Epidemiología y variabilidad patogénica del virus del mosaico del pepino dulce (Pepino mosaic virus). Nuevas enfermedades asociadas a su presencia (torrao o cribado)

El Pepino mosaic virus (PepMV) es un Potexvirus que fue descrito por primera vez en pepino dulce (Solanum muricatum Ait.) en Perú (Jones et. Al., 1980). En 1999 se detectó en Holanda, infectando a tomate (Van der Vlugt et al., 2000) mostrando una variada sintomatología. Desde entonces, el PepMV se ha expandido rápidamente por las principales áreas productoras de tomate del Mundo. Este virus se ha convertido en uno de los principales problemas en la producción de tomate en Europa donde produce importantes pérdidas económicas. La rápida expansión del PepMV en las zonas afectadas ha estado facilitada por su eficaz transmisión mecánica con las operaciones de cultivo y mediante los insectos polinizadores (Lacasa et al., 2003), sin embargo no se ha detectado la existencia de ninguna especie de insecto capaz de actuar como vector del mismo. El PepMV podría permaneces en el campo de una cosecha a la siguiente infectando a la flora arvense que podría actuar como reservorio de la enfermedad. Asimismo se ha comprobado la transmisión por semilla que podría constituir la forma de dispersión del virus a la larga distancia (Córdoba et. A., 2007). Actualmente se conocen diferentes aislados del virus que afectan al tomate. En primer momento, debido a las diferencias biológicas y moleculares observadas entre el aislado del PepMV que infectaba a tomate y el original de pepino dulce, distintos autores consideraron el aislado de tomate como un aislado diferente, denominándolo aislado tipo tomate (Van der Vlugt y Beredsen, 2002). Estudios posteriores demostraron que los aislados de PepMV de Europa, América del Norte y Canadá comparados con el aislado original de pepino dulce presentaban diferencias evidentes en sintomología, así como estructura poblacional del PepMV en cultivo de tomate en España, analizando y secuenciando tres zonas distintas del genoma del virus.Alfaro Fernández, AO. (2009). Epidemiología y variabilidad patogénica del virus del mosaico del pepino dulce (Pepino mosaic virus). Nuevas enfermedades asociadas a su presencia (torrao o cribado) [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/7027Palanci

Virosis en tomate transmitidas por semilla y su control

[ES] Las virosis transmitidas por semilla en el cultivo del tomate crean gran preocupación entre los productores, y son de especial atención en aquellos que se dedican al cultivo de variedades locales donde las semillas se extraen durante la campaña y son empleadas para cultivos posteriores con lo que la infección y dispersión de estos virus es mucho más frecuente. Entre los virus transmitidos por semilla en tomate destacan el virus del mosaico del tomate (ToMV) y el virus del mosaico del pepino dulce (PepMV). Ambos virus se caracterizan por transmitirse, además de por semilla, de manera mecánica fácilmente y son muy estables manteniéndose en los restos del cultivo anterior y en las infraestructuras empleadas durante el manejo del cultivo. Sin embargo, la localización de estos virus en las semillas contaminadas difiere, mientras que PepMV se localiza únicamente de manera superficial, ToMV puede encontrarse además en zonas más internas como en el endospermo. Esto hace que los tratamientos empleados para la desinfección de semillas infectadas con cada uno de estos virus sea distinto: mientras que PepMV puede ser inactivado con tratamientos químicos superficiales, el tratamiento para descontaminar semillas con ToMV debe ser térmico a elevadas temperaturas.