The occurrence of <i>Aerococcus urinaeequi</i> and non-aureus staphylococci in raw milk negatively correlates with <i>Escherichia coli</i> clinical mastitis

<i>Escherichia coli</i> is a common environmental pathogen associated with clinical mastitis (CM) in dairy cattle. There is an interest in optimizing the udder microbiome to increase the resistance of dairy cattle to <i>E. coli</i> CM; however, the details of which members of the healthy udder microbiome may play a role in antagonizing <i>E. coli</i> are unknown. In this study, we characterized the bacterial community composition in raw milk collected from quarters of lactating Holstein dairy cows that developed <i>E. coli</i> CM during lactation, including milk from both healthy and diseased quarters (<i>n</i> = 1,172). The milk microbiome from infected quarters was compared before, during, and after CM. A combination of 16S rRNA gene amplicon and metagenomic sequencing was used generate data sets with a high level of both depth and breadth. The microbial diversity present in raw milk significantly decreased in quarters experiencing <i>E. coli</i> CM, indicating that <i>E. coli</i> displaces other members of the microbiome. However, the diversity recovered very rapidly after infection. Two genera, <i>Staphylococcus</i> and <i>Aerococcus</i>, and the family Oscillospiraceae were significantly more abundant in healthy quarters with low inflammation. Species of these genera, <i>Staphylococcus auricularis, Staphylococcus haemolyticus,</i> and <i>Aerocussus urinaeequi</i>, were identified by metagenomics. Thus, these species are of interest for optimizing the microbiome to discourage <i>E. coli</i> colonization without triggering inflammation.IMPORTANCEIn this study, we show that <i>E. coli</i> outcompetes and displaces several members of the udder microbiome during CM, but that microbial diversity recovers post-infection. In milk from quarters which remained healthy, the community composition was often highly dominated by <i>S. auricularis, S. haemolyticus, A. urinaeequi,</i> and <i>S. marcescens</i> without increases in somatic cell count (SCC). Community dominance by these organisms, without inflammation, could indicate that these species might have potential as prophylactic probiotics which could contribute to colonization resistance and prevent future instances of <i>E. coli</i> CM.