Capsular polysaccharide mediates Streptococcus agalactiae to resist Nile tilapia macrophage phagocytosis

Streptococcus agalactiae (S. agalactiae) is a highly pathogenic gram-positive bacterium for farmed tilapia. The surface capsular polysaccharide (cps) containing terminal sialic acid (sia) residues is a crucial virulence factor for S. agalactiae. However, there is limited knowledge regarding the effects of cps and sia on the phagocytosis of S. agalactiae by teleost macrophages (MФ). In this study, we investigated the role of cps and sia in tilapia MФ phagocytosis of S. agalactiae using cps-deficient mutant ∆cps and sia-deficient mutant ∆neuA. Nonopsonic phagocytosis of wild type (WT), ∆cps and ∆neuA was initially analyzed by flow cytometry. The results indicated that MФ phagocytosis of ∆cps and ∆neuA was significantly higher than that of WT. The proportion of MФ with higher intracellular reactive oxygen species (ROS) or nitric oxide (NO) levels, and overall mean fluorescence intensity (MFI) of the ROS or NO in MФ increased higher during nonopsonic phagocytosis in the Δcps and ΔneuA. The expression levels of pro-inflammatory factors (IL-1β, IL-6, TNF-α) during S. agalactiae infection revealed that S. agalactiae could dampen the inflammatory response of MФ through the cps and sia. Increased susceptibility of the ∆cps and ∆neuA to phagocytic killing indicated that S. agalactiae could impair the phagocytic clearance of MФ through the cps or sia. Furthermore, opsonic phagocytosis of the ∆cps and ∆neuA by MФ was dramatically enhanced compared to the WT under complement or antibody mediation. Moreover, relative to ∆cps and ∆neuA, nonopsonic and opsonic phagocytic receptors were significantly down-regulated during MФ phagocytosis of WT, indicating that S. agalactiae might resist phagocytosis of MФ by directly or indirectly inhibiting the expression of phagocytic receptors through the cps or sia. Our data suggest that cps plays a crucial role in the interference of S. agalactiae on the phagocytosis of tilapia MФ, and sia may be a significant immune escape site. These results provide valuable insights into the mechanism of S. agalactiae attempting to escape the defenses of teleost innate immune system and offer clues for S. agalactiae vaccine development.