Geographical distribution, genetic studies and vector transmission of alfalfa enamovirus-1 infecting alfalfa crop in Argentina.

Alfalfa (Medicago sativa L.) growing areas of Argentina were surveyed between 2010 and 2018 to determine the geographical distribution and analyse the genetic diversity among alfalfa enamovirus-1 (AEV-1) isolates. The virus was detected in all 17 surveyed alfalfa-producing provinces, with a prevalence of 64%. The plant virus AEV-1 is widely distributed in the country, and its transmission vector has been unknown until now. Here we show that the black aphid Aphis craccivora can transmit AEV-1. The CP sequence identity among 16 AEV-1 isolates from Argentina was from 98 to 100% and from 98.9 to 100% at nucleotide and amino acid levels, respectively, indicating a low level of sequence variation among these isolates. The phylogenetic analysis based on the complete nucleotide sequence of the CP gene indicated that AEV-1 isolates are closely related and clustered in a monophyletic group. These results suggest that AEV-1 has spread very recently in Argentina. In the present study, we report the geographical distribution of AEV-1 in the main alfalfa-growing areas of Argentina and, for the first time, identify an insect vector and describe the CP gene diversity of an enamovirus worldwide.

Genome characterization of an Argentinean isolate of alfalfa leaf curl virus.

We investigated the molecular characteristics of an Argentinean isolate of alfalfa leaf curl virus (ALCV-Arg), a virus of the genus Capulavirus in the family Geminiviridae that was isolated from alfalfa plants showing dwarfism. The genome was found to be 2,750 nucleotides in length. In pairwise comparisons, this ALCV isolate shared 83.2% to 92.6% sequence identity with European ALCV isolates. Sequence comparisons and phylogenetic analysis showed that this isolate combines features of strains A and B of ALCV. Recombination analysis showed that ALCV-Arg is a recombinant isolate that was generated by intraspecific recombination between ALCV strains A and B. The results of this study not only show that ALCV-Arg is unique because it combines features of strains A and B but also show that ALCV naturally infects this forage crop on the American continent.

Complete genome sequence of sunflower ring blotch virus, a new potyvirus infecting sunflower in Argentina.

The complete genome sequence of sunflower ring blotch virus (SuRBV), a previously undescribed potyvirus infecting sunflower in Argentina, is reported. The SuRBV genome comprises 9555 nucleotides (nt) and encodes a polyprotein of 3061 amino acids, flanked by 5' and 3' untranslated regions of 117 and 255 nt, respectively. Phylogenetic analysis showed that SuRBV belongs to the potato virus Y (PVY) subgroup and clusters together with sunflower chlorotic mottle virus and bidens mosaic virus. Percentage nucleotide identity between the whole genomes of SuRBV and BiMV was 70.6%, suggesting SuRBV should be considered a distinct species in the genus Potyvirus.

Complete genome sequence of a new enamovirus from Argentina infecting alfalfa plants showing dwarfism symptoms.

Alfalfa dwarf disease, probably caused by synergistic interactions of mixed virus infections, is a major and emergent disease that threatens alfalfa production in Argentina. Deep sequencing of diseased alfalfa plant samples from the central region of Argentina resulted in the identification of a new virus genome resembling enamoviruses in sequence and genome structure. Phylogenetic analysis suggests that it is a new member of the genus Enamovirus, family Luteoviridae. The virus is tentatively named "alfalfa enamovirus 1" (AEV-1). The availability of the AEV-1 genome sequence will make it possible to assess the genetic variability of this virus and to construct an infectious clone to investigate its role in alfalfa dwarfism disease.

Analysis of the coding-complete genomic sequence of groundnut ringspot virus suggests a common ancestor with tomato chlorotic spot virus.

Groundnut ringspot virus (GRSV) and tomato chlorotic spot virus (TCSV) share biological and serological properties, so their identification is carried out by molecular methods. Their genomes consist of three segmented RNAs: L, M and S. The finding of a reassortant between these two viruses may complicate correct virus identification and requires the characterization of the complete genome. Therefore, we present for the first time the complete sequences of all the genes encoded by a GRSV isolate. The high level of sequence similarity between GRSV and TCSV (over 90 % identity) observed in the genes and proteins encoded in the M RNA support previous results indicating that these viruses probably have a common ancestor.

