Impact of metritis on the generation of reactive oxygen species by circulating phagocytes and plasma lipopolysaccharide concentration in peripartum dairy cows

This study examined the relationship between postpartum metritis, reactive oxygen species (ROS) generation, and plasma lipopolysaccharide (LPS) concentration in peripartum dairy cows. Blood was collected twice weekly from 2 weeks prepartum through 6 weeks postpartum. Whole blood chemiluminescence (WBCL) was measured using the luminol‐enhanced zymosan‐stimulated chemiluminescence assay. Cows were examined for uterine health disorders and classified into two groups, healthy (n = 11) and metritis (n = 5). Metritis had a significant effect on WBCL, with cows with metritis having a higher WBCL. Plasma LPS concentrations in cows with metritis were significantly higher than in healthy cows. To examine the effect of LPS on WBCL, blood was sampled in healthy peripartum cows (1 to 2 weeks prepartum, n = 8; 0 to 3 weeks postpartum, n = 11; and 4 to 8 weeks postpartum, n = 8) and incubated with LPS. At 1 endotoxin units/mL of LPS, similar to the plasma LPS concentration in cows with metritis, the WBCL increased in cows at 0 to 3 weeks postpartum. Results indicate that the increase in ROS generation and plasma LPS concentration are associated with metritis, and LPS may be responsible for enhanced ROS generation in early postpartum dairy cows.     

Periparturient insulin secretion and whole-body insulin responsiveness in dairy cows showing various forms of ketone pattern with or without puerperal metritis

To study the effect of time and different forms of hyperketonemia, with or without puerperal metritis, on insulin and glucose responses, 31 Holstein cows were subjected to glucose (GTT) and insulin tolerance tests (ITT) between 18 and 22 d before, and on days 7 and 60-70 after calving. Plasma concentrations of beta-hydroxybutyrate (BHB), nonesterified fatty acids, glucose, insulin, insulin-like growth factor I and leptin were measured from 18 d before until 70 d after calving. The revised quick insulin sensitivity index (RQUICKI) was calculated at each time point. First postpartum (PP) ovulation was monitored by milk progesterone. Based on BHB patterns and clinical findings, animals were classified as 1) Normoketonemic (NK, n = 9); 2) Transiently hyperketonemic (tHK, n = 7); 3) Continuously HK (cHK, n = 7); and 4) Continuously HK, with signs of puerperal metritis (cHK + PM, n = 6). Insulin area under the curve (AUC) and insulin response to glucose were significantly lower in the early PP period than in late-pregnancy (P < 0.001), and on day 7 after calving in cHK and cHK + PM groups compared to NK and tHK groups (P < 0.001). On day 7, insulin stimulated a decrease in plasma glucose in cHK, cHK + PMthan NK, and tHK groups. Normoketonemic cows (group 1) ovulated earlier than all other groups (P = 0.002). There was no correlation between GTT and ITT variables and the RQUICKI. Time had a significant effect on RQUICKI. Long-term hyperketonemia, especially combined with puerperal metritis, interacts with secretion of insulin and whole-body IR, and results in a significant delay in PP ovarian activity in dairy cows.     

阴离子盐对围产期奶牛的作用机理及其应用

本论文从阴离子盐的作用机理出发,综述了阴离子盐对奶牛采食量、尿液pH值、产奶性能与乳成分、血液离子浓度和血液生化指标、奶牛健康等的影响.     

Comparison of bacterial endotoxin lipopolysaccharide concentrations in the blood,ovarian follicular fluid and uterine fluid: a clinical case of bovine metritis

We investigated the concentration of the bacterial endotoxin lipopolysaccharide (LPS) in the blood, ovarian follicular fluid and uterine fluid of a clinical case of bovine metritis. A 2-year-old lactating Holstein cow exhibited continuous fever >39.5°C for more than 2 weeks after normal calving. The cow produced a fetid, watery, red-brown uterine discharge from the vagina and was diagnosed with metritis. The LPS concentrations in plasma and uterine fluid were 0.94 and 6.34 endotoxin units (EU)/ml, respectively. One of seven follicles showed an extremely high level of LPS (12.40 EU/ml) compared to the other follicles (0.62–0.97 EU/ml). These results might suggest the presence of high concentration of LPS in follicles in cows with postpartum metritis.     

