scholarly journals First Report of Celery mosaic virus Infecting Celery in Venezuela

Plant Disease ◽  
2006 ◽  
Vol 90 (8) ◽  
pp. 1111-1111 ◽  
Author(s):  
T. Fernández ◽  
O. Carballo ◽  
K. Zambrano ◽  
M. Romano ◽  
E. Marys
Keyword(s):  
Mosaic Virus ◽  
Microscopic Analysis ◽  
Apium Graveolens ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Leaf Extracts ◽  
Electron Microscopic ◽  
First Report ◽  
Three Samples ◽  
Topo Vector

Celery mosaic virus (CeMV) is a significant pathogen of celery (Apium graveolens) worldwide (1). In 2005, in a produce market located in Los Salias, Miranda, celery plants with mottling and leaf malformation were noticed. Electron microscopic analysis of leaf-dip preparations from three symptomatic samples revealed flexuous viral particles that were 750 nm long. Infected cells contained pinwheel inclusions typical of those associated with potyvirus infection. Inoculation of healthy celery plants with leaf extracts from four symptomatic plants produced symptoms identical to those first observed. A survey of five produce markets in Miranda was conducted to determine the prevalence of virus infection in celery using serological and molecular analyses. Mottling and malformation of celery leaflets were observed in all the markets visited. Symptoms were noted in all five markets in each of three visits during a 3-month period. A total of 125 postharvested symptomatic plants were collected from five markets on March 29, 2005 and tested for CeMV using double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with antiserum provided by F. Rabenstein, BAX, Aschersleben, Germany. Of the 125 samples collected during the survey, 53% were ELISA positive. Twenty ELISA-positive samples were also tested using reverse transcription-polymerase chain reaction (RT-PCR) with general primers for the family Potyviridae (2). All 20 samples produced an amplicon of the expected size (1.7 kbp) after RT-PCR. Amplicons from three samples were cloned into the pCR-TOPO vector (Invitrogen, Carlsbad, CA). Sequence analysis of one clone revealed more than 98% nt to a CeMV isolate from Australia (GenBank Accession No AF203532). To our knowledge, this is the first report of CeMV in Venezuela. Our results suggest that the disease may be widely spread on celery crops growing in the areas surrounding produce markets in Miranda State. References: (1) A. Brunt et al. Viruses of Plants. CAB International, Wallingford, Oxon, UK. 1996. (2) J. Chen et al. Arch. Virol. 146:757, 2001.

scholarly journals First Report of Tomato Brown Rugose Fruit Virus Infecting Tomato Crop in Saudi Arabia

Plant Disease ◽  
2021 ◽  
Author(s):  
Ahmed Sabra ◽  
Mohammed Ali Al Saleh ◽  
I. M. Alshahwan ◽  
Mahmoud A. Amer
Keyword(s):  
Saudi Arabia ◽  
Mosaic Virus ◽  
Solanum Lycopersicum ◽  
Tomato Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
First Report ◽  
Pcr Products ◽  
Dark Green ◽  
Leaf Symptoms