Complete nucleotide sequence of Alfalfa mosaic virus isolated from alfalfa (Medicago sativa L.) in Argentina.

The complete nucleotide sequence of an Alfalfa mosaic virus (AMV) isolate infecting alfalfa (Medicago sativa L.) in Argentina, AMV-Arg, was determined. The virus genome has the typical organization described for AMV, and comprises 3,643, 2,593, and 2,038 nucleotides for RNA1, 2 and 3, respectively. The whole genome sequence and each encoding region were compared with those of other four isolates that have been completely sequenced from China, Italy, Spain and USA. The nucleotide identity percentages ranged from 95.9 to 99.1 % for the three RNAs and from 93.7 to 99 % for the protein 1 (P1), protein 2 (P2), movement protein and coat protein (CP) encoding regions, whereas the amino acid identity percentages of these proteins ranged from 93.4 to 99.5 %, the lowest value corresponding to P2. CP sequences of AMV-Arg were compared with those of other 25 available isolates, and the phylogenetic analysis based on the CP gene was carried out. The highest percentage of nucleotide sequence identity of the CP gene was 98.3 % with a Chinese isolate and 98.6 % at the amino acid level with four isolates, two from Italy, one from Brazil and the remaining one from China. The phylogenetic analysis showed that AMV-Arg is closely related to subgroup I of AMV isolates. To our knowledge, this is the first report of a complete nucleotide sequence of AMV from South America and the first worldwide report of complete nucleotide sequence of AMV isolated from alfalfa as natural host.

Occurrence of Yam Mosaic Virus and Yam Mild Mosaic Virus on Dioscorea spp. Germplasm Collection in Cuba-Epidemiology of Associated Diseases.

Potyvirus diseases are one of the main challenges facing the production of yam (Dioscorea spp.). The objective of this study was to identify the potyviruses present in the Dioscorea spp. germplasm collection at Instituto de Investigaciones de Viandas Tropicales (INIVIT) to establish methodologies for the characterization of the associated diseases. For this purpose, immunochemical and molecular methods were used to identify the potyviruses present. The symptomatology of Dioscorea spp. at INIVIT's germplasm collection was described. In addition, the severity and incidence in the germplasm collection and production areas were evaluated. As a result, the first report of yam mosaic virus (Potyvirus yamtesselati) and yam mild mosaic virus (Potyvirus yamplacidum) in Cuba is presented. The existence of resistant, tolerant, and susceptible cultivars to potyvirus-associated diseases in the germplasm collection was detected, and the incidence of these diseases was higher than 64% in the production areas evaluated. This study represents a step forward in the establishment of certification programs for propagating material of Dioscorea spp. in Cuba.

An Unwanted Association: The Threat to Papaya Crops by a Novel Potexvirus in Northwest Argentina.

An emerging virus isolated from papaya (Carica papaya) crops in northwestern (NW) Argentina was sequenced and characterized using next-generation sequencing. The resulting genome is 6667-nt long and encodes five open reading frames in an arrangement typical of other potexviruses. This virus appears to be a novel member within the genus Potexvirus. Blast analysis of RNA-dependent RNA polymerase (RdRp) and coat protein (CP) genes showed the highest amino acid sequence identity (67% and 71%, respectively) with pitaya virus X. Based on nucleotide sequence similarity and phylogenetic analysis, the name papaya virus X is proposed for this newly characterized potexvirus that was mechanically transmitted to papaya plants causing chlorotic patches and severe mosaic symptoms. Papaya virus X (PapVX) was found only in the NW region of Argentina. This prevalence could be associated with a recent emergence or adaptation of this virus to papaya in NW Argentina.

Phylodynamics of sunflower chlorotic mottle virus, an emerging pathosystem.