[EN] Viral diseases transmitted through seed create a great concern among the tomato producers, especially those who use local varieties that harvest their own seeds from the previous growing season fruits. In this case the infection and spread of seed-transmitted viruses is more usual. ToMV and PepMV are the two main seed-transmitted viruses which affect tomato crops. Both viruses are easily mechanically and seed transmitted, and remain infective in the plant debris of the previous crop and in the crop structures. However, the location of the virus in the contaminated seed is different. PepMV is present only externally in the seed coat, but ToMV could be also found in the endosperm. Therefore seed treatments to inactivate these two viruses are different; while PepMV could be inactivated by external chemical treatments, ToMV infected seeds should be thermal treated in order to eliminate further seedling infections.Alfaro Fernández, AO.; Font San Ambrosio, MI. (2020). Virosis en tomate transmitidas por semilla y su control. En I Congrés de la Tomaca Valenciana: La Tomaca Valenciana d'El Perelló. Editorial Universitat Politècnica de València. 97-114. https://doi.org/10.4995/TOMAVAL2017.2017.6524OCS9711

Fine mapping of wmv1551, a resistance gene to Watermelon mosaic virus in melon

[EN] Recessive resistance to Watermelon mosaic virus (WMV) in melon has previously been reported in the African accession TGR-1551. Using a population of recombinant inbred lines (RIL), derived from a cross between TGR-1551 and the susceptible Spanish cultivar Bola de Oro' (BO), a major quantitative trait locus (QTL) controlling the resistance was previously mapped to a region of approximately 760kb in chromosome 11. Minor QTLs were also reported with lower effects, dependent on the environmental conditions. A genotyping by sequencing (GBS) analysis of the RIL population has provided new information that allowed the better location of the major QTL in chromosome 11. Moreover, three minor QTLs in chromosomes 4, 5, and 6 were identified. Generations derived from the RIL population were subsequently phenotyped for resistance and genotyped with SNP markers to fine map the resistance derived from TGR-1551. The results obtained have allowed to narrow the position of the resistance gene on chromosome 11, designated as wmv(1551), to a 141-kb region, and the confirmation of a minor QTL in chromosome 5. The effect of the minor QTL in chromosome 5 was significant in heterozygote plants for the introgression in chromosome 11. The SNP markers linked to both QTLs will be useful in breeding programs aimed at the introgression of WMV resistance derived from TGR-1551. Future work will be directed to identifying the resistance gene, wmv(1551), in the candidate region on chromosome 11.This study was partially supported by the Spanish Ministerio de Economia y Competitividad grants AGL2014-53398-C2 (1-R and 2-R), by the Spanish Ministerio de Ciencia, Innovacion y Universidades grants AGL2017-85563-C2 (1-R and 2-R) and BIO2017-83184-R, and by the PROMETEO project 2017/078 (to promote excellence groups) by the Conselleria d'Educacio, Investigacio, Cultura i Esports (Generalitat Valenciana).Pérez De Castro, AM.; Esteras Gómez, C.; Alfaro Fernández, AO.; Daròs, J.; Monforte Gilabert, AJ.; Picó Sirvent, MB.; Gómez-Guillamón, ML. (2019). Fine mapping of wmv1551, a resistance gene to Watermelon mosaic virus in melon. Molecular Breeding. 39(7):1-15. https://doi.org/10.1007/s11032-019-0998-zS115397Abreu-Neto JB, Turchetto-Zolet AC, Valter de Oliveira LF, Bodanese Zanettini MH, Margis-Pinheiro M (2013) Heavy metal-associated isoprenylated plant protein (HIPP): characterization of a family of proteins exclusive to plants. FEBS J 280:1604–1616Aragonés V, Pérez-de-Castro A, Cordero T, Cebolla-Cornejo J, López C, Picó B, Daròs JA (2018) A Watermelon mosaic virus clone tagged with the yellow visual maker phytoene synthase facilitates scoring infectivity in melon breeding programs. Eur J Plant Pathol 153:1317–1323. https://doi.org/10.1007/s10658-018-01621-xBachlava E, Bertrand F, De Vries J, Joobeur T, King J, Kraakman P (2014) Patent No. US20140059712.Multiple-virus-resistant melonChen S, Li F, Liu D, Jiang C, Cui L, Shen L, Liu G, Yang A (2017) Dynamic expression analysis of early response genes induced by potato virus Y in PVY-resistant Nicotiana tabacum. Plant Cell Rep 36:297–311Colcombet J, Hirt H (2008) Arabidopsis MAPKs: a complex signalling network involved in multiple biological processes. Biochem J 413:217–226Cordero T, Cerdán L, Carbonell A, Katsarou