Lactate Dehydrogenase In Milk Research Articles

295 Effects of a Supraphysiological Oxytocin Injection After Parturition on Blood-Milk-Barrier Closure and Igg Transfer into Milk in Dairy Cows

Abstract Oxytocin (OT) can affect the blood-milk-barrier integrity by modulating the closure of tight junctions. Earlier research demonstrated a transfer of circulating immunoglobulins (Ig) into milk upon OT application during an LPS-induced mastitis. The role of OT in colostrogenesis, i.e., IgG transfer into colostrum, is not fully understood. Our study aimed at retarding the postpartum closure of the blood-milk-barrier and concomitantly prolonging the transfer of IgG into colostrum of dairy cows by a supraphysiological OT injection. Multiparous Holstein cows (n = 28) were enrolled. Blood and milk samples were collected at the first 2 milkings after parturition and milk yields were recorded. Ten cows received 50 IU OT i.v. at 30 min after the first milking (OT+30), 9 cows were injected 50 IU OT i.v. at 120 min before the second milking (OT-120), and 9 cows remained untreated (CTRL). Cows were milked at 6.7 ± 0.7 h after parturition (first milking) and 11.5 ± 0.3 h later (second milking). Milk was analyzed for fat, protein, lactose and IgG, serum albumin (SA), and electrolytes (Na, K, Cl) concentrations and for lactate dehydrogenase (LDH) activity. Concentration of α-lactalbumin (α-LA) was measured in blood plasma. Data were evaluated using a mixed model (SAS) with group, milking number, and group x milking number as fixed effects, and cow as random subject. In all groups, concentrations of protein, IgG, SA, LDH in milk decreased from first to second milking, while lactose content in milk increased (P < 0.05). Milk yield and milk fat content did not differ between first and second milking (P > 0.05). Concentration of α-LA in plasma tended to decrease from first to second milking. Treatments did not affect changes in blood and milk from first to second milking. In conclusion, a single injection of supraphysiological OT either at 30 min after the first milking postpartum or 2 h before the second milking postpartum did not retard the closure of the blood-milk-barrier in dairy cows. Furthermore, OT application did not prolong the active transport of IgG into milk after the first milking.

Assessment of milk and blood enzymes in Holstein cows with subclinical mastitis

The objective of this study was to determine the activities of lactate dehydrogenase (LDH), alkaline phosphatase (ALP), gamma-glutamyl transferase (GGT), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in the milk and blood of lactating Holstein cows and investigate their association with subclinical mastitis (SCM). This study also aims to evaluate the impact of SCM and its severity based on California mastitis test (CMT) scores on the above mentioned enzymes, as well as to test the efficiency of their use as an early predictive diagnostic tool. Thirty multiparous, lactating dairy Holstein cows were examined using CMT and according to the obtained results classified into four groups; control group 1 (SCM-free = 10, score = 0), group 2 (weak positive = 8, score +1), group 3 (positive = 7, score +2) and group 4 (strong positive = 5, score +3). The milk LDH, ALP, and GGT activities were significantly higher in groups 3 and 4 compared to group 2 and the control group. LDH activity was significantly higher in group 2 compared to the control group. The mean blood LDH and ALP activities in SCM groups were significantly higher than in the control group. ALP blood activity was significantly higher in group 4 compared to group 2. The blood GGT activity was significantly higher in group 4 compared to other groups. There was a significant positive association between milk LDH, ALP, and GGT enzymes as well as the blood enzymes (ALP and GGT) in SCM cows. The results showed that subclinical mastitis causes considerable changes in the activities of enzymes in milk and blood of infected cows.

