FIG 3.

Immunolocalization and developmental expression of N. bombycis SWP9 and SWP7. (A) Immunoelectron microscopy localization and developmental expression of SWP9 and SWP7. (A1 and A3) Images of 30% and 45% Percoll gradient centrifugation fractions showing strong enrichment of sporonts with a visible cell coat and internal colloidal gold particles (18 nm) of SWP9 and SWP7 (white arrowheads). (A2 and A4) The 60% and 75% Percoll gradient centrifugation fractions are rich in sporoblasts, with internal and surface coat localization of SWP9 and SWP7. The sporoblast-containing portions have a scattered developing polar tube and a thick surface coat (large black arrowheads). A portion of the SWP9 gold particles are secreted and are present in the thickened surface coat (red arrowheads). The cytoplasm still has some diffuse gold particles of SWP9 in it (white arrowheads). However, the vast majority of the SWP7 colloidal gold particles are distributed in the cytoplasm. (B1 and B2) The 90% Percoll gradient centrifugation fraction shows nearly mature N. bombycis spores with localization of the exospore (black arrows), endospore (red arrowheads), and polar tube (white arrows). The surface of the mature N. bombycis spore plasmalemma is enclosed by a relatively thick electron-lucent endospore that is in turn covered by an electron-dense exospore coat. Colloidal gold particles of SWP9 and SWP7 are primarily dispersed in the exospore (black arrows), endospore (red arrowheads), and polar tube (white arrows) of the mature N. bombycis spores. (B3) Note the absence of gold particles in the control sections of the mature N. bombycis spore (using negative-control rabbit serum). Sp, sporonts; Sb, sporoblasts; S, mature spores; PT, polar tube; N, nucleus; Ex, exospores; En, endospore. (B) SWP9 and SWP7 of N. bombycis also localized at the polar tubes of germinated spores. N. bombycis spore germination was induced in 0.1 mol K₂CO₃ for 30 min at 28°C and then incubation with mouse anti-PTP1 (A2 and B2) and rabbit anti-SWP7 (A3) and anti-SWP9 (B3) antibodies. DAPI was used to stain the nucleus (A1 and B1). (A4 and B4) Merged images.