Objective: Ayu (Plecoglossus altivelis) vibriosis threatens ayu aquaculture seriously caused by mass mortality due to severe infections. We characterized the vibriosis pathogen of ayu in Ninghai country., Methods: A dominant strain was isolated and identified by a series of biochemical and physiological tests. The lethal dose 50% (LD50) was calculated by the modified Karber's method. PCR amplification and sequence analysis were used to further identify the pathogen., Results: LD50 of ayu-H080701 was 1.2 x 10(4) CFU to ayu. PCR amplification showed that the bacterial universal primers for 16S rRNA gene and the specific primers for the metalloprotease (MP) gene of Listonella anguillarum worked. 16S rRNA gene analysis showed that ayu-H080701 shared 99.4%-99.5% nucleotide identical to L. anguillarum isolates, while 94.3% and 91.9% nucleotide identical to L. pelagius and Photobacterium damselae respectively. Metalloprotease analysis showed that ayu- H080701 shared 97.6%-98.8% amino acid sequence identical to L. anguillarum isolates, while lower than 75.6 % to other bacteria. Phylogenetic analysis showed that ayu-H080701 grouped constantly with L. anguillarum isolates., Conclusion: The biochemical, physiological tests and sequence analysis all strongly supported the identification of the pathogen causing ayu vibriosis in Ninghai country, China, as an isolate of L. anguillarum.