Associations of Automatically Recorded Body Condition Scores with Measures of Production, Health, and Reproduction

In the present study, we hypothesize that an automated body condition scoring system could be an indicator of health and pregnancy success in cows. Therefore, the objective of this study is to determine the relationship of the automated registered body condition score (BCS) with pregnancy and inline biomarkers such as milk beta-hydroxybutyrate (BHB), milk lactate dehydrogenase (LDH), milk progesterone (mP4), and milk yield (MY) in dairy cows. Indicators from Herd NavigatorTM were grouped into classes based on their arithmetic means. Values were divided into various classes: MY: ≤31 kg/day (first class—67.3% of cows) and >31 kg/day (second class—32.7%); BHB in milk: ≤0.06 mmol/L (first class—80.7% of cows) and >0.06 mmol/L (second class—16.9%); milk LDH activity: ≤27 µmol/min (first class—69.5% of cows) and >27 µmol/min (second class—30.5%); milk progesterone value: ≤15.5 ng/mL (first class—28.8% of cows) and >15.5 ng/mL (second class—71.2%); and BCS: 2.5–3.0 (first class—21.4% of cows), >3.0–3.5 (second class—50.8%), and >3.5–4.0 (third class—27.8%). According to parity, the cows were divided into two groups: 1 lactation (first group—38.9%) and ≥2 lactations (second group—61.1%). Based on our investigated parameters, BCS is associated with pregnancy success because the BCS (+0.29 score) and mP4 (10.93 ng/mL) of the pregnant cows were higher compared to the group of non-pregnant cows. The MY (−5.26 kg, p < 0.001) and LDH (3.45 µmol/min) values were lower compared to those in the group of non-pregnant cows (p < 0.01). Statistically significant associations of BCS and mP4 with the number of inseminations were detected. The number of inseminations among cows with the highest BCS of >3.5–4.0 was 42.41% higher than that among cows with the lowest BCS of 2.5–3.0 (p < 0.001). BCS can also be a health indicator. We found that the LDH content was greatest among cows with the highest BCS of >3.5–4.0; this value was 6.48% higher than that in cows with a BCS of >3.0–3.5 (p < 0.01). The highest MY was detected in cows with the lowest BCS of 2.5–3.0, which was 29.55% higher than that in cows with the highest BCS of >3.5–4.0 (p < 0.001). BCS was the highest in the group of cows with mastitis (4.96% higher compared to the group of healthy cows), while the highest statistically significant mean differences in body condition score (9.04%) were estimated between the mastitis and metritis groups of cows (p < 0.001).

Effect of controlled-release monensin on automatically registered body condition score, milk β-hydroxybutyrate, milk yield and milk lactate dehydrogenase in fresh dairy cows.

The aim of this study was to compare the effect of controlled-release monensin on the automatic registered body condition score (BCS), and biomarkers registered using a fully automated inline analyzer, such as milk β-hydroxybutyrate (BHB), milk yield (MY) and milk lactate dehydrogenase (LDH). Two experimental groups were formed: (1) monensin group (GK) supplemented with monensin (a monensin controlled release capsule (MCRC) of 32.4 g, n = 42) and (2) control group (GO) (capsule containing no monensin, n = 42). Treatment began 21 days before calving, and the experiment was finished one month after calving. In order to gather data about MY, BHB, and LDH, Herd Navigator a real-time analyzer (Lattec I/S, Hillerød, Denmark) was used together with a DeLaval milking robot (DeLaval Inc., Tumba, Sweden). BCS was measured using 3D BCS cameras (DeLaval, DeLaval International AB). All data were registered at one, 15 and 30 days after calving. The statistical analysis was performed using SPSS 26.0 (SPSS Inc., Chicago, USA) package. It was concluded that in the group of cows with monensin supplement (a monensin controlled release capsule of 32.4 g,), the body condition score was statistically significantly higher at the 15th (+0.24, p=0.003) and 30th (+0.52, p⟨0.001) days after calving, the productivity of cows in this group increased by 10.25% from the 1st to the 15th day and by 22.49% from the beginning of the experiment to the 30th day (p⟨0.001), lactate dehydrogenase activities at the 15th and 30th days after calving in this group were lower (p⟨0.001), and also in this group, the number of cows with a value of β-hydroxybutyrate of 0.06 mmol/L decreased from the beginning of the experiment to 30 days after calving by 4.70% (from 19.00% to 14.30%) compared with the control group.

