Nucleotide sequence and genomic organization of an ophiovirus associated with lettuce big-vein disease

The complete nucleotide sequence of an ophiovirus associated with lettuce big-vein disease has been elucidated. The genome consisted of four RNA molecules of approximately 7ò8, 1ò7, 1ò5 and 1ò4 kb. Virus particles were shown to contain nearly equimolar amounts of RNA molecules of both polarities. The 5'- and 3'-terminal ends of the RNA molecules are largely, but not perfectly, complementary to each other. The virus genome contains seven open reading frames. Database searches with the putative viral products revealed homologies with the RNA-dependent RNA polymerases of rhabdoviruses and Ranunculus white mottle virus, and the capsid protein of Citrus psorosis virus. The gene encoding the viral polymerase appears to be located on the RNA segment 1, while the nucleocapsid protein is encoded by the RNA3. No significant sequence similarities were observed with other viral proteins. In spite of the morphological resemblance with species in the genus Tenuivirus, the ophioviruses appear not to be evolutionary closely related to this genus nor any other viral genus

The control of PVY in Dutch seed potato culture

Over the recent years Potato virus Y presents a growing problem in Dutch seed potato culture. In recent years a significant % of seed potato lots was de-classified due to PVY infections. This apparent increase in PVY infections was unexpected since no increase in field symptoms were observed and the numbers of aphids caught in the yellow water traps and high suction traps showed a clear decline over the last 10 years http://www.aab.org.uk/images/VIRO_CONF_PROG.pd

Tomato marchitez virus, a new plant picorna-like virus from tomato related to tomato torrado virus

A new virus was isolated from a tomato plant from the state of Sinaloa in Mexico. This plant showed symptoms locally known as `marchitez disease¿: severe leaf necrosis, beginning at the base of the leaflets, and necrotic rings on the fruits. A virus was isolated from the infected plant consisting of isometric particles with a diameter of approximately 28¿nm. The viral genome consists of two (+)ssRNA molecules of 7221 (RNA1) and 4898¿nts (RNA2). The viral capsid contains three coat proteins of 35, 26 and 24¿kDa, respectively. The abovementioned characteristics: symptoms, morphology, number and size of coat proteins, and number of RNAs are similar to those of the previously described tomato torrado virus (ToTV). Sequence analysis of the entire viral genome shows that this new virus is related to, but distinct from, ToTV and that these members of two obviously new virus species belong to the recently proposed plant virus genus Torradovirus. For this new virus, the name tomato marchitez virus (ToMarV) is proposed

Sequence analysis and genomic organization of Aphid lethal paralysis virus: a new member of the family Dicistroviridae

The complete nucleotide sequence of the genomic RNA of an aphid-infecting virus, Aphid lethal paralysis virus (ALPV), has been determined. The genome is 9812 nt in length and contains two long open reading frames (ORFs), which are separated by an intergenic region of 163 nt. The first ORF (5' ORF) is preceded by an untranslated leader sequence of 506 nt, while an untranslated region of 571 nt follows the second ORF (3' ORF). The deduced amino acid sequences of the 5' ORF and 3' ORF products respectively showed similarity to the non-structural and structural proteins of members of the newly recognized genus Cripavirus (family Dicistroviridae). On the basis of the observed sequence similarities and identical genome organization, it is proposed that ALPV belongs to this genus. Phylogenetic analysis showed that ALPV is most closely related to Rhopalosiphum padi virus, and groups in a cluster with Drosophila C virus and Cricket paralysis virus, while the other members of this genus are more distantly related. Infectivity experiments showed that ALPV can not only infect aphid species but is also able to infect the whitefly Trialeurodes vaporariorum, extending its host range to another family of the order Hemipter

Het genus Torradovirus, een nieuw geslacht van plantenvirussen

Het genus Torradovirus is een nieuw geslacht van plantenvirussen. Hiertoe behoren het tomatentorradovirus (ToTV), het tomatenmarchitezvirus (ToMarV) en het tomatenchocolàtevirus (ToChV)

