Cortisol levels in skimmed milk during the first 22 weeks of lactation and response to short-term metabolic stress and lameness in dairy cows.

BACKGROUND: Cortisol is secreted into blood in reaction to acute stress, but also in phases of diminished feed intake and changed animal behavior. As cows do not always show clear signs of discomfort, reliable diagnostic markers could be used to provide information regarding individual cows' distress. The objective of this study was to establish an ether free immunoassay for the detection of cortisol and to determine values during the first 22 weeks of lactation. Furthermore, the response in milk cortisol levels was assessed during times of metabolic stress and pain associated symptoms of lameness. METHODS: Milk yield and composition, blood serum glucose, NEFA and BHBA as well as milk cortisol were determined in 24 multiparous Holstein-Friesian cows over the course of the first 22 weeks of lactation. Animals were further checked for signs of clinical diseases on a daily basis. Two feed restrictions over three days (FR; 70 % of precious ad libitum intake) were performed during the 4(th) wk and the 21(st) wk, respectively. An ELISA for cortisol measurement in easily accessible bovine skimmed milk was established and applied. RESULTS: On the last day of FR in early lactation, a reduction in milk yield and changes in serum metabolites compared to respective previous values were detected. The FR in mid-lactation resulted in no changes in milk production and serum metabolites. Milk cortisol was highest during first wk of lactation and remained on comparable levels thereafter. Milk yield and composition were not influenced by FR. Lameness resulted in enhanced milk cortisol levels. CONCLUSION: Milk cortisol could be used as an indicator of painful symptoms such as lameness. Higher values of milk cortisol levels during first wk of lactation should be taken into account for interpretation.

Risk, knowledge and preventive measures of smallholder dairy farmers in northern Malawi with regard to zoonotic brucellosis and bovine tuberculosis.

Milk production using local cattle breed-types is an age-old practice in Malawi. Although dairy farming is becoming more common as a result of the increasing population and demand for milk and milk products, there is limited knowledge of the farmers' awareness of zoonotic disease risks, their preventative practices and the disease burden in animals. This study determined dairy farmers' general knowledge of zoonoses, assessed their risks for infection with zoonotic bovine tuberculosis (bTB) and brucellosis, and evaluated farm practices to prevent disease transmission. A questionnaire was drawn up and administered by the authors. It was used to collect information about the knowledge and preventive practices of 140 out of 684 registered dairy farmers at Mzuzu Agricultural Development Division, northern Malawi. During a second visit to 60 out of the 140 farms, a total of 156 and 95 cattle were tested for brucellosis and tuberculosis, respectively. Most farmers (77.1%) knew or had heard of zoonotic diseases, whilst 75.0% correctly named at least one zoonotic disease. More survey participants named tuberculosis as a zoonotic disease compared to brucellosis (74.3% versus 2.9%). The most commonly named means of transmission were milk (67.0%) and meat (56.0%). Almost all survey participants (96.4%) practised at least one farm activity that could lead to potential transmission of brucellosis or bTB, including sale (67.0%) and consumption (34.0%) of unpasteurised milk. Antibodies against brucellosis were found in 12 cattle (7.7%), whilst one animal (1.1%) reacted to the tuberculin skin test. General knowledge about possible transmission of diseases between humans and animals was high, although most farmers practised risk behaviours that could potentially expose the public to milk-borne zoonotic diseases such as brucellosis and bTB. Furthermore, some animals had positive results for brucellosis and tuberculosis tests. Therefore, improvement of zoonotic disease prevention programmes, as well as further investigation into the prevalence and risk factors for zoonoses, is recommended.

Effect of feeding level of pregnant dairy heifers sired by one bull on maternal metabolism, placental parameters and birth weight of their female calves.