Can unsupervised learning methods applied to milk recording big data provide new insights into dairy cow health?

Among the dairy sector's current concerns, the assessment of global animal health status is a complex challenge. Its multidimensionality means that global monitoring tools are rarely considered. Instead, specific disease detection is often studied separately and, due to financial and ethical issues, uses small-scale data sets focusing on few biomarkers. Several studies have already been conducted using milk Fourier transform mid-infrared (FT-MIR) spectroscopy to detect mastitis and lameness or to quantify health-related biomarkers in milk or blood. Those studies are relevant but they focus mainly on one biomarker or disease. To solve this issue and the small-scale data set, in this study, we proposed a holistic approach using big data obtained from milk recording, including milk yield, somatic cell count, and 27 FT-MIR-based predictors related to milk composition and animal health status. Using 740,454 records collected from 114,536 first-parity Holstein cows in southern Belgium, we performed repeated unsupervised learning algorithms based on Ward's agglomerative hierarchical clustering method to find potential interesting patterns. A divide-and-conquer approach was used to overcome the limitation of computational resources in clustering a relatively large data set. Five groups of records were identified. Differences observed in the fourth group suggested a relationship to metabolic disorders. The fifth group seemed to be related to mastitis. In a second step, we performed a partial least squares discriminant analysis (PLS-DA) to predict the probability of belonging to those specific groups for the entire data set. The obtained global accuracy was 0.77 and the balanced accuracy (i.e., the mean between sensitivity and specificity) of discriminating the fourth and fifth groups was 0.88 and 0.96, respectively. Then, a validation of the interpretation of those groups was performed using 204 milk and blood reference records. The predicted probability associated with the metabolic disorders issue had significant correlations of 0.54 with blood β-hydroxybutyrate, 0.44 with blood nonesterified fatty acids, -0.32 with blood glucose, -0.23 with milk glucose-6-phosphate, and 0.38 with milk isocitrate. In contrast, the predicted probability of belonging to the mastitis group had correlations of 0.69 with milk lactate dehydrogenase, 0.46 with milk N-acetyl-β-d-glucosaminidase, -0.18 with milk free glucose, and 0.16 with milk glucose-6-phosphate. Consequently, these results suggest that the obtained quantitative traits indirectly reflect some of the main health disorders in dairy farming and could be used to monitor dairy cows on a large scale. By using unsupervised learning on large-scale milk recording data and then validating the pattern using reference laboratory measures, we propose a new approach to quickly assess dairy cow health status.

Feeding supplemental 25-hydroxyvitamin D3 increases serum mineral concentrations and alters mammary immunity of lactating dairy cows