Onderzoek naar mogelijke TBV-reservoirs in onkruid

In tulpen veroorzaakt het potyvirus tulpenmoza¿ekvirus (TBV) van alle virussen de meeste directe schade (visuele virussymptomen en opbrengstverlies) en indirecte schade (keuringsmaatregelen, beheersingsmaatregelen, etc.). Dit virus wordt door bladluizen verspreid. Om een virusvrije teelt optimaal mogelijk te maken is kennis over het beperken van virusverspreiding binnen een partij / veld van eerste belang (dit blijkt uit onderzoek van laatste decennia). Dit onderzoeksproject geeft aandacht aan een ontbrekend puzzelstukje, kennis over de potentiële gevaren die vanuit de omliggende groene ruimte / natuur de teelt van tulp schade aan kunnen brengen. In dit project is onderzoek gedaan naar aanwezigheid van TBV in bladluisvluchten en onkruiden welke verzameld zijn in teeltregio’s van tulp waar in het verleden aanwijzingen/suggesties waren voor onverklaarbare toename van TBV in partijen tulp. In dit project zijn in 2012 op twee locaties gedetailleerde bladluisvangsten uitgevoerd en op 9 locaties zijn onkruidmonsters verzameld. Daarnaast is een collectie bladluizen gevangen in 2011 in de omgeving van Lelystad voor dit onderzoek gebruikt. De gevangen bladluizen en de onkruidmonsters zijn onderzocht op aanwezigheid van TBV. Dit project heeft tot de volgende conclusies geleid: • Pas bij de echte temperatuurstijging in de tweede helft van mei 2012 kwam een grote bladluizenvlucht op gang. Per dag werden er met één fuik tientallen tot zelfs een paar honderd bladluizen gevangen. • Er is aangetoond dat er grote verschillen in omvang van bladluizenvluchten kunnen zijn. Stel dat op beide locaties een vergelijkbare partij met 1% TBV werd geteeld, dan was het risico op virusverspreiding door bladluizen vanwege verschil in aantallen bladluizen op locatie B 2,5x zo groot als op locatie A. • Tientallen bladluizen gevangen in de omgeving van Lelystad, 290 bladluizen gevangen in Dirkshorn en ruim 700 bladluizen gevangen in Aartswoud, zijn met PCR-diagnostiek onderzocht op aanwezigheid van TBV. Met behulp van de toegepaste onderzoeksmethoden zijn er geen aanwijzingen gevonden dat gevangen bladluizen TBV bij zich hadden. • Analyse op aanwezigheid van TBV in onkruiden op percelen die in het verleden in verband zijn gebracht met onverwachte toename aan TBV heeft niet geresulteerd in de identificatie van TBV-geïnfecteerde onkruiden. Het feit dat er geen TBV in de gevangen bladluizen is aangetoond, en dat er geen TBV in onkruidmonsters is aangetoond, doet vermoeden dat er vooralsnog geen infectiebron is voor TBV anders dan tulp. Voor de tulpensector is dit een hoopgevend resultaat

A new virus infecting Myzus persicae has a genome organization similar to the species of the genus Densovirus

The genomic sequence of a new icosahedral DNA virus infecting Myzus persicae has been determined. Analysis of 5499 nt of the viral genome revealed five open reading frames (ORFs) evenly distributed in the 5' half of both DNA strands. Three ORFs (ORF1-3) share the same strand, while two other ORFs (ORF4 and ORF5) are detected in the complementary sequence. The overall genomic organization is similar to that of species from the genus Densovirus. ORFs 1-3 most likely encode the non-structural proteins, since their putative products contain conserved replication motifs, NTP-binding domains and helicase domains similar to those found in the NS-1 protein of parvoviruses. The deduced amino acid sequences from ORFs 4 and 5 show sequence similarities with the structural proteins of the members of the genus Densovirus. These data indicate that this virus is a new species of the genus Densovirus in the family Parvoviridae. The virus was tentatively named Myzus persicae densovirus. The nucleotide sequence reported in this study appears in the EMBL, GenBank and DDBJ nucleotide sequence databases under accession number AY148187