Association of increased serum acute‐phase protein concentrations with reproductive performance in dairy cows with postpartum metritis

Background: It is known that quantification of bovine acute‐phase proteins could routinely provide early diagnostic and prognostic information for monitoring herd health status. Objective: The aim of this study was to test the hypothesis that haptoglobin (Hp) and serum amyloid A (SAA) have the potential for indicating uterine infection, mainly due to subclinical endometritis, which can seriously influence the reproductive performance in dairy cows. Methods: A total of 264 serum samples were collected from 24 Holstein dairy cows at defined intervals from 1 week prepartum to 6 months postpartum. Clinically healthy animals (Group 1, n=6) were compared with 24 animals having acute puerperal metritis (Group 2, n=18) and with concentrations obtained from healthy heifers (baseline values, n=10). The concentrations of serum Hp and SAA were measured using a hemoglobin‐binding assay and Western blot analysis, respectively. Reproductive performance was assessed as the number of days open and the conception rate at the end of the study. Results: Upper cut‐off values for Hp and SAA concentrations in heifers were 130.9 μg/L and 51.9 μg/mL, respectively. Hp concentrations in Group 2 were significantly higher than those in Group 1 at all time points (P<.001). Additionally, among the successfully pregnant animals, the number of days open was significantly higher in 6 cows with Hp >130.9 μg/mL (median=316 days) than in 11 cows with Hp ≤130.9 μg/mL (median=120 days) (P=.0024). Conclusions: Serum Hp and SAA concentrations can be used to recognize uterine infection in postpartum cows. The association between elevated Hp values and number of days open suggests Hp may also be a useful indicator of poor prognosis for reproductive performance.     

乳牛产后期生理性免疫状态的研究

为了研究乳牛在产后期的生理性免疫状态，检测了产后期乳牛淋巴细胞及其亚群数量和淋巴细胞增殖活化功能及其变化。结果发现，乳牛淋巴细胞及其亚群数量和淋巴细胞增殖活化功能在分娩后第1d最低，然后表现出缓慢上升的总体趋势。CD3细胞数在产后第15d低于产后第1d，到产后第30d时达到最高值；CD4细胞数一直多于CD8细胞数；CD21细胞数在产后第15d时达到最高值，以后缓慢下降；淋巴细胞的增殖活化能力在产后第30d最高，未观察到淋巴细胞有明显的凋亡和增生现象。结果表明，乳牛免疫状态在产后第1d最低，以后逐渐升高，第15d时细胞免疫最低而体液免疫最高，第30d时细胞免疫最强。     

Feed‐derived volatile basic nitrogen increases reactive oxygen species production of blood leukocytes in lactating dairy cows

The present study investigated over 9 months the changes of fermentative quality of total mixed rations (TMR) containing grass silage (GS) as a major component, associated with changes in the volatile basic nitrogen (VBN) levels in an experimental dairy farm. Effects of VBN levels in TMR on metabolic parameters, reactive oxygen species (ROS) production by blood polymorphonuclear leukocytes (PMNs) and conception rates for dairy cows were analyzed. According to VBN levels in TMR during survey periods, three distinct phases were identified; phase A with low VBN; phase B with high VBN; and phase C with mid‐VBN. Metabolic parameters in blood were all within normal range. However, during phases B and C, nitrogen metabolic indices such as blood urea nitrogen and milk urea nitrogen showed higher levels compared to those in phase A, and a simultaneous increase in ROS production by blood PMNs and the load on hepatic function in metabolic parameters was observed in the cows with a lower conception rate. This suggests that feeding TMR with elevated VBN levels due to poor fermented GS results in stimulation of ROS production by PMNs by ammonia, and negatively affects metabolism and reproductive performance in lactating dairy cow.     