This study aimed to evaluate the effect of different phases of feed restriction on the metabolism and placental indices of growing heifers inseminated with sexed semen of one bull and on the birth weights of their calves. Red-Holstein heifers were randomly divided into three groups. C-group animals (N=17) daily received recommended energy and crude protein (standard diet). ER-Group animals (N=14) were fed 60% of recommended energy and crude protein for the first two pregnancy months followed by the standard diet. LR-group animals (N=13) were provided with the standard diet throughout the first seven months and with 60% energy and crude protein for the last two months of pregnancy. Blood metabolites and weights of dams were assessed regularly during pregnancy. Placenta weight, area of placentomes and calves' birth weights were examined directly after birth. The physiological levels of blood metabolites varied in C-group animals during the different pregnancy stages. Both restriction periods resulted in reduced weight gain of the dams. ER-group animals showed a marked compensatory growth during mid-pregnancy. Serum glucose, cholesterol and beta-hydroxybutyrate were lower in ER-group animals compared with C-group animals during early restriction. During late restriction, only non-esterified fatty acids increased in LR-group animals. Placental parameter and calves' birth weights did not differ between groups. Results indicate only minor effects of a 40% energy and protein restriction during early or late pregnancy in growing heifers on maternal metabolic and placental indices as well as on foetal development, but further studies might show long-term consequences of offspring.

Correlations between milk and plasma levels of amino and carboxylic acids in dairy cows.

The objective of this study was to investigate the relationship between the concentrations of 19 amino acids, glucose, and seven carboxylic acids in the blood and milk of dairy cows and their correlations with established markers of ketosis. To that end, blood plasma and milk specimens were collected throughout lactation in two breeds of dairy cows of different milk yield. Plasma concentrations of glucose, pyruvate, lactate, α-aminobutyrate, ß-hydroxybutyrate (BHBA), and most amino acids, except for glutamate and aspartate, were on average 9.9-fold higher than their respective milk levels. In contrast, glutamate, aspartate, and the Krebs cycle intermediates succinate, fumarate, malate, and citrate were on average 9.1-fold higher in milk than in plasma. For most metabolites, with the exception of BHBA and threonine, no significant correlations were observed between their levels in plasma and milk. Additionally, milk levels of acetone showed significant direct relationships with the glycine-to-alanine ratio and the BHBA concentration in plasma. The marked decline in plasma concentrations of glucose, pyruvate, lactate, and alanine in cows with plasma BHBA levels above the diagnostic cutoff point for subclinical ketosis suggests that these animals fail to meet their glucose demand and, as a consequence, rely increasingly on ketone bodies as a source of energy. The concomitant increase in plasma glycine may reflect not only the excessive depletion of protein reserves but also a potential deficiency of vitamin B6.

NMR metabolomic analysis of dairy cows reveals milk glycerophosphocholine to phosphocholine ratio as prognostic biomarker for risk of ketosis.

Ketosis is a common metabolic disease in dairy cows. Diagnostic markers for ketosis such as acetone and beta-hydroxybutyric acid (BHBA) are known, but disease prediction remains an unsolved challenge. Milk is a steadily available biofluid and routinely collected on a daily basis. This high availability makes milk superior to blood or urine samples for diagnostic purposes. In this contribution, we show that high milk glycerophosphocholine (GPC) levels and high ratios of GPC to phosphocholine (PC) allow for the reliable selection of healthy and metabolically stable cows for breeding purposes. Throughout lactation, high GPC values are connected with a low ketosis incidence. During the first month of lactation, molar GPC/PC ratios equal or greater than 2.5 indicate a very low risk for developing ketosis. This threshold was validated for different breeds (Holstein-Friesian, Brown Swiss, and Simmental Fleckvieh) and for animals in different lactations, with observed odds ratios between 1.5 and 2.38. In contrast to acetone and BHBA, these measures are independent of the acute disease status. A possible explanation for the predictive effect is that GPC and PC are measures for the ability to break down phospholipids as a fatty acid source to meet the enhanced energy requirements of early lactation.

Smallholder dairy production in Northern Malawi: production practices and constraints.

