Fig. 4: Change in the ploidy levels of the nucleus, mitochondrion, nucleomorph, and chloroplast of Chroomonas sp. after ingestion by N. aeruginosum.

a Schematic diagrams showing the coculture of N. aeruginosum and Chroomonas sp. Dc01 used to investigate the change in the ploidy level of Chroomonas sp. Dc01 ingested by N. aeruginosum in (b–e). Chroomonas sp. Dc01 was removed from the coculture for N. aeruginosum cells to digest most of the kleptoplasts derived from Chroomonas sp. (~95 and 60% of N. aeruginosum cells had completely digested the nucleus and kleptoplast derived from Chroomonas sp., respectively). Then, the starved N. aeruginosum cells were again fed Chroomonas sp. until approximately 75% of the cells had ingested Chroomonas sp. After this, free-living Chroomonas sp. was removed from the coculture (day 0) and N. aeruginosum cells with kleptoplasts were cultured for 14 days under continuous light (10 µmol photons m⁻² s⁻¹). b Micrographs showing the change in the nucleus and kleptoplast derived from Chroomonas sp. in N. aeruginosum cells. Cells were stained with SYBR Green. Images of differential interference contrast (DIC), SYBR Green staining, and kleptoplast red fluorescence (Chl) are shown. The arrowhead indicates a nucleus derived from Chroomonas sp. The cell in the image from day 3 ingested two nuclei. Scale bar = 10 μm. c Change in N. aeruginosum cell density. d Change in the ratio of N. aeruginosum cells retaining the nucleus derived from Chroomonas sp. and kleptoplast fluorescence intensity per N. aeruginosum cell. For kleptoplast fluorescence, the average intensity at day 0 was defined as “1.0.” The kleptoplast fluorescence intensity was quantified both in N. aeruginosum cells with and without the nucleus derived from Chroomonas sp. The error bars represent standard deviation (n = 30 cells for each time point). e Change in SYBR Green fluorescence intensity per Chroomonas sp. nucleus ingested by N. aeruginosum. The average at day 0 was defined as “1.0.” The error bars represent standard deviation (n = 30 nuclei). f Schematic diagrams showing the coculture of N. aeruginosum and Chroomonas sp. used to investigate the change in the ploidy level of Chroomonas sp. Dc01 ingested by N. aeruginosum in (g, h). To avoid qPCR amplification of residual DNA from Chroomonas sp. Dc01 after starvation of N. aeruginosum, Chroomonas sp. HrL01 instead of Dc01 was fed to N. aeruginosum before starvation and then Dc01 was added to the culture. Others are the same as in (a). g Micrographs showing that described in (b) except that they show the cells from (f) instead of (a). h qPCR showing changes in ploidy of nuclear (rpl8 gene), mitochondrial (cox1 gene), nucleomorph (gidA gene), and kleptoplast (rpoB gene) genomes of Chroomonas sp. Dc01 ingested by N. aeruginosum. The values were normalized with values of N. aeruginosum 18S rDNA. The average at day 0 was defined as “1.0” for respective genomes. The error bars represent standard deviation (three independent cultures).