A total of 258 bovine-associated Staphylococcus aureus isolates from the United States, Chile, and the United Kingdom, plus the reference isolate S. aureus Newbould 305 (NCIMB 702892), were analyzed by multilocus sequence typing (MLST). A collection of previously characterized United Kingdom isolates were also included in the analysis. The results demonstrated that MLST is suitable for the differentiation of bovine S. aureus isolates from various sites (milk, teat skin, milking machine unit liners, hands, and bedding) and countries. The theory of the host specificity of S. aureus is supported by the detection of a previously undescribed clonal complex that comprised 87.4% of the isolates studied, with representatives from all geographic locations investigated. This suggests that a single clonal group has achieved a widespread distribution and is responsible for the majority of infections. Some sequence types (STs; ST25, ST115, ST124, and ST126) demonstrated site specificity, as they were significantly (P < 0.05) associated with milk or teat skin.Staphylococcus aureus is a major cause of bovine mastitis and is spread from cow to cow (skin or milk) via the milking machine (35, 57). Environmental spread may also occur, since strains of S. aureus have been isolated from the environment of dairy farms and from other species that are present on dairy farms (32,39).A number of studies have identified potential sources of the pathogen and have investigated strain-specific differences (19, 40). The major potential sources identified were milk, body sites, and, to a lesser extent, the environment (40). Studies investigating the global population structure of bovine S. aureus suggest that a relatively few specialized clones are responsible for the majority of intramammary infections (IMIs) (26, 55), although some authors did not report between-farm genetic homogeneity (25,46). Most of these studies have used techniques such as phage typing (19, 40) and pulsed-field gel electrophoresis (PFGE) (7, 25) to compare isolates. These methods lack intercenter reproducibility (55). Library typing systems such as binary typing (BT) (53) and multilocus sequence typing (MLST) (31) have been developed to overcome these problems by producing results that are repeatable between laboratories and over time.The aim of this study was to investigate the effectiveness of MLST as a method for the typing of S. aureus isolates of bovine origin from a number of distinct geographical sources. A collection of isolates previously characterized by phage typing (19) and by PFGE and binary typing (55) was used to compare these methods to MLST. The data were then used in a preliminary analysis of the evolutionary and population biology of S. aureus isolates of bovine origin.

The purposes of this study were to identify sources of Staphylococcus aureus on dairies and to determine whether S. aureus colonization of heifer body sites increases the risk of S. aureus IMI at parturition. In herds with high (> 10%) or low (< 3%) prevalence of S. aureus IMI, S. aureus was isolated from heifer teat skin, heifer external orifices, housing, feedstuffs, humans, nonbovine animals, air, and equipment. Additionally, in herds with high prevalence, S. aureus was isolated from bedding, insects, and water. The predominant sources of S. aureus for both groups were other IMI and heifer body sites. Heifers with prepartum lacteal secretions with S. aureus were at greater risk of S. aureus IMI at parturition than were prepartum heifers with lacteal secretions that were negative for S. aureus. Heifers with teat skin colonized by S. aureus were 3.34 times more likely to have S. aureus IMI at parturition than were noncolonized heifers. Overall, 35% of 700 heifers were colonized with S. aureus on a body site at least once. Although colonizations of most body sites appeared to be transient, a few heifers were colonized on the same site for 1 yr. Persistently colonized heifers may represent the primary reservoirs of S. aureus for other heifers.

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Relationships between genetic measures of mastitis (somatic cell score, score for clinical mastitis, and scores for IMI with major or minor pathogens) and immunological parameters (physiological and molecular markers) were examined for periparturient Holstein cows. Physiological markers included 11 in vitro immunological assays. Molecular markers included the second exon of the DRB3 locus of the bovine major histocompatibility complex, the IgG2 isotype genotype, and the CD18 genotype (the locus responsible for bovine leukocyte adhesion deficiency). A gene substitution model was used to estimate the additive genetic effects of alleles of the three molecular markers on estimated breeding value (EBV) for mastitis measures. Pearson correlation coefficients between EBV for immunological assays and EBV for mastitis measures were computed. Molecular markers explained up to 40% of the variation in EBV for measures of mastitis. The presence of allele DRB3.2*16 was associated with higher EBV for SCS. Allele DRB3.2*8 was associated with increased EBV for clinical mastitis, as was the IgG2b allele and the normal CD18 allele. Alleles DRB3.2*11, *23, IgG2a, and the recessive allele for bovine leukocyte adhesion deficiency were associated with decreased clinical mastitis. A positive genetic association was found between allele DRB3.2*24 and EBV for IMI by major pathogens and between DRB3.2*3 and IMI by minor pathogens. Several correlations between EBV for immunological assays and EBV for mastitis measures were significantly different from 0. Cows with low EBV for SCS tended to have neutrophils that had greater functional ability at maximal immunosuppression, low serum IgG1, and high numbers of circulating mononuclear cells. Immunological parameters, including physiological and molecular markers, are useful aids to understand the genetics of resistance to mastitis.

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