Potential mechanism of action of J5 vaccine in protection against severe bovine coliform mastitis.

Coliform mastitis is one of the most difficult diseases to treat in the modern dairy industry. Curative therapy with antibiotics remains only moderately effective and depends on the stage at which the disease is treated, The most successful strategies for combating coliform. mastitis appear to be prevention by hygienic management or prophylactic immunization. The severity of clinical symptoms of coliform mastitis has been shown to be reduced by immunization with the Escherichia coli J5 vaccine. However, although the J5 vaccine has been licensed in the United States for about 10 years, the immunological basis of its mechanism of action is still unknown. Until now, protection by J5 vaccination has often been explained by a straight forward mechanism of enhanced antibody production resulting in increased opsonization of coliform bacteria and lipopolysaccharides (LPS). The possibility that J5 vaccination could decrease risk factors for coliform mastitis such as impaired blood polymorphonuclcar neutrophil leukocyte (PMN) diapedesis has never been investigated. This review provides arguments to support the hypothesis that J5 vaccination may reduce the severity of coliform mastitis by inducing a condition of mammary gland hyper-responsiveness, characterized by a T helper 1 (Th1) response and mediated by memory cells inside the mammary gland, finally resulting in enhanced PMN diapedesis upon an intramammary infection

Effect of somatotropin on changes in milk-production and composition during coliform mastitis in periparturient cows.

The potential protective and therapeutic effects of bST during coliform mastitis in periparturient cows were evaluated. In a first study, 19 cows, classified as moderate or severe responders based on the respiratory burst activity of blood neutrophils, were treated with recombinant bST or vehicle 48 h after intramammary inoculation of Escherichia coli. Clinical status and changes in milk production and composition were compared in the four groups. In a second study, 8 cows received bST or vehicle 7 d before bacterial challenge. During mastitis, losses in milk production and compositional changes were most pronounced in infected glands and in severe responders. Milk production of bST cows recovered better than that of placebo cows. Recovery of milk components was accelerated in severe responders treated with bST, but not in moderate responders. Pretreatment of severe responders with bST enhanced milk production before infection, protected the mammary glands from excessive loss of milk during the subsequently induced coliform mastitis, and accelerated normalization of milk composition. In conclusion, the beneficial effects of bST upon normalization of milk production and composition in periparturient cows suffering from coliform mastitis seem to be restricted to the severe responders. In severe responders that had been treated with bST, changes observed during mastitis resembled those in moderate responders treated with the placebo

Local and systemic effects of endotoxin mastitis on the chemiluminescence of milk and blood neutrophils in dairy cows.

The local and systemic effects of intramammary lipopolysaccharide (LPS) injection on the chemiluminescence (CL) of milk and blood polymorphonuclear leukocytes (PMN) were investigated in six healthy early lactation cows. Clinical signs of acute mastitis such as fever, increased heart rate and a decreased milk production were observed in all cows. Before LPS challenge, the CL activity of milk PMN was significantly lower than that of blood PMN (P < 0.01). A significant negative correlation was found between pre-challenge milk and blood PMN CL and, the decreased milk production in unchallenged quarters. The CL activity of milk PMN from LPS-injected quarters increased following LPS challenge, whereas it remained unchanged in control quarters. The CL activity of blood PMN showed a biphasic increase, with two peaks and a valley below pre-challenge CL activity (P < 0.01). At post-challenge hours (PCH) 6 and 12, the CL activity of milk PMN from LPS-injected quarters exceeded that of blood PMN (P < 0.05 and P < 0.001, respectively). The decreased CL activity of blood PMN and the enhanced CL activity of milk PMN during endotoxin-induced mastitis was reflected by changes in the shape of the CL curve. In blood PMN, a decrease of the second peak of the CL curve suggests that the myeloperoxidase (MPO)-H2O2 system is impaired during endotoxin-induced mastitis. In contrast, the MPO-H2O2 system was enhanced in milk PMN from challenged quarters. The highest duration and intensity of reactive oxygen intermediate (ROI) production was observed in milk PMN from LPS-injected quarters at PCH 12. The increased viability of PMN in LPS-injected quarters and to a lesser extent in control quarters suggests possible effects of both facilitated diapedesis and inflammatory mediators on milk PMN survival. In conclusion, our results suggest that a combination of local and systemic action of E. coli endotoxin is involved in the priming of milk PMN during mastitis

