Cyclical heat stress during lactation influences the microstructure of the bovine mammary gland.

This study aimed to evaluate the impact of heat stress on mammary epithelial cell (MEC) losses into milk, secretory mammary tissue structure, and mammary epithelial cell activity. Sixteen multiparous Holstein cows (632 ± 12 kg BW) approximately 100 d in milk housed in climate-controlled rooms were paired by body weight and randomly allocated to one of 2 treatments, heat stress (HS) or pair feeding thermoneutral (PFTN) using 2 cohorts. Each cohort was subjected to 2 periods of 4 d each. In period 1, both treatments had ad libitum access to a common total mixed ration and were exposed to a controlled daily temperature-humidity index (THI) of 64. In period 2, HS cows were exposed to controlled cyclical heat stress (THI: 74 to 80), while PFTN cows remained at 64 THI and daily dry matter intake was matched to HS. Cows were milked twice daily, and milk yield was recorded at each milking. Individual milk samples on the last day of each period were used to quantify MEC losses by flow cytometry using butyrophilin as a cell surface marker. On the final day of period 2, individual bovine mammary tissue samples were obtained for histomorphology analysis, assessment of protein abundance, and evaluation of gene expression of targets associated with cellular capacity for milk and milk component synthesis, heat response, cellular proliferation, and autophagy. Statistical analysis was performed using the GLIMMIX procedure of SAS. Milk yield was reduced by 4.3 kg by HS (n = 7) compared with PFTN (n = 8). Independent of treatment, MEC in milk averaged 174 cells/mL (2.9% of total cells). There was no difference between HS vs. PFTN cows for MEC shed or concentration in milk. Alveolar area was reduced 25% by HS, and HS had 4.1 more alveoli than PFTN. Total number of nucleated MEC per area were greater in HS (389 ± 1.05) compared with PFTN (321 ± 1.05); however, cell number per alveolus was similar between groups (25 ± 1.5 vs. 26 ± 1.4). There were no differences in relative fold expression for GLUT1, GLUT8, CSN2, CSN3, LALBA, FASN, HSPA5, and HSPA8 in HS compared with PFTN. Immunoblotting analyses showed a decrease abundance for phosphorylated STAT5 and S6K1, and an increase in LC3 II in HS compared with PFTN. These results suggest that even if milk yield differences and histological changes occur in the bovine mammary gland after 4 d of heat exposure, MEC loss into milk, nucleated MEC number per alveolus, and gene expression of nutrient transport, milk component synthesis, and heat stress related targets are unaffected. In contrast, the abundance of proteins related to protein synthesis and cell survival decreased significantly, while an upregulation of proteins associated with autophagy in HS compared with PFTN.

Relationships between gastrointestinal permeability, heat stress, and milk production in lactating dairy cows.

Heat stress (HS) is a global issue that decreases farm profits and compromises animal welfare. To distinguish between the direct and indirect effects of HS, 16 multiparous Holstein cows approximately 100 DIM were assigned to one of 2 treatments: pair fed to match HS cow intake, housed in thermoneutral conditions (PFTN, n = 8) or cyclical HS (n = 8). All cows were subjected to 2 experimental periods. Period 1 consisted of a 4 d thermoneutral period with ad libitum intake. During period 2 (P2), the HS cows were housed in cyclical HS conditions with a temperature-humidity index (THI) ranging from 76 to 80 and the PFTN cows were exposed to a constant THI of 64 for 4 d. Dry matter intake of the PFTN cows was intake matched to the HS cows. Milk yield, milk composition, rectal temperature, and respiration rate were recorded twice daily, blood was collected daily via a jugular catheter, and cows were fed twice daily. On d 3 of each period, Cr-EDTA and sucralose were orally administered and recovered via 24 h total urine collection to assess gastrointestinal permeability. All data were analyzed using the GLIMMIX procedure in SAS. The daily data collected in P1 was averaged and used as a covariate if deemed significant in the model. Heat stress decreased voluntary feed intake by 35% and increased rectal temperature and respiration rate (38.4°C vs. 39.4°C and 40 vs. 71 respirations/min, respectively). Heat stress reduced DMI by 35%, which accounted for 66% of the decrease in milk yield. The yields, and not concentrations, of milk protein, fat, and other solids were lower in the HS cows on d 4 of P2. Milk urea nitrogen was higher and plasma urea nitrogen tended to be higher on d 3 and d 4 of HS. Glucose was 7% lower in the HS cows and insulin was 71% higher in the HS cows than the PFTN cows on d 4 of P2. No difference in lipopolysaccharide-binding protein was observed. Heat stress cows produced 7 L/d more urine than PFTN cows. No differences were detected in the urine concentration or percentage of the oral dose recovered for Cr-EDTA or sucralose. In conclusion, HS was responsible for 34% of the reduction of milk yield. The elevated MUN and the tendency for elevated plasma urea nitrogen indicate a whole-body shift in nitrogen metabolism. No differences in gastrointestinal permeability or lipopolysaccharide-binding protein were observed. These results indicate that, under the conditions of this experiment, activation of the immune system by gut-derived lipopolysaccharide was not responsible for the decreased milk yield observed during HS.

