Contribution of sortase SrtA2 to <i>Lactobacillus casei</i> BL23 inhibition of <i>Staphylococcus aureus</i> internalization into bovine mammary epithelial cells

<div><p>Probiotics have been considered as a promising strategy to prevent various diseases in both humans and animals. This approach has gained interest in recent years as a potential means to control bovine mastitis. In a previous study, we found that several <i>L</i>. <i>casei</i> strains, including BL23, were able to inhibit the internalization of <i>S</i>. <i>aureus</i>, a major etiologic agent of mastitis, into bovine mammary epithelial cells (bMEC). This antagonism required a direct contact between <i>L</i>. <i>casei</i> and bMEC or <i>S</i>. <i>aureus</i>, suggesting the inhibition relied on interactions between <i>L</i>. <i>casei</i> cell surface components and bMEC. In this study, we have investigated the impact of some candidates which likely influence bacteria host cell interactions. We have shown that <i>L</i>. <i>casei</i> BL23 <i>fbpA</i> retained its inhibitory potential, indicating that <i>L</i>. <i>casei</i> BL23 antagonism did not rely (solely) on competition between <i>S</i>. <i>aureus</i> and <i>L</i>. <i>casei</i> fibronectin-binding proteins for adhesion to bMEC. We have then investigated the impact of four sortase mutants, <i>srtA1</i>, <i>srtA2</i>, <i>srtC1</i> and <i>srtC2</i>, and a double mutant (<i>srtA1-srtA2</i>) on <i>L</i>. <i>casei</i> BL23 inhibitory potential. Sortases are responsible for the anchoring on the bacterial cell wall of LPXTG-proteins, which reportedly play an important role in bacteria-host cell interaction. All the <i>srt</i> mutants tested presented a reduced inhibition capacity, the most pronounced effect being observed with the <i>srtA2</i> mutant. A lower internalization capacity of <i>L</i>. <i>casei srtA2</i> into bMEC was also observed. This was associated with several changes at the surface of <i>L</i>. <i>casei</i> BL23 <i>srtA2</i> compared to the wild type (wt) strain, including altered abundance of some LPXTG- and moonlighting proteins, and modifications of cell wall structure. These results strongly support the role of sortase A2 in <i>L</i>. <i>casei</i> BL23 inhibition against <i>S</i>. <i>aureus</i> internalization. Deciphering the contribution of the cell surface components altered in <i>srtA2</i> strain in the inhibition will require further investigation.</p>