Polyembryony, brood-chamber initiation and sperm utilization in cyclostome bryozoans.

Monozygotic polyembryony is a form of asexual reproduction that proceeds by division of post-zygotic stages to produce clonal broods sharing the same sexually derived genotype. The persistence of polyembryony has puzzled evolutionary biologists because it seems to combine the contrasted fitness disadvantages of cloning and sexual reproduction while compromising the respective benefits. Cyclostomata is a phylogenetically ancient order of bryozoans, which in the Late Triassic began to develop voluminous brood chambers. In Recent forms, individual brood chambers accommodate multiple larvae. Based on the evidence of late 19th and early 20th century microscopy, each brood originates by iterative budding of a primary embryo. Recently, molecular genotyping of brooded embryos and maternal colonies in one cyclostome species, Crisia denticulata, has conclusively confirmed monozygotic polyembryony and indicated that the single genotype cloned within each brood chamber arises from outcrossing via water-borne sperm. Harmer's original histological inference of polyembryony in three suborders was corroborated by the extensive work of Borg, with evidence of the same process in the two remaining extant suborders. It is reasonable to infer that the enlarged brood chamber, recorded in all living families of cyclostomes except the Cinctiporidae, is associated with embryonic budding. Nevertheless, the possibility of multiple fertilizations within the brood chamber, or of self-fertilization, or of parthenogenesis cannot be dismissed without knowing the genotypic composition of mother and embryos. Such possibilities are significant in understanding the retention of this arguably paradoxical reproductive mode and identifying its possible adaptive significance in cyclostomes. Also significant in this respect is whether brood chamber development is induced by the presence of water-borne allosperm, as has been found in the cheilostome Celleporella hyalina (and some other spermcast maters), where female investment is triggered by allosperm uptake. Hypothesis 1: Individual brood chambers in all major groups of cyclostomes brood non-selfed, non-parthenogenetic embryos of identical genotype. To date, polyembryony in cyclostomes has only been proven genetically in Crisia denticulata, using microsatellites developed for that species. The studentship will employ molecular markers (ISSRs) to investigate the genetic composition of broods in a range of cyclostome species chosen to represent all extant clades, utilizing the latest understanding of cyclostome phylogeny being developed at the Natural History Museum. Laboratory rearing in reproductive isolation will test the possibility of self-fertilization. Hypothesis 2: Brood chamber development in cyclostomes is triggered by the presence of allosperm. The sporadic distribution of brood chambers within and between cyclostome colonies raises important questions about the control of their development. In particular, does the uptake of allosperm trigger brood chambers to form, as in some cheilostomes? The effect of the presence of conspecific sperm upon gonozooid development during colony growth will be elucidated in laboratory cultures, compared with control colonies in continuing isolation. Supplementary investigation: Onward culture of colonies through multiple reproductive cycles with additional matings (exposure to allosperm through to cessation of larval release) will determine duration and numerical extent of embryonic cloning, and whether gonozooids can be used for successive broods of different genotype. Hypothesis 3: Do cyclostomes regulate inbreeding by differential acceptance of sperm|? The cheilostome (non-polyembryonic) bryozoan Celleporella hyalina utilizes sperm differentially depending on the relatedness between source and recipient. Does the same pattern occur in the polyembryonic cyclostomes? Trial laboratory matings between colonies of varying relatedness will test this.