Thryssa of the family Engraulidae are widely distributed in the Atlantic Ocean, the Pacific Ocean and the Indian Ocean, with important commercial values in fishery and aquaculture of many countries. In order to achieve unambiguous species recognition and define species status, 652 bp segments of mtDNA cytochrome oxidase subunit I from 62 specimens representing 6 species of genus Thryssa were used as DNA barcoding to examine genetic distances within and among all reported morphological species in coastal waters of China mainland. The results showed that the nucleotide compositions were T: 29.0%, C: 26.3%, A: 25.3%, G: 19.4%, A+T(54.3%) is higher than C+G(45.7%), and the average value of transition/transversion ratios was 3.76. All species were monophyletic and formed into 5 clades with 100% bootstrap supports, of which T.vitirostris and T.mystax were in the same clade, and the average genetic distances within and among clades were 0.2%(0.0%-0.4%), 17.7%(15.7%-19.0%) respectively. Genetic distances among and within T.kammalensis, T.hamiltoni, T.dussumieri and T.setirostris were 15.7%-18.6%, 0.0%-0.3% respectively, i.e., inter-species distances were 10 times greater than intra-species divergence, which confirmed their species status. Although distances within T.mystax and T.vitirostris were both 0.1%, which were close to the 4 congeneric species, and the genetic distance between T.vitirostris and T.mystax was 0.6%, which fell within the scope of most reported intra-species distances, suggesting their closer relationship. Due to some distinguishable morphological and molecular differences, T.mystax and T.vitrirostris should be treated as 2 subspecies or recently diverged species. In conclusion, DNA barcoding based on COⅠ sequences is useful for identifying correctly 4 Thryssa species in coastal waters of China, yet the data also revealed possible cases of unrecognized subspecies or recently diverged species. T.mystax and T.vitrirostris should be managed as 2 significant units, highlighting the importance of synergy between molecular, and biological researches in understanding and documenting Chinese marine fish biodiversity.