Identifying genetic factors in Streptococcus uberis that enable evasion of the host immune response

Mastitis—the inflammation of the mammary tissue—is a major endemic disease in dairy cattle that often results from intramammary infection, causing significant losses in the dairy industry in both profit and product quality. With the continued development of mastitis control programs, the incidence of infections associated with contagious pathogens in commercial dairies has declined, yet these measures have had limited success in controlling environmental mastitis pathogens, which remain an issue even in well-managed herds. The pathogen Streptococcus uberis, in particular, accounts for a substantial proportion of clinical and subclinical mastitis cases; furthermore, its various strains differ in terms of their virulence and resistance to the host immune response, often able to survive and proliferate despite substantial neutrophil activity, which normally would kill pathogens. To better understand the nature of this resistance, six mutated clones of the S. uberis clinical isolate UT888 were subjected to a bactericidal assay using fresh bovine neutrophils, and their growth at designated time points was compared, with the objective of identifying mutants that were susceptible to being killed by neutrophils. These mutants were selected from the previously generated mutant library based on the results of a preliminary MTT screening that indicated that the six chosen mutants produced significantly greater reactive oxygen species (ROS) in the presence of neutrophils (PMN+) than without (PMN-). As greater ROS production is considered an indicator of cell stress, it was expected that these mutants would be more susceptible to neutrophil killing and thus would exhibit less cell growth in PMN+ versus PMN- conditions in the bactericidal assay. However, five of the six mutants presented with a greater ratio of cell growth of PMN+: PMN-; a single mutant (12E2) exhibited a lower ratio and thus may present an area of interest for future investigations.