Abstract: Objective To understand the virulence genes and antibiotic resistance of Vibrio parahaemolyticus serotype O3:K6 in southeastern coastal area of China. Methods Antibiotic susceptibility testing was performed for 77 strains of O3:K6 V. parahaemolyticus collected in Shanghai,Shenzhen,Zhejiang and Jiangsu provinces(municipality) during 2002-2016. Results Up to 94.81% and 1.30% of the tested strains were positive for tdh and trh respectively. The sensitive rates of the strains to some antibiotics were low,i. e. 7.79% to ampicillin,18.18% to streptomycin,31.17% to kanamycin,49.35% to sulfamethoxazole and 54.55% to ciprofloxacin,but the resistance rates were high to ampicillin(84.42%),sulfamethoxazole (36.36%) and streptomycin(32.47%). The sensitive rates to ceftriaxone(96.10%),cefoxitin(87.01%),cefepime(81.82%), gentamicin (97.40%) and azithromycin (93.51%) were high. All the strains were sensitive to imipenem,trimethoprim/sulfamethoxazole,nalidixic acid,doxycycline and chloramphenicol. Eleven strains(14.29%) were multidrug resistant,and all these strains were pathogenic. Conclusion Most O3:K6 V. parahaemolyticus isolated from southeastern coastal area of China were tdh positive and trh negative. Except ampicillin,sulfamethoxazole,aminoglycoside and ciprofloxacin,the O3:K6 V. parahaemolyticus isolates were sensitive to common used antibiotics,however,some strains showed multidrug resistance.