Milk production in Malawi is still unsatisfactory despite efforts by different stakeholders to boost the dairy sector. To investigate the roots of the problem, a survey on the current production practices and constraints on smallholder dairy farming was conducted in the Northern Region of the country. A total of 210 farmers were interviewed. The results revealed that farmers had small herd sizes in the region with an average of 2.2 ± 0.6 cattle per farmer. Average herd size was larger in male-managed farms than in female-managed farms (2.6 ± 2.8 vs. 1.8 ± 1.3), farmers with more than 5 years of dairy farming experience had larger herds than those with less experience (2.6 ± 2.8 vs. 1.9 ± 1. 2) and farmers who grazed their animals tended to have larger herds than those that stall-fed their animals (4.4 ± 5.1 vs. 1.9 ± 1.3). Average milk production was 8.2 ± 6.5 l per cow per day. Higher average daily milk production was observed in farmers with above primary school education (10.3 ± 8.3 vs. 7.7 ± 5.6), those with dairy farming as main activity (9.3 ± 6.6 vs. 6. 5 ± 6.1) and farmers with more than 2 years of experience in dairy farming (9.3 ± 6.3 vs. 6.1 ± 6.4). Unreliable supply of improved animal genetics, poor animal health, feed shortage and poor prices for milk were considered to be the most important constraints to smallholder dairy farming in descending order.

Rumen-protected conjugated linoleic acid supplementation to dairy cows in late pregnancy and early lactation: effects on milk composition, milk yield, blood metabolites and gene expression in liver.

BACKGROUND: Conjugated linoleic acid (CLA) is a collective term for isomers of octadecadienoic acid with conjugated double-bond system. Thus, it was the objective to investigate whether milk composition and metabolic key parameters are affected by adding CLA to the diet of dairy cows in the first four weeks of lactation. METHODS: A study was carried out with five primiparous cows fed a CLA supplemented diet compared to five primiparous cows without CLA supplementation. CLA supplemented cows received 7.5 g CLA/day (i.e. 50% cis(c)9,trans(t)11- and 50% t10,c12-CLA) starting two weeks before expected calving and 20 g CLA/day (i.e. 50% c9,t11- and 50% t10,c12-CLA) throughout day 1 to 28 of lactation. RESULTS: The CLA supplement was insufficiently accepted by the animals: only 61.5% of the intended amount was ingested. Fed CLA were detectable in milk fat, whereas contents of c9,t11-CLA and t10,c12-CLA in milk fat were higher for CLA supplemented cows compared to the control group. On average over the entire treatment period, there was a decrease of saturated fatty acids (FA) in milk fat of CLA supplemented cows, combined with a higher content of monounsaturated and trans FA.Our study revealed no significant effects of c9,t11- and t10,c12-CLA supplementation either on milk yield and composition or on metabolic key parameters in blood. Furthermore the experiment did not indicate significant effects of c9,t11- and t10,c12-CLA-supplementation on gene expression of peroxisome proliferator-activated receptor-alpha (PPARalpha), PPARgamma, sterol regulatory element-binding protein-1 and tumor necrosis factor-alpha in liver tissue. CONCLUSIONS: Feeding c9,t11- and t10,c12-CLA during the first weeks after calving did not affect metabolic key parameters of blood serum or milk composition of fresh cows. Milk fatty acid composition was changed by feeding c9,t11- and t10,c12-CLA resulting in higher contents of these isomers in milk fat. High contents of long chain FA in milk fat indicate that CLA supplementation during the first four weeks of lactation did not affect massive peripheral lipomobilization.

Degradation of Cry1Ab protein from genetically modified maize (MON810) in relation to total dietary feed proteins in dairy cow digestion.