K, Kalantidis K, Daròs JA (2017) Dicer-like 4 is involved in restricting the systemic movement of Zucchini yellow mosaic virus in Nicotiana benthamiana. Mol Plant-Microbe Interact 30:63–71Desbiez C, Joannon B, Wipf-Scheibel C, Chandeysson C, Lecoq H (2009) Emergence of new strains of Watermelon mosaic virus in South-eastern France: evidence for limited spread but rapid local population shift. Virus Res 141:201–208Desbiez C, Lecoq H (2008) Evidence for multiple intraspecific recombinants in natural populations of Watermelon mosaic virus (WMV, Potyvirus). Arch Virol 153:1749–1754Díaz-Pendón JA, Fernández-Muñoz R, Gómez-Guillamón ML, Moriones E (2005) Inheritance of resistance to Watermelon mosaic virus in Cucumis melo that impairs virus accumulation, symptom expression, and aphid transmission. Phytopathology 95:840–846Díaz-Pendón JA, Mallor C, Soria C, Camero R, Garzo E, Fereres A, Alvarez JM, Gómez-Guillamón ML, Luis-Arteaga M, Moriones E (2003) Potential sources of resistance for melon to nonpersistently aphid-borne viruses. Plant Dis 87:960–964Díaz-Pendón JA, Truniger V, Nieto C, Garcia-Mas J, Bendahmane A, Aranda MA (2004) Advances in understanding recessive resistance to plant viruses. Mol Plant Pathol 5:223–233Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15Esteras C, Formisano G, Roig C, Díaz A, Blanca J, Garcia-Mas J, Gómez-Guillamón ML, López-Sesé AI, Lázaro A, Monforte AJ, Picó B (2013) SNP genotyping in melons: genetic variation, population structure, and linkage disequilibrium. Theor Appl Genet 126:1285–1303Fereres A, Moreno A (2011) Integrated control measures against viruses and their vectors. In: Caranta C, Aranda MA, Tepfer M, López-Moya J (eds) Recent Advances in Plant Virology, Caister Academic Press, Norfolk, pp 237–262Fernández-Silva I, Eduardo I, Blanca J, Esteras C, Picó B, Nuez F, Arús P, García-Mas J, Monforte A (2008) Bin mapping of genomic and EST-derived SSRs in melon (Cucumis melo L.). Theor Appl Genet 118:139–150Fukino N, Sakata Y, Kunihisa M, Matsumoto S (2007) Characterization of novel simple sequence repeat (SSR) markers for melon (Cucumis melo L.) and their use for genotyping identification. J Hort Sci Biotechnol 82:330–334 Details about primer sequences: http://cse.naro.affrc.go.jp/nbk/List_CMN.xlsGilbert RZ, Kyle MM, Munger HM, Gray SM (1994) Inheritance of resistance to Watermelon mosaic virus in Cucumis melo L. HortSci 29:107–110González VM, Aventín N, Centeno E, Puigdomènech P (2013) High presence/absence gene variability in defense-related gene clusters of Cucumis melo. BMC Genomics 14:782González-Ibeas D, Blanca J, Donaire L, Saladié M, MArcarell-Creus A, Cano-Delgado A, García-Mas J, Llave C, Aranda MA (2011) Analysis of the melon (Cucumis melo) small RNAome by high-throughput pyrosequencing. BMC Genomics 12:393González-Ibeas D, Cañizares J, Aranda MA (2012) Microarray analysis shows that recessive resistance to Watermelon mosaic virus in melon is associated with the induction of defense response genes. Mol Plant-Microbe Interact 25:107–118Hashimoto M, Neriya Y, Yamaji Y, Namba S (2016) Recessive resistance to plant viruses: potential resistance genes beyond translation initiation factors. Front Microbiol 7:1695JUAN A. DIAZ-PENDON, VERONICA TRUNIGER, CRISTINA NIETO, JORDI GARCIA-MAS, ABDELHAFID BENDAHMANE, MIGUEL A. 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Inoculation of cucumber, melón and zucchini varieties with Tomato leaf curl New Delhi virus (ToLCNDV) and evaluation of infection using different methods

