Comparative embryology and muscle development of polyclad flatworms (Platyhelminthes: Rhabditophora)

Polyclads belong to the phylum Platyhelminthes, lineage Rhabdotophora, are simple Bilateria, and represent an interesting and useful group for research in developmental biology. Although polyclads, together with catenulids and macrostomids, have been argued to be most closely related to the ancestral flatworm and hold the key to understand the relationship between development and evolution, knowledge of their embryonic development is still scarce and most of the work on spiralian development has focused on mollusks and annelids. In view of polyclad embryonic significance, a comparative study of their embryonic development including several species of direct and indirect developers was performed. Developing embryos of 16 species representing 10 families were examined and followed through embryogenesis until hatching. Considerable differences in egg plates, egg capsule morphology, size and number of eggs, and developmental time lines were found among the analyzed species. A correlation between developmental times and morphology of egg capsules was found; likewise it was possible to link larger eggs to direct developing species and longer developmental time and for most but not all, indirect developing species to smaller egg sizes. The number of eggs per egg capsule does not appear to be of systematic value, instead the morphology of the female reproductive system may play a significant role in determining the number of embryos per egg capsule. The influence of parental care on hatching success was also determined for two local species. Covering of egg masses by the adult was observed for individuals of both species and although this parental care is not necessary for egg development or hatching, it plays a significant role in the hatching success of Pleioplana atomata embryos. For individuals of Imogine zebra, parental covering of recently laid egg masses may play a role in egg capsule formation. In the polyclad Pericelis cata a case of developmental dimorphism was found. Larvae and juveniles from the same parent hatched simultaneously throughout the three-day hatching period. This represents the first case of true poecilogony reported for polyclad flatworms which may be a bet-hedging strategy in which benthic juveniles are recruited to the parental habitat, and concurrently, siblings disperse as larvae. The most notable feature of the development of P. cata was the unusual appearance of extra-embryonic yolk inside the egg capsules. Similarly, almost all the larvae of this species had only one eye, whereas three eyes characterize typical Muller\u27s and Gotte\u27s larvae. Understanding the origin of the muscular system may have implications for the understanding of bilaterian evolution. Hence, a comparative analysis of body wall formation and muscle organization during embryogenesis was performed. Fluorescent dye-conjugated phalloidin was used to characterize the musculature of Maritigrella crozieri and Melloplana ferruginea which represent an indirect and a direct developing species, respectively. In both species, the first myoblasts were localized in the periphery of the egg. Progressively, myoblasts formed unorganized and rudimentary muscle fibers that further differentiated during development. Muscle differentiation was similar between the two species; however, the process of muscle development progressed quite differently in larvae and juveniles. These results provide additional support that the orthogonal muscle pattern is a symplesiomorphy of Spiralia and it may have been present in the stem species of all Bilateria