Development of an international, multidisciplinary, patient-centered Standard Outcome Set for Multiple Sclerosis: The S.O.S.MS project.

BACKGROUND: Currently, outcomes of Multiple Sclerosis (MS) are not standardized and it is unclear which outcomes matter most to people living with MS. A consensus between patients and healthcare professionals on which outcomes to measure and how, would facilitate a move towards value-based MS care. OBJECTIVE: to develop an internationally accepted, patient-relevant Standard Outcome Set for MS (S.O.S.MS). METHODS: A mixed-method design was used, including a systematic literature review, four patient focus groups (n=30) and a RAND-modified Delphi process with seventeen MS experts of five disciplines from seven countries (the Netherlands, United States of America, Portugal, Ireland, India, New Zealand, Switzerland and Turkey). RESULTS: A standard outcome set for MS was defined, consisting of fourteen outcomes divided in four domains: disease activity (n=3), symptoms (n=4), functional status (n=6), and quality of life (n=1). For each outcome, an outcome measure was selected and the measurement protocol was defined. In addition, seven case-mix variables were selected. CONCLUSION: This standard outcome set provides a guideline for measuring outcomes of MS in clinical practice and research. Using this set to monitor and (inter)nationally benchmark real-world outcomes of MS can support improvement of patient value and ultimately guide the transition towards value-based MS care.

Persistence of sugars used for intestinal permeability measures in an in vitro rumen environment.

Orally administered synthetic sugars are routinely used as markers of intestinal permeability in nonruminants and young calves, but not adult ruminants, likely because of uncertainty surrounding degradation of such sugar markers (e.g., d-mannitol, sucralose, lactulose) in the rumen. The objective was to evaluate persistence of d-mannitol, sucralose, and lactulose in a closed in vitro rumen fermentation system over 48 h. The null hypothesis was that sugar concentration would not be affected by time. Rumen contents were collected and processed under anerobic conditions a total of 12 times from a ruminally cannulated lactating Holstein cow. These 12 rumen samplings reflect 4 in vitro experiments (d-mannitol, sucralose, lactulose, and d-glucose as a positive methodological control), each replicated 3 times. For each replication, filtered rumen contents and rumen buffer (1:3; vol/vol) were added to a series of six 500-mL flasks, each containing 3 filter bags. Each filter bag contained 500 mg of ground total mixed ration (94.2% dry matter; 15.2% crude protein, 40.9% neutral detergent fiber, 3.9% fat, and 6.2% ash, dry matter basis) and three 5-mm glass beads. The 6 flasks represented 0, 6, 12, 24, and 48 h time points, and a 48-h negative control flask. A single sugar was tested during each experimental replicate. Final flask concentrations of each sugar were 4.07 mg/mL d-glucose, 1.99 mg/mL d-mannitol, 2.17 mg/mL sucralose, or 3.10 mg/mL lactulose. Flasks were incubated under anerobic conditions at 39°C where they remained undisturbed until the designated time of removal (0, 6, 12, 24, or 48 h). At removal, an aliquot of each flask was removed and sugar concentration was quantified by HPLC-mass spectrometry. Data for each experiment were analyzed using an ANOVA model that included the single fixed effect of time (0, 6, 12, 24, or 48 h); flask within replicate was the random term. Lactulose was not resolved in any samples due to interfering components within the sample matrix; no lactulose data are presented. As expected, positive methodological control of glucose decreased to negligible concentrations by 6 h of in vitro incubation. d-Mannitol followed the same pattern as glucose, which was different from our hypothesis. The interpretation is that d-mannitol is degraded in the in vitro rumen culture system and, by extension, is therefore not a viable choice to use in in vivo intestinal permeability tests in adult ruminants when dosed orally. As hypothesized, sucralose concentration did not change over 48 h of incubation in a closed in vitro rumen fermentation system. This suggests feasibility of orally dosed sucralose in adult ruminants as a rumen-inert marker of intestinal permeability with subsequent analysis of biological samples (e.g., urine, blood) by HPLC-mass spectrometry.

Redesigning value-based hospital structures: a qualitative study on value-based health care in the Netherlands.

