This study aimed at investigating gender difference in the coordination of the subdivisions of the trapezius muscle during a repetitive box-folding movement task. Twenty-two healthy volunteers (11 males and 11 females) performed the repetitive box-folding task for 34 min. During the task, perceived exertion and surface electromyographic (EMG) signals from the upper, middle, and lower trapezius subdivisions were recorded. Absolute and normalised root mean square (RMS) values as well as normalised mutual information (NMI) values were calculated to assess, respectively, activation levels within muscle subdivisions and functional connectivity among subdivisions. Females compared with males were characterised by higher normalised RMS values in the upper trapezius (P < 0.05) and higher NMI values within the upper-middle subdivision pair (P < 0.05) during repetitive box-folding. The elevated normalised level of activation of the upper trapezius as well as the enhanced functional connectivity among upper-middle trapezius subdivisions underlined that females adopted a different motor strategy than males did during a dynamic repetitive task. Such differences within and among muscle subdivisions may not be a favourable trait and could partly contribute to the higher prevalence of musculoskeletal disorders reported in females.