Immunoproteomic analysis of the serum IgG response to cell wall-associated proteins of Staphylococcus aureus strains belonging to CC97 and CC151.

CC97 and CC151 are two of the most common Staphylococcus aureus lineages associated with bovine intramammary infection. The genotype of the infecting S. aureus strain influences virulence and the progression of intramammary disease. Strains from CC97 and CC151 encode a distinct array of virulence factors. Identification of proteins elaborated in vivo will provide insights into the molecular mechanism of pathogenesis of these lineages, as well as facilitating the development of tailored treatments and pan-lineage vaccines and diagnostics. The repertoire of genes encoding cell wall-anchored (CWA) proteins was identified for S. aureus strains MOK023 (CC97) and MOK124 (CC151); MOK023 encoded more CWA proteins than MOK124. Serum collected during an in vivo challenge trial was used to investigate whether the humoral response to cell wall proteins was strain-specific. Immunoproteomic analysis demonstrated that the humoral response in MOK023-infected cows predominantly targeted high molecular weight proteins while the response in MOK124-infected cows targeted medium or low molecular weight proteins. Antigenic proteins were identified by two-dimensional serum blotting followed by mass spectometry-based identification of immunoreactive spots, with putative antigens subsequently validated. The CWA proteins ClfB, SdrE/Bbp and IsdA were identified as immunogenic regardless of the infecting strain. In addition, a number of putative strain-specific imunogens were identified. The variation in antigens produced by different strains may indicate that these strains have different strategies for exploiting the intramammary niche. Such variation should be considered when developing novel control strategies including vaccines, therapeutics and diagnostics.

Immunoproteomic analysis of the secretome of bovine-adapted strains of Staphylococcus aureus demonstrates a strain-specific humoral response.

Staphylococcus aureus is a common pathogen associated with bovine intramammary infection. A number of distinct S. aureus lineages are associated with such infections although there is a dearth of knowledge regarding the major immunogenic antigens associated with each lineage and whether these antigens provide protection against heterologous strains. Identification of the major immunogenic antigens of the predominant bovine-adapted S. aureus lineages would assist in the design of effective vaccines and diagnostic tests to control intramammary infections caused by S. aureus. The aim of this study was to characterise the serum IgG response to S. aureus extracellular proteins in cows infected with strains from different lineages, as well as to identify antigenic proteins produced by these strains. Genotypic characterisation found that strain MOK124 (CC151) encoded more toxins, including the ruminant-specific leukocidin LukMF, compared to strain MOK023 (CC97). In addition, MOK124 secreted more toxins in vitro, compared to MOK023. Immunoproteomic analysis was performed using sera from cows infected with either MOK023 or MOK124. One-dimensional serum blotting revealed that cows infected with MOK023 predominantly generated a humoral response against high molecular weight proteins while cows infected with MOK124 primarily generated a humoral response against low molecular weight proteins. Two-dimensional serum blotting demonstrated that antibodies produced by an MOK023 infected cow could cross react with some of the extracellular proteins produced by MOK124 and vice versa. Mass spectrometry analysis of immunoreactive proteins identified common candidate immunogens produced by both strains, including α-hemolysin and ß-hemolysin. In addition, strain-specific candidate immunogens were also identified. This study demonstrates that genes encoding important S. aureus secreted virulence factors, the production of cognate gene products, and the humoral immune response to infection is, to an extent, strain-dependent. However, the identification of some common candidate immunogens suggests that there are proteins that can be exploited for further vaccine or diagnostic research that targets S. aureus strains from a variety of lineages.