BACKGROUND: A crucial component of value-based health care concerns the redesign of organizational structures. In theory, hospital structures should follow value creation: addressing medical conditions for specific groups of patients over full cycles of care. In practice, however, it remains unclear how hospitals can reorganize themselves into value-based structures. The purpose of this study is to explore the ways in which Dutch hospitals are currently implementing and pursuing value-based redesign. METHODS: This qualitative exploratory study used semi-structured interviews and a focus group for data collection. Transcripts were analyzed through deductive coding, for which we used Mintzberg's theory on organizational structures, particularly his work on design parameters. RESULTS: In their efforts to create more value-based structures, Dutch hospitals often employ a variety of liaison devices, such as project teams and committees. By contrast, the actual formation of units around medical conditions is much rarer. Outcome data are widely used within planning and control systems, and some hospitals partake in external benchmarking. Not all hospitals use cost indicators for monitoring performance. CONCLUSIONS: Value-based redesign is not necessarily a matter of radical changes or binary choices. Instead, as Dutch hospitals show, it can be an incremental process, with a variety of potential knobs to turn to various degrees. Health care executives, managers, and professionals thus have a wide range of options when they aim for more value-based structures. Our conceptualization of "value-based design parameters" can help guide the selection and implementation of strategies and mechanisms for further coordination around medical conditions over full cycles of care.

Algorithm development for individualized precision feeding of supplemental top dresses to influence feed efficiency of dairy cattle.

Individualized, precision feeding of dairy cattle may contribute to profitable and sustainable dairy production. Feeding strategies targeted at optimizing efficiency of individual cows, rather than groups of animals with similar characteristics, is a logical goal of individualized precision feeding. However, algorithms designed to make feeding recommendations for specific animals are scarce. The objective of this study was to develop and test 2 algorithms designed to improve feed efficiency of individual cows by supplementing total mixed rations (TMR) with varying types and amounts of top-dressed feedstuffs. Twenty-four Holstein dairy cows were assigned to 1 of 3 treatment groups as follows: a control group fed a common TMR ad libitum, a group fed individually according to algorithm 1, and a group fed individually according to algorithm 2. Algorithm 1 used a mixed-model approach with feed efficiency as the response variable and automated measurements of production parameters and top-dress type as dependent variables. Cow was treated as a random effect, and cow by top-dress interactions were included if significant. Algorithm 2 grouped cows based on top-dress response efficiency structure using a principal components and k-means clustering. Both algorithms were trained over a 36-d experimental period immediately before testing, and were updated weekly during the 35-d testing period. Production performance responses for dry matter intake (DMI), milk yield, milk fat percentage and yield, milk protein percentage and yield, and feed efficiency were analyzed using a mixed-effects model with fixed effects for feeding algorithm, top dress, week, and the 2- and 3-way interactions among these variables. Milk protein percentage and feed efficiency were significantly affected by the 3-way interaction of top dress, algorithm, and week, and DMI tended to be affected by this 3-way interaction. Feeding algorithm did not affect milk yield, milk fat yield, or milk protein yield. However, feeding costs were reduced, and hence milk revenue increased on the algorithm-fed cows. The efficacy of feeding algorithms differed by top dress and time, and largely relied on DMI shifts to modulate feed efficiency. The net result, for the cumulative feeding groups, was that cows in the algorithm 1 and 2 groups earned over $0.45 and $0.70 more per head per day in comparison to cows on the TMR control, respectively. This study yielded 2 candidate approaches for efficiency-focused, individualized feeding recommendations. Refinement of algorithm selection, development, and training approaches are needed to maximize production parameters through individualized feeding.

Effects of sodium salicylate and time postpartum on mammary tissue proliferation, gene transcript profile, and DNA methylation.

Previous studies have demonstrated nonsteroidal antiinflammatory drug treatment in early lactation had a positive impact on whole-lactation milk production in older cows. The objective of this study was to evaluate proliferative, transcriptional, and epigenetic changes in the mammary gland that could explain increased production responses due to nonsteroidal antiinflammatory drug treatment. Sodium salicylate (SAL; 125 g/d) or water (CON) were administered via oral drench to multiparous Holstein cows (n = 8/treatment) once daily for 3 d beginning approximately 24 h after parturition, and mammary tissue was collected on d 1, 4, and 45 postpartum. Day 1 tissue was collected immediately preceding the initial drench, and d 4 tissue was collected 24 h following the final drench. Blood was collected twice weekly and analyzed for plasma glucose, insulin, ß-hydroxybutyrate, free fatty acids, and prolactin. Cows were milked twice daily until d 7 of lactation, and thrice daily for the remainder of the study. Total RNA extracted from tissue was deep-sequenced and analyzed for differential gene expression using DESeq2. We detected no treatment effect on milk yield or plasma metabolites through 45 d of lactation; additionally, no change in mammary epithelial cell proliferation was detected when assessed by Ki67 labeling. Comparison of SAL versus CON revealed that only 16 of 18,286 genes were differentially expressed (false discovery rate <0.1) in mammary tissue collected on d 45, whereas no differentially expressed genes due to treatment were detected on d 1 or 4. Analysis of transcriptional differences over time showed downregulation of pathways related to immune cell recruitment and differentiation, and extensive overlap with pathways related to cholesterol synthesis and liver X receptor signaling. Global DNA methylation of mammary tissue was decreased for CON compared with SAL. Transcriptome analysis emphasized extensive involvement of immune-related signaling pathways in the switch from lactogenesis to galactopoiesis, and changes in methylation with SAL treatment merit future investigation into epigenetic effects on milk production.