八珍汤对乳牛产后免疫状态的影响

为了探索提高乳牛产后期免疫状态的新方法，对试验乳牛分4组（产前第30d至产前第1d灌喂组、产前第15d至产后第15d灌喂组、产后第1d至第30d灌喂组、不灌喂中药对照组）灌喂中药八珍汤，检测产后期淋巴细胞及其亚群数量和淋巴细胞增殖活化功能。结果发现，产前组CD3细胞数量在产后第1d升高；产前产后组CD3、CD4和CD8细胞数量在产后第1d和第15d升高；产后组CD3细胞数量在产后第15d和第30d升高，CD4细胞数量也在产后第30d升高。淋巴细胞对ConA的反应能力，产前产后组在产后第1～30d明显提高，产后组在产后第15d和第30d明显提高。各组乳牛产后期CD21细胞数量的变化相近。结果表明，从产前第15d开始到产后第15d每日喂八珍汤，能明显提高乳牛产后期T细胞及其亚群数量和增加淋巴细胞增殖活化功能，而对B细胞数量增加的作用不明显。     

Alterations in biochemical profiles and reproduction performance in postpartum dairy cows with metritis

The aim of the present study was to evaluate the biochemical profile and reproductive performance in postpartum dairy cows with clinical metritis. The research was carried out using performed on 213 pluriparous lactating cows from two dairy herds in southeast Brazil. Dairy cows were assigned into a group of healthy cows (n = 147) and cows with metritis (n = 66). Blood samples to assess serum albumin, cholesterol, fibrinogen, calcium, gamma-glutammyltransferase (GGT), aspartate aminotransferase (AST) and β-hydroxybutyrate (BHBA) concentrations were performed on day 8 ± 2 postpartum. Metritic cows had lower (p < .05) serum albumin, cholesterol and calcium concentration, and higher (p < .05) GGT and AST activity, and BHBA and fibrinogen concentrations when compared to the healthy group. The calving-to-conception interval and services per pregnancy were higher in metritic cows (p < .01) than in healthy cows and the conception rate at first service was lower in metric group (p < .01). Variable importance in projection plots demonstrated that cholesterol was the main discriminator between metritic and healthy cows. Our results indicated alterations in the biochemical profile and impaired reproduction performance in metritic cows.     

Effect of additional vitamin E and zinc supplementation on immunological changes in peripartum Sahiwal cows

This study was conducted to exploit ameliorative effect of additional vitamin E and/or zinc supplementation on immune response of peripartum Sahiwal cows. Thirty‐two pregnant dry Sahiwal cows were blocked into four treatment groups (n = 8), namely control, zinc (Zn), vitamin E (Vit E) and zinc + vitamin E (Zn + Vit E). Feeding regimen was same in all the groups except that the Sahiwal cows in the zinc‐, vitamin E‐ and zinc + vitamin E‐fed groups were additionally supplemented with 60 mg Zn/kg DM, 1000 IU vitamin E and 60 mg/kg + 1000 IU Zn + vitamin E, respectively, from day 60 pre‐partum to day 90 post‐partum. Blood samples were collected on days ?60, ?45, ?30, ?15, ?7, ?3, 0, 3, 7, 15, 30, 45, 60, 90 and 120 with respect to day of parturition and analysed for total immunoglobulin (TIG), immunoglobulin G (IgG), interleukin‐2 (IL‐2), vitamin E (Vit E) and zinc (Zn) status. Before calving, cows showed a decrease in plasma TIG, IgG, IL‐2, Vit E and Zn levels. However, increased levels of plasma TIG, IgG, IL‐2, Vit E and Zn were observed after calving. After calving, Sahiwal cows supplemented with Zn + Vit E had higher plasma TIG, IgG and IL‐2 in comparison with cows of control and Zn + Vit E‐fed groups. In the present study, plasma vitamin E level was higher in Vit E‐fed and Zn + Vit E‐fed cows; however, zinc level was higher in Zn‐ and Zn + Vit E‐supplemented cows. In conclusion, a reduced immune response during peripartum period in Sahiwal cows was ameliorated by dietary vitamin E and zinc supplementation.     