Tomato (Solanum lycopersicum L.) is the most economically important member of family Solanaceae and cultivated worldwide and one of the most important crops in Saudi Arabia. The aim of this study is screening of the most common viruses in Riyadh region and identified the presence of tomato brown rugose fruit virus (ToBRFV) in Saudi Arabia. In January 2021, unusual fruit and leaf symptoms were observed in several greenhouses cultivating tomatoes commercially in Riyadh Region, Saudi Arabia. Fruit symptoms showed irregular brown spots, deformation, and yellowing spots which render the fruits non-marketable, while the leaf symptoms included mottling, mosaic with dark green wrinkled and narrowing. These plants presented the symptoms similar to those described in other studies (Salem et al., 2015, Luria et al., 2017). A total 45 Symptomatic leaf samples were collected and tested serologically against suspected important tomato viruses including: tomato chlorosis virus, tomato spotted wilt virus, tomato yellow leaf curl virus, tomato chlorotic spot virus, tomato aspermy virus, tomato bushy stunt virus, tomato black ring virus, tomato ringspot virus, tomato mosaic virus, pepino mosaic virus and ToBRFV using Enzyme linked immunosorbent assay (ELISA) test (LOEWE®, Biochemica, Germany), according to the manufacturers' instructions. The obtained results showed that 84.4% (38/45) of symptomatic tomato samples were infected with at least one of the detected viruses. The obtained results showed that 55.5% (25/45) of symptomatic tomato samples were found positive to ToBRFV, three out of 25 samples (12%) were singly infected, however 22 out of 45 (48.8%) had mixed infection between ToBRFV and with at least one of tested viruses. A sample with a single infection of ToBRFV was mechanically inoculated into different host range including: Chenopodium amaranticolor, C. quinoa, C. album, C. glaucum, Nicotiana glutinosa, N. benthamiana, N. tabacum, N. occidentalis, Gomphrena globosa, Datura stramonium, Solanum lycopersicum, S. nigrum, petunia hybrida and symptoms were observed weekly and the systemic presence of the ToBRFV was confirmed by RT-PCR and partial nucleotide sequence. A Total RNA was extracted from DAS-ELISA positive samples using Thermo Scientific GeneJET Plant RNA Purification Mini Kit. Reverse transcription-Polymerase chain reaction (RT-PCR) was carried out using specific primers F-3666 (5´-ATGGTACGAACGGCGGCAG-3´) and R-4718 (5´-CAATCCTTGATGTG TTTAGCAC-3´) which amplified a fragment of 1052 bp of Open Reading Frame (ORF) encoding the RNA-dependent RNA polymerase (RdRp). (Luria et al. 2017). RT-PCR products were analyzed using 1.5 % agarose gel electrophoresis. RT-PCR products were sequenced in both directions by Macrogen Inc. Seoul, South Korea. Partial nucleotide sequences obtained from selected samples were submitted to GenBank and assigned the following accession numbers: MZ130501, MZ130502, and MZ130503. BLAST analysis of Saudi isolates of ToBRFV showed that the sequence shared nucleotide identities ranged between 98.99 % to 99.50 % among them and 98.87-99.87 % identity with ToBRFV isolates from Palestine (MK881101 and MN013187), Turkey (MK888980, MT118666, MN065184, and MT107885), United Kingdom (MN182533), Egypt (MN882030 and MN882031), Jordan (KT383474), USA (MT002973), Mexico (MK273183 and MK273190), Canada (MN549395) and Netherlands (MN882017, MN882018, MN882042, MN882023, MN882024, and MN882045). To our knowledge, this is the first report of occurrence of ToBRFV infecting tomato in Saudi Arabia which suggests its likely introduction by commercial seeds from countries reported this virus and spread in greenhouses through mechanical means. The author(s) declare no conflict of interest. Keywords: Tomato brown rugose fruit virus, tomato, ELISA, RT-PCR, Saudi Arabia References: Luria N, et al., 2017. PLoS ONE 12(1): 1-19. Salem N, et al., 2015. Archives of Virology 161(2): 503-506. Fig. 1. Symptoms caused by ToBRFV showing irregular brown spots, deformation, yellowing spots on fruits (A, B, C) and bubbling and mottling, mosaic with dark green wrinkled and narrowing on leaf (D).

scholarly journals First Report of Tomato torrado virus Infecting Tomato in Colombia

Plant Disease ◽  
2012 ◽  
Vol 96 (4) ◽  
pp. 592-592 ◽  
Author(s):  
M. Verbeek ◽  
A. M. Dullemans
Keyword(s):  
Mosaic Virus ◽  
Potato Virus ◽  
Polyclonal Antibodies ◽  
Sandwich Elisa ◽  
Potato Virus X ◽  
Rt Pcr ◽  
Leaf Extracts ◽  
Serological Tests ◽  
First Report ◽  
Initial Symptoms

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 continent. References: (1) C. F. Gambley et al. Plant Dis. 94:486, 2010. (2) M. Verbeek et al. Arch. Virol. 152:881, 2007. (3) M. Verbeek et al. Arch. Virol. 155:751, 2010.

scholarly journals First Report of Apium virus Y on Cilantro, Celery, and Parsley in California

