Function of milk polymorphonuclear neutrophil leukocytes in bovine mammary glands infected with Corynebacterium bovis

Abstract

Corynebacterium bovis is one of the most commonly isolated bacteria from aseptically collected bovine milk samples. The objective of the current study was to characterize the bovine innate immune response by evaluating milk polymorphonuclear neutrophilic leukocytes (PMNL) in mammary glands infected with C. bovis. Twenty quarters infected with C. bovis and 28 culture-negative quarters (with milk somatic cell count <1×10(5) cells/mL) were used. The percentages of milk PMNL and the PMNL expression of L-selectin (CD62L), β2-integrin (CD11b), and one of the endothelial-selectin ligands (CD44), as well as the levels of intracellular reactive oxygen species (ROS) and the phagocytosis of Staphylococcus aureus, were evaluated by flow cytometry. The apoptosis and necrosis rates of the PMNL were quantified using dual-color flow cytometry with fluorescein-labeled annexin and propidium iodide. The present study revealed a higher percentage of PMNL in the milk from C. bovis-infected quarters, although no significant differences were found in levels of CD44, CD62L, or CD11b expression among the PMNL. A lower percentage of apoptotic PMNL was observed in C. bovis-infected quarters, as well as higher percentages of viable PMNL and of PMNL that produced intracellular ROS. However, no alterations were observed in phagocytosis of Staph. aureus by the PMNL or in intensity of intracellular ROS production by PMNL. Thus, results from this investigation of the PMNL function support, at least in part, the fact that intramammary infections by C. bovis may offer protection against intramammary infections by other bacteria.

Effects of bovine leukemia virus infection on milk neutrophil function and the milk lymphocyte profile

“…The present study used 57 quarters of the mammary glands of 19 dairy cows in a commercial herd, at different lactation stages. Due to the effects of bacterial mastitis pathogens on neutrophil function [ 36 – 40 ], the following exclusion criteria were applied: 1) bacteriologically positive quarters; 2) quarters with abnormal secretions in the strip cup test; and 3) quarters with high somatic cell counts (SCCs) based on the previously proposed threshold for SCC [ 41 , 42 ].…”

Expression of CD14 and toll-like receptors 2 and 4 by milk neutrophils in bovine mammary glands infected with Corynebacterium bovis

“…Lastly, a labeled secondary antibody was added and incubated for 30 min at room temperature in the dark to visualize CH138A binding. Neutrophils were identified using flow cytometry based on their cytoplasmic granularity and CH138A positivity (Fig.1), as previously described by Piepers et al (2009) and Blagitz et al (2013). The secondary labeled mAb consisted of 1µL of fluorescein isothiocyanate (FITC) (M31501; Invitrogen, Carlsbad, USA).…”

“…bovis is of interest to mastitis researchers because of the observation that quarters infected with this bacterium are significantly less likely to become infected with other, more pathogenic bacteria (Brooks & Barnum 1984, Sordillo et al 1988, Lam et al 1997, Blagitz et al 2013, although no consensus has been reached (Zadoks et al 2001, Reyher et al 2012. With this concept in mind, it should be noted that polymorphonuclear neutrophil leukocytes (PMNLs) provide the first cellular line of defense against invading mastitis pathogens and are the main source of resistance during the early stages of bacterial invasion (Schukken et al 2011.…”

Transmission dynamics of intramammary infections caused by Corynebacterium species

“…Although Corynebacterium spp. are classified as minor pathogens (Brooks and Barnum, 1984b;Harmon, 1994;Blagitz et al, 2013), the increased prevalence of Corynebacterium spp. IMI in some modern dairy farms (Pitkälä et al, 2004) warrants further investigation into the specific properties and roles of the bacteria.…”