Rumen volatile fatty acid molar proportions, rumen epithelial gene expression, and blood metabolite concentration responses to ruminally degradable starch and fiber supplies.

The objective of this work was to characterize rumen volatile fatty acid (VFA) concentrations, rumen epithelial gene expression, and blood metabolite responses to diets with different starch and fiber sources. Six ruminally cannulated yearling Holstein heifers (body weight = 330 ± 11.3 kg) were arranged in a partially replicated Latin square experiment with 4 treatments consisting of different starch [barley (BAR) or corn (CRN)] and fiber [timothy hay (TH) or beet pulp (BP)] sources. Treatments were arranged as a 2 × 2 factorial. Beet pulp and TH were used to create relative changes in apparent ruminal fiber disappearance, whereas CRN and BAR were used to create relative changes in apparent ruminal starch disappearance. Each period consisted of 3 d of diet adaptation and 15 d of dietary treatment. In situ disappearance of fiber and starch were estimated from bags incubated in the rumen from d 10 to 14. From d 15 to 17, rumen fluid was collected every hour from 0500 to 2300 h. Rumen fluid samples were pooled by animal/period and analyzed for pH and VFA concentrations. On d 18, 60 to 80 papillae were biopsied from the epithelium and preserved for gene expression analysis. On d 18, one blood sample per heifer was collected from the coccygeal vessel. In situ ruminal starch disappearance rate (7.30 to 8.72%/h for BAR vs. 7.61 to 10.5%/h for CRN) and the extent of fiber disappearance (22.2 to 33.4% of DM for TH vs. 34.4 to 38.7% of DM for BP) were affected by starch and fiber source, respectively. Analysis of VFA molar proportions showed a shift from propionate to acetate, and valerate to isovalerate on TH diets compared with BP. Corn diets favored propionate over butyrate in comparison to BAR diets. Corn diets also had higher molar proportions of valerate. Expression of 1 gene (SLC9A3) were increased in BP diets and 2 genes (BDH1 and SLC16A4) tended to be increased in TH diets. Plasma acetate demonstrated a tendency for a starch by fiber interaction with BAR-BP diets having the highest plasma acetate, but other metabolites measured were not significant. These results suggest that TH has the greatest effect on shifts in VFA molar proportions and epithelial transporters, but does not demonstrate shifts in blood metabolite concentrations.

Clinical Impact of Accelerate Pheno Rapid Blood Culture Detection System in Bacteremic Patients.

BACKGROUND: Accelerate Pheno blood culture detection system (AXDX) provides rapid identification and antimicrobial susceptibility testing results. Limited data exist regarding its clinical impact. Other rapid platforms coupled with antimicrobial stewardship program (ASP) real-time notification (RTN) have shown improved length of stay (LOS) in bacteremia. METHODS: A single-center, quasi-experimental study of bacteremic inpatients before and after AXDX implementation was conducted comparing clinical outcomes from 1 historical and 2 intervention cohorts (AXDX and AXDX + RTN). RESULTS: Of 830 bacteremic episodes, 188 of 245 (77%) historical and 308 (155 AXDX, 153 AXDX + RTN) of 585 (65%) intervention episodes were included. Median LOS was shorter with AXDX (6.3 days) and AXDX + RTN (6.7 days) compared to historical (8.1 days) (P = .001). In the AXDX and AXDX + RTN cohorts, achievement of optimal therapy (AOT) was more frequent (93.6% and 95.4%, respectively) and median time to optimal therapy (TTOT) was faster (1.3 days and 1.4 days, respectively) compared to historical (84.6%, P ≤ .001 and 2.4 days, P ≤ .001, respectively). Median antimicrobial days of therapy (DOT) was shorter in both intervention arms compared to historical (6 days each vs 7 days; P = .011). Median LOS benefit during intervention was most pronounced in coagulase-negative Staphylococcus bacteremia (P = .003). CONCLUSIONS: LOS, AOT, TTOT, and total DOT significantly improved after AXDX implementation. Addition of RTN did not show further improvement over AXDX and an already active ASP. These results suggest that AXDX can be integrated into healthcare systems with an active ASP even without the resources to include RTN.

