Background: The so-called ventral organs are amongst the most enigmatic structures in Onychophora (velvet worms). They were described as segmental, ectodermal thickenings in the onychophoran embryo, but the same term has also been applied to mid-ventral, cuticular structures in adults, although the relationship between the embryonic and adult ventral organs is controversial. In the embryo, these structures have been regarded as anlagen of segmental ganglia, but recent studies suggest that they are not associated with neural development. Hence, their function remains obscure. Moreover, their relationship to the anteriorly located preventral organs, described from several onychophoran species, is also unclear. To clarify these issues, we studied the anatomy and development of the ventral and preventral organs in several species of Onychophora.Results: Our anatomical data, based on histology, and light, confocal and scanning electron microscopy in five species of Peripatidae and three species of Peripatopsidae, revealed that the ventral and preventral organs are present in all species studied. These structures are covered externally with cuticle that forms an internal, longitudinal, apodeme-like ridge. Moreover, phalloidin-rhodamine labelling for f-actin revealed that the anterior and posterior limb depressor muscles in each trunk and the slime papilla segment attach to the preventral and ventral organs, respectively. During embryonic development, the ventral and preventral organs arise as large segmental, paired ectodermal thickenings that decrease in size and are subdivided into the smaller, anterior anlagen of the preventral organs and the larger, posterior anlagen of the ventral organs, both of which persist as paired, medially-fused structures in adults. Our expression data of the genes Delta and Notch from embryos of Euperipatoides rowelli revealed that these genes are expressed in two, paired domains in each body segment, corresponding in number, position and size with the anlagen of the ventral and preventral organs.Conclusions: Our findings suggest that the ventral and preventral organs are a common feature of onychophorans that serve as attachment sites for segmental limb depressor muscles. The origin of these structures can be traced back in the embryo as latero-ventral segmental, ectodermal thickenings, previously suggested to be associated with the development of the nervous system.