Dynamics of Leukocytes and Cytokines During Experimentally-Induced Streptococcus uberis Mastitis

Environmental mastitis is of increasing prevalence in well managed dairy herds throughout the world. Of the environmental pathogens, Streptococcus uberis is one of the most prevalent, accounting for a significant proportion of clinical and subclinical intramammary infections in lactating and nonlactating dairy cows and heifers. In spite of this, the pathogenesis of S. uberis mastitis is not well understood. The objective of this study was to determine dynamics of leukocytes and cytokines during experimentally-induced S. uberis mastitis. Five Jersey and 5 Holstein cows were challenged via intramammary inoculation with S. uberis into 2 uninfected mammary glands. One uninfected unchallenged mammary gland from each cow served as a negative control. Rectal temperatures, blood and milk samples were collected frequently for one week after challenge. Clinical status of mammary glands was recorded twice daily during milking. Milk samples were collected to enumerate bacteria and somatic cells, and to determine concentrations of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-8 (IL-8). Neutrophils were isolated from blood to determine expression of CD11b, CD18 and CD62 during the course of experimental infection. Sixteen of 20 challenged mammary glands developed clinical mastitis between 84 and 108 hours after intramammary challenge, and peak clinical signs of mastitis were observed at 144 hours. The number of S. uberis in milk from challenged mammary glands increased significantly (P\u3c0.05) by 48 hours after challenge, in spite of a massive leukocyte infiltration from the bloodstream, evidenced by an increase in the number of somatic cells in milk which began at 18 hours after challenge (P\u3c0.001) and remained elevated throughout the study period. Significant amounts of TNF-α, IL-1β and IL-8 in milk were detected 66 hours after challenge (P\u3c0.05). Peak concentrations of TNF-α were observed at 120 hours after challenge, preceding peak clinical signs. Concentrations of IL-1β in mammary secretions fluctuated considerably from 66 to 144 hours after challenge, but were significantly higher (P\u3c0.01) than prechallenge values and values found in milk from control mammary glands. Similar to TNF-α, peak IL-8 concentrations were observed at 120 hours after challenge. The 4 challenged mammary glands that did not become infected exhibited an increase in somatic cells and IL-1β concentrations in a pattern that appeared to be similar to that of challenged infected mammary glands, but of lesser magnitude. Expression of CD11b was upregulated at 84, 90 and 114 hours after challenge (P\u3c0.05), and expression of CD18 was increased 20% at 48 hours after challenge (P\u3c0.05). Expression of CD18 decreased sharply 96 hours after challenge and remained depressed until the end of the sampling period (P\u3c0.01). Results from this experiment demonstrated that S. uberis experimental intramammary infection induced local production of TNF-α, IL-1β and IL-8, which may play a role in the pathogenesis of S. uberis mastitis. Furthermore, an increase in somatic cells in milk occurred earlier than increases in adhesion molecule expression or cytokine production, suggesting that other mediators may be involved in initial leukocyte recruitment into the mammary gland after infection