Abstract
The genetic variability among 13 isolates of Olive mild mosaic virus (OMMV) and of 11 isolates of Tobacco necrosis virus D (TNV-D) recovered from Olea europaea L. samples from various sites in Portugal, was assessed through the analysis of the coat protein (CP) gene sequences. This gene was amplified through reverse transcriptase polymerase chain reaction (RT-PCR), cloned, and 5 clone sequences of each virus isolate, were analysed and compared, including sequences from OMMV and TNV-D isolates originally recovered from different hosts and countries and available in the GenBank, totalling 131 sequences. The encoded CP sequences consisted of 269 amino acids (aa) in OMMV and 268 in TNV-D. Comparison of the CP genomic and amino acid sequences of the isolates showed a very low variability among OMMV isolates, 0.005 and 0.007, respectively, as well as among TNV-D isolates, 0.006 and 0.008. The maximum nucleotide distances of OMMV and TNV-D sequences within isolates were also low, 0.013 and 0.031, respectively, and close to that found between isolates, 0.018 and 0.034, respectively. In some cases, less variability was found in clone sequences between isolates than in clone sequences within isolates, as also shown through phylogenetic analysis. CP aa sequence identities among OMMV and TNV-D isolates ranged from 84.3% to 85.8%. Comparison between the CP genomic sequences of the two viruses, showed a relatively low variability, 0.199, and a maximum nucleotide distance between isolates of 0.411. Analysis of comparative models of OMMV and TNV-D CPs, showed that naturally occurring substitutions in their respective sequences do not seem to cause significant alterations in the virion structure. This is consistent with a high selective pressure to preserve the structure of viral capsid proteins.
Highlights
Olive mild mosaic virus (OMMV) and Tobacco necrosis virus D (TNV-D) originally placed in the Necrovirus genus, were recently divided and included into the new genera Alphanecrovirus and Betanecrovirus, respectively, based on the level of sequence diversity in their polymerases.Complete genomic sequences of TNV-D were obtained from isolates recovered from bean in England [1], tobacco in Hungary [2] and, more recently, from an olive tree in Portugal [3].In 2004, an olive isolate initially identified as a TNV-D isolate based on the sequencing of the coat protein (CP) gene [4] was later considered a distinct species, OMMV [5], following the complete sequencing of its genome
The genetic diversity in OMMV and TNV-D coat protein was evaluated by sequence analysis of the CP of thirteen OMMV isolates and eleven TNV-D isolates, eight of which were obtained from double infected tissues
Additional eight OMMV and three TNV-D CP sequences were retrieved from Genbank and included in this study
Summary
Complete genomic sequences of TNV-D were obtained from isolates recovered from bean in England [1], tobacco in Hungary [2] and, more recently, from an olive tree in Portugal [3]. In 2004, an olive isolate initially identified as a TNV-D isolate based on the sequencing of the coat protein (CP) gene [4] was later considered a distinct species, OMMV [5], following the complete sequencing of its genome. Based on the deduced amino acid (aa) CP sequence, OMMV showed 85.1% identity with that of TNVD [3], which explains its initial diagnosis as a TNV-D isolate. Differentiation between OMMV and TNV-D is only possible through PCR based assays using specific primers [8] or through complete genome sequencing. TNV isolates referred in earlier literature may reveal themselves to be either TNV-D or OMMV
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