Graduate Student Literature Review: Potential mechanisms of interaction between bacteria and the reproductive tract of dairy cattle.

Although the presence of bacteria has been characterized throughout the reproductive tracts of multiple species, how these bacteria may interact with the host has yet to be described. Previous reviews have described how pathogenic bacteria interact with the reproductive tract to cause infections such as metritis. This review aimed to summarize the knowledge related to pathogenic and nonpathogenic bacteria in various locations of the bovine reproductive tract and the possible mechanisms underlying host-microbe interactions during gametogenesis and early pregnancy. Lactic acid bacteria such as Lactobacillus seem to be beneficial in multiple areas of the reproductive tract: they have been associated with increased oocyte quality when in follicular fluid and secrete reactive oxygen species that are beneficial during placental angiogenesis. However, other bacteria, including Enterococcus, Staphylococcus, and Streptococcus, may modulate T helper cells that inhibit maternal recognition of pregnancy. Available data on the reproductive microbiome focus on variations in microbial communities and their associations with reproductive performance. However, research on these host-microbiome interactions may provide more insight on how bacteria affect fertility.

Onset of the magnetized arc and its effect on the momentum of a low-power two-stage pulsed magneto-plasma-dynamic thruster.

A new type of plasma accelerator-a low-power (<30W), miniature (cm-sized), two-stage pulsed magneto-plasma-dynamic thruster-has been proposed. Being magnetized by an axially symmetric dc magnetic field of ∼200 mT, the vacuum arc discharge demonstrates a threshold behavior: Parameters such as thrust and the thrust-to-power ratio rapidly jump after a certain dc voltage (∼30 V) is applied on the accelerating electrode. We show that such an effect improves the thrust (from ∼2 to ∼210 µN), efficiency (from ∼1% to 50%), and thrust-to-power ratio (from ∼0.5 to ∼18 µN/W).

Real-World Results of Ocrelizumab Treatment for Primary Progressive Multiple Sclerosis.

BACKGROUND: Recently, ocrelizumab (Ocrevus®) was approved for the treatment of primary progressive multiple sclerosis (PPMS) based on data from the ORATORIO clinical trial. Real-world data about the clinical effectiveness of ocrelizumab has yet to be gathered. OBJECTIVE: The aim of this study was to provide data about the clinical effectiveness of ocrelizumab for patients diagnosed with PPMS in a real-world setting. METHODS: We conducted a retrospective cohort study of all patients with PPMS who started ocrelizumab treatment (n = 21) in St. Antonius Hospital (Utrecht/Nieuwegein, the Netherlands) between April 2018 and December 31, 2018. Primary outcome was pre- versus post-ocrelizumab disability worsening rate (from 96 weeks prior to first ocrelizumab administration up to 24 weeks post first ocrelizumab administration). RESULTS: Disability worsening rate while on treatment significantly differed (lower) from disability worsening rate in pre-treatment period (Z = -2.81, p ≤ .01). Three out of 17 patients showed a clinically relevant improvement in disability status after treatment start. CONCLUSION: Ocrelizumab can stabilize disability progression in patients with PPMS. Some patients even showed a clinically relevant improvement in disability status. Further research should help to identify which patients benefit most from ocrelizumab.

Form of calf diet and the rumen. II: Impact on volatile fatty acid absorption.

Diet is known to affect rumen growth and development. Calves fed an all-liquid diet have smaller and less developed rumens and a decreased ability to absorb volatile fatty acids (VFA) compared to calves fed both liquid and dry feed. However, it is unknown how rumens respond when challenged with a defined concentration of VFA. The objective of this study was to assess the effects of 2 different feeding programs on VFA absorption in preweaned calves. Neonatal Holstein bull calves were individually housed and randomly assigned to 1 of 2 diets. The diets were milk replacer only (MRO; n = 5) or milk replacer with starter (MRS; n = 6). Diets were isoenergetic (3.87 ± 0.06 Mcal of metabolizable energy per day) and isonitrogenous (0.17 ± 0.003 kg/d of apparent digestible protein). Milk replacer was 22% crude protein, 21.5% fat (dry matter basis). The textured calf starter was 21.5% crude protein (dry matter basis). Feed and ad libitum water intakes were recorded daily. Calves were exposed to a defined concentration of VFA buffer (acetate 143 mM, propionate 100 mM, butyrate 40.5 mM) 6 h before euthanasia on d 43 ± 1. Rumen fluid samples were obtained every 15 to 30 min for 6 h to measure the rate of VFA absorption. Rumen tissues were obtained from the ventral sac region and processed for morphological and immunohistochemical analyses of the VFA transporters monocarboxylate transporter 1 (MCT1) and 4 (MCT4). Body growth did not differ between diets, but empty reticulorumens were heavier in MRS than MRO calves (0.67 vs. 0.39 ± 0.04 kg) and MRS calves had larger papillae areas (0.76 vs. 15 ± 0.08 mm2). We observed no differences between diets in terms of the abundance of MCT1 and MCT4 per unit area. These results indicate that the extrapolated increase in total abundance of MCT1 or MCT4 in MRS calves was not due to increased transporter density per unit area. Modeled VFA absorption metrics (flux, mmol/h, or 6 h absorbed VFA in mmol) were not different across diets. These results demonstrate that the form of calfhood diet, whether solely MR or MR and starter, does not alter VFA absorption capacity when the rumen is exposed to a defined concentration of VFA at 6 wk of age.

