Characterization of Cell Division in the Tissues of the Calanoid Copepod, Neocalanus flemingeri from Diapause through Early Oogenesis

Unlike most calanoid copepods, females of the diapausing copepod, Neocalanus flemingeri (Miller, 1988) fuel oogenesis entirely through stored energy. Due to the reliance on stored energy, N. flemingeri females manage their internal lipid stores to maximize oocyte production which differs from the reproductive program of most calanoids that couple food availability with oogenesis. In this study, both lipid content and cell division within the reproductive structures in females were examined as diapause was terminated and oogenesis began. In June and September 2019, diapausing females were collected from depth in Prince William Sound, Alaska. Incubation experiments in 5-Ethynyl-2´-deoxyuridine (EdU) were conducted to quantify and pinpoint the location of cell division within the body from diapause through early oogenesis. Imaging of EdU-treated females using confocal microscopy revealed evidence of cell division in the ovary within 24 hours after collection. Both oogonia and oocytes incorporated EdU based on the location of cells in the posterior end of the ovary. Dividing cells in the ovary peaked in number at 72 hours, remained high over two weeks, and decreased thereafter with no staining detected at four weeks after collection. Thus, the production of new oocytes stopped two to four weeks before females release their first clutch of eggs. The pattern of cell division in the ovary parallels the up- and down-regulation of early germline development genes reported in an earlier transcriptomics study. These results suggest that oogenesis is sequential in N. flemingeri which synchronizes egg maturation, unlike other calanoid copepods where most oocyte stages are observed concurrently within the ovary. The magnitude of cell division in the ovary of individual females were compared with their respective total lipid contents and prosome lengths. Numbers of dividing cells in the ovaries were positively correlated with both prosome length and lipid content, suggesting that total fecundity is higher in copepods with longer prosome lengths and more lipid. In this study, duration of the period of active cell division appeared to be similar in all females independent of prosome size or lipid content. Understanding the internal physiological process of reproduction in lipid-rich copepods like N. flemingeri is an important step in knowing how and to what magnitude egg production can be affected by climate change. This is the first study that tracked cell division in post-diapause N. flemingeri. This capital breeder meters its energy sources by varying the number of dividing cells and limiting cell division in the ovary to the first three to four weeks post-diapause. As waters continue to warm, predictions of shorter diapause lengths, and both smaller lipid reserves and prosome lengths have been hypothesized. Negative impacts on egg production in this species could lead to a decrease in population numbers and thus a decrease in a vital food source for many birds and fishes