Abstract: In order to expand the reservoir of highly active agarase-producing strains and to mine agarase-related gene pools, this study isolated strains from abalone viscera. Primary screening was conducted using a single carbon source medium combined with Lugo's iodine staining, followed by rescreening via the 3,5-dinitrosalicylic acid (DNS) method to determine enzyme activity. Agarase-producing strains were identified by 16S rDNA sequencing, and their genomic sequences and enzymatic properties were compared. Four strains of Vibrio mediterranei (2DB7, 2DB15, 2DBB20 and 2DBB51) producing highly active agarases were screened. The activities of the crude enzyme solutions from these strains were 315.17, 340.99, 293.88, and 324.37 U/mL, respectively. Whole genome sequencing revealed that 2DB15, 2DBB20, and 2DBB51 encode relatively abundant agarase genes and related enzyme genes, including β-agarases (GH16 and GH50 families), β-galactosidase, and α-neoagarobiose hydrolase. The agarases produced by these four strains (2DB7, 2DB15, 2DBB20 and 2DBB51) reached their peak relative activity at 45, 40, 45, and 45 ℃, and at pH 8.0, 7.0, 8.0, and 7.0, respectively. Furthermore, all enzymes maintained good stability below 40 ℃ and within a pH range of 5.0 to 9.0. The research results could enrich the theoretical foundation for agarase production by Vibrio sp. and provide high-quality enzyme resources for the eco-friendly production of agar oligosaccharides.