Form of calf diet and the rumen. I: Impact on growth and development.

Preweaning diet is known to affect rumen tissue appearance at the gross level. The objectives of this experiment were to investigate effects of different preweaning diets on the growth and development of the rumen epithelium and on putative rumen epithelial stem and progenitor cell measurements at the gene and cell levels. Neonatal Holstein bull calves (n = 11) were individually housed and randomly assigned to 1 of 2 diets. The diets were milk replacer only (MRO; n = 5) or milk replacer with starter (MRS; n = 6). Diets were isoenergetic (3.87 ± 0.06 Mcal of metabolizable energy per day) and isonitrogenous (0.17 ± 0.003 kg/d of apparent digestible protein). Milk replacer was 22% crude protein, 21.5% fat (dry matter basis). The textured calf starter was 21.5% crude protein (dry matter basis). Water was available ad libitum and feed and water intake were recorded daily. Putative stem and progenitor cells were labeled by administering a thymidine analog (5-bromo-2'-deoxyuridine, BrdU; 5 mg/kg of body weight in sterile saline) for 5 consecutive days and allowed a 25-d washout period. Calves were killed at 43 ± 1 d after a 6 h exposure to a defined concentration of volatile fatty acids. We obtained rumen tissue from the ventral sac and used it for immunohistochemical analyses of BrdU (putative stem and progenitor cells) and Ki67 (cell proliferation), gene expression analysis, and morphological measurements via hematoxylin and eosin staining. Epithelial stem and progenitor cell gene markers of interest, analyzed by real-time quantitative PCR, were ß1-integrin, keratin-14, notch-1, tumor protein p63, and leucine-rich repeat-containing G protein-coupled receptor 5. Body growth did not differ by diet, but empty reticulorumens were heavier in MRS calves (MRS: 0.67 ± 0.04 kg; MRO: 0.39 ± 0.04 kg). The percentage of label-retaining BrdU basale cells was higher in MRO calves than in MRS calves (2.0 ± 0.3% vs. 0.3 ± 0.2%, respectively). We observed a higher percentage of basale cells undergoing proliferation in MRS calves than in MRO calves (18.4 ± 2.6% vs. 10.8 ± 2.8%, respectively). Rumen epithelial gene expression was not affected by diet, but the submucosa was thicker in MRO calves and the epithelium and corneum/keratin layers were thicker in MRS calves. Presumptive stem and progenitor cells in the rumen epithelium were identifiable by their ability to retain labeled DNA in the long term, changed proliferative status in response to diet, and likely contributed to observed treatment differences in rumen tissue thickness.

Technical note: Infusion, sampling, and vacuum-assisted collection devices for use in ruminally cannulated calves.

Calves can be ruminally cannulated at young ages, but equipment size limitations preclude use of an infusion and sampling device in these small animals. Likewise, a procedure to easily evacuate rumen contents in young calves has not been described. Overcoming these technical complications related to assessment of ruminal passage kinetics, nutrient digestion, and volatile fatty acid absorption would aid in future studies advancing our knowledge of dairy calf nutrition. The first objective was to design and fabricate 2 devices (one device for infusion and sampling, and another for vacuum-assisted collection) suitable for use in young ruminally cannulated dairy calves. The second objective was to test the utility of these tools when performing procedures commonly used in ruminant nutrition research. A single weaned 62-d-old ruminally cannulated calf was used to evaluate the ability to infuse a solution of LiCoEDTA and sample rumen contents through the cannula cap over a period of 2 h to assess the rumen liquid passage rate (procedure 1). The device was capable of infusing the LiCoEDTA and sampling the rumen fluid, as evidenced by the presence of elevated Co concentrations in the sampled rumen fluid. Using the fluid samples obtained, liquid passage rate within the calf was estimated to be 40.2% of ruminal fluid/h. The second procedure tested the vacuum-assisted collection device and consisted of evacuating and weighing the rumen contents, which is considered a key preparatory step in washed reticulorumen technique experiments that aim to measure nutrient absorption. In agreement with existing literature, evacuated rumen contents represented approximately 4% of the calf's body weight. In conclusion, custom-built devices for infusion, sampling, and vacuum-assisted collection were efficacious when tested in a 62-d-old ruminally cannulated calf fed a diet of 100% texturized starter (18% crude protein, as-fed). Fellow scientists may employ and further modify these techniques to suit their needs when assessing passage kinetics, nutrient digestion, and volatile fatty acid absorption in calves.