Safety of standardized Macleaya cordata extract in an eighty‐four‐day dietary study in dairy cows

The objective of this study was to evaluate the safety of a standardized Macleaya cordata Extract Product (MCEP) containing the quaternary benzophenanthridine alkaloids, sanguinarine and chelerythrine, when fed to dairy cows. Thirty‐six dairy cows were randomized into three groups with twelve cows/treatment in two replica pens for each treatment group: control (C) without MCEP added to feed, treatment 1 (SANG‐1000) with MCEP added to feed at 1,000 mg/animal/day (1.5 mg/kg bw/day) and treatment 2 (SANG‐10000) with MCEP added to feed at 10,000 mg/animal/day (15.5 mg MCEP/kg bw/day). After two weeks of acclimation, animals were observed for an 84‐day experimental period, with body weight, feed intake and milk production measured daily. Milk composition was analysed every two weeks. Haematological analyses were performed on Day 0 and Day 84, and clinical chemistry analyses were performed on Day 84 of the study. There was no statistically significant difference (p > .10) among the three groups on body condition score, milk production or milk composition over the study period. There were no significant differences in body weight gain or feed consumption among the three groups. Animals in the SANG‐10000 group had significantly higher mean corpuscular volume (MCV) than the C group (p < .1) and lower mean corpuscular haemoglobin concentration (MCHC) than the SANG‐1000 group (p < .1). Concentrations of sanguinarine and chelerythrine in milk samples collected on Day 84 were below the detection limit (LOD) as measured by high‐performance liquid chromatography‐mass spectrometry (HPLC‐MS/MS). In conclusion, this study presents compelling data supporting the hypothesis that the test product MCEP, when included in the TMR at up to 10,000 mg/animal/day (15.5 mg MCEP/kg bw/day), is well tolerated by dairy cows.     

Preanalytical factors affecting blood inorganic phosphate concentration in dairy cows

BACKGROUND: Hypophosphatemia is a condition that affects feed intake and milk production in dairy cows, and, to assess it, a precise biological parameter is needed. Plasma inorganic phosphate (Pi) can be used as a potential indicator of bioavailability of feed phosphorus in dairy cows. OBJECTIVE: The purpose of this study was to evaluate the effects of sample type (serum vs. plasma), blood sampling site (jugular vs. coccygeal), and morning feeding on blood Pi concentration in dairy cows. METHODS: Blood samples from 28 clinically healthy multiparous Holstein cows were analyzed for Pi concentration. The animals were sampled twice by coccygeal vessel into Vacutainer tubes both with and without sodium heparin, 3 hours after the morning feeding. In 4 additional cows, the concentration of Pi was measured in blood samples obtained simultaneously by jugular and coccygeal venipuncture into heparinized tubes before the morning feeding and 2, 4, and 6 hours after the feeding. The results were analyzed using parametric statistical tests. RESULTS: Serum and plasma concentrations of Pi were correlated (r(2) = 0.999, P < .0001), but serum Pi concentration was higher than plasma Pi (P < .02). Plasma Pi concentration 2 hours after feeding tended to be higher than that before feeding in jugular samples, but the difference was not significant. There was no significant variation in plasma Pi during the morning postfeeding times, although jugular concentrations were 19% lower than coccygeal (P = .04). CONCLUSION: To increase the precision of plasma Pi measurements as an indicator of bioavailability of feed phosphorus, sample type, sampling time, and site must be taken into account, to correctly interpret plasma Pi concentrations.     

Effect of nutrient levels during the far‐off period on postpartum productivity in dairy cows

The present study aimed to determine the effects of differing nutrient levels during the far‐off period on postpartum metabolism and milk production in lactating cows. Twenty‐six multiparous cows were assigned to three dietary treatments in the far‐off period: a low‐energy diet (L, n = 9, 80% intake of the total digestible nutrients requirement), a moderate‐energy diet (M, n = 8, 105%) and a high‐energy diet (H, n = 9, 130%). During the close‐up period, all cows were provided with 105% intake. After parturition, all cows were fed a lactation diet. The BCS recovery was slow, and low milk yield was found in the H group. In the L group, BCS recovery was favorable after parturition, and lactation persistence was increased. The L group had low rumen endotoxin activity and a high initial ovulation rate after parturition. These findings indicate that a high‐energy diet during the far‐off period has a deleterious effect on milk production. In contrast, the restricted diet in the far‐off period increased adaptability with respect to peri‐parturition metabolic changes, improved the post‐parturition nutritional state, and increased milk production. Furthermore, it suggests that the nutrient levels in the far‐off period affect rumen endotoxin activity and reproductive function after parturition.     

