BACKGROUNDGross human anatomy is essential in undergraduate programs across biological and health sciences. While extensive literature explores medical students' knowledge in this area, studies on non-medical students, particularly those in physical education, are scarce.OBJECTIVEThis study assessed the anatomy knowledge among Brazilian physical education students and explored differences based on employment status, type of class instruction (face-to-face vs. online), and involvement in academic activities. Additionally, we investigated students' perceptions of the assessment instrument and the gross human anatomy course itself.METHODSWe conducted a cross-sectional study involving physical education students of both sexes, aged 18 and older, from four public Brazilian universities. Participants completed a 15-question multiple-choice survey on human anatomy systems. Correct answers received one point, with a total potential score of 15 points (100%). We categorized scores as excellent (≥ 90%), good (71-89%), sufficient (50-70%), and insufficient (< 50%). Participants had 90minutes to complete the survey. We presented data as median and interquartile range [IQR], median difference (Δ), and 95% confidence intervals. Scores were compared against the median absolute (7.5 points) and relative (50%) values. We used rank-biserial correlation for effect size and set a significance level 0.05. The study included 216 students (143 males) with a median age of 22.4 years [IQR: 4.0].RESULTSThe final scores were significantly above the cut-offs, with a median absolute score of 9.0 [IQR: 3.0] and a relative score of 60% [IQR: 20%], showing large effects (Δ: +2.0 [1.5-2.0], P<0.001; Δ: +13.33% [10.0-13.34%], P<0.001). There were no significant differences in either the absolute or relative final scores (P≥0.05) between students who participated in academic activities and those who did not, nor between students who attended face-to-face versus online gross human anatomy classes. However, we found a significant difference between non-working students and their employed counterparts, with non-workers scoring higher both absolutely (P=0.002) and relatively (P=0.002) on the gross human anatomy questionnaire. Most of the participants described the difficulty of the gross human anatomy questionnaire as "average" (51.39%, n=111) and rated the gross human anatomy course difficulty as "difficult" (52.8%, n=114).CONCLUSIONBrazilian physical education students demonstrated only sufficient knowledge in gross human anatomy (60%). Therefore, strategies to enhance or maintain their knowledge throughout the educational program are vital.