This is the peer reviewed version of the following article: Figás-Moreno, MDR.; Alfaro Fernández, AO.; Font San Ambrosio, MI.; Borràs Palomares, D.; Casanova-Calancha, C.; Hurtado Ricart, M.; Plazas Ávila, MDLO.... (2017). Inoculation of cucumber, melón and zucchini varieties with Tomato leaf curl New Delhi virus (ToLCNDV) and evaluation of infection using different methods. Annals of Applied Biology. 170(3):405-414. doi:10.1111/aab.12344, which has been published in final form at http://doi.org/10.1111/aab.12344. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.[EN] The disease caused by Tomato leaf curl New Delhi virus (ToLCNDV), which is naturally transmitted by the whitefly Bemisia tabaci, causes important economic losses in cucurbit crops. The availability of simple and efficient inoculation protocols and detection methods is necessary for screening varieties and germplasm collections as well as for breeding populations. We evaluated the infectivity of ToLCNDV inocula prepared using three different buffers for mechanical sap inoculation in a susceptible variety of zucchini. We found that inoculum prepared with buffer III, which contains polyvinylpyrrolidone, is highly efficient for mechanical inoculation, with 100% of plants displaying severe symptoms 21 days post-inoculation. Using this buffer, we mechanically inoculated 19 commercial varieties of cucurbit crops (six of cucumber, six of melon and seven of zucchini), evaluated the evolution of symptoms and diagnosed infection using nine different ToLCNDV detection methods (four based on serology, four based on molecular hybridization and one based on PCR detection). The results revealed that all varieties are susceptible, although cucumber varieties display less severe symptoms than those of melon or zucchini. All detection methods were highly efficient (more than 85% of plants testing positive) in melon and zucchini, but in cucumber, the percentage of positive plants detected with serology and molecular hybridization methods ranged from 20.4% with Squash leaf curl virus (SLCV) antiserum, to 78.5% with DNA extract hybridization. Overall, the best detection results were obtained with PCR, with 92.6%, 92.4% and 98.4% cucumber, melon and zucchini plants, respectively, testing positive. When considering the overall results in the three crops, the best serology and molecular hybridization methods were those using Watermelon chlorotic stunt virus (WmCSV) antiserum and DNA extract, respectively. The inoculation methodology developed and the information on detection methods are of great relevance for the selection and breeding of varieties of cucurbit crops that are tolerant or resistant to ToLCNDV.Figás-Moreno, MDR.; Alfaro Fernández, AO.; Font San Ambrosio, MI.; Borràs Palomares, D.; Casanova-Calancha, C.; Hurtado Ricart, M.; Plazas Ávila, MDLO.... (2017). Inoculation of cucumber, melón and zucchini varieties with Tomato leaf curl New Delhi virus (ToLCNDV) and evaluation of infection using different methods. Annals of Applied Biology. 170(3):405-414. doi:10.1111/aab.12344S405414170

Assessment of Multilocus Sequence Analysis (MLSA) for Identification of Candidatus Liberibacter Solanacearum from Different Host Plants in Spain

[EN] Liberibacteris a bacterial group causing different diseases and disorders in plants. Among liberibacters,CandidatusLiberibacter solanaceraum (CLso) produces disorders in several species mainly within Apiaceae and Solanaceae families. CLso isolates are usually grouped in defined haplotypes according to single nucleotide polymorphisms in genes associated with ribosomal elements. In order to characterize more precisely isolates of CLso identified in potato in Spain, a Multilocus Sequence Analysis (MLSA) was applied. This methodology was validated by a complete analysis of ten housekeeping genes that showed an absence of positive selection and a nearly neutral mechanism for their evolution. Most of the analysis performed with single housekeeping genes, as well as MLSA, grouped together isolates of CLso detected in potato crops in Spain within the haplotype E, undistinguishable from those infecting carrots, parsnips or celery. Moreover, the information from these housekeeping genes was used to estimate the evolutionary divergence among the different CLso by using the concatenated sequences of the genes assayed. Data obtained on the divergence among CLso haplotypes support the hypothesis of evolutionary events connected with different hosts, in different geographic areas, and possibly associated with different vectors. Our results demonstrate the absence in Spain of CLso isolates molecularly classified as haplotypes A and