Neu1-deficient zebrafish cells exhibit reduced Edwardsiella piscicida infection due to altered lysosomal exocytosis and membrane dynamics

Edwardsiella piscicida is a Gram-negative intracellular pathogen causing Edwardsiellosis, leading to economic losses in aquaculture. While phagocytosis is its primary infection route, alternative entry pathways remain largely unexplored. Neu1 sialidase, a lysosomal enzyme in glycoconjugate degradation, was investigated for its role in E. piscicida infection using primary cultured cells derived from Neu1-KO zebrafish fin (Neu1-KO cells). Compared to wild-type (WT) cells, Neu1-KO cells exhibited lower infection rates, which were associated with enhanced lysosomal exocytosis. Infection was restored by the intracellular calcium chelator BAPTA-AM, highlighting the role of exocytosis. Leupeptin, a cysteine/serine protease inhibitor, increased E. piscicida infection in Neu1-KO cells. Neu1-KO cells exhibited lower rab10 expression and reduced membrane ruffling, which was restored by BAPTA-AM and leupeptin. Given the role of epidermal growth factor receptor (EGFR) signaling, we assessed its phosphorylation, which was reduced in Neu1-KO cells but restored by treatment with BAPTA-AM and leupeptin. This suggests that inhibiting lysosome exocytosis or extracellular protease activity may enhance EGFR phosphorylation. These findings indicate that the decreased E. piscicida infection in Neu1-KO cells resulted from enhanced lysosomal exocytosis, leading to increased extracellular protease secretion, subsequent EGFR inactivation by extracellular protease, and reduced EGFR-regulated ruffling. This study provides novel insights into the regulatory mechanisms of bacterial infection and lysosomal exocytosis, informing potential therapeutic strategies against intracellular pathogens.