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Escherichia coli is a frequent cause of clinical mastitis in dairy cows. It has been shown that a prompt response of the mammary gland after E. coli entry into the lumen of the gland is required to control the infection, which means that the early detection of bacteria is of prime importance. Yet, apart from lipopolysaccharide (LPS), little is known of the bacterial components which are detected by the mammary innate immune system. We investigated the repertoire of potential bacterial agonists sensed by the udder and bovine mammary epithelial cells (bMEC) during E. coli mastitis by using purified or synthetic molecular surrogates of bacterial agonists of identified pattern-recognition receptors (PRRs). The production of CXCL8 and the influx of leucocytes in milk were the readouts of reactivity of stimulated cultured bMEC and challenged udders, respectively. Quantitative PCR revealed that bMEC in culture expressed the nucleotide oligomerization domain receptors NOD1 and NOD2, along with the Toll-like receptors TLR1, TLR2, TLR4, and TLR6, but hardly TLR5. In line with expression data, bMEC proved to react to the cognate agonists C12-iE-DAP (NOD1), Pam3CSK4 (TLR1/2), Pam2CSK4 (TLR2/6), pure LPS (TLR4), but not to flagellin (TLR5). As the udder reactivity to NOD1 and TLR5 agonists has never been reported, we tested whether the mammary gland reacted to intramammary infusion of C12-iE-DAP or flagellin. The udder reacted to C12-iE-DAP, but not to flagellin, in line with the reactivity of bMEC. These results extend our knowledge of the reactivity of the bovine mammary gland to bacterial agonists of the innate immune system, and suggest that E. coli can be recognized by several PRRs including NOD1, but unexpectedly not by TLR5. The way the mammary gland senses E. coli is likely to shape the innate immune response and finally the outcome of E. coli mastitis.

Staphylococcus aureus is a major cause of mastitis in ruminants. In ewe mastitis, symptoms range from subclinical to gangrenous mastitis. S. aureus factors or host-factors contributing to the different outcomes are not completely elucidated. In this study, experimental mastitis was induced on primiparous ewes using two S. aureus strains, isolated from gangrenous (strain O11) or subclinical (strain O46) mastitis. Strains induced drastically distinct clinical symptoms when tested in ewe and mice experimental mastitis. Notably, they reproduced mild (O46) or severe (O11) mastitis in ewes. Ewe sera were used to identify staphylococcal immunoreactive proteins commonly or differentially produced during infections of variable severity and to define core and accessory seroproteomes. Such SERological Proteome Analysis (SERPA) allowed the identification of 89 immunoreactive proteins, of which only 52 (58.4%) were previously identified as immunogenic proteins in other staphylococcal infections. Among the 89 proteins identified, 74 appear to constitute the core seroproteome. Among the 15 remaining proteins defining the accessory seroproteome, 12 were specific for strain O11, 3 were specific for O46. Distribution of one protein specific for each mastitis severity was investigated in ten other strains isolated from subclinical or clinical mastitis. We report here for the first time the identification of staphylococcal immunogenic proteins common or specific to S. aureus strains responsible for mild or severe mastitis. These findings open avenues in S. aureus mastitis studies as some of these proteins, expressed in vivo, are likely to account for the success of S. aureus as a pathogen of the ruminant mammary gland.

International audience; The objective of this study was to review the possible relationships between hyperketonemia and the function of phagocytes with respect to the bovine udder defense mechanism. We hypothesize that an increased incidence of clinical mastitis in high-producing cows is caused by the impairment of the udder defense mechanism during hyperketonemia. First, we review the acute phase of udder defense mechanisms after intramammary infection. The physiological changes of cows in negative energy balance are subsequently discussed. Finally, possible relationships between udder defense and physiological changes during negative energy balance, especially hyperketonemia, are reviewed. The three stages of an acute phase of udder defense are: (1) immediately eliminating invading pathogens by phagocytes, (2) releasing inflammatory substances, especially chemoattractants, and (3) migration of polymorphonuclear leukocytes into the infected udder. Leukocytes from hyperketonemia subjects show a lower capacity of the phagocytic defense mechanism. In addition, the phagocytic and bactericidal capacities of neutrophils are reduced when these cells are acting in the presence of high concentrations of ketone bodies. Lower amounts of cytokine production after bacterial infection are observed in ketotic subjects. The chemotactic capacity of blood leukocytes is impaired in leukocytes obtained from ketotic cows. Lower numbers of blood leukocytes are observed in ketotic cows. In conclusion, the impairment of the udder defense mechanism in negative energy balance cows seems related to hyperketonemia.; Hypercétonémie et diminution des défenses mammaires : une synthèse. L'objectif de cette étude est de présenter des relations possibles entre l'hypercétonémie et la fonction des phagocytes pour ce qui est des mécanismes de défense de la mamelle. Notre hypothèse est qu'une augmentation des mammites cliniques chez des vaches laitières à haute production serait causée par une détérioration des mécanismes de défense de la mamelle suite à une hypercétonémie. Premièrement, le stade aigu des mécanismes de défense de la mamelle après une infection a été étudié. Ensuite, les changements physiologiques chez les vaches ayant un bilan énergétique négatif ont été présentés. Finalement, les relations entre les mécanismes de défense de la mamelle et les changements physiologiques au cours d'un bilan énergétique négatif, notamment une hypercétonémie, ont été revus. Les trois stades d'une phase aiguë de défense mammaire sont les suivants : (1) des organismes pathogènes envahisseurs sont phagocytés immédiatement, (2) des substances inflammatoires sont produites, en particulier des substances chimio-tactiques, (3) des leucocytes polynucléaires migrent vers la mamelle infectée. La capacité phagocytaire des leucocytes ainsi que celle des neutrophiles et la capacité bactéricide de ceux-ci sont apparemment détériorés dans les conditions d'hypercétonémie. La quantité de cytokines produites après une infection bactérienne est diminuée chez les sujets hypercétonémiques. Le chimiotactisme des leucocytes sanguins est détérioré dans les leucocytes provenant de vaches hypercétonémiques. Des quantités plus faibles de leucocytes sanguins sont observés dans les vaches hypercétonémiques. En conclusion, la détérioration des mécanismes de défense de la mamelle au cours de la période du bilan énergétique négatif semble liée à l'hypercétonémie.

International audience; Data from a prospective longitudinal study carried out from 1986 to 1990 in 47 commercial Holstein dairy herds from western France were used to quantify the effects of udder health disorders on the risk of culling. Logistic regression was used to assess the relationship between 4 udder health disorders and early and late culling. Teat injuries were associated with an increased risk of early culling in the current and following lactations. Mastitis and high milk cell count were associated with an increased risk of late culling in the current and following lactations, respectively