Plant Disease ◽  
2008 ◽  
Vol 92 (8) ◽  
pp. 1254-1254 ◽  
Author(s):  
T. Tian ◽  
H.-Y. Liu ◽  
S. T. Koike
Keyword(s):  
Mosaic Virus ◽  
Consensus Sequence ◽  
Amino Acid Sequences ◽  
Apium Graveolens ◽  
Petroselinum Crispum ◽  
Rt Pcr ◽  
First Report ◽  
Virus Particles ◽  
Pcr Products ◽  
Reverse Primer

Recently, Apium virus Y (ApVY) was detected in field-grown cilantro (Coriandrum sativum), celery (Apium graveolens), and parsley (Petroselinum crispum) in California. In 2003, cilantro plants growing in three different fields in California (Monterey, San Joaquin, and San Luis Obispo counties) expressed symptoms of mosaic, vein clearing, and stunting. When plant sap was examined by transmission electron microscopy, flexuous, rod-shaped virus particles were observed. Total RNA was extracted from the symptomatic cilantro plants and used as a template in reverse transcription (RT)-PCR using universal potyvirus primers according to Chen et al. (1). The RT-PCR product was cloned into pGEM-T (Promega, Madison, WI) and the insert of 1,713 bp was sequenced (GenBank Accession No. EU515125). Nucleotide sequences from clones derived from three different infected cilantro plants were 89 to 97% identical to ApVY sequences encoding partial sequence of polyprotein in GenBank (Accession Nos. AY049716, EU127499, AF207594, AF203529, and EU255632). In 2007, celery plants showing necrotic line patterns and necrotic lesions on lower leaves and petioles were observed in several fields in two coastal counties in California (Monterey and Santa Clara counties). Flexuous, rod-shaped virus particles were also observed in the sap of those plants. ELISA for Cucumber mosaic virus and RT-PCR for Celery mosaic virus were negative. ApVY specific primers were designed on the basis of a consensus sequence of ApVY identified from cilantro in 2003; reverse primer 5′-GGCTCTTGCTATAGACAAATAGT-3′ and forward primer 5′-GAAGACCAAGCCAATGTGTGTA-3′. The sequence of RT-PCR products (GenBank Accession No. EU515126) amplified from infected celery had 90 to 98% nucleotide identity to ApVY. When the deduced amino acid sequences of NIb and CP regions from both cilantro and celery were used for comparison, they showed 95 to 99% identity with the known ApVY GenBank sequences mentioned above. More than 10 asymptomatic parsley plants growing in fields adjacent to the infected celery were also tested for ApVY and found to be infected. ApVY was previously identified in three Apiaceae weeds in Australia (2) and in celery in New Zealand (3). To our knowledge, this is the first report of ApVY on cilantro, celery, and parsley in California. References: (1) J. Chen et al. Arch. Virol. 146:757, 2001. (2) J. Moran et al. Arch. Virol. 147:1855, 2002. (3) J. Tang et al. Plant Dis. 91:1682, 2007.

scholarly journals First Report of Pepino Mosaic Virus on Tomato

Plant Disease ◽  
2000 ◽  
Vol 84 (1) ◽  
pp. 103-103 ◽  
Author(s):  
R. A. A. van der Vlugt ◽  
C. C. M. M. Stijger ◽  
J. Th. J. Verhoeven ◽  
D.-E. Lesemann
Keyword(s):  
Mosaic Virus ◽  
Natural Infection ◽  
Microscopic Analysis ◽  
Potato Virus X ◽  
Viral Disease ◽  
Electron Microscopic ◽  
Serological Tests ◽  
Pepino Mosaic Virus ◽  
First Report ◽  
Nucleotide Sequence Alignment

Early in 1999 a new viral disease occurred in protected tomato (Lycopersicon esculentum) crops in the Netherlands. Infected plants showed yellow leaf spots and mosaic. Transmission electron microscopic analysis revealed particles typical of potexviruses. Only three potexviruses have been reported to infect solanaceous crops: Pepino mosaic virus (PepMV), Potato aucuba mosaic virus (PAMV), and Potato virus X (PVX). Inoculation of test plants and serological tests showed that the new virus clearly differed from PAMV and PVX. Immuno-electron microscopy with antiserum to PepMV (1), the original PepMV isolate, and the virus from tomato showed decoration titers of 1:800 (homologous) and 1:400, respectively. Neither virus reacted with antiserum to PVX, nor did PVX react with antiserum to PepMV. Results of host plant analysis with 17 plant species mostly resembled those expected for PepMV. Nucleotide sequence alignment of DNA fragments obtained by reverse-transcriptase polymerase chain reaction with a specific primer set for potexviruses, directed against the RNA polymerase region, showed 93% identity between PepMV and the virus from tomato, while homologies with PVX, PAMV, and other potexviruses were <60%. Results indicate that the potexvirus in tomato is PepMV. PepMV was first found in pepino (Solanum muricatum) in Peru in 1974 and described by Jones et al. in 1980 (1). This is the first report of a natural infection of tomato by PepMV. Reference: (1) R. Jones et al. Ann. Appl. Biol. 94:61, 1980.