Technical note: p40 antibody as a replacement for p63 antibody in bovine mammary immunohistochemistry.

Tumor protein 63 (p63) is a nuclear antigen found in basal epithelial cells. To date, 10 isoforms of p63 have been identified, falling into 2 major groups identified by presence or absence of an N-terminal transactivation domain (TAp63 and ΔNp63, respectively). Literature suggests that ΔNp63 is the predominant form expressed in basal epithelial cells and myoepithelial cells (MYEC). The mouse anti-p63 antibody, clone 4B1E12, has been used as a specific nuclear marker for bovine MYEC. Unfortunately, this antibody is no longer commercially available. A new mouse monoclonal antibody, clone BC28, specific to ΔNp63 (designated p40) has been developed. We hypothesized that the p40 antibody would be an appropriate substitution as a MYEC and epithelial basal cell marker. An array of archived formalin-fixed, paraffin-embedded bovine tissues were subjected to immunohistochemical staining for either p40 or p63, with a subset being dual stained for direct comparison. Positive staining for p40 and p63 was observed in serial sections of mammary, skin, rumen, salivary gland, ureter, and bladder. As predicted, negative staining for p40 and p63 was observed in testis and intestine. Dual staining for p40 and p63 in calf mammary (n = 4), lactating mammary (n = 4), rumen (n = 4), and skin (n = 4) showed nearly 100% agreement. Thus, we established that the mouse monoclonal antibody, clone BC28, is a suitable replacement for anti-p63, clone 4B1E12, as a marker of MYEC and basal epithelial cells in bovine tissues.

We wanted to choose us: how embryo donors choose recipients for their surplus embryos.

OBJECTIVE: This study aimed to explore factors affecting donors' choice of recipients for their surplus embryos in the New Zealand context of conditional, known donations. BACKGROUND: Internationally, embryo donation has a low uptake in spite of large numbers of cryopreserved embryos. Possible reasons include a lack of knowledge about and concern for the future welfare of the resultant offspring. In New Zealand, donors and recipients meet prior to donation and legislation supports disclosure and access to genetic knowledge. METHOD: Twenty-two embryo donors (10 couples, two individuals) were interviewed between March 2012 and February 2013 about their experiences of donation and factors affecting their donation. Interview data were analysed thematically. RESULTS: In the interests of the welfare of the child resulting from donation, donors were invested in choosing recipients who would make suitable parents. They attempted to choose recipients similar to themselves, as well as those that they trusted to disclose the manner of conception and facilitate agreed-upon information exchange and contact. CONCLUSION: The interest of donors in ensuring offspring well-being may lend support to conditional forms of open donation, allowing for assessment of recipients' suitability to parent, and for negotiation around information exchange and contact.

Short communication: Does early-life administration of a Megasphaera elsdenii probiotic affect long-term establishment of the organism in the rumen and alter rumen metabolism in the dairy calf?

Megasphaera elsdenii is a bacterial species of the rumen that can utilize lactate to produce butyrate, a key volatile fatty acid often implicated in driving calf rumen development. Because lactate is abundant in the rumen of young calves, administration of M. elsdenii to increase butyrate production and thus promote calf rumen development is an appealing possibility. The main objective of this study was to determine whether M. elsdenii administration to calves via oral drench at 14 d of age affected its long-term establishment at 70 d postadministration. Ruminal volatile fatty acid and lactate profiles and blood glucose and ß-hydroxybutyrate concentrations were also examined to determine potential influence on rumen metabolism. Six neonatal Holstein heifer calves were blocked on d 1 by body weight (41.3 ± 1.8 kg) and total serum protein (5.23 ± 0.16 g/dL) and assigned to either the M. elsdenii (n = 3) or control (n = 3) treatment groups. On d 14, calves in the M. elsdenii group orally received 25 mL of a commercially available M. elsdenii suspension, whereas calves in the control group received 25 mL of the same product that had been autoclaved. Rumen contents and blood samples were collected weekly from each animal until 84 d of age. The oral administration of M. elsdenii at 14 d did not increase the abundance of M. elsdenii 70 d postdosing, alter rumen fermentation, or change blood metabolites associated with butyrate. These results suggest that a single administration of the M. elsdenii probiotic may not affect the rumen establishment of the organism.

