Bactericidal activity of macrophages against Streptococcus uberis is different in mammary gland secretions of lactating and drying off cows

Abstract

The aim of this study was to compare the ability of milk macrophages and macrophages from the mammary gland secretions during the mid-dry period for their interaction with the mastitis-causing Streptococcus uberis. We also aimed to determine if S. uberis induced the release of the cytokine tumour necrosis alpha (TNF-α) and the bactericidal moiety nitric oxide (NO) from milk macrophages of lactating cows and macrophages from the mammary gland secretions at the mid-dry period. Macrophages were isolated from the mammary gland secretions of cows during the mid-lactation or mid-dry period, and compared with blood monocytes for their interaction with the important mastitis-causing pathogen S. uberis. When infected in vitro with S. uberis, milk macrophages from lactating cows with S. uberis released modest amounts of the cytokine tumour necrosis factor alpha (TNF-α) (139 pg/ml) and the bactericidal moiety nitric oxide (NO) (3–4 μM of nitrite). Blood monocytes from lactating cows released significantly higher amounts of TNF-α (345 ± 143 pg/ml) and NO (7 ± 2 μM of nitrite) after interaction with S. uberis, compared to milk macrophages ( P < 0.01 for both TNF-α and NO). Stimulation of blood monocytes with the cytokine interferon-gamma (IFN-γ) enhanced significantly the release of NO and TNF-α, but IFN-γ did not significantly enhance the production of NO and TNF-α by milk macrophages from lactating cows. Milk macrophages from all lactating cows failed to kill S. uberis efficiently, and this lack of killing was unaffected by prior treatment with gamma interferon (IFN-γ) ( P > 0.05). Rather, S. uberis multiplied significantly inside infected milk macrophages from lactating cows, with a two-fold increase in bacterial numbers at 2 h post-infection. Milk macrophages from lactating cows were able however, to kill a significant proportion (50–60%, P < 0.01) of phagocytosed Staphylococcus aureus. Blood monocytes from all cows were found to exert significant bactericidal activity against S. uberis. There were no significant differences in the bactericidal activity of milk macrophages obtained from lactating cows with low somatic cell counts (SCC; <10 5 ml −1) compared with those with a mildly elevated SCC (>10 5 ml −1) ( P > 0.05). In contrast, mammary gland secretion macrophages isolated from the same cows in the mid-dry period killed a significant proportion of phagocytosed S. uberis (50–65% of ingested S. uberis killed, P < 0.01) although cytokine production in response to in vitro bacterial infection was low. We conclude that the bactericidal activity of mammary gland secretion macrophages against a virulent strain of S. uberis is low during the lactation period. In addition, our data indicate that S. uberis is not a strong inducer of NO and TNF-α in macrophages from the milk or mammary gland secretions of cows during the drying off period. Finally, IFN-γ does not activate milk macrophages or macrophages from cows during the lactating period or mammary gland secretions during the drying off period.