Identification of Three Distinct Classes of Satellite RNAs Associated With Two Cucumber mosaic virus Serotypes from the Ornamental Groundcover Vinca minor

2012 ◽  
Vol 13 (1) ◽  
pp. 18 ◽  
Author(s):  
John R. Fisher
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
The United States ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Pcr Cloning ◽  
Cloning And Sequencing ◽  
First Report ◽  
Vinca Minor ◽  
Satellite Rnas

Cucumber mosaic virus (CMV) is a cosmopolitan virus which may also have small satellite RNAs (satRNA) associated with it affecting symptom development. Vinca minor (periwinkle) plants exhibiting subtle mosaic symptoms tested positive for CMV by enzyme linked immunosorbent assay (ELISA). Double-stranded ribonucleic acid (dsRNA) analysis of CMV-Vinca field isolates in Nicotiana tabacum ‘Glurk’ suggested two sizes of putative satRNA associated with CMV. Immunocapture RT-PCR, cloning, and sequencing of the movement protein, coat protein, and satRNAs demonstrated serogroup 1A and serogroup 2 CMV helper strains and three distinct classes of satRNAs of four sizes. Further, two classes of satRNAs could be distinguished by their necrosis domains. Previously CMV was reported in V. minor in New Jersey. This is the first report of CMV in V. minor in Ohio and the first report of satRNA associated with CMV in V. minor in the United States. Accepted for publication 1 February 2012. Published 12 April 2012.

scholarly journals Natural Infection of Canavalia ensiformis with Tobacco mosaic virus in Venezuela

Plant Disease ◽  
2004 ◽  
Vol 88 (6) ◽  
pp. 681-681 ◽  
Author(s):  
E. Marys ◽  
E. Ortega ◽  
O. Carballo ◽  
C. Ramis
Keyword(s):  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Natural Infection ◽  
Amino Acid Sequences ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Electron Microscopic ◽  
Crop Species ◽  
Jack Bean ◽  
Canavalia Ensiformis

Jack bean (Canavalia ensiformis) is a valuable green manure and cover-crop species. In late summer of 2002, jack bean plants showing severe stunting, leaf mosaic, mottling, distortion, and general yellowing were observed in fields located in Maracay, Aragua State, Venezuela. Sap from symptomatic leaves was used to mechanically inoculate healthy jack bean, and field symptoms were successfully reproduced. Similar inoculations on Nicotiana tabacum var. Sansum resulted in mosaic symptoms and leaf distortion. Electron microscopic examination of leafdip preparations showed filamentous rods resembling those of a tobamovirus. Tobacco mosaic virus (TMV) was specifically identified with TMV-specific polyclonal antibody (PVAS-958, ATTC) in enzyme-linked immunosorbent assay. Sequence analysis of a coat protein gene (CP) fragment amplified using reverse transcription-polymerase chain reaction (RT-PCR) with primers TMV-CP-F and TMV-CP-R (1) from total RNA confirmed the diagnosis. Nucleotide and deduced amino acid sequences of a 450-bp region of the RT-PCR product were 96 to 99% and 98 to 100% identical, respectively, to the TMV CP gene in GenBank Accession Nos. J02415 and X68110. On the basis of foliar symptoms, incidence of TMV in jack bean was more than 50% in this experimental field. The source of infection is not known. Because TMV is reported to be seedborne in many other plant species, testing jack bean seed stocks for TMV infection could have important implications on the future control of the virus. To our knowledge, this is the first report of natural infection of jack bean by a tobamovirus. Reference: (1) N. J. Spence et al. Eur. J. Plant Pathol. 107:633, 2001.

