complex (BRDC). As their name suggests, interferons (IFNs) are produced by cells to interfere with viral replication. Paramyxoviruses have previously been shown to block production and downstream signaling of type I IFNs. For the first time, it is shown here that PIV-3 can induce protective type III IFNs in epithelial cells, the primary site of PIV-3 infection. However, we found that PIV-3 modulates signaling pathways downstream of the type III IFN receptor to block production of several specific molecules that aid in a productive antiviral response. Importantly, this work expands our understanding of how PIV-3 effectively evades host innate immunity.

Review of the health effects of wildland fire smoke on wildland firefighters and the public.

Each year, the general public and wildland firefighters in the US are exposed to smoke from wildland fires. As part of an effort to characterize health risks of breathing this smoke, a review of the literature was conducted using five major databases, including PubMed and MEDLINE Web of Knowledge, to identify smoke components that present the highest hazard potential, the mechanisms of toxicity, review epidemiological studies for health effects and identify the current gap in knowledge on the health impacts of wildland fire smoke exposure. Respiratory events measured in time series studies as incidences of disease-caused mortality, hospital admissions, emergency room visits and symptoms in asthma and chronic obstructive pulmonary disease patients are the health effects that are most commonly associated with community level exposure to wildland fire smoke. A few recent studies have also determined associations between acute wildland fire smoke exposure and cardiovascular health end-points. These cardiopulmonary effects were mostly observed in association with ambient air concentrations of fine particulate matter (PM2.5). However, research on the health effects of this mixture is currently limited. The health effects of acute exposures beyond susceptible populations and the effects of chronic exposures experienced by the wildland firefighter are largely unknown. Longitudinal studies of wildland firefighters during and/or after the firefighting career could help elucidate some of the unknown health impacts of cumulative exposure to wildland fire smoke, establish occupational exposure limits and help determine the types of exposure controls that may be applicable to the occupation.

Vitamin D status of dairy cattle: Outcomes of current practices in the dairy industry.

The need for vitamin D supplementation of dairy cattle has been known for the better part of the last century and is well appreciated by dairy producers and nutritionists. Whether current recommendations and practices for supplemental vitamin D are meeting the needs of dairy cattle, however, is not well known. The vitamin D status of animals is reliably indicated by the concentration of the 25-hydroxyvitamin D [25(OH)D] metabolite in serum or plasma, with a concentration of 30ng/mL proposed as a lower threshold for sufficiency. The objective of this study was to determine the typical serum 25(OH)D concentrations of dairy cattle across various dairy operations. The serum 25(OH)D concentration of 702 samples collected from cows across various stages of lactation, housing systems, and locations in the United States was 68±22ng/mL (mean ± standard deviation), with the majority of samples between 40 and 100ng/mL. Most of the 12 herds surveyed supplemented cows with 30,000 to 50,000 IU of vitamin D3/d, and average serum 25(OH)D of cows at 100 to 300 DIM in each of those herds was near or above 70ng/mL regardless of season or housing. In contrast, average serum 25(OH)D of a herd supplementing with 20,000 IU/d was 42±15ng/mL, with 22% below 30ng/mL. Cows in early lactation (0 to 30d in milk) also had lower serum 25(OH)D than did mid- to late-lactation cows (57±17 vs. 71±20ng/mL, respectively). Serum 25(OH)D of yearling heifers receiving 11,000 to 12,000 IU of vitamin D3/d was near that of cows at 76±15ng/mL. Serum 25(OH)D concentrations of calves, on the other hand, was 15±11ng/mL at birth and remained near or below 15ng/mL through 1mo of age if they were fed pasteurized waste milk with little to no summer sun exposure. In contrast, serum 25(OH)D of calves fed milk replacer containing 6,600 and 11,000 IU of vitamin D2/kg of dry matter were 59±8 and 98±33ng/mL, respectively, at 1mo of age. Experimental data from calves similarly indicated that serum 25(OH)D achieved at approximately 1mo of age would increase 6 to 7ng/mL for every 1,000 IU of vitamin D3/kg of dry matter of milk replacer. In conclusion, vitamin D status of dairy cattle supplemented with vitamin D3 according to typical practices, about 1.5 to 2.5 times the National Research Council recommendation, is sufficient as defined by serum 25(OH)D concentrations. Newborn calves and calves fed milk without supplemental vitamin D3, however, are prone to deficiency.

