Effect of subclinical intramammary infection on somatic cell counts and chemical composition of goats' milk.

We investigated effects of subclinical intramammary infection (IMI) on milk somatic cell count (SCC) and milk composition in udder halves of dairy goats. A total of 35 mixed-age Alpine does (70 udder halves; approximately 55 kg body weight) were rotationally grazed on a mixture of vegetative forages (wheat/berseem clover, sudan grass and cowpeas). Milk samples for bacterial analysis and SCC were collected monthly from both halves from April to September, 2001. Across stages of lactation, 19-31% of udder halves became infected. The prevalence of IMI exhibited quadratic patterns through multi-peaked responses within each stage of lactation. Higher rates of IMI were observed during the early stage of lactation (19% in May) and in the late stage of lactation (31% in September). Coagulase negative Staphylococcus (CNS, 43.7%), Staph. aureus (35.4%), and Pseudomonas aeruginosa (12.4%) were the most prevalent pathogens. Within single-strain IMI, log SCC (6.24) was lower (P<0.01) for CNS than those derived from IMI by Staph. aureus (6.49), Ps. aeruginosa (6.53) or Serratia spp. (6.90). Infected udder halves had a higher average SCC (4761 v. 2259 x 10(3) cells/ml; P<0.01) than uninfected halves, but uninfected halves often had similar levels of SCC to infected halves. Daily average milk production was not significantly different between infected and non-infected goats and the relationship between IMI and SCC was not always correlated. Effective mastitis screening requires bacteriological culture since SCC was not highly correlated.

Functional analysis of recombinant bovine CD14.

Studies in mice and humans indicate that membrane CD14 (mCD14) on the surface of monocytes, macrophages and polymorphonuclear neutrophils (PMN) mediate activation of these cells by lipopolysaccharide (LPS). Soluble CD14 (sCD14), in the circulation, binds to LPS and blocks LPS binding to mCD14. The role of bovine CD14 in cellular activation by LPS is undefined. Changes in CD18 expression on PMN and steady state levels of mRNA for tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and IL-8, sensitive markers for activation of leukocytes by LPS, were used to measure functional activity of recombinant bovine sCD14 (rbosCD14). Whole blood (n=3 cows) treated with LPS alone caused CD18 expression on PMN to increase by 12% (P<0.02), whereas pre-incubation of LPS with 10 or 100 microg/mL of rbosCD14 completely inhibited increase in CD18 expression. After treating whole blood with LPS at concentrations of 1, 100 or 10(4) ng/mL for 2 h, level of mRNA for TNF-alpha, IL-6 and IL-8 in leukocytes and concentration of TNF-alpha in plasma increased. However, pre-incubation of LPS with rbosCD14 inhibited the increase in TNF-alpha mRNA, but not the increase in IL-6 and IL-8 mRNA. Excess amount of anti-human CD14 monoclonal antibody (MAB) also inhibited LPS-induced increase in TNF-alpha mRNA. Preincubation of LPS with rbosCD14, or rbosCD14 plus MAB did not affect LPS-induced increase in TNF-alpha in plasma. Collectively, results indicate that rbosCD14 inhibit LPS-induced increase in CD18 expression and TNF-alpha mRNA. However, secretion of TNF-alpha was not inhibited by pre-incubation of LPS with rbosCD14. The TNF-alpha in plasma may partially induce transcription of IL-6 and IL-8, which contribute to the CD14-independent increase in level of mRNA for IL-6 and IL 8.

Apoptosis of bovine neutrophils during mastitis experimentally induced with Escherichia coli or endotoxin.

OBJECTIVE: To determine whether apoptosis of neutrophils was accelerated during mastits experimentally induced by use of Escherichia coli or E coli endotoxin and whether differences were apparent in the response to E coli or endotoxin. ANIMALS: 11 healthy lactating Holstein cows. PROCEDURE: Blood samples were collected from cows at various intervals after intramammary inoculation with E coli or endotoxin. Percentage of apoptotic neutrophils detected after in vitro incubation for 3 hours was determined. Fluorescein isothiocyanate-labeled annexin-V in combination with propidium iodide was used to distinguish apoptosis and necrosis of neutrophils. Total and differential circulating leukocyte counts and rectal temperature were determined at the time of collection of blood samples. Milk yield and milk somatic cell counts were determined at the time of milking. RESULTS: Inoculation of endotoxin did not accelerate in vitro induction of neutrophil apoptosis. However, inoculation of E coli increased the percentage of apoptotic neutrophils. At 18 hours after inoculation, 20% of the neutrophils were apoptotic, compared with 5% before inoculation. Milk somatic cell count and rectal temperature increased, milk production and total leukocyte count decreased, and percentage of immature neutrophils increased after inoculation with E coli or endotoxin. However, kinetics of the responses were more rapid, more severe, and of shorter duration during endotoxin-induced mastitis. CONCLUSIONS AND CLINICAL RELEVANCE: In vitro induction of apoptosis of neutrophils was accelerated only during E coli-induced mastitis and not during endotoxin-induced mastitis. Endotoxin inoculation as a model for studying coliform mastitis in dairy cows should be viewed with caution.