Objectives were to determine the effects of feeding supplemental 25-hydroxyvitamin D3 [25(OH)D3] on concentrations of vitamin D metabolites and minerals in serum, mammary immune status, and responses to intramammary bacterial infection in dairy cows. Sixty multiparous, pregnant lactating Holstein cows with somatic cell count <200,000/mL were blocked by days in milk and milk yield and randomly assigned to receive a daily top-dressed dietary supplement containing 1 or 3 mg of vitamin D3 (1mgD or 3mgD), or 1 or 3 mg 25(OH)D3 (1mg25D or 3mg25D) for 28 d (n = 15/treatment). Cows were kept in a freestall barn and fed a total mixed ration in individual feeding gates. Individual dry matter intake (DMI) and milk yield were recorded daily, and milk and blood samples were collected at 0, 7, 14, and 21 d relative to the start of treatment. At 21 d, cows fed 1mgD and 3mg25D received an intramammary challenge with Streptococcus uberis. Cows were observed for severity of mastitis, and blood and milk samples were collected every 12 h to measure inflammation. The 1mg25D and 3mg25D cows had greater serum 25(OH)D3 concentrations at 21 d compared with 1mgD and 3mgD cows (62 ± 7, 66 ± 8, 135 ± 15, and 232 ± 26 ng/mL for 1mgD, 3mgD, 1mg25D, and 3mg25D, respectively). The 3mg25D cows had greater concentrations of Ca and P in serum at 21 d compared with other treatments (Ca = 2.38, 2.4, 2.37, and 2.48 ± 0.02 mM, 1.87, 1.88, and 2.10 ± 0.08 mM for 1mgD, 3mgD, 1mg25D, and 3mg25D, respectively). Yields of milk and milk components, DMI, body weight, and concentrations of 1,25-dihydroxyvitamin D and Mg in serum did not differ among treatments. Abundance of mRNA transcripts for interleukin-1β (IL1B) and inducible nitric oxide synthase (iNOS) in milk somatic cells before S. uberis challenge were increased in cows fed 25(OH)D3 compared with cows fed vitamin D3. Furthermore, IL1B, iNOS, β-defensin 7, and β-defensin 10 in milk somatic cells increased as concentrations of 25(OH)D3 increased in serum. Cows fed 3mg25D had less severe mastitis at 60 and 72 h after challenge with S. uberis compared with cows fed 1mgD. Concentrations of bacteria, somatic cells, and serum albumin in milk after challenge did not differ between treatments; however, an interaction between treatment and day was detected for lactate dehydrogenase in milk. Expression of adhesion protein CD11b on milk neutrophils after the S. uberis challenge was greater among 3mg25D cows compared with 1mgD cows. Transcripts of CYP24A1 and iNOS in milk somatic cells during mastitis also were greater in 3mg25D cows compared with 1mgD cows. Feeding 25(OH)D3 increased serum 25(OH)D3 more effectively than supplemental vitamin D3, resulting in increased serum mineral concentrations, increased expression of vitamin D-responsive genes, and altered immune responses to intramammary bacterial challenge.

Ketoprofen affects the mammary immune response in dairy cows in vivo and in vitro

Nonsteroidal anti-inflammatory drugs are commonly administered parenterally in addition to antimicrobial mastitis therapy to increase the well-being of the diseased animal. As mastitis is usually a localized infection of mammary tissue, we tested the hypothesis that a local administration of nonsteroidal anti-inflammatory drugs through the teat canal could have anti-inflammatory effects on the affected area. We investigated the effects of intramammarily administered ketoprofen (KET) during an LPS-induced immune response on somatic cell count (SCC) and blood-milk barrier integrity. In addition, we investigated the effects of KET on the mRNA abundance of immune factors and their prostaglandin E2 secretion in primary bovine mammary epithelial cells in vitro. Six cows received 0.2 µg of LPS (serotype O26:B6) together with 50 mg of KET into one quarter and LPS only in the opposing quarter. The increase of SCC and of serum albumin (SA) and IgG concentrations and the increase of lactate dehydrogenase (LDH) activity in milk induced by LPS were lower in quarters that received KET in addition. In 3 cows, intramammary KET (50 mg) without additional LPS did not affect SCC, SA, IgG, and LDH in milk. Effects of KET on the immune response of mammary epithelial cells in vitro were investigated in cells from 3 cows challenged with or without LPS (0.2 µg/mL) and with or without additional KET in 2 concentrations (1.25 or 2.5 mg/mL). Ketoprofen reduced the LPS-induced increase of mRNA abundance of tumor necrosis factor α, IL-8, serum amyloid A, and cyclooxygenase-2. The mRNA abundance of cyclooxygenase-1 and prostaglandin E synthase was reduced in cells without LPS challenge by addition of KET at 2.5 mg/mL. Furthermore, the LPS-induced secretion of prostaglandin E2 of mammary epithelial cells into the supernatant could not be detected if KET was added. The results demonstrate that intramammary KET diminishes the increase of SCC and reduces the impairment of the blood-milk barrier (based on SA and LDH in milk), leading to a reduced IgG concentration in milk during LPS-induced mastitis. In mammary epithelial cells, KET limits the expression of several immune factors that are increased during an immune response. In summary, intramammary administration of KET reduces the inflammatory response in the mammary gland. However, it remains unclear whether the inhibited transfer of immune cells and IgG from blood into milk after KET administration would reduce the success of the immune defense in infectious mastitis.

