Persistent Staphylococcus aureus Chronic Bovine Mastitis

Staphylococcus aureus is a versatile opportunistic bacterial pathogen in humans and animals. S. aureus is known for its great capability to specifically adapt to different hosts and multiple host sites expressing a large set of virulence factors to evade attacks of the immune system. S. aureus host adaptation often results in the emergence of so-called persistent phenotypes, which are difficult to eradicate, such as in chronic bovine mastitis or chronic osteomyelitis and cystic fibrosis. To date, S. aureus chronic bovine mastitis still represents a major health problem for dairy cattle accounting for great economic losses in dairy industries. The mechanism by which S. aureus persist and spread within a herd is incompletely understood. In this study, we aim to identify S. aureus surface-exposed factors and secreted proteins/ genes capable to establish a persistent intramammary infection. The proposed project is based on the availability of a unique set of S. aureus host adapted isolates from the same clonal ancestry, where key factors of persistence can be exclusively assigned to the natural selection process within the host. To this end, we will analyze the initial and host-adapted isolates in an ex vivo bovine mammary gland explant model using transcriptome, metabolome and proteome-based methods to identify changes of the composition of bacterial cell-surface associated and secreted factors utilized by the host- adapted isolate to establish a persistent infection. Loss (gain)-of-function mutations of selected candidate molecules will be generated and their contribution to forming biofilms, adhering to and invading bovine epithelial cells will be determined. As a future perspective, newly identified candidate molecules of persistence could lead to the development of tests that would enable us to predict the chronic potential of an isolated strain and would allow the development of novel targeted treatment regimens, which is of utmost importance to effectively combat chronic/ persistent intramammary infections.

We investigated the adaptation of the bacterial pathogen Staphylococcus aureus (S. aureus) within its host. This bacterium is a master in adaptation and can colonize and infect different hosts and multiple host sites. We demonstrate that bacterial in-host evolution opens a new, alternative route to persistence leading away from intracellular towards extracellular persistence. This finding is relevant as S. aureus is one of the six so-called ESKAPE pathogens, a group of emerging, highly virulent and antibiotic-resistant bacterial pathogens. This new bacterial route to persistence has been first described in a dairy cow with a long-lasting, chronic udder infection with implications for animal welfare, food safety and milk yield. S. aureus is a common colonizer of human and animal skin. However, host adaptation often results in the emergence of so-called persistent phenotypes, which are difficult to eradicate, such as in chronic infections of the udder in dairy cows (bovine mastitis). We investigated the adaptation of S. aureus within the animal host over three months in a naturally infected dairy cow with chronic, subclinical mastitis. The analysis of the bacterial genomes revealed a complete replacement of the original bacterial strain by another genetic variant - caused by an adaptation through mutation and selection. This study was the first to observe a genetic and phenotypical adaptation of S. aureus within its bovine host. We could demonstrate that the bacterial genome mutation involves only a single base exchange in one gene. The newly evolved variant exhibits significant changes in infection-relevant phenotypes, such as higher proteolytic activity, biofilm formation and bacterial growth. The project's outcome significantly improved our understanding of the dynamic adaptive behaviour of this pathogen during chronic, persistent infections. It could help develop targeted intervention strategies against bovine S. aureus or other mastitis pathogens.