Distribution and epidemiological patterns of sunflower chlorotic mottle virus (SCMoV) in sunflower (Helianthus annuus L.) growing areas in Argentina were studied from 2006 to 2017. The virus was detected exclusively in the Pampas region (Entre Ríos, Santa Fe, Córdoba, La Pampa and Buenos Aires provinces). Phylodynamic analyses performed using the coat protein gene of SCMoV isolates from sunflower and weeds dated the most recent common ancestor (MRCA) back to 1887 (HPD95% = 1572-1971), which coincides with the dates of sunflower introduction in Argentina. The MRCA was located in the south of Buenos Aires province and was associated with sunflower host (posterior probability for the ancestral host, ppah = 0.98). The Bayesian phylodynamic analyses revealed the dispersal patterns of SCMoV, suggesting a link between natural host diversity, crop displacement by human activities and virus spread.

Redefining the medicago sativa alphapartitiviruses genome sequences.

In alfalfa samples analyzed by hightroughput sequencing, four de novo assembled contigs encoding gene products showing identities to alphapartitiviruses proteins were found based on BlastX analysis. The predicted amino acid (aa) sequences of two contigs presented 99-100% identity to the RNA-dependent RNA polymerase (RdRp) and the capsid protein (CP) of the recently reported medicago sativa alphapartitivirus 1 (MsAPV1). In addition, the remaining two contigs shared only 56% (CP) and 70% (RdRp) pairwise aa identity with the proteins of MsAPV1, suggesting that these samples presented also a novel Alphapartitivirus species. Further analyses based on complete genome segments termini and the presence/absence of alphapartitivirus RNA in several samples and public alfalfa RNA datasets corroborated the identification of two different alphapartitivirus members. Our results likely indicate that the reported MsAPV1 genome was previously reconstructed with genome segments of two different alphapartitiviruses. Overall, we not only revisited the MsAPV1 genome sequence but also report a new tentative alphapartitivirus species, which we propose the name medicago sativa alphapartitivirus 2. In addition, the RT-PCR detection of both MsAPV1 and MsAPV2 in several alfalfa cultivars suggests a broad distribution of both viruses.

The Westward Journey of Alfalfa Leaf Curl Virus.

Alfalfa leaf curl virus (ALCV), which causes severe disease symptoms in alfalfa (Medicago sativa L.) and is transmitted by the widespread aphid species, Aphis craccivora Koch, has been found throughout the Mediterranean basin as well as in Iran and Argentina. Here we reconstruct the evolutionary history of ALCV and attempt to determine whether the recent discovery and widespread detection of ALCV is attributable either to past diagnostic biases or to the emergence and global spread of the virus over the past few years. One hundred and twenty ALCV complete genome sequences recovered from ten countries were analyzed and four ALCV genotypes (ALCV-A, ALCV-B, ALCV-C, and ALCV-D) were clearly distinguished. We further confirm that ALCV isolates are highly recombinogenic and that recombination has been a major determinant in the origins of the various genotypes. Collectively, the sequence data support the hypothesis that, of all the analyzed locations, ALCV likely emerged and diversified in the Middle East before spreading to the western Mediterranean basin and Argentina.

Complete genome sequence and integrated protein localization and interaction map for alfalfa dwarf virus, which combines properties of both cytoplasmic and nuclear plant rhabdoviruses.

We have determined the full-length 14,491-nucleotide genome sequence of a new plant rhabdovirus, alfalfa dwarf virus (ADV). Seven open reading frames (ORFs) were identified in the antigenomic orientation of the negative-sense, single-stranded viral RNA, in the order 3'-N-P-P3-M-G-P6-L-5'. The ORFs are separated by conserved intergenic regions and the genome coding region is flanked by complementary 3' leader and 5' trailer sequences. Phylogenetic analysis of the nucleoprotein amino acid sequence indicated that this alfalfa-infecting rhabdovirus is related to viruses in the genus Cytorhabdovirus. When transiently expressed as GFP fusions in Nicotiana benthamiana leaves, most ADV proteins accumulated in the cell periphery, but unexpectedly P protein was localized exclusively in the nucleus. ADV P protein was shown to have a homotypic, and heterotypic nuclear interactions with N, P3 and M proteins by bimolecular fluorescence complementation. ADV appears unique in that it combines properties of both cytoplasmic and nuclear plant rhabdoviruses.