A study on IL8RB gene polymorphism as a potential immuno-compromised adherent in exaggeration of parenteral and mammo-crine oxidative stress during mastitis in buffalo

The genetic markers in inflammatory responses during mastitis afford a reasonable way for improving milk production in the Egyptian buffalo breed. Among them is the interleukin 8 Receptor Gene (IL8RB) (CXCR2); a chemokine receptor gene augments the neutrophil migration during infection. To understand its role better during mastitis in Egyptian buffalos, twenty-five dairy animals representing the normal, sub-clinically, clinically and chronically affected buffalos were randomly selected from different districts. Screening criteria for mastitis were based on somatic cell count and California mastitis test assays on their milk samples. Biochemically, mastitis induced an increase in milk lactate dehydrogenase, alkaline phosphatase and catalase activities and serum malanoaldehyde concentration. The total antioxidant concentrations, however, decreased in serum and milk during mammary inflammation. The protein profiling of milk whey proved an accelerated mammary inflammatory influx of blood-borne proteins during mastitis. The genomic DNAs were extracted from blood samples and the CXCR2 sequence of 1246bp covering a part of intron 1, exon 2 and a part of 3′UTR were submitted to Genbank (accession # KY399457.1). The study clearly defined the presence of four SNPs. Three were detected as synonymous substitutions in coding region and one in the 3′UTR region. Only SNP C/A at c.127 was found to be highly associated with mastitis. In conclusion, the results warrant the potential correlation between the genetic SNP variance for certain genes and the incidence of mastitis in buffalo breed.

Differential glucocorticoid-induced closure of the blood-milk barrier during lipopolysaccharide- and lipoteichoic acid-induced mastitis in dairy cows

Bacteria invading the mammary gland can cause pathogen-dependent differences in the permeability of the blood-milk barrier leading to the differential paracellular transfer of blood and milk components. Glucocorticoids such as prednisolone (PRED) are known to increase the integrity of the blood-milk barrier and quickly restore the decreased milk quality associated with mastitis. The objective of this study was to examine the effect of intramammary PRED on the differential permeability of the blood-milk barrier during mastitis induced by lipopolysaccharide (LPS) from Escherichia coli or lipoteichoic acid (LTA) from Staphylococcus aureus. Thirty-one dairy cows, divided into 6 groups, were injected via a teat canal with LPS, LTA, LPS and PRED, LTA and PRED, saline (control), or PRED. Milk and blood samples were collected 0 to 8h after challenge and analyzed for somatic cell count, IgG, serum albumin, and lactate dehydrogenase in milk, or α-lactalbumin in plasma. Somatic cell count was similarly elevated in LPS- and LTA-challenged quarters and was reduced to control quarter levels only in LTA-challenged quarters with PRED administration. Lactate dehydrogenase activity was highly elevated in LPS quarters and only slightly elevated in LTA quarters, but decreased to control quarter levels with PRED administration. For serum albumin and IgG, only LPS quarters showed an elevation in concentration and PRED treatment reduced the concentration to control quarter level. We found no differences in α-lactalbumin concentrations in plasma in PRED-treated cows compared with cows that only received LPS or LTA. In conclusion, the pathogen-specific appearance of blood constituents in milk during mastitis demonstrates a differential activation of the blood-milk barrier that, in turn, can be manipulated by intramammary glucocorticoids. The results show that the administration of PRED during mastitis increases the blood-milk barrier integrity but has implications in reducing the transfer of IgG that specifically occurs during E. coli mastitis. In addition, it can also reduce the number of migrating immune cells dependent on the mastitis-inducing pathogen. Potential effects of PRED on the cure of naturally occurring mastitis have to be taken into consideration.

