Localization of ribosomal RNA and Pbs21-mRNA in the sexual stages of Plasmodium berghei using electron microscope in situ hybridization.

A reproducible technique for the ultrastructural localization of RNAs in malaria parasites has been developed which combines excellent structural preservation with high hybridization signals. Signals obtained following in situ hybridization with an antisense rRNA probe which recognizes all forms of small subunit (SSU) rRNA correlate with the density of ribosomes in the parasite cytoplasm and show that a) the male gametocyte has only 12 to 25% the ribosomes found in the female cell and asexual parasite and b) the probe did not hybridize with an electron-dense nuclear body previously called a nucleolus. We suggest this structure is either a transcription-, or a replication-factory. Using a probe for the sexual stage-specific protein Pbs21 mRNA, signal was found only in female gametocytes, zygotes and ookinetes and showed a non-random, clumped cytoplasmic distribution. It is not known at present whether the non-random distribution of the Pbs 21 mRNA is critical to the very delayed translation of the Pbs21 message into protein, which occurs only in the zygote and ookinete.