scholarly journals First Report of Barley yellow mosaic virus in Barley in Spain

Plant Disease ◽  
2005 ◽  
Vol 89 (1) ◽  
pp. 105-105 ◽  
Author(s):  
M. A. Achon ◽  
M. Marsiñach ◽  
C. Ratti ◽  
C. Rubies-Autonell
Keyword(s):  
Mosaic Virus ◽  
Yellow Mosaic Virus ◽  
Complete Agreement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
First Report ◽  
Barley Yellow Mosaic Virus ◽  
Pcr Products ◽  
Mild Mosaic Virus ◽  
Polymerase Chain

Recently, the presence of Barley mild mosaic virus (BaMMV) and the weakly serological detection of Barley yellow mosaic virus (BaYMV) were reported in Spain (1); both viruses are members of the genus Bymovirus (family Potyviridae). Random and symptomatic surveys were conducted during February and March of 2003 in barley fields in northeastern Spain to determine the occurrence of BaMMV and BaYMV. Leaves from 316 samples collected in 15 fields were analyzed using enzyme-linked immunosorbent assay (ELISA) with commercial antisera specific for BaYMV and BaMMV (Loewe Biochemica, Munich) as well as antisera against both viruses (provided by T. Klumen). Positive ELISA samples were further analyzed using reverse transcription-polymerase chain reaction (RT-PCR) with specific primers that amplify 445 bp of BaMMV and 433 bp of BaYMV (2). Complete agreement was observed between the ELISA and RT-PCR results. Mixed infections of BaYMV and BaMMV were detected in 10 samples, BaYMV in 5 samples and BaMMV in 3 samples. Samples positive for both viruses that exhibited clear mosaic symptoms were collected in two fields. RT-PCR products from five BaYMV-infected samples were cloned and sequenced and showed 96 to 98% identity to BaYMV isolates previously reported from Europe (Genbank Accession Nos. AJ1515479-85 and X95695-7) and 92 to 95% identity with isolates reported from Asia (GenBank Accession Nos. AB023585-96, AJ132268, AJ224619-22, AJ224624-28, AF536944-46, AF536948-58, D01091, D00544, and Z24677). Sequence identity of Spanish isolates were 96 to 99%. To our knowledge, this is the first report of BaYMV infecting barley in Spain and illustrates the association of both Bymoviruses infecting barley. References: (1) M. A. Achon et al. Plant Dis. 87:1004, 2003. (2) D. Hariri et al. Eur. J. Plant Pathol. 106:365, 2000.

scholarly journals First report of papaya ring spot virus (PRSV) infecting jute (Corchorus olitorius) in India

Plant Disease ◽  
2021 ◽  
Author(s):  
C. Biswas ◽  
P. Dey ◽  
Veegala Ramesh Babu ◽  
N. M. Alam ◽  
Gouranga Kar
Keyword(s):  
Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Post Inoculation ◽  
Rt Pcr ◽  
Potato Leaf ◽  
Spot Virus ◽  
First Report ◽  
Leaf Tissues ◽  
Papaya Ring Spot Virus ◽  
Ring Spot