Concomitant lipopolysaccharide-induced transfer of blood-derived components including immunoglobulins into milk

During a mammary immune response, the integrity of the blood-milk barrier is negatively affected and becomes leaky. The aim of the present study was to demonstrate the blood origin, and to investigate changes in the concentration, of various constituents including immunoglobulins in blood and milk during the early phase of lipopolysaccharide (LPS)-induced mastitis. Five lactating dairy cows received continuous β-hydroxybutyrate (BHBA) clamp infusions to maintain elevated BHBA blood concentrations (1.5 to 2.0mmol/L) from 48h before and 8h after LPS administration. One udder quarter was infused with 200μg of Escherichia coli LPS. A second quarter served as control. Milk and blood samples were taken hourly for 8h postchallenge (PC). The somatic cell count in LPS-challenged quarters was increased from 4h PC to the end of the experiment compared with control quarters. In LPS-challenged quarters, l-lactate, BHBA, lactate dehydrogenase (LDH), IgG1, and IgG2 were increased at 3h PC and remained elevated until the end of experiment (8h PC) compared with control quarters. In addition, the optical density values in milk in a nonquantitative ELISA for antibodies directed against bluetongue virus (used as a measure of nonspecific antibody transfer; all animals were vaccinated) increased and, thus, indicates an increase in these antibodies in response to LPS treatment. l-Lactate concentration also increased in blood 2h PC and in the milk of control quarters during the experiment from 3h PC. A second experiment was conducted in vitro to investigate a possible contribution from destructed milk cells to l-lactate concentration and activity of LDH in milk. Aliquots of milk samples (n=8) were frozen (−20°C) or disrupted with ultrasound, respectively. Freeze thawing and ultrasound treatment increased LDH in milk samples, but had no effect on l-lactate concentrations. Results suggest that intramammary infusion of LPS induces a systemic response, as evidenced by an elevation of blood l-lactate concentration. The concomitant changes of all investigated components suggest that they were blood derived. However, the increase in blood components in the milk is not necessarily supportive of the mammary immune system, and likely a side effect of reduced blood-milk barrier integrity.

Lipopolysaccharide and lipoteichoic acid induce different immune responses in the bovine mammary gland

Different pathogens, such as Escherichia coli and Staphylococcus aureus, can be responsible for different outcomes of mastitis; that is, acute and severe or chronic and subclinical. These differences in the disease could be related to different mammary responses to the pathogens. The objective of this study was to determine if intramammary challenge with the endotoxins lipopolysaccharide (LPS), from E. coli, and lipoteichoic acid (LTA), from Staph. aureus, induce different immune responses in vivo in milk cells and mammary tissue. To provide a reference level for comparing the challenge and to show the different stimulation of the mammary immune system on a quantitatively similar level, dosages of LPS and LTA were chosen that induced an increase of somatic cells in milk to similar maxima. One udder quarter in each of 21 lactating dairy cows was challenged with 0.2μg of LPS or 20 μg of LTA. From these quarters and from respective control quarters, milk cells or tissue biopsies were obtained at 0, 6, and 12h relative to the challenge to measure mRNA expression of tumor necrosis factor-α (TNFα), IL-1β, IL-8, lactoferrin, and RANTES (regulated upon activation, normal T-cell expressed and secreted). Furthermore, if no biopsies were performed, hourly milk samples were taken for measurement of somatic cell count, lactate dehydrogenase (LDH), and TNFα. Somatic cell count increased in all treatments to similar maxima with LPS and LTA treatments. Concentrations of TNFα in milk increased with LPS but not with LTA. The activity of LDH in milk increased in both treatments and was more pronounced with LPS than with LTA. The mRNA expression of TNFα, IL-1β, IL-8, and RANTES showed increases in milk cells, and LPS was a stronger inducer than LTA. Lactoferrin mRNA expression decreased in milk cells with LPS and LTA treatments. The measured factors did not change in either treatment in mammary tissue. Challenge of udder quarters with dosages of LPS and LTA that induce similar increases in SCC stimulate the appearance of different immune factor patterns. This dissimilar response to LPS and LTA may partly explain the different course and intensity of mastitis after infection with E. coli and Staph. aureus, respectively.