Evidence synthesis for health policy and systems: a methods guide

Over the last two decades, major gains have been made in global health: life expectancy has increased dramatically; polio eradication is tantalizingly within reach; six million more children survived until their fifth birthday; malaria deaths halved, while more than 20 million people living with HIV gained access to lifesaving treatment. These are all colossal achievements, which amount to millions of lives saved. Despite these successes, progress has often been uneven, both between and within countries. There remains a 31-year discrepancy between the countries with the shortest and longest life expectancies, while more than half of the world's population is unable to access health services without incurring financial hardship. The implications of this are profound. Lack of access to affordable, quality health care perpetuates a vicious cycle of poverty and ill health, and every year millions of people, mostly in the world's poorest countries, die from sicknesses that we know how to prevent and treat. Tackling these challenges will require modern health systems that ensure services reach the poorest, the most vulnerable and those who are most often left behind. These systems must be more dynamic and multisectoral; they must move beyond a focus on diseases and curative care, and place the needs of people and communities at their core. They must also empower people to take charge of their health, with a lifelong focus on preventing the major causes of disease and ill health. Realizing this vision calls for health system reforms and policies grounded in tangible evidence for 'what works' and how. This is a fundamental contribution of the research community - but it means providing research evidence that is synthesized, accessible and contextualized, to enhance its applicability in different health systems. That's what this guide is all about. Evidence Synthesis for Health Policy and Systems: A Methods Guide aims to support researchers and decision-makers, wherever they may live in the world, to generate and use high-quality evidence synthesis on health policy and systems. It outlines well-conducted, applied examples of relevant methods and techniques. I trust it will prove useful to researchers and decision-makers everywhere as they seek to play their part in promoting health, keeping the world safe, and serving the vulnerable.

Performance strategies affect mammary gland development in prepubertal heifers.

In Brazil, the majority of dairy cattle are Holstein × Gyr (H×G). It is unknown whether excessive energy intake negatively affects their mammary development to the same extent as in purebred Holsteins. We hypothesized that mammary development of H×G heifers can be affected by dietary energy supply. We evaluated the effect of different average daily gains (ADG) achieved by feeding different amounts of a standard diet during the growing period on biometric measurements, development of mammary parenchyma (PAR) and mammary fat pad (MFP), and blood hormones. At the outset of this 84-d experiment, H×G heifers (n = 18) weighed 102.2 ± 3.4 kg and were 3 to 4 mo of age. Heifers were randomly assigned to 1 of 3 ADG programs using a completely randomized design. Treatments were high gain (HG; n = 6), where heifers were fed to gain 1 kg/d; low gain (LG; n = 6), where heifers were fed to gain 0.5 kg/d; and maintenance (MA; n = 6), where heifers were fed to gain a minimal amount of weight per day. Heifers were fed varying amounts of a single TMR to support desired BW gains. Over the 84 d, periodic biometric and blood hormone measurements were obtained. On d 84, all heifers were slaughtered and carcass and mammary samples were collected. At the end, HG heifers weighed the most (181 ± 7.5 kg), followed by LG (146 ± 7.5 kg) and MA (107 ± 7.5 kg) heifers. The ADG were near expected values and averaged 0.907, 0.500, and 0.105 ± 0.03 kg/d for HG, LG, and MA, respectively. In addition, body lengths, heart girths, and withers heights were affected by dietary treatment, with MA heifers generally being the smallest and HG heifers generally being the largest. Body condition scores differed by treatment and were highest in HG and lowest in MA heifers; in vivo subcutaneous fat thickness measurement and direct analysis of carcass composition supported this. The HG heifers had the heaviest MFP, followed by LG and then MA heifers. Amount of PAR was highest in LG heifers and was the same for HG and MA heifers. The percentage of udder mass occupied by PAR was lowest in HG heifers, differing from LG and MA heifers. Composition of MFP was not evaluated. Regarding PAR composition, no differences in ash or DM were found. On the other hand, CP concentration of PAR for HG heifers was lower than that for LG heifers, which was lower than that for MA heifers. Regarding the fat content, HG treatment was higher than LG and MA treatment, which did not differ from each other. In PAR, differences in relative abundance of genes related to both stimulation and inhibition of mammary growth were observed to depend on dietary treatment, sampling day, or both. The same can be said for most of the blood hormones that were measured in this experiment. In this experiment, high ADG achieved by feeding different amounts of a standard diet during the growing period negatively affected mammary development.