MicroRNA expression profiles of bovine milk exosomes in response to Staphylococcus aureus infection.

BACKGROUND: Milk exosomes are a rich source of microRNAs (miRNAs) that are protected from degradation. Ingestion of milk and subsequent absorption of miRNAs into recipient cells by endocytosis may play a role in the regulation of neonatal innate and adaptive immunity. In contrast, the miRNA content of milk exosomes may also be indicative of a lactating animal's health; whereby, the presence or absence of specific miRNAs could serve as biomarkers for early detection of bacterial infection that can lead to mastitis. In the present study, we therefore analyzed and compared miRNA expression profiles of milk exosomes from four Holstein cows obtained during mid-lactation prior to and after infection (48 h) of the mammary gland with Staphylococcus aureus. METHODS: Milk exosomes, purified from control and S. aureus infected cows, were extracted for RNA. Following preparation indexed libraries from both groups the samples were subjected to next generation sequencing. RESULTS: Next generation sequencing of eight, unpooled small RNA libraries derived from milk exosomes produced about 60.5 million high-quality, bovine-specific sequence reads for comparison of miRNA expression between treatments. Sequence identity analysis showed the miRNAs make up about 13 % of the average RNA content of these exosomes. Although 417 known bovine miRNAs were identified, miRNAs represented the least diverse class of RNA accounting for only 1 % of all unique sequences. The 20 most prevalent unique sequences within this class accounted for about 90 % of the total miRNA-associated reads across samples. Non-annotated, unique reads provided evidence for another 303 previously unknown bovine miRNAs. Expression analyses found 14 known bovine microRNAs significantly differed in frequency between exosomes from infected and control animals. CONCLUSIONS: Our survey of miRNA expression from uninfected milk exosomes and those produced in response to infection provides new and comprehensive information supporting a role for delivery into milk of specific miRNAs involved in immune response. In particular, bta-miR-142-5p, and -223 are potential biomarkers for early detection of bacterial infection of the mammary gland. Additionally, 22 mammary-expressed genes involved in regulation of host immune processes and response to inflammation were identified as potential binding targets of the differentially expressed miRNAs.

Cellular calcium dynamics in lactation and breast cancer: from physiology to pathology.

Breast cancer is the second leading cause of cancer mortality in women, estimated at nearly 40,000 deaths and more than 230,000 new cases diagnosed in the U.S. this year alone. One of the defining characteristics of breast cancer is the radiographic presence of microcalcifications. These palpable mineral precipitates are commonly found in the breast after formation of a tumor. Since free Ca(2+) plays a crucial role as a second messenger inside cells, we hypothesize that these chelated precipitates may be a result of dysregulated Ca(2+) secretion associated with tumorigenesis. Transient and sustained elevations of intracellular Ca(2+) regulate cell proliferation, apoptosis and cell migration, and offer numerous therapeutic possibilities in controlling tumor growth and metastasis. During lactation, a developmentally determined program of gene expression controls the massive transcellular mobilization of Ca(2+) from the blood into milk by the coordinated action of calcium transporters, including pumps, channels, sensors and buffers, in a functional module that we term CALTRANS. Here we assess the evidence implicating genes that regulate free and buffered Ca(2+) in normal breast epithelium and cancer cells and discuss mechanisms that are likely to contribute to the pathological characteristics of breast cancer.

The Ca(2+)/H(+) antiporter TMEM165 expression, localization in the developing, lactating and involuting mammary gland parallels the secretory pathway Ca(2+) ATPase (SPCA1).

