Sea urchin, Strongylocentrotus intermedius, transplanted from Japan in 1989, has been widely cultured along the coasts of Liaoning and Shandong Provinces and has become the dominant and most economically important maricultured species in North China. However, a lesion syndrome symptom of S. intermedius broke out frequently these years, showing lethargy in activities, blackish peristomial membrane and body well lesions, and brought about high mortality eventually. Six representative prominent bacterial strains were isolated from diseased sea urchin from September 2009 to January 2010. By means of API 20NE and 16S rRNA gene sequences analysis, isolates were identified as Shewanella aquimarina, Pseudoalteromonas tetraodonis, Vibrio shilonii, V. harveyi, V. fortis and V. splendidus. Bacterial challenge tests showed that their representative isolates were virulent to S. intermedius with LD50 values ranging from 9.2 × 104 to 3.4 × 106 CFU/g body weight, among which S. aquimarina , V. fortis and P. tetraodonis were highly virulent, and the other three isolates showed moderate virulence. The results indicated that a variety of bacteria including Shewanella, Pseudoalteromonas and Vibrio were involved in the mortality of S. intermedius, and the six isolates were opportunistic pathogens of sea urchins. All isolates reported herein were sensitive to ampicillin, enrofloxacin, ofloxacin, doxycycline and florfenicol.