Differences in rumen fermentation characteristics between low‐yield and high‐yield dairy cows in early lactation

Relationship between rumen fermentation parameters, blood biochemical profiles and milk production traits in different yielding dairy cows during early lactation was investigated. Twelve dairy cows were divided into two groups based on their milk yield, that is low‐yield (LY) and high‐yield (HY) groups. Rumen fluid and blood were collected at 3 weeks prepartum and 4, 8 and 12 weeks postpartum. Results showed that proportions of acetate, propionate to total short chain fatty acids and acetate : propionate ratio were changed (P < 0.05) in both groups during the peripartum period, whereas butyrate and acetate : butyrate ratio were only altered in the HY group. Blood cholesterol, beta‐hydroxybutyric acid (BHBA) and glutamic oxaloacetic transaminase in the HY group were higher (P < 0.01) than those in the LY group. Principal component analysis revealed that milk yield and milk compositions were differently clustered between groups. These parameters showed similar direction with dry matter intake in the HY group and adverse direction in the LY group. Linear regression analysis indicated that butyrate was positively correlated with BHBA (P < 0.05) in the HY group. This study suggests that cows in the HY group seem to accommodate appropriately to negative energy balance in early lactation through rumen fermentation.     

Hair cortisol levels of lactating dairy cows in cold‐ and warm‐temperate regions in Japan

We compared the hair cortisol levels of lactating dairy cows in a cold‐ and a warm‐temperate region out of four climatic zones in Japan. We simultaneously investigated the effects of calving number, lactation period and month of hair sampling. Hair of nine Holstein lactating cows chosen from each region (i.e. 18 cows per sampling) was sampled in March, June, September and December. Number of calvings (1, 2, ≥3) and lactation duration (early: <100, middle: 101–200, and late: >201 days) were balanced between regions. Cortisol was extracted from hair by methanol, and its level was determined with a cortisol immunoassay kit. A multi‐way analysis of variance revealed that the effects of month of hair sampling (P < 0.001) and its combination with region (P < 0.05) were significant. In a multiple comparison test, significant differences (P < 0.01) in hair cortisol level (pg/mg of hair) were found between June (13.0 ± 1.0) and the other 3 months, and between September (1.6 ± 0.2) and December (4.5 ± 0.3). The rise in cortisol level from March to June was more intense in the cold‐temperate region. These results demonstrate the necessity of considering seasonal variations in each climatic region when we use hair cortisol level as an indicator of stress.     

Quantification of pericaudal adipocyte diameter in dairy cattle during peripartum: a complementary method to study energetic status using conventional histology

The energetic status of high‐yielding Holstein‐Friesian dairy cattle was studied during peripartum under field conditions using body condition score (BCS), glycemia, seric ß–hydroxybutyrate and adipose tissue cellularity. This last method was tested as a complementary tool for energetic status assessment. Biopsies of pericaudal subcutaneous adipose tissue were obtained from 25 multiparous animals at 28 days before and 21 days after parturition. Samples were routinely processed for histological examination and stained with haematoxylin and eosin (H&E). The mean diameter of adipocytes (MDA) was measured with the aid of a digital image processor. During the same period, blood samples were collected weekly for metabolite determinations. The MDA at 28 days pre‐partum and 21 days post‐partum were 72.1 vs. 66.2 μm respectively (p = 0.055), and the corresponding BCS at these moments was 3.32 vs. 3.19 (p = 0.068). At ?28 days pre‐partum, the BCS was positively correlated with MDA (Pearson’s r = 0.521, p = 0.016) and with glycemia (Pearson’s r = 0.404, p = 0.056). Correlations between BCS and MDA, and between BCS and glycemia, with ß–hydroxybutyrate although not significant, suggest that routine histological preparations of biopsies from subcutaneous adipose tissue could be included as an easy and valuable tool for research purposes to evaluate metabolic adaptation of dairy cows to peripartum, as well as the incidence of metabolic disorders and productive performance.     