Plasma membrane Ca(2+)-ATPase 2 (PMCA2) knockout mice showed that ~60% of calcium in milk is transported across the mammary cells apical membrane by PMCA2. The remaining milk calcium is thought to arrive via the secretory pathway through the actions of secretory pathway Ca(2+)-ATPase's 1 and/or 2 (SPCA1 and 2). However, another secretory pathway calcium transporter was recently described. The question becomes whether this Golgi Ca(2+)/H(+) antiporter (TMEM165) is expressed sufficiently in the Golgi of lactating mammary tissue to be a relevant contributor to secretory pathway mammary calcium transport. TMEM165 shows marked expression on day one of lactation when compared to timepoints prepartum. At peak lactation TMEM165 expression was 25 times greater than that of early pregnancy. Forced cessation of lactation resulted in a rapid ~50% decline in TMEM165 expression at 24h of involution and TMEM165 expression declined 95% at 96 h involution. It is clear that the timing, magnitude of TMEM165 expression and its Golgi location supports a role for this Golgi Ca2(+)/H(+) antiporter as a contributor to mammary Golgi calcium transport needs, in addition to the better-characterized roles of SPCA1&2.

Differential chemokine and cytokine production by neonatal bovine γδ T-cell subsets in response to viral toll-like receptor agonists and in vivo respiratory syncytial virus infection.

γδ T cells respond to stimulation via toll-like receptors (TLR). Bovine γδ T cells express TLR3 and TLR7, receptors that are key for the recognition of viruses such as bovine respiratory syncytial virus (BRSV); however, responses of γδ T cells to stimulation via these receptors, and their role during viral infections, remains unclear. Here, we demonstrate that neonatal bovine γδ T cells exhibit robust chemokine and cytokine production in response to the TLR3 agonist, Poly(I:C), and the TLR7 agonist, Imiquimod. Importantly, we observe a similar phenotype in γδ T-cell subsets purified from calves infected with BRSV. Bovine γδ T cells are divided into subsets based upon their expression of WC1, and the response to TLR stimulation and viral infection differs between these subsets, with WC1.1(+) and WC1(neg) γδ T cells producing macrophage inflammatory protein-1α and granulocyte-macrophage colony-stimulating factor, and WC1.2(+) γδ T cells preferentially producing the regulatory cytokines interleukin-10 and transforming growth factor-ß. We further report that the active vitamin D metabolite 1,25-dihydroxyvitamin D3 does not alter γδ T-cell responses to TLR agonists or BRSV. To our knowledge, this is the first characterization of the γδ T-cell response during in vivo BRSV infection and the first suggestion that WC1.1(+) and WC1(neg) γδ T cells contribute to the recruitment of inflammatory populations during viral infection. Based on our results, we propose that circulating γδ T cells are poised to rapidly respond to viral infection and suggest an important role for γδ T cells in the innate immune response of the bovine neonate.

Dataset of bovine mammary gland dry secretion proteome from the end of lactation through day 21 of the dry period.

This dataset is a label-free quantitation of proteins milk and dry secretions from the end of lactation through day 21 of the dry period using liquid chromatography with tandem mass spectrometry (LC-MS/MS). The data supplied in this article supports the accompanying publication entitled "Characterization of bovine mammary gland dry secretions and their proteome from the end of lactation through day 21 of the dry period" [1]. The Thermo mass spectrometry raw files and MaxQuant files have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset number PXD017837.

Characterization of bovine mammary gland dry secretions and their proteome from the end of lactation through day 21 of the dry period.

