Viroids are small, nontranslatable pathogenic RNAs that replicate autonomously and traffic systemically in their host plants. We have used in situ hybridization to analyze the trafficking pattern of Potato spindle tuber viroid (PSTVd) in tomato and Nicotiana benthamiana. When PSTVd was inoculated onto the stem of a plant, it replicated and trafficked to sink, but not source, leaves. PSTVd was absent from shoot apical meristems. In the flowers of infected plants, PSTVd was present in the sepals, but was absent in the petals, stamens, and ovary. The replicative form of PSTVd was detected in the phloem. Our data demonstrate that (i) PSTVd traffics long distance in the phloem and this trafficking is likely sustained by replication of the viroid in the phloem, and (ii) PSTVd trafficking is governed by plant developmental and cellular factors. The dependency of PSTVd and other viroids on cellular mechanisms for RNA trafficking makes them excellent tools to study such mechanisms.