No abstract

To investigate the role of individual structural domains in viroid pathogenicity and replication, a series of iuterspeific chimeras was constructed by ex ging the terminal left (TL) and/or Pathogenicity (P) domains between tomato apical stunt (TASVd) and citrus exocortis (CEVd) virolds. AU six chimeras tested were replicated stably in tomato, and the symptoms exhibited by infected plants were intermediate between those induced by the parental viroids. Quantitative comparisons of symptom development and progehy accumulation revealed that: (a) the TL domain of TASVd contains a determinant required for appearance of severe veinal necrosis in tomato, (is) the severe epinasty and stunting chatacteristic of TASVd requires the presence of its TL and P domains, and (id) the variable (V) and terminal right (Til) domains comprising the right side of the native structure also play an important role in virold pathogenicity. Chimeras containing the right side ofTASVd accumulated to higher levels early in infection, and infected plants developed more severe symptoms than those whose right halves were derived from CEVd. Although the Individual contributions of the TL and P domains to symptom induction could not be completely separated from that of virold titer, the TL domain appears to exert a greater effect upon symptom severity than does the P domain.The TL, P. V, and TR domains of TASVd and CEVd contain three discrete regions ofsequence and/or structural variability that may correspond to the pathogenicity determinants uncovered by our genetic analysis.Viroids are the smallest known agents of infectious diseasesmall [246-375 nucleotides (nt)], highly structured, singlestranded RNA molecules that lack a protein capsid and detectable mRNA activity (1, 2). The apparent absence of pathogen-specified proteins implies that the disease process is initiated by the direct interaction of one or more structural determinants with as yet unidentified host cell constituents. Thus, viroid replication and pathogenesis provide an attractive model system in which to investigate the molecular mechanisms controlling host-pathogen interaction as well as gene expression in the uninfected host cell.Based upon sequence similarities between potato spindle tuber viroid (PSTVd) and several other viroid species, Keese and Symons (3) have proposed that viroids contain five structural domains-i.e., a conserved central region (CCR), flanking pathogenicity (P) and variable (V) domains, and left (TL) and right (TR) terminal loops (Fig. 1) virulence is determined by the interaction of nucleotides within a portion of the PSTVd P domain known as the "virulence modulating" region with as yet unidentified host factor(s), but the possible role of structural elements outside the P domain in regulating viroid symptom expression is unclear. Many of the changes in host metabolism associated with viroid (or virus) disease are probably secondary consequences of infection, and symptom expression need not be directly related to replication. Indeed, symptom ...

The Columnea latent viroid (CLV) occurs latently in certain Columnea erythrophae plants grown commercially. In potato and tomato, CLV causes potato spindle tuber viroid (PSTV)-like symptoms. Its nucleotide sequence and proposed secondary structure reveal that CLV consists of a single-stranded circular RNA of 370 nucleotides which can assume a rod-like structure with extensive base-pairing characteristic of all known viroids. The electrophoretic mobility of circular CLV under nondenaturing conditions suggests a potential tertiary structure. CLV contains extensive sequence homologies to the PSTV group of viroids but contains a central conserved region identical to that of hop stunt viroid (HSV). CLV also shares some biological properties with each of the two types of viroids. Most probably, CLV is the result of intracellular RNA recombination between an HSV-type and one or more PSTV-type viroids replicating in the same plant.

49Recent works in plant virus evolution are revealing that genetic structure and behavior of 50 virus and viroid populations can explain important pathogenic properties of these agents, such 51 as host resistance breakdown, disease severity, and host shifting among others. Genetic 52 variation is essential for the survival of organisms. The exploration of how these subcellular 53 parasites generate and maintain a certain frequency of mutations at the intra-and inter-host 54 levels is revealing novel molecular virus-plant interactions. They emphasize the role of host 55 environment in the dynamic genetic composition of virus populations. Functional genomics has 56 identified host factors that are transcriptionally altered after virus infections. The analyses of 57 these data by means of systems biology approaches are uncovering critical plant genes 58 specifically targeted by viruses during host adaptation. Also, a next-generation re-sequencing 59 approach of a whole virus genome is opening new avenues to study virus recombination and 60 the relationships between intra-host virus composition and pathogenesis. Altogether, the 61 analyzed data indicate that systematic disruption of some specific parameters of evolving virus 62 populations could lead to more efficient ways of disease prevention, eradication, or tolerable 63 virus-plant coexistence. 64 65 66 Acosta-Leal, R., Duffy, S., Xiong, Z., Hammond, R. W., and Elena, S. F. 2011. Advances in 67 plant virus evolution: Translating evolutionary insights into better disease management. 68 Phytopathology (accepted). 69 Acosta--Leal et al (2011). Mini--review in Phytopathology 4 70Viruses and viroids appear to be the fastest-evolving plant pathogens (39), and cause 71 tremendous economical crop losses annually. Some, such as the single-stranded DNA 72 begomoviruses, are emergent problems worldwide (117, 126). These subcellular pathogens 73 have higher mutation rates than, and distinct evolutionary dynamics from, bacterial and fungal 74 phytopathogens. Understanding their reproductive and transmission strategies -their biology, 75 ecology, and evolution -can lead to insights and interventions for effective crop disease 76 management. This review highlights how viruses and viroids achieve and maintain their unique 77 parasitic lifestyles and how evolutionary virology and systems biology approaches to virus-plant 78 interactions have implications for pathogen control. 79The ability of viruses and viroids to change, and to change rapidly, underlies many 80 disease management concerns. Excepting migration from distant locations and other countries, 81 variability in plant pathogen populations is the necessary initial step in adaptation to new plants 82 (host shifting), resistance breaking (RB), and changes in symptoms and virulence. Many times 83 the rise and fall of different genotypes in a population is due to the effects of natural selection: 84 variant genomes that generate more viable descendants become more frequent over time. This 85 process can be sped up ...

Infectivity studies with highly infectious RNA inocula generated by ribozyme cleavage were used to compare the biological properties of three apparently nonviable mutants of potato spindle tuber viroid (PSTVd). One of these mutants (PSTVd-P) contains three nucleotide substitutions in the left terminal loop, and mechanical inoculation of tomato seedlings with RNA transcripts at levels equivalent to 10 3 -10 5 times the ID 50 for PSTVd-Intermediate failed to result in systemic infection. Viable progeny containing a spontaneous C G change at position 4 could, however, be recovered from transgenic Nicotiana benthamiana plants that constitutively expressed PSTVd-P RNA. The initial mutations in