Immunoproteomics to identify Staphylococcus aureus antigens expressed in bovine milk during mastitis

Abstract

Staphylococcus aureus is an opportunistic pathogen affecting both human and animal species. An effective vaccine to prevent S. aureus bovine disease and transmission would have positive effects on animal well-being, food production, and human health. The objective of this study was to identify multiple antigens that are immunoreactive during udder colonization and disease for exploration as vaccine antigens to prevent bovine mastitis. Staphylococcus aureus produces several cell wall-anchored and surface-associated virulence factors that play key roles in the pathogenesis of mastitis. Many of these proteins are conserved between different strains of S. aureus and represent promising vaccine candidates. We used an immunoproteomics approach to identify antigenic proteins from the surface of S. aureus. The expression of cell wall and surface proteins from S. aureus was induced under low iron conditions, followed by trypsin extraction and separation by 2-dimensional electrophoresis. The separated proteins were blotted with antibodies from mastitic bovine milk and identified by liquid chromatography-mass spectrometry. Thirty-eight unique proteins were identified, of which 8 were predicted to be surface exposed and involved in S. aureus virulence. Two surface proteins, iron-regulated surface determinant protein C (IsdC) and ESAT-6 secretion system extracellular protein (EsxA), were cloned, expressed, and purified from Escherichia coli for confirmation of immune reactivity by ELISA. A PCR of 37 bovine S. aureus isolates indicated that the presence of esxA and isdC is conserved, and amino acid alignments revealed that IsdC and EsxA sequences are highly conserved. The immunoproteomics technique used in this study generated reproducible results and identified surface exposed and reactive antigens for further characterization.

Evaluation of Chromogenic Culture Media for Rapid Identification of Gram-Positive Bacteria Causing Mastitis

“…The ability to rapidly identify Staph. aureus infected cows are essential for the control of SCM in problem-herds, considering the potential of contagious transmission and low cure risk (24). The results of Staph.…”

Comparison of antibody repertories against Staphylococcus aureus in healthy and infected dairy cows with a distinct mastitis history and vaccinated with a polyvalent mastitis vaccine

“…Immunoproteomics approaches have been used to identify relevant immunoreactive candidate antigens from bovine-associated S. aureus (Tedeschi et al, 2009;Misra et al, 2018). Nevertheless, none of the previous studies have addressed the heterogeneity of anti-staphylococcal antibody production by dairy cows with a Cunha et al: STAPHYLOCOCCUS AUREUS MASTITIS VACCINE Figure 3.…”

“…Although surface proteins are considered crucial antigens in the prevention of new infections (Anderson et al, 2012;Foster et al, 2014;Misra et al, 2018), cytosolic S. aureus proteins are also important antigens that should not be neglected. Some cytoplasmic proteins possess moonlighting functions, displaying distinct functions at different stages; these proteins may also appear extracellularly, and might play an important role in the virulence of the microorganism during infection (Henderson and Martin, 2013).…”

“…In the present study, we used blood serum samples for a serological proteome approach to study a disease of the mammary gland. Misra et al (2018) used pooled milk samples from acutely clinical mastitic cows and bacteriologically negative healthy cows. However, the antibody titers in milk from healthy mammary glands in dairy cows correlated with blood titers, due to the transudation of plasma antibodies and preferential transport of IgG 1 (Fox and Adams, 2000) and IgG 2 (Paape et al, 2003).…”

“…Staphylococcus aureus is primarily an extracellular pathogen; as a result, host immune responses to this species depend mainly on neutrophil defense mechanisms supported by anti-staphylococcal adaptive humoral responses. Thus, the development of an effective vaccine for combating IMI from S. aureus would be most welcome; an effective vaccine that can prevent new IMI has not yet been developed (Misra et al, 2018;Rainard et al, 2018). A great challenge for vaccine research and development is to identify protective antigens (Rainard et al, 2018) that include multiple conserved antigens and afford cross-protection against distinct S. aureus strains (Misra et al, 2018).…”