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Antibiotic-resistant bacteria are severe threats to aquaculture industry. Boosting and modulating host immune responses has been proved to be an effective strategy to combat with bacterial infections and there is an urgent need for novel immunomodulators. Cathelicidins is an important family of host defense peptides (HDPs) that possess direct antimicrobial activities and potent immunomodulatory properties. Several cathelicidins have been identified and characterized from diverse fish species. Considering the relatively conserved immune systems between different fish species, it is reasonable to speculate that cathelicidins from different fish species possess immunomodulating functions on the other fish species. In the present study, two fish-derived cathelicidins (CATH_BRALE and codCath1) were selected to investigate their protective effect on zebrafish with bacterial infections. They exhibited potent and broad-spectrum antimicrobial activities against the tested aquatic Gram-positive and Gram-negative pathogenic bacteria, with MIC values ranging 2.34–18.75 μg/ml for CATH_BRALE and 2.34–37.5 μg/ml for codCath1. And their antimicrobial effect is so rapid that they killed the bacteria within 60 min. Unlike conventional antibiotics, they kill bacteria by inducing bacterial membrane permeabilization and cell disruption. Besides direct antimicrobial activity, CATH_BRALE and codCath1 exhibited potent immunomodulatory functions by both inhibiting bacteria induced zebrafish pro-inflammatory cytokine gene (TNF-α, IL-1β, and IL-6) expression and stimulating zebrafish chemokine gene IL-8 expression. In vivo challenge test proved that they could significantly decrease the bacterial numbers and enhance the survival rates of zebrafish. All the results above imply the great potential of CATH_BRALE and codCath1 as novel peptide immunomodulators in fish aquaculture industry. • Two fish-derived cathelicidins exhibited potent antimicrobial activities against aquatic pathogenic bacteria. • CATH_BRALE and codCath1 kill bacteria by inducing bacterial membrane permeabilization and cell disruption. • CATH_BRALE and codCath1 exhibited potent immunomodulatory function. • CATH_BRALE and codCath1 significantly decreased the bacterial number and enhanced the survival rate of zebrafish. • CATH_BRALE and codCath1 possess great potential to be novel peptide immunomodulators in fish aquaculture industry. [ABSTRACT FROM AUTHOR]