The dry period in dairy cows is critical to mammary health. We used label-free mass spectroscopy to quantify whey proteins from milk/dry secretions collected on days 0, 3,10, and 21 of the dry period. The number of proteins identified was 776. The number of proteins upregulated at one or more times in the dry period was 109. The most significant enriched Gene Ontology (GO) terms for the upregulated proteins were immune function and stress-related. The number of proteins that are downregulated in the dry period was 68. The most significant enriched GO terms for the down-regulated proteins were stress and immune function related. In other assays, total protein and lactoferrin concentrations rose significantly by days 10 and 21 of the dry period. We then examined day 21 dry secretions for their ability to inhibit bacterial growth. Bacterial growth inhibition did not correlate with lactoferrin for 4 coliforms tested nor for all 7 mastitis pathogens tested. Eleven proteins out of the total number of proteins identified were significantly associated with positive or negative coliform growth in day 21 dry secretions. The dry secretion protein composition changes we found may provide new information for understanding the dry period and indirectly involution. BIOLOGICAL SIGNIFICANCE: The dry period in a cow is critical to the success of a cow's subsequent lactation for many reasons. Of particular importance is the rate of establishment of new intramammary infections (IMI), which is significant in the dry period. IMI susceptibility is most significant in the first few weeks of the dry period. There are both physical and mechanical reasons for increased IMI susceptibility in the first few weeks of the dry period. We also know that dry secretions have high levels of antimicrobial proteins and bacterial growth is diminished in dry secretions compared to milk. However, we found that by day 21 of the dry period, antimicrobial proteins, such as lactoferrin, are no longer correlated with reduced bacterial growth in dry secretions. We established a dry secretion proteome for the first 3 weeks of the dry period. The library of proteins in dry secretions presented provides a better understanding of the critical dry period in dairy cow management.

Lactation stage impacts the glycolytic function of bovine CD4+ T cells during ex vivo activation.

Dairy cattle undergo dynamic physiological changes over the course of a full lactation into the dry period, which impacts their immunocompetence. During activation, T cells undergo a characteristic rewiring to increase the uptake of glucose and metabolically reprogram to favor aerobic glycolysis over oxidative phosphorylation. To date it remains to be completely elucidated how the altered energetic demands associated with lactation in dairy cows impacts T cell metabolic reprogramming. Thus, in our ex vivo studies we have examined the influence of stage of lactation (early lactation into the dry period) on cellular metabolism in activated bovine CD4+ T cells. Results showed higher rates of glycolytic function in activated CD4+ T cells from late lactation and dry cows compared to cells from early and mid-lactation cows. Similarly, protein and mRNA expression of cytokines were higher in CD4+ T cells from dry cows than CD4+ T cells from lactating cows. The data suggest CD4+ T cells from lactating cows have an altered metabolic responsiveness that could impact the immunocompetence of these animals, particularly those in early lactation, and increase their susceptibility to infection.

Genome Sequence of a Staphylococcus aureus Strain Isolated from a Dairy Cow That Was Nonresponsive to Antibiotic Treatment.

Staphylococcus aureus can cause mastitis in dairy cattle. We report the genome sequence of a Staphylococcus aureus strain isolated from a dairy cow with a chronic case of mastitis. The infection with this strain of Staphylococcus aureus was not cleared from the animal with antibiotic treatment.

Vitamin D status and its relationship to body fat, final height, and peak bone mass in young women.

CONTEXT: Vitamin D insufficiency has now reached epidemic proportions and has been linked to low bone mineral density, increased risk of fracture, and obesity in adults. However, this relationship has not been well characterized in young adults. OBJECTIVE: The objective of the study was to examine the relationship between serum 25-hydroxyvitamin D (25OHD), anthropometric measures, body fat (BF), and bone structure at the time of peak bone mass. DESIGN: This was a cross-sectional study. OUTCOME MEASURES AND SUBJECTS: Anthropometric measures, serum 25OHD radioimmunoassay values, and computed tomography and dual-energy x-ray absorptiometry values of BF and bone structure in 90 postpubertal females, aged 16-22 yr, residing in California were measured. RESULTS: Approximately 59% of subjects were 25OHD insufficient (or=30 ng/ml). Strong negative relationships were present between serum 25OHD and computed tomography measures of visceral and sc fat and dual-energy x-ray absorptiometry values of BF. In addition, weight, body mass, and imaging measures of adiposity at all sites were significantly lower in women with normal serum 25OHD concentrations than women with insufficient levels. In contrast, no relationship was observed between circulating 25OHD concentrations and measures of bone mineral density at any site. Unexpectedly, there was a positive correlation between 25OHD levels and height. CONCLUSIONS: We found that vitamin D insufficiency is associated with increased BF and decreased height but not changes in peak bone mass.