The sheep is a commonly used animal model for human lumbar spine surgery, but only in vitro investigations comparing the human and ovine spine exist. Spinal musculature has previously not been compared between man and sheep. This additional knowledge could further indicate to what extent these species are biomechanically similar. Therefore, the purpose of the study was to investigate spinal muscle morphometric properties using magnetic resonance imaging (MRI) in different age groups of healthy human participants and sheep in vivo. Healthy human participants (n = 24) and sheep (n = 17) of different age groups underwent T1-weighted MRI of the lumbar spine. Regions of interest of the muscles erector spinae (ES), multifidus (M) and psoas (PS) were identified. The ratio of flexor to extensor volume, ratio of M to ES volume, and muscle fat relative to an area of intermuscular fat were calculated. Sheep M to ES ratio was significantly smaller than in the human participants (sheep 0.16 ± 0.02; human 0.37 ± 0.05; P < 0.001), although flexor to extensor ratio was not significantly different between species (human 0.39 ± 0.08; sheep 0.43 ± 0.05; P = 0.06). Age did not influence any muscle ratio outcome. Sheep had significantly greater extensor muscle fat compared with the human participants (M left human 40.64%, sheep 53.81%; M right human 39.17%, sheep 51.33%; ES left human 40.86%, sheep 51.29%; ES right human 35.93%, sheep 44.38%; all median values; all P < 0.001), although PS did not show any significant between-species differences (PS left human 36.89%, sheep 33.67%; PS right human 32.78%, sheep 30.09%; P < 0.05). The apparent differences in the size and shape of sheep and human lumbar spine muscles may indicate dissimilar biomechanical and functional demands, which is an important consideration when translating to human surgical models.