On multiple separate occasions, ancestral male/female nematode species have convergently evolved self-fertile hermaphroditism. In C. elegans, hermaphrodites achieve self-fertility by producing sperm as larval L4s, storing their sperm, and then making a one-time switch to oogenesis. Here we report that hermaphrodites of the same family but in the Rhabditis clade, specifically Rhabditis sp. SB347, JU1782, and JU1783, achieve self-fertility by continuously producing both sperm and oocytes from L4 and throughout the adult lifespan.
In the hermaphrodite germlines of these male/female/hermaphrodite Rhabditis species, we find distinct clusters of cells alongside the developing oocytes. In contrast, these cell clusters are not present in females, which implies their sperm-related function. The number of cells per cluster increases in a distal to proximal fashion most often in a pattern of 1, 2, 4, or 8 cells. In addition, many clusters contain dividing cells with tubulin spindles, suggesting that they proliferate along the germline. Immunocytochemistry shows that the expression of major sperm protein (MSP) is found only in the late clusters (often in the vicinity of the -1 to -3 oocyte). Our analysis suggests that the early cell clusters are mitotic progenitors of meiotic spermatocytes in the late clusters. The mitotic, undifferentiated character of the progenitor cells leads us to conclude that they are analogous to the recently clarified transit amplifying (TA) cells in the C. elegans mitotic region. However, unlike in C. elegans, the presumed TA cells in Rhabditis hermaphrodites exist in isolated clusters, physically distant from both the stem cell niche and other cell clusters. Such mitotic cluster formation presents a unique opportunity to study TA cell development. Ongoing studies of this divergent mechanism of spermatogenesis are characterizing both protein and RNA expression of conserved regulators of the cell cycle and the germ cell differentiation. By understanding the cellular and genetic pathways in non-C. elegans nematode species, we hope to elucidate germ cell development and differentiation in a novel evolutionary context.