Fig. 4
High-resolution immunofluorescent identification of Plasmodium liver stage (LS) parasites. a High-resolution images of P. vivax hypnozoites demonstrate these forms have minimal nuclear material and are negative for schizogony markers EXP1, EXP 2 and MSP1. Hypnozoites stain positive for cytosolic markers MIF, HSP70, and GAPDH and reveal a functioning apicoplast. b By day 8, P. vivax schizonts are several times larger than the host cell hepatic nucleus, feature genome replication and segmentation, and stain positive for EXP1, EXP2, ACP, and MSP1. c The LS of P. falciparum is shorter than that of P. vivax schizonts; developing parasites are correspondingly less small. By day 5 merozoite segmentation has begun (as noted by ACP staining of separate apicoplasts) but not complete (as noted by diffuse staining of MSPs). d Immunofluorescent staining of day 8 P. vivax LS schizonts with anti-Pvs16, a sexual stage-specific biomarker for immature gametocytes, showed co-localization with developing LS merozoites indicated by segmented DNA. However, anti-Pvs16 signal does not appear in every LS. e The PHH system successfully supports complete maturation of Plasmodium LS schizonts measured by breakthrough into blood stage using reticulocyte (P. vivax, days 9–11) or RBC (P. falciparum, days 7–8) overlays with initial giemsa staining every 6 h. P. vivax overlays show formation of merozoite packages which rupture into the reticulocyte culture leading to invasion. Alternatively, no merosomes were captured in P. falciparum overlays but early rings were present within the first 12 h and continued culture progressed to an asynchronous population at > 1% parasitemia. White scale bars represent 5 µm, gray scale bars represent 10 µm