B, traditionally considered causal agents of zebra chip in potato, as well as the uncertain possibility of the present haplotype to produce major disease outbreaks in potato that may depend on many factors that should be further evaluated in future worksThis research was funded by Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria (INIA), grant numbers AT2016-007 and RTA2014-00008-C04-03-E, co-financed by FEDER.Ruiz-Padilla, A.; Redondo, C.; Asensio, A.; Garita-Cambronero, J.; Martinez, C.; Perez-Padilla, V.; Marquinez, R.... (2020). Assessment of Multilocus Sequence Analysis (MLSA) for Identification of Candidatus Liberibacter Solanacearum from Different Host Plants in Spain. Microorganisms. 8(9):1-19. https://doi.org/10.3390/microorganisms8091446S11989Haapalainen, M. (2014). Biology and epidemics ofCandidatusLiberibacter species, psyllid-transmitted plant-pathogenic bacteria. Annals of Applied Biology, 165(2), 172-198. doi:10.1111/aab.12149Raddadi, N., Gonella, E., Camerota, C., Pizzinat, A., Tedeschi, R., Crotti, E., … Alma, A. (2010). ‘Candidatus Liberibacter europaeus’ sp. nov. that is associated with and transmitted by the psyllid Cacopsylla pyri apparently behaves as an endophyte rather than a pathogen. Environmental Microbiology, 13(2), 414-426. doi:10.1111/j.1462-2920.2010.02347.xWang, N., Pierson, E. A., Setubal, J. C., Xu, J., Levy, J. G., Zhang, Y., … Martins, J. (2017). The Candidatus Liberibacter–Host Interface: Insights into Pathogenesis Mechanisms and Disease Control. Annual Review of Phytopathology, 55(1), 451-482. doi:10.1146/annurev-phyto-080516-035513Morris, J., Shiller, J., Mann, R., Smith, G., Yen, A., & Rodoni, B. (2017). Novel ‘Candidatus Liberibacter’ species identified in the Australian eggplant psyllid, Acizzia solanicola. Microbial Biotechnology, 10(4), 833-844. doi:10.1111/1751-7915.12707Alfaro-Fernández, A., Hernández-Llopis, D., & Font, M. I. (2017). Haplotypes of ‘Candidatus Liberibacter solanacearum’ identified in Umbeliferous crops in Spain. European Journal of Plant Pathology, 149(1), 127-131. doi:10.1007/s10658-017-1172-2Haapalainen, M., Wang, J., Latvala, S., Lehtonen, M. T., Pirhonen, M., & Nissinen, A. I. (2018). Genetic Variation of ‘Candidatus Liberibacter solanacearum’ Haplotype C and Identification of a Novel Haplotype from Trioza urticae and Stinging Nettle. Phytopathology®, 108(8), 925-934. doi:10.1094/phyto-12-17-0410-rHaapalainen, M., Latvala, S., Wickström, A., Wang, J., Pirhonen, M., & Nissinen, A. I. (2019). A novel haplotype of ‘Candidatus Liberibacter solanacearum’ found in Apiaceae and Polygonaceae family plants. European Journal of Plant Pathology, 156(2), 413-423. doi:10.1007/s10658-019-01890-0Mauck, K. E., Sun, P., Meduri, V. R., & Hansen, A. K. (2019). New Ca. Liberibacter psyllaurous haplotype resurrected from a 49-year-old specimen of Solanum umbelliferum: a native host of the psyllid vector. Scientific Reports, 9(1). doi:10.1038/s41598-019-45975-6Teixeira, D. C., Eveillard, S., Sirand-Pugnet, P., Wulff, A., Saillard, C., Ayres, A. J., & Bove, J. M. (2008). The tufB-secE-nusG-rplKAJL-rpoB gene cluster of the liberibacters: sequence comparisons, phylogeny and speciation. INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 58(6), 1414-1421. doi:10.1099/ijs.0.65641-0Glaeser, S. P., & Kämpfer, P. (2015). Multilocus sequence analysis (MLSA) in prokaryotic taxonomy. Systematic and Applied Microbiology, 38(4), 237-245. doi:10.1016/j.syapm.2015.03.007Gevers, D., Cohan, F. M., Lawrence, J. G., Spratt, B. G., Coenye, T., Feil, E. J., … Swings, J. (2005). Re-evaluating prokaryotic species. Nature Reviews Microbiology, 3(9), 733-739. doi:10.1038/nrmicro1236Swisher Grimm, K. D., & Garczynski, S. F. (2019). Identification of a New Haplotype of ‘CandidatusLiberibacter solanacearum’ inSolanum tuberosum. Plant Disease, 103(3), 468-474. doi:10.1094/pdis-06-18-0937-reLin, H., Lou, B., Glynn, J. M., Doddapaneni, H., Civerolo, E. L., Chen, C., … Vahling, C. M. (2011). The Complete Genome Sequence of ‘Candidatus Liberibacter solanacearum’, the Bacterium Associated with Potato Zebra Chip Disease. PