Abstract
Invertebrates lack adaptive immunity and rely exclusively on innate defenses to combat pathogens. Recent studies have shown that invertebrates exhibit immune priming, a form of innate immune memory in which prior pathogen exposure enhances protection against reinfections. Despite its ecological and applied relevance, immune priming remains poorly explored in marine invertebrates. Most studies adopt a host-centric view, focusing on the immune response while overlooking that immunity occurs within a complex microbial context in which the host, its microbiota, and pathogens interact. This gap limits our understanding of how microbiota and pathogens behave under priming pressure. Here, we investigated the effects of immune priming on host survival, pathogen dynamics, and microbiota composition in the Manila clam (Ruditapes philippinarum), a key species for global aquaculture, challenged with the bacterial pathogen Vibrio europaeus. Immune priming was induced orally using live V. europaeus at a sublethal dose, followed by a lethal secondary challenge. Primed clams exhibited a significant increase in survival after the second challenge (87% survival vs. 0% in non-primed clams) and rapidly reduced the pathogen concentration within 48 h post-challenge below the mortality threshold observed in non-primed clams (~10(5) copies mg(-1)). Notably, the pathogen was detected at low but non-harmful concentrations (~10(2) copies mg(-1)) in primed clams during both challenges. Full-length 16S rRNA amplicon sequencing analyses revealed that immune priming reshaped the host microbiota. Beta diversity analyses suggest the establishment of a specific bacterial community in primed clams. Clustering analyses identified a priming-associated microbiota dominated by Acinetobacter, Brevundimonas, Sphingobium, and Psychrobacter, which persisted at the end of the priming and the secondary challenge but was absent or depleted in non-primed clams. In contrast, members of Arcobacteraceae emerged only during the second challenge, decreasing in primed clams but increasing in non-primed clams in association with high mortality. This study provides the first phenotypic evidence of oral immune priming in the Manila clam, identifies a specific microbiota and detects the pathogen despite effective host protection. This offers a new perspective to address immune priming as an emergent property of dynamic interactions between the host, its microbiota, and the pathogen.