The neutrophil function and lymphocyte profile of milk from bovine mammary glands infected with Streptococcus dysgalactiae

Abstract

Streptococcus dysgalactiae is a bacterium that accounts for a notable proportion of both clinical and subclinical intramammary infections (IMIs). Thus, the present study explores the function of milk neutrophils and the lymphocyte profile in mammary glands naturally infected with Streptococcus dysgalactiae. Here, we used 32 culture-negative control quarters from eight clinically healthy dairy cows with low somatic cell counts and 13 S. dysgalactiae-infected quarters from six dairy cows. Using flow cytometry, we evaluated the percentage of milk monocytes/macrophages and neutrophils, expression of CD62L, CD11b and CD44 by milk neutrophils, the levels of intracellular reactive oxygen species (ROS) production and phagocytosis of Staphylococcus aureus by milk neutrophils, and neutrophil viability. Furthermore, the percentages of B cell (CD21(+)) and T lymphocyte subsets (CD3(+)/CD4(+)/CD8(-); CD3(+)/CD8(+)/CD4(-); and CD3(+)/CD8(-)/CD4(-)), and the expression of CD25 by T milk lymphocytes (CD3(+)) and T CD4(+) milk cells were also assessed by flow cytometry using monoclonal antibodies. The present study showed a higher SCC and percentage of milk neutrophils, and a decrease in the percentage of milk monocytes/macrophages from S. dysgalactiae-infected quarters when compared to uninfected ones. We also observed a higher expression of CD11b by milk neutrophils and a tendency toward a decrease in neutrophil apoptosis rate in S. dysgalactiae-infected quarters. In addition, the S. dysgalactiae-infected quarters had higher percentages of milk T cells (CD3(+)) and their subset CD3(+)CD8(+)CD4(-) cells. Overall, the present study provided new insights into S. dysgalactiae IMIs, including distinct lymphocyte profiles, and a tendency toward an inhibition of apoptosis in milk neutrophils.

Diagnosing mastitis in early lactation: use of Somaticell®, California mastitis test and somatic cell count

“…Thus, milk quarter samples with CMT score higher than zero is suggestive to be infected. Mononuclear leukocytes are known to predominate in colostrum and milk samples from healthy cows Blagitz et al 2015). Moreover, higher CMT scores are not due simply to an increase in SCC, but also to alteration of the proportion of leukocytes in mammary gland secretions, which may be a result of increased neutrophils in infected mammary glands (Della ).…”

Intracellular reactive oxygen species production and phagocytosis of Staphylococcus aureus by milk neutrophils as tool to diagnose mastitis and identify susceptible dairy cows

“…The MFI of S. aureus phagocytosis was estimated from the median value of PI fluorescence (Blagitz et al , 2015. At least 20,000 cells were examined per sample.…”

“…The median fluorescence intensity (MFI) of ROS production was estimated from the median of DCF fluorescence (Blagitz et al , 2015. For this assay, 10,000 gated neutrophils were examined per sample.…”

“…The phagocytosis assay was performed using flow cytometry of PI-labeled S. aureus (Hasui et al 1989, 2015. Briefly, 2×10…”

“…In a healthy cow's milk the majority of somatic cells are macrophages and some polymorphonuclear neutrophils (PMN), epithelial and lymphocytes are also presented (Blagitz et al 2015). In a case of intrammamary infection, as in mastitis, the level of neutrophils increases from 11% (healthy quarter) up to 90% (Pyörälä 2003, Mira et al 2013, which makes the percentage of neutrophils in the SCC a potential mastitis indicator.…”