Jute is the most important bast fibre crop of the world, which is mainly cultivated in India, Nepal, Bangladesh, China, Indonesia and South American countries. The fibre is utilized for making apparels, ropes, bags, carpets etc (Biswas et al. 2014). This bio-fibre is gaining importance due to growing environmental consciousness worldwide. In June 2019, we noticed jute plants (less than 2%) showing virus like symptoms viz., downward curling, puckering, angular brownish to yellowish spots etc in a farmer’s field at Amdanga Block, North 24 Paraganas, West Bengal, India. To identify the virus, five symptomatic leaves from five different plants were used for high throughput sequencing (HTS). We extracted total RNA from each leaf which was subjected to construction of cDNA libraries. Sequencing was done on Illumina Hiseq 4000 (CytoScan, Thermo Fisher). Approximately 46 million 105 nt paired end reads were generated. Raw reads were trimmed and filtered to perform de novo assembly as described previously by (Grabherr et al. 2013). The obtained contig was 10,326 bp nucleotides (nt) long and in BLASTn against GenBank showed highest identity with papaya ring spot virus (PRSV) with the contig covering 99.6% of the viral genome. The obtained contig shared 99.33% sequence similarity with PRSV strain P (Accession No. MT470188). The selected leaf samples were also tested by double-antibody sandwich (DAS)- enzyme linked immunosorbent assay (ELISA) for papaya ring spot virus (PRSV) along with some common viruses, viz., Potato leaf roll virus (PLRV), Watermelon mosaic virus, Cowpea mosaic virus and Cucumber mosaic virus with the help of commercial diagnostic kits (Agdia). However, only the test with PRSV gave positive reaction for the symptomatic samples. The major symptoms of PRSV on papaya are severe mosaic, chlorosis, reduced lamina with curling and puckering (Gonsalves et al. 2010). To confirm PRSV infection, five symptomatic leaf samples (used for HTS) were collected and whole RNA was extracted from the samples using RNeasy plant minikit (Qiagen, USA). Reverse transcriptase polymerase chain reaction (RT-PCR) was conducted by using isolated RNA. One pair of PRSV specific primer (PSRV1F: 5' TTAAATCTGATTCGTC 3' PRSV 1R: 5'GAAATTCACGCAAAGTCGA3') was developed by using primer BLAST software and was used in RT-PCR assays. Amplified fragments were cloned and sequenced and all the fragments shared 98% sequence identity with PRSV. One of the amplicons was deposited in NCBI (Accession No. MN615832). Crude sap was prepared by homogenizing PRSV-infected jute leaf tissues in 0.1 M sodium phosphate buffer and 2% carborundum dust was added as abrasive (Holkar et al. 2018). The sap was then gently rubbed on to the healthy papaya leaves for inoculation. Typical PRSV like symptoms appeared in inoculated leaves 10 days post inoculation which confirmed the presence of PRSV-P. PRSV was detected by RT-PCR as well as (DAS)-ELISA from all inoculated infected papaya leaf tissues, but could not be detected from uninoculated healthy papaya tissues. To the best of our knowledge, this is the first report of PRSV-P infecting jute in India. References: Biswas, C. et al. 2014. Plant Dis. 98(4): 565. https:// doi.org/10.1094/PDIS-08-13-0826-PDN. Gonsalves, D. et al. 2010. The Plant Health Instructor. https:// doi.org/10.1094/PHI-I-2010-1004-01 Holkar, S. K. et al. 2018. Crop Protection. 108:110-119. https://doi.org/10.1016/j.cropro.2017.12.013 Grabherr, M. et al. 2011. Nat Biotechnol. 29(7): 644-652.

scholarly journals First report of groundnut ringspot virus infecting Zinnia sp. in Brazil

Plant Disease ◽  
2021 ◽  
Author(s):  
Felipe Franco de Oliveira ◽  
Gabriel Madoglio Favara ◽  
Camila Geovana Ferro ◽  
Heron Delgado Kraide ◽  
Eike Yudi Nishimura Carmo ◽  
...  
Keyword(s):  
Nucleotide Sequence ◽  
Nucleotide Sequences ◽  
Ringspot Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Concentric Ring ◽  
Leaf Extracts ◽  
Total Rna ◽  
First Report ◽  
Two Samples

