Autophagy Dually Induced by AMP Surplus and Oxidative Stress Enhances Hemocyte Survival and Bactericidal Capacity via AMPK Pathway inCrassostrea hongkongensis

Crassostrea hongkongensis(Hong Kong oyster) is an ecologically and economically valuable shellfish endemic to South/Southeast Asia. Due to ocean acidification and warming waters, they have become increasingly vulnerable to invading microbes includingVibrio parahaemolyticus, a significant foodborne human pathogen. In recent years, outbreaks ofV. parahaemolyticushave emerged as a perennial phenomenon in parts of the world, necessitating to better understand the biology of host-pathogen interactions in this under-examined marine invertebrate. Although an immunologically relevant autophagy apparatus has been identified inCrassostrea gigas, an evolutionarily close mollusk cousin, the precise mechanistic details ofC. hongkongensisautophagy duringV. parahaemolyticusinfection are still wanting. Here, we compellingly demonstrated thatin vivo V. parahaemolyticuschallenge robustly triggered autophagic signaling inC. hongkongensishemocytes peaking at 6 h post-infection, which subsequently promoted bacterial clearance and dampened premature apoptosis. Simultaneously, a large surplus of adenosine monophosphate (AMP) and elevations in reactive oxygen species (ROS, specifically mitochondrial O(2)(-)and cellular H2O2) formation were observed post-infection. Extrinsically applied AMP and ROS could synergistically induce AMP-activated protein kinase (AMPK) phosphorylation to stimulate downstream autophagic events.V. parahaemolyticusinfection-induced autophagy was pharmacologically shown to be AMPK-dependentin vivo. Overall, our results establish autophagy as a crucial arm of host defense againstVibrioinfections in mollusks, and provide new insights into the underappreciated roles of ROS and AMP as co-regulators of autophagy.