Abstract: The mussel is one of the main cultivated species in the world. A significant challenge faced by suspension-cultured mussels is the high incidence of mussel fall-off from cultivation ropes, adversely impacting harvest yields, which have been documented at commercial mussel farms in the United Kingdom, the United States of America, Canada, Spain, New Zealand and China. Byssus is an important attachment structure for marine mussels, and weakness in byssal thread attachment is a major factor leading to mussel detachment from ropes. To investigate the relationship between genetic variability and byssal thread phenotypic characteristics in the hard-shelled mussel (Mytilus coruscus), we collected three wild populations of M. coruscus from different latitudes in the East China Sea, including the Shengsi (SS), Jiaojiang (JJ), and Fuding (FD) populations. The genetic diversity and structure of these populations were investigated using 10 microsatellite loci. The mean observed heterozygosity (H_o) in the SS population was 0.44, higher than the mean H_o values of the JJ (0.40) and FD (0.39) populations. The mean inbreeding coefficients (F_is) in the SS population was 0.20, lower than the mean F_is values of the JJ (0.33) and FD populations (0.40). These results revealed that the SS population exhibited higher genetic diversity compared to the other two populations. The different numbers of private alleles (P_a) in the three populations, ranging from 10 to 17, suggest that these populations have experienced selective pressures from various environments. Moreover, genetic differentiation was observed in the genetic distance between the SS population and the other two populations. We also examined the phenotypic characteristics of their byssal threads. There were significant differences in byssus attachment strength among the three populations, with the SS population located at the highest latitude secreting more byssal threads and exhibiting greater byssal breaking force and plaque adhesion strength, while the Fuding (FD) population located at the lowest latitude had the weakest byssal attachment. The observed differentiation in private alleles and byssus phenotypes might suggest that the three wild populations have experienced different environmental selective pressures. This study provides insight for future genetic enhancement programs aimed at improving byssus attachment in M. coruscus.