The Diversity of Torque Teno Viruses: In Vitro Replication Leads to the Formation of Additional Replication-Competent Subviral Molecules

The family Anelloviridae comprises torque teno viruses (TTVs) diverse in genome structure and organization. The isolation of a large number of TTV genomes (TTV Heidelberg [TTV-HD]) of 26 TTV types is reported. Several isolates from the same type indicate sequence variation within open reading frame 1 (ORF1), resulting in considerably modified open reading frames. We demonstrate in vitro replication of 12 full-length genomes of TTV-HD in 293TT cells. Propagation of virus was achieved by several rounds of infections using supernatant and frozen whole cells of initially infected cells. Replication of virus was measured by PCR amplification and transcription analyses. Subgenomic molecules (mu TTV), arising early during propagation and ranging in size from 401 to 913 bases, were cloned and characterized. Propagation of these mu TTV in in vitro cultures was demonstrated in the absence of full-length genomes.