Factors associated with concentrations of select cytokine and acute phase proteins in dairy cows with naturally occurring clinical mastitis

The objective of the current observational study was to determine the potential associations between cow factors, clinical mastitis (CM) etiology, and concentrations of select acute phase proteins and cytokines in milk from affected quarters of cows with CM. Cows with CM (n=197) were grouped based on systemic disease severity, milk culture result, parity, days in milk (DIM), previous CM occurrence, and season of the year when CM occurred. Concentrations of lipopolysaccharide-binding protein (LBP), haptoglobin (Hp), BSA, IFN-γ, tumor necrosis factor-α (TNF-α), IL-1β, IL-8, IL-10, IL-12, transforming growth factor (TGF)-α, and TGF-β and activity of lactate dehydrogenase (LDH) were evaluated. Differences in the least squares means log10 transformed concentrations of these proteins were compared using multiple linear regression mixed models. The milk concentrations of LBP, Hp, IL-1β, IL-10, and IL-12, and activity of LDH in milk were higher in cows with moderate to severe versus mild systemic disease. The concentrations of Hp, BSA, IL-1β, and IL-10 in milk were higher in cows with a gram-negative versus gram-positive milk culture result. Season of the year when CM occurred was associated with the concentration of all proteins evaluated except for IL-1β and IL-12. Concentrations were higher in the winter versus summer except for Hp and TGF-β, for which the opposite was true. Concentrations of LBP, IL-10, and IL-12, and LDH activity in milk were associated with DIM group. Except for LBP, these proteins were lower in cows with CM during the first 60 DIM versus those in mid or later lactation. Interferon-γ, TNF-α, and IL-8 were undetectable in 67, 31, and 20% of samples, respectively. Detection of IFN-γ and IL-8 was associated with season, and detection of TNF-α and IL-8 was associated with systemic disease severity. The current study provides the most comprehensive report of milk concentrations of innate immune response proteins in cows with naturally occurring CM and identifies factors that potentially influence those concentrations. Further investigation into the seasonal variation of cytokine production and its potential effect on the outcome of CM is warranted. Furthermore, the results of this study provide useful data for planning future studies examining the role of the innate immune response in CM.

Haptoglobin and lactate dehydrogenase measurements in milk for the identification of subclinically diseased udder quarters

Diagnosis of subclinical mastitis is of increasing importance and appropriate detection methods are needed. Both haptoglobin (Hp), an acute phase protein in cattle, as well as lactate dehydrogenase (LDH), an ubiquitous enzyme, can be successfully used to detect clinical mastitis. The present paper describes quantification of Hp and LDH in milk samples from healthy and subclinically diseased udder quarters. Hp was analysed in the laboratory using an ELISA. The activity of LDH was measured in raw milk directly in the milking parlor. Both parameters were suitable to distinguish between sterile samples and bacteriologically positive samples. The ability to differentiate between minor and major pathogens was better for Hp in skim milk than for LDH in raw milk. Hp and somatic cell count (SCC) as well as LDH and SCC were positively correlated (&lt;i&gt;r&lt;/i&gt; = 0.8 and &lt;i&gt;r&lt;/i&gt; = 0.76, respectively). Subclinical mastitis was defined as follows: SCC &gt; 100 &amp;times; 10&lt;sup&gt;3&lt;/sup&gt; cells/ml and bacteriological positive findings in two out of three weekly samples. Sensitivity and specificity were above 85% for Hp and above 81% for LDH. Using a less rigid classification to define mastitis, i.e. SCC &amp;lt; 200 &amp;times; 10&lt;sup&gt;3&lt;/sup&gt; cells/ml and two out of three weekly samples bacteriologically positive, sensitivity for Hp improved (89%) and remained unchanged for LDH. Both parameters are useful parameters for the diagnosis of subclinical mastitis. LDH activity in raw milk was less sensitive and specific than Hp but the method described herein offers the opportunity to measure LDH activity directly in the milking parlor and might therefore be suitable for on-line system developments.