The experimental verification of viroid-host adaptation

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All pathogen has host specificity. Viroid, the smallest known pathogen, is an excellent model to analyze which part of the RNA molecule is responsible for host specificity. Hop stunt viroid (HSVd) spreads widely among various crops, and the unique sequence variations are found depending on the host species. To verify experimentally how unique variants has adapted to the varieties of hosts, (1) we have constructed random HSVd-variants consisting of various mutations found in the nature, inoculated them to several hosts, and analyzed how they adapt to each hosts, and (2) we have infected hops with natural HSVd isolates, cultivated them for 10 years, and examined the progeny sequences to analyze how each isolates adapted to hop.The first, using random cDNA shuffling, we constructed random HSVd-variants. They were inoculated to cucumber, grapevine, peach, citrus, hop; the host for HSVd, and A. thaliana; non-host for HSVd. HSVd progeny accumulated in cucumber, grapevine, peach, citrus and hop, but no in A. thaliana. The nucleotide sequence analysis of progeny propagated in each species revealed that only HSVd-grapevine type was adaptable for grapevine but several types of chimeras between HSVd-grapevine and -citrus, as well as HSVd-grapevine type, were dominant in the other hosts. The variations characteristic for HSVd-plum were detected in hop and peach in the early stage of infection, but most of them disappeared in the later stage.The second, hops were infected with four natural HSVd-isolates (HSVd-grapevine, -hop, -plum and -citrus) and cultivated for 10 years. All isolates were extracted every autumn for sequencing to analyze the rate of sequence variation. The rate of variation after the 10 years of persistent infection was the highest in HSVd-citrus and -plum, but significantly lower in HSVd-hop and -grapevine. The result indicated that the fitness of the four isolates to hop was different.