To investigate the relative degradation and fragmentation pattern of the recombinant Cry1Ab protein from genetically modified (GM) maize MON810 throughout the gastrointestinal tract (GIT) of dairy cows, a 25 months GM maize feeding study was conducted on 36 lactating Bavarian Fleckvieh cows allocated into two groups (18 cows per group) fed diets containing either GM maize MON810 or nearly isogenic non-GM maize as the respective diet components. All cows were fed a partial total mixed ration (pTMR). During the feeding trial, 8 feed (4 transgenic (T) and 4 non-transgenic (NT) pTMR) and 42 feces (26 T and 18 NT) samples from the subset of cows fed T and NT diets, and at the end of the feeding trial, digesta contents of rumen, abomasum, small intestine, large intestine and cecum were collected after the slaughter of six cows of each feeding group. Samples were analyzed for Cry1Ab protein and total protein using Cry1Ab specific ELISA and bicinchoninic acid assay, respectively. Immunoblot analyses were performed to evaluate the integrity of Cry1Ab protein in feed, digesta and feces samples. A decrease to 44% in Cry1Ab protein concentration from T pTMR to the voided feces (9.40 versus 4.18 mug/g of total proteins) was recorded. Concentrations of Cry1Ab protein in GIT digesta of cows fed T diets varied between the lowest 0.38 mug/g of total proteins in abomasum to the highest 3.84 mug/g of total proteins in rumen. Immunoblot analysis revealed the extensive degradation of recombinant Cry1Ab protein into a smaller fragment of around 34 kDa in GIT. The results of the present study indicate that the recombinant Cry1Ab protein from MON810 is increasingly degraded into a small fragment during dairy cow digestion.

Cholesterol synthesis in the lactating cow: Induced expression of candidate genes.

Despite the extensive knowledge for other species, cholesterol metabolism in ruminants is nowadays still not clear. Huge differences in milk cholesterol concentration are observed between breeds, managing strategies, individuals and moment of the lactating cycle, but the genetic actors working in the process of cholesterol secretion into milk have not been identified. As ruminant diet contains no cholesterol, understanding the mechanisms and regulation of synthesis, transport and secretion into milk is crucial when trying to reduce the amount of this metabolite in dairy products. The present work aims to study the expression of candidate genes for these processes in the liver of Bos taurus during the lactating cycle. Liver biopsies were obtained from 16 adult brown Swiss cows at different time points (2 weeks pre-partum and 0, 2, 4 and 8 weeks post-partum). After RNA extraction and reverse transcription, gene expression of candidate genes was studied using quantitative RT-PCR. Key enzymes of the cholesterol synthesis (3-hydroxy-methyglutaryl-coenzyme-A (HMG-CoA) synthase, HMG-CoA reductase and farnesyldiphosphat-farnesyltransferase (FDFT)) and gene expression feed-back regulators involved in lipid metabolism (sterol regulatory element binding proteins (SREBP1and 2) SREBP-cleavage activating protein (Scap) were selected as candidate genes. HMG-CoA-reductase and FDFT showed a huge expression increase until week 2 post-partum (p<0.01), most probably in response to the new requirements in the mammary gland. As well, and as a possible explanation for such modifications, an increase in the expression of the regulators SREBP1 and Scap was observed (p<0.01 and p<0.05 respectively). Most important, the whole synthesis machinery showed a coordinated regulation, as highly significant positive correlations were found between the expression levels of the above mentioned enzymes (p<0.01). The increase of milk and blood cholesterol levels in B. taurus after parturition might be the result of a coordinated induction in the expression of key liver enzymes and their regulating factors.

Degradation of Cry1Ab protein from genetically modified maize in the bovine gastrointestinal tract.

Immunoblotting assays using commercial antibodies were established to investigate the unexpected persistence of the immunoactive Cry1Ab protein in the bovine gastrointestinal tract (GIT) previously suggested by enzyme-linked immunosorbent assay (ELISA). Samples of two different feeding experiments in cattle were analyzed with both ELISA and immunoblotting methods. Whereas results obtained by ELISA suggested that the concentration of the Cry1Ab protein increased during the GIT passage, the immunoblotting assays revealed a significant degradation of the protein in the bovine GIT. Samples showing a positive signal in the ELISA consisted of fragmented Cry1Ab protein of approximately 17 and 34 kDa size. Two independent sets of gastrointestinal samples revealed the apparent discrepancy between the results obtained by ELISA and immunoblotting, suggesting that the antibody used in the ELISA reacts with fragmented yet immunoactive epitopes of the Cry1Ab protein. It was concluded that Cry1Ab protein is degraded during digestion in cattle. To avoid misinterpretation, samples tested positive for Cry1Ab protein by ELISA should be reassessed by another technique.