Genotyping for better knowledge on spread, virulence and resistance of coagulase negattive staphylococci from cases of mastitis in dairy cows including zoonotic aspects

The project objectives were to increase the knowledge on the epidemiology of udder infections (IMI) in dairy cows caused by three common species of coagulase negative staphylococci (CoNS), i.e. Staphylococcus (S.) chromogenes  (SCH) S. simulans (SSI) and S. epidermidis (SEP). We also investigated zoonotic aspects concerning SEP infections. CoNS are part of the normal skin flora in both animals and humans, but are also a common cause of infections. In dairy cows CoNS are common bacterial findings in milk from udder quarters with subclinical mastitis, but CoNS are also a common finding in human healthcare associated infections. 

In a previous study, we collected CoNS isolates from milk of dairy cows with subclinical mastitis. Amon those 105 isolates of SCH, 118  SSI isolates and 130 SEP isolates were selected. We also collected 60 SEP-isolates from milk filters on farm and 13 SEP isolates from nasal samples of farmers and personnel (NS). All bacterial isolates were analyzed using whole genome sequencing (WGS) to establish relatedness and genetic variation within species, and between/within herds. We also investigated prevalences of genes coding for antimicrobial resistance (AMR) and virulence as well as associations between genotypes and degree of inflammation as well as persitency of IMI. To investigate zoonotic aspects of SEP we compared sequencing data from milk and milk filter isolates with SEP-isolates from NS and with already existing data for 88 human isolates from healthcare associated infections. 

Among milk isolates 47 sequence types (STs) of SCH were identified, ST6 and ST109 being the most common. Among the SEP-isolates 31 STs were identified and ST99, ST100 and ST570 were most common. It was not possible to identify STs for SSI but several clusters were identified. Each dominating SEP-ST was found in several herds and in several cows within herd which indicates spread between and within herds. Similar patterns were also seen for some SCH-STs. Some spread within, but not between, herds was also observed for some SSI-genotypes. The results indicate botj differences and similarities in spread and variation between and within species which may be associated with the variation in AMR and virulence that was observed between and within species. 

In total, only two AMR genes per species were found for SCH and SSI while the corresponding number for SEP was higher (14). The occurrence per gene was, however, mostly very low. The gene blaZ, which indicates resistance to penicillin, was, however, found in 22% of SCH and 45% of SEP but in 0% of SSI isolates.  These results highlights the importance to investigate this trait before treatment of SCH and SEP mastitis. It was also clear that the occurrence of blaZ was more common among some types of SCH and SEP. variation between and within species was also observed in occurrence of potential virulence factors (pVFs). The number of pVFs per isolate was similar between species but the occurrence of specific pVFs or function groups of pVFs varied somewhat between STs and clusters within species. Differences between SCH-STs was found in prevalence of persistent IMI and degree of inflammation. For SEP comparisons were also made between milk isolates and isolated from milk filters collected 5 years later in 18 herds. In 12 of those the same ST was found at both samplings which indicates that the same SEP can remain over a long time period. 

Finally, SEP isolates from milk/milk filters from cows were compared with human isolates (NP and VRI). Of a total of 61 STs only 4 were found both in cows and humans. None of those STs were common. ST2 and ST215 dominated among isolates from VRI but were not found among the milk isolates. AMR genes were more common among VRI isolates.

The project gave new knowledge on the epidemiology of IMI with SCH, SEP and SSI which can be of importance for preventive measures for mastitis in dairy cows. The results also indicate that overlapping of SEP between cows and humans are uncommon. Epidemiologic associations and genome characteristics for the four STs that did overlap need further investigation.