Functional maturation during bovine granulopoiesis

Granulocytic precursor cells undergo morphologic changes in the nucleus and the cytoplasm during the process of granulopoiesis, which takes place in the bone marrow. These changes are associated with the development of stage-specific proteins necessary for the highly specialized roles of polymorphonuclear leukocytes in phagocytosis, bacterial killing, and in mediating the inflammatory process. The objective of the current study was to sequence the various events that occur upon functional development of granulocytic bone marrow cells in the bovine species. Cells were obtained from the bone marrow of clinically healthy cows and separated into different stages of maturation using density gradient centrifugation. Three cellular fractions were obtained that were enriched for either early immature, late immature or mature granulocytic cells. Functions and receptor expressions assessed in the three maturation stages were: Fc-IgG(2) receptor and CD11b expression, phagocytosis of Escherichia coli, respiratory burst activity, and cellular myeloperoxidase activity. Immature cells expressed already Fc-IgG(2) receptor and CD11b on their cytoplasma membrane. Phagocytic ability was acquired in the myelocytic stage, but only the more mature forms were readily capable of phagocytosis. Promyelocytes, myelo-cytes and metamyelocytes showed no respiratory burst activity. Only band and segmented cells produced reactive oxygen species. Myeloperoxidase was present at all stages of maturity. Thus, each of the maturation stages was characterized by a selective expression of one or more functions and receptors. Therefore, sequential biochemical maturation is postulated during bovine granulopoiesis

Decreased neutrophil bactericidal activity during phagocytosis of a slime-producing Staphylococcus aureus strain.

Phagocytosis and intracellular killing by bovine polymorphonuclear leukocytes (PMN) are important host defence mechanisms against mastitis caused by Staphylococcus aureus. We compared the phagocytosis and overall killing of a non slime-producing (NSP) S. aureus and its slime-producing (SP) variant by blood PMN, using an in vitro bacteriological assay. Seven clinically healthy Holstein-Friesian dairy cows in mid-lactation stage were used for this purpose. The percentages of overall killing for the NSP and SP variant were 34 +/- 3% and 21 +/- 4% (P < 0.05) and the corresponding percentages of phagocytosis were 40 +/- 4% and 31 +/- 4%, respectively. A significant positive correlation (r = 0.79; P < 0.001) was found between phagocytosis and overall killing. These results suggest that the presence of slime was responsible for a decreased phagocytic ingestion and overall killing

Influence of 17 beta-estradiol, progesterone, and dexamethasone on diapedesis and viability of bovine blood polymorphonuclear leukocytes

The aim of the current study was to investigate whether polymorphonuclear leukocyte (PMN) diapedesis and viability are influenced by steroid hormones. Using an in vitro model with different types of cell layers ( bovine mammary epithelial cells and fibroblasts), we investigate whether steroid hormone treatments (17beta-estradiol, progesterone, and dexamethasone) have an influence on the diapedesis capacity and viability of PMN. In addition, we studied apoptosis of PMN in the in vitro model and evaluated the influence of different types of cell layers and steroid hormone treatments on this process. A significant decrease in the number of viable PMN in the lower compartment of the in vitro model (i.e., number of migrated PMN x viability after migration) was found after 17beta-estradiol treatment, whereas no influence was detected after progesterone or dexamethasone treatment. The effect of 17beta-estradiol was not due to a lower viability before migration as none of the treatments caused a significant effect on the viability before diapedesis. This treatment effect was not influenced by endogenous 17beta-estradiol or progesterone levels before isolation because there was no correlation between these plasma levels and PMN diapedesis capacity or viability. Furthermore, migration through epithelial cells caused a significant decrease in viability of PMN due to increased apoptosis but not necrosis

Apoptosis and necrosis of blood and milk polymorphonuclear leukocytes in early and midlactating healthy cows.

Increased milk somatic cell counts (SCC) are used as an indicator for bovine mastitis. During mastitis, polymorphonuclear leukocytes (PMN) become the predominant cell type. Shortly after parturition, the severity of mastitis is increased and several PMN functions are downregulated. Apoptotic and necrotic processes of PMN could influence SCC and PMN functions. In this study, the percentages of apoptotic and necrotic PMN in blood and milk from early and midlactating healthy cows were compared. Apoptosis and necrosis of PMN were quantified using a dual-color flow cytometric procedure with fluorescein labeled annexin-V (green) and propidium iodide (red). Using this technique three different subpopulations of bovine PMN could be detected: apoptotic cells (high intensive green fluorescence), necrotic cells (high intensive green and high intensive red fluorescence) and viable cells (low intensive green and low intensive red fluorescence). Following a 4 h incubation of blood from both groups of cows at 37 degreesC to induce apoptosis, the mean percentage of apoptotic blood PMN was significantly higher (P<0.01) in early lactating cows (15.1%, n=9) compared with midlactating cows (5.3%, n=10). The mean percentage of necrotic PMN remained lower than 5% in all cows. In contrast to blood, no significant difference was found between the percentage of apoptotic PMN in milk from early (41.2%, n=7) and midlactating cows (34.0%, n=8). The percentage of necrotic PMN in milk from early lactating cows (25.9%, n=7) was significantly higher than that in midlactating cows (14.2%, n=8) (P<0.05). Higher percentages of apoptotic as well as necrotic PMN were consistently found in milk compared to blood in all cows. From these results, it can be concluded that spontaneously induced apoptosis was higher in blood PMN from early lactating cows than in blood PMN from midlactating cows. The higher percentage of necrotic milk PMN in early lactating cows than in midlactating cows could be explained by the induction of secondary necrosis