Zinnia sp. is a genus belonging to Asteraceae family, originated in Mexico and adapted to a warm-hot climate (Hemmati and Mehrnoosh, 2017). Several types of zinnias with different flower color and forms are cultivated in Brazil (Min et al., 2020 and Souza Jr. et al., 2020). Characteristic symptoms of infection caused by orthotospovirus, including chlorotic spots and concentric rings on the leaves, were observed in two plants of Zinnia sp. of a florist located in the city of Piracicaba, State of São Paulo, Brazil. Orthotospovirus-like particles were observed by transmission electron microscope in leaf extracts from both plants, stained negatively with 1% uranyl acetate. By analyzing ultrathin sections of infected leaf tissues, particles of 80-100 nm in diameter were found in the lumen of the endoplasmic reticulum and nucleocapsid aggregates in the cytoplasm. Total RNA extracted separately from the leaves of both samples, using the Purelink Viral DNA / RNA kit (Thermo Fisher Scientific), was used to detect the virus by reverse transcription polymerase chain reaction (RT-PCR), using the universal primers for orthotospovirus BR60, complementary to the 3’ end of the non-translated region of the S RNA (position 1 to 15 nt), and BR65, matching the nucleocapsid gene (N) (position 433 to 453 nt), generating and amplicon of 453 nt (Eiras et al., 2001). Amplicons of the expected size were obtained for the two samples. An amplicon was purified with the Wizard SV Gel and PCR Clean-Up System kit (Promega) and sequenced in both directions at Macrogen Inc (South Korea). The nucleotide sequence (GenBank MW629018) showed 99.29-99.76% identity with nucleotide sequences of the orthotospovirus groundnut ringspot virus (GRSV) isolates (GenBank MH686229 and KY400110). Leaf extracts from symptomatic plants were also analyzed by plate-trapped antigen-enzyme-linked immunosorbent assay (PTA-ELISA), using polyclonal antiserum produced against the GRSV nucleocapsid protein (Esquivel et al., 2019). The absorbance values obtained for the extracts of the two symptomatic plants of Zinnia sp. (1.3 and 1.7) were twice as high as the value obtained for the healthy plant extract (0.5). Leaf extract of symptomatic Zinnia sp. was inoculated mechanically onto leaves of healthy plants of Zinnia sp., Capsicum annuum cv. Dara, Cucumis sativus, Cucurbita pepo cv. Caserta, Chenopodium amaranticolor, Datura stramonium, Nicotiana tabacum cv. Turkish and Solanum lycopersicum cv. Compack. At 5 days post inoculation (dpi), inoculated leaves of D. stramonium reacted with local lesions, and at 9 dpi, newly developed leaves of inoculated S. lycopersicum plants showed necrotic spot and concentric ring symptoms, whereas C. annuum exhibited concentric rings at 10 dpi. Inoculated zinnia plants showed systemic chlorotic spot and concentric ring symptoms at 20 dpi, indistinguishable from those observed under natural infection. The other inoculated plant species were not symptomatic, nor the virus was detected. PTA-ELISA and RT-PCR confirmed infection with GRSV in symptomatic plants. The amplicons generated by RT-PCR of total RNA extracted from an experimentally infected plant of C. annuum and D. stramonium, and two plants of Zinnia sp. were sent for nucleotide sequencing. The obtained nucleotide sequences (MW629019, MW629020, MW629021, MW629022) shares 100% identity with the nucleotide sequence corresponding to the original GRSV isolate (MW629018) identified in Zinnia sp. This is the first report of the natural occurrence of GRSV in Zinnia sp. in Brazil. Studies on incidence and damage are needed to recommend alternatives for management.

scholarly journals Identification, Characterization, and Molecular Detection of Alfalfa mosaic virus in Potato

2006 ◽  
Vol 96 (11) ◽  
pp. 1237-1242 ◽  
Author(s):  
H. Xu ◽  
J. Nie
Keyword(s):  
Mosaic Virus ◽  
Gene Sequence ◽  
Alfalfa Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Polymorphism Analysis ◽  
Rt Pcr ◽  
Potato Leaf ◽  
Simple Method ◽  
Cp Gene ◽  
Nucleotide Sequence Alignment

Alfalfa mosaic virus (AMV) was detected in potato fields in several provinces in Canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (RT-PCR). The identity of eight Canadian potato AMV isolates was confirmed by sequence analysis of their coat protein (CP) gene. Sequence and phylogenetic analysis indicated that these eight AMV potato isolates fell into one strain group, whereas a slight difference between Ca175 and the other Canadian AMV isolates was revealed. The Canadian AMV isolates, except Ca175, clustered together among other strains based on alignment of the CP gene sequence. To detect the virus, a pair of primers, AMV-F and AMV-R, specific to the AMV CP gene, was designed based on the nucleotide sequence alignment of known AMV strains. Evaluations showed that RT-PCR using this primer set was specific and sensitive for detecting AMV in potato leaf and tuber samples. AMV RNAs were easily detected in composite samples of 400 to 800 potato leaves or 200 to 400 tubers. Restriction analysis of PCR amplicons with SacI was a simple method for the confirmation of PCR tests. Thus, RT-PCR followed by restriction fragment length polymorphism analysis may be a useful approach for screening potato samples on a large scale for the presence of AMV.

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