Molecular Epidemiology Of Staphylococcus Aureus Isolated From Bulk Tank Milk Samples Of U.S. Dairy Farms

Abstract

Staphylococcus aureus (S. aureus) is the most common cause of contagious bovine mastitis. Colonization of dairy cows by S. aureus, especially those expressing multi-drug resistance (MDR) and toxin production, is a concern for animal-health and well-being as well as public health. The primary objectives of this study were to investigate the prevalence, antibiotic resistance, virulence determinants and genetic diversity of S. aureus in bulk tank milk samples (BTM) using whole genome sequencing. A total of 365 pooled BTM samples (189 during winter and 176 during summer) were collected in 2016 from 189 herds from 19 different states in the United States. Staphylococcus aureus was cultured from 170 BTM samples, indicating a prevalence of 46.6% in the BTM sample set and a herd prevalence of 62.4% (118 out of 189 herds). The most commonly identified antibiotic resistance gene was norA followed by aph(3’)-Ia. The mecA gene responsible for methicillin resistance was observed in only one isolate. Twenty-six isolates were identified as multi-drug resistant. In general, 100 % of the sequenced S. aureus isolates harbored at least one virulence gene and the aur, hlgB, hlgA, hlgC, hlb, lukE, lukD and splA genes were the most frequently detected. In the 53 staphylococcus enterotoxin gene positive isolates, sen, sem, sei, seg, seo and seu were the most frequently detected putative virulence genes. Fourteen different sequence types (ST) based on muti-locus sequences and 18 spa types were identified. The most common ST type was ST2187 and the most common spa type identified was t529. The most predominant clone was CC5 represented by multiple ST types and CC151 was the second most predominant clone. The single isolate identified as MRSA belonged to ST72-CC5 and spa type t126. A secondary study objective was to determine herd characteristics and management practices associated with the presence of S. aureus, as well as MDR strains of S. aureus, BTM samples of U. S. dairy herds. Majority of the dairies adopted important mastitis-prevention practices such as fore-striping (86.9%), pre-milking teat disinfection (94%), post-milking teat disinfection (98.8%), drying teats before applying the milking unit (87.5%) and infusing long acting intramammry antibiotics at dry-off (95.8%). Mixed logistic regression modelling indicated that herd characteristics and management practices associated with a lower odds for isolating S. aureus from BTM included larger herd size (>500 lactating cows), herds with Jersey or other as the predominant breed (vs Holstein) and herds using non-manure organic bedding materials (e.g. straw, shavings) for lactating cows versus reclaimed sand. The practices of housing lactating cows on a bedded pack (vs free stalls) and herds that routinely examined foremilk as a part of their milking routine were associated with higher odds of isolating S. aureus from BTM. Mixed logistic regression models did not identify any herd level risk factors associated with the presence of MDR strains of S. aureus in BTM. Our findings indicated a high prevalence of S. aureus in BTM, with little antibiotic resistance but often carrying genes for the various enterotoxins identified in S. aureus. This study could help identify the predominant clones (CC5 & CC151) and provides continued surveillance of S. aureus strains found in U.S. dairy herds. It also provides an insight into the level of adoption of various recommended management practices for control of S. aureus mastitis by U.S. dairy herds. Furthermore, with ongoing changes in housing, bedding and milking practices, it is important to examine newly adopted practices as potential risk factors for S. aureus transmission within and between herds.