414
Vasa localization during embryonic development is similar across solitary ascidi-
ans. Vasa mRNA is expressed in the posterior lineage and late the germ cells of the
gonad in another solitary species, Boltenia villosa (Brown and Swalla 2007 ), similar to
Ciona intestinalis. Vasa localized in two posterior cells at the eight-cell stage and later
cleavage stages. A strong signal was detected in posterior cells of the cleaving embryo,
which gave rise to B7.6 cells. Expression of vasa later became localized to two cells
posterior to the endodermal strand of the tailbud embryo. These putative germ cells are
most likely derived from the B8.12 cells, as reported for Ciona (Fig. 8.8).
Embryos in colonial species are brooded and lineage-tracing studies lag behind
those in the more accessible free-spawning solitary species. Nevertheless, determina-
tive development is a conserved characteristic in ascidians. As observed in solitary
species, vasa-mRNA concentrates in granular structures of the fertilized egg cortex in
the colonial ascidian Botryllus schlosseri (Brown et al. 2009 ). During the first cleav-
age, vasa-mRNA localized to the vegetal pole of the embryo. Vasa is then segregated
in the cleavage furrow of the posterior-most B4.1 pair of cells of the eight-cell embryo
and in the descendant B5.1 pair of cells of the 16-cell embryo. During gastrulation,
vasa-mRNA was expressed in a pair of cells found posterior to the blastopore that are
likely to correspond to the precursor germ line B7.6 pair, reminiscent of results in
Ciona (Shirae-Kurabayashi et al. 2006 ). So far, only one study investigated the devel-
opmental stages of colonial ascidians later than gastrula. In Botryllus primigenus,
both vasa-mRNA and protein were detected in embryos until the tailbud stage, but no
signals were detected in the tail nor the trunk of tadpole larva (Kawamura et al. 2010 ).
More studies are needed to investigate the fate of germ cells in tailbud stage embryos,
tadpole larvae and during metamorphosis of colonial ascidians.
Taken together, all the available data strongly suggest that in both solitary and
colonial ascidians the process of germ cell segregation during embryonic develop-
ment is very similar, and that the number of germ line blastomeres and their descen-
dants is constant during embryogenesis to the tailbud stage.
8.4.2 Gonad Regeneration During Asexual Reproduction
in Colonial Ascidians
In colonial ascidians such as Botryllus and Botrylloides, asexual growth by budding
is a continuously occurring process. Each new generation is interconnected to each
other and the parent by a common vasculature, and germ cells are passed on between
generations through this vascular network (Fig. 8.9a and b).
Vasa is expressed in small cells within the blood vessels, as well as in small,
developing oocytes, maturing oocytes and testes both in Botryllus schlosseri and
in another, related species, Botrylloides violaceus (Brown and Swalla 2007 ;
Brown et al. 2009 ). Vasa-positive germ cell precursors are mobile, and can
migrate to a niche in the developing new bodies (buds), then expand and differ-
entiate into gametes (Sunanaga et al. 2006 ; Brown and Swalla 2007 ; Brown
T. Aguero et al.