Detectie van plantenpathogenen en –aantasters

Nederland speelt internationaal een belangrijke rol in productie en handel van plantaardig materiaal. Hierin zijn vaak ongewenste veroorzakers van quarantaine- (Q) en kwaliteitsziekten (K) en -plagen een storende factor. Om de (fytosanitaire) kwaliteit van verhandeld (met name ook geëxporteerd) plantmateriaal ook in de toekomst te kunnen blijven garanderen, en eigen teelten te vrijwaren van ongewenste organismen is het van groot belang vroegtijdig deze plantenpathogenen te kunnen detecteren en te identificeren. Hiervoor zijn gevalideerde onderzoekmethoden noodzakelijk, die geschikt zijn voor routinematig gebruik door het bedrijfsleven, de keuringsdiensten en de NVWA. De hier voorgestelde intensieve samenwerking tussen kennisinstellingen, overheid en keuringslaboratoria is noodzakelijk om de agrarische sector toekomstbestendig te houden. Daarnaast dient naast aandacht voor nieuwe ontwikkelingen ook aandacht besteedt te worden aan kwaliteitsborging en efficiëntie verbetering van bestaande technieken zodat de taken van de keuringsdiensten beter uitgevoerd kunnen worden

First Report of Tomato torrado virus Infecting Tomato in Colombia

Tomato (Solanum lycopersicum L.) plants grown in plastic greenhouses near Villa de Leyva, northeast of Bogota, Colombia showed necrotic spots on the leaves in September 2008. Initial symptoms were necrosis beginning at the base of leaflets that were surrounded by yellow areas. These symptoms resembled those described for Tomato torrado virus (ToTV; family Secoviridae, genus Torradovirus), which was first found in Spain (2). Other (tentative) members of the genus Torradovirus, Tomato marchitez virus (ToMarV), Tomato chocolate spot virus (ToChSV), and Tomato chocolàte virus (ToChV) (3) induce similar symptoms on tomato plants. One sample, coded T418, was stored in the freezer and brought to our lab in 2011. Serological tests (double-antibody sandwich-ELISA) using polyclonal antibodies (Prime Diagnostics, Wageningen, The Netherlands) on leaf extracts showed the absence of Pepino mosaic virus (PepMV), Tobacco mosaic virus (TMV), Tomato spotted wilt virus (TSWV), Cucumber mosaic virus (CMV), Potato virus X (PVX), and Potato virus Y (PVY). Leaf extracts were mechanically inoculated onto the indicator plants Physalis floridana, Nicotiana hesperis ‘67A’, and N. occidentalis ‘P1’ (six plants in total) and were kept in a greenhouse at 20°C with 16 h of light. Necrotic symptoms appeared 4 to 5 days postinoculation and resembled those described for ToTV (2). Two dip preparations of systemically infected P. floridana and N. occidentalis leaves were examined by electron microscopy, which revealed the presence of spherical virus particles of approximately 30 nm. To confirm the presence of ToTV, total RNA was extracted from the original leaf material and an inoculated P. floridana and N. occidentalis plant using the Qiagen Plant Mini Kit (Qiagen, Hilden, Germany) following manufacturer's instructions. ToTV-specific primer sets ToTV-Dp33F/ToTV-Dp20R (5'-TGCTCAATGTTGGAAACCCC-3'/5'-AGCCCTTCATAGGCTAGCC-3', amplifying a fragment of the RNA1 polyprotein with an expected size of 751 bp) and ToTV-Dp1F/ToTV-Dp2R (5'-ACAAGAGGAGCTTGACGAGG-3'/5'-AAAGGTAGTGTAATGGTCGG-3', amplifying a fragment on the RNA2 movement protein region with an expected size of 568 bp) were used to amplify the indicated regions in a reverse transcription (RT)-PCR using the One-Step Access RT-PCR system (Promega, Madison, WI). Amplicons of the predicted size were obtained in all tested materials. The PCR products were purified with the Qiaquick PCR Purification Kit (Qiagen) and sequenced directly. BLAST analyses of the obtained sequences (GenBank Accession Nos. JQ314230 and JQ314229) confirmed the identity of isolate T418 as ToTV, with 99% identity to isolate PRI-ToTV0301 in both fragments (GenBank Accession Nos. DQ388879 and DQ388880 for RNA1 and RNA 2, respectively). To our knowledge, this is the first report of ToTV in Colombia, and interestingly, since ToTV has been found only in Europe and Australia (1) so far, this is the first report of ToTV on the American continen