Sleep and lying behavior of milking Holstein cows at commercial tie‐stall dairy farms

The aim of this study was to clarify cow's sleep behavior on farm and to examine the influence of some environment and management factors on cow's sleep. Total 85 behavior data (summer 41, autumn 44) were corrected from 12 commercial tie‐stall dairy farm in northern Tohoku area. Daily duration, bout frequency, and averaged bout duration of sleep and lying were measured. Even though there was no significant difference in any lying measurements, daily duration, bout frequency, and averaged bout duration of sleep was significantly different among farms (p < 0.01, p < 0.01, and p < 0.05, respectively). However, we could not find that any managements influence on sleep. There was significant seasonal difference in daily sleep duration (p < 0.01) and sleep bout frequency (p < 0.05). On the other hand, daily lying duration and averaged lying bout duration were significantly different between seasons (p < 0.001 and p < 0.05, respectively). Primiparous cows slept longer and more frequent than multiparous (both p < 0.001). Milk solid‐non‐fat content was positively correlated with daily sleep duration (p < 0.05) and bout frequency (p < 0.01). These results might imply the productive importance of sleep related with anabolic hormones release.     

Variation in selected blood plasma constituents during the post-partum and breeding periods in dairy cows

Concentrations of insulin-like growth factor-I, albumin, beta hydroxybutyrate, glucose and urea were measured in 12 plasma samples collected over 9 weeks from mid-August from each of 40 cows in four seasonal herds which calved from mid-July. These herds comprised Jersey or Holstein-Friesian animals, with each breed grazed at two stocking rates. Daily dry matter intakes (kg dry matter/cow/day), which were estimated from pasture assessments before and after grazing, ranged from 4% (low stocked Jersey herd) to 38% (high stocked Friesian herd) below recommended intakes for each breed of cow in peak lactatipn. Stocking rate was associated with a 4% reduction in liveweight for cows in the high stocked herds and a 20% reduction in daily milk production (as a proportion of the low stocked herds' liveweight and milk production).

Eight cows produced at least one blood sample with a beta hydroxybutyrate concentration of more than 1.0 mmol/l. There were 42% of samples with glucose concentrations greater than 4.1 mmol/l. These two concentrations were the maximum values classified as normal for these two metabolites by the Ruakura Animal Health Laboratory. No concentrations of the other metabolites were outside their respective normal ranges. Breed, age and individual animals usually had greater effects on the average metabolite concentrations than did stocking rate.

The greatest effect was associated with sample day, especially for insulin-like growth factor-I and urea. A significant breed x stocking rate x sample day interaction in urea concentrations indicated that changes in the feeding value of the grazed pastures available to each herd were sufficient to produce separate fluctuating patterns in the daily average concentrations.

These results showed that frequent sampling on a herd basis would be necessary to identify the significance of factors which may be contributing to sample day variation in the concentration of metabolites in the plasma of pasture-fed cows.     

Characterization of ruminal dynamics in Holstein dairy cows during the periparturient period

We used four pregnant Holstein cows to delineate ruminal adaptations as cows transitioned from one lactation to the next. Cows were fed typical diets through far-off and close-up dry periods and lactation. We measured ruminal characteristics on day 72 (late lactation), 51 (far-off dry), 23 and 9 (close-up dry) prepartum and on days 6, 20, 34, 48, 62, 76 and 90 postpartum (early lactation). Measurements included: ruminal fill (weight of actual contents), ruminal capacity (volume of rumen when fully filled), digestibilities and ruminal passage rates. Ruminal capacity tended to increase linearly during early lactation but was stable during dry and transition periods. Both total and liquid fill decreased linearly during the dry period, increased across parturition, and increased linearly through early lactation. Dry matter fill decreased as cows were fed the close-up diet at day 23 prepartum then increased near parturition and continued to increase across early lactation. Solid passage rate was greatest when cows were fed the close-up diet, and decreased throughout the transition period. In lactation, solid passage rate responded quadratically with peak at day 48 followed by decreases through day 90 postpartum. Liquid passage increased linearly across the transition period. Total tract organic matter digestibilities increased linearly over the dry period with significant increases prior to or immediately after parturition, then they remained relatively stable over early lactation until they increased at day 90. Fibre digestibilities demonstrated quadratic responses over early lactation, being higher on day 6 and day 90 than at other times. Starch digestibilities decreased linearly across both the dry and transition periods with decreases in lactation until day 62 followed by increases until day 90. High producing lactating dairy cows go through a multitude of ruminal adaptations, in terms of digestion, passage, capacity and fill, as they transition from one lactation to the next.