Accuracy of qPCR and bacterial culture for the diagnosis of bovine intramammary infections and teat skin colonisation with Streptococcus agalactiae and Staphylococcus aureus using Bayesian analysis

Streptococcus agalactiae (Strep. agalactiae) and Staphylococcus aureus (Staph. aureus) are originally regarded as contagious mastitis pathogens, however, both pathogens have recently been isolated from extramammary and environmental sites, indicating that other sites than the udder might contribute to the spread of these pathogens potentially causing intramammary infections. Diagnostic tools to identify pathogens at extramammary sites are available but still needs to be validated. The objective of this cross-sectional field study was to estimate the diagnostic sensitivity (Se) and specificity (Sp) of the commercially available Mastit4 qPCR assay and bacterial culture (BC) in identifying Strep. agalactiae and Staph. aureus from milk and teat skin samples. We randomly selected 30-40 cows with high somatic cell counts from eight Danish Strep. agalactiae-positive dairy herds with automatic milking systems. Teat skin samples and aseptic milk samples were collected from right rear quarters (n = 287) for BC and PCR analysis. Se and Sp were estimated in a Bayesian latent class analysis. For milk samples, the Se and Sp of qPCR for Strep. agalactiae were estimated to 0.97 and 0.99, respectively, whereas the Se and Sp of BC were 0.41 and 1.00, respectively. The Se and Sp of qPCR for Staph. aureus were estimated to 0.95 and 0.99, respectively, whereas the Se and Sp of BC were 0.54 and 0.77, respectively. For teat skin samples, the Se and Sp of qPCR for Strep. agalactiae were estimated to be 0.97 and 0.96, respectively, whereas the Se and Sp of BC were 0.33 and 1.00, respectively. The Se and Sp of qPCR for Staph. aureus were estimated to 0.94 and 0.98, respectively, whereas the Se and Sp of BC were 0.44 and 0.74, respectively. In conclusion, the Se for diagnosing Strep. agalactiae and Staph. aureus IMI was higher for qPCR than BC, suggesting that qPCR is a valuable method for detecting both pathogens from quarter-level milk samples. The performance of BC in the detection of Strep. agalactiae and Staph. aureus on teat skin was poor compared to qPCR, indicating that differences in the target condition of the two methods should be considered when implementing them as routine diagnostic tests for detecting teat skin colonisers. The low Se of BC may preclude the use of BC for skin testing, and qPCR is better for this task.info:eu-repo/semantics/publishedVersio

Genomic analysis of European bovine Staphylococcus aureus from clinical versus subclinical mastitis

Abstract: Intramammary infections (IMI) with Staphylococcus aureus are a common cause of bovine mastitis and can result in both clinical (CM) or subclinical mastitis (SCM). Although bacterial isolates of S. aureus differ in their virulence potential it is largely unclear which bacterial virulence factors are responsible for increased clinical severity. We performed a genome wide association study and used a generalized linear mixed model to investigate the correlation between gene carriage, lineage and clinical outcome of IMI in a collection of S. aureus isolates from cattle with CM (n = 125) and SCM (n = 151) from 11 European countries. An additional aim was to describe the genetic variation of bovine S. aureus in Europa. The dominant lineages in our collection were clonal complex (CC) 151 (81/276, 29.3%), CC97 (54/276, 19.6%), CC479 (32/276, 11.6%) and CC398 (19/276, 6.9%). Virulence and antimicrobial resistance (AMR) gene carriage was highly associated with CC. Among a selection of nine virulence and AMR genes, CC151, CC479 and CC133 carried more virulence genes than other CCs, and CC398 was associated with AMR gene carriage. Whereas CC151, CC97 were widespread in Europe, CC479, CC398 and CC8 were only found in specific countries. Compared to CC151, CC479 was associated with CM rather than SCM (OR 3.62; 95% CI 1.38–9.50) and the other CCs were not. Multiple genes were associated with CM, but due to the clustering within CC of carriage of these genes, it was not possible to differentiate between the effect of gene carriage and CC on clinical outcome of IMI. Nevertheless, this study demonstrates that characterization of S. aureus CC and virulence genes helps to predict the likelihood of the occurrence of CM following S. aureus IMI and highlights the potential benefit of diagnostics tools to identify S. aureus CC during bovine mastitis