Typical harvesting baskets (TB) are used in various agricultural workplaces; however, no study to date has reported their effect on the musculoskeletal system. Therefore, this study aimed to evaluate the effects of a novel basket with attached rotational handles (RHB) to help alleviate the work-related physical burden of farmers. We analyzed the surface electromyograms (EMGs) of seven muscles, evaluated the subjective discomfort levels and locally perceived discomfort (LPD) scores to investigate the discomfort in the whole body and seven hand muscles, respectively. The EMGs showed that muscle activity decreased in five muscles (flexor carpi ulnaris, extensor carpi radialis, lateral triceps, middle deltoid, and upper trapezius) and increased in two (biceps brachii and erector spinae) when the RHB was used (p < 0.05). The subjective discomfort score for the RHB decreased compared to that for TB (p < 0.001). The LPD scores also decreased, and the RHB and TB scores ranged from 1.25–1.40 and 3.1–3.25, respectively. The use of the RHB may prevent wrist bending, and reduce the activity of certain muscles while increasing the activity of other muscles. Therefore, it is necessary to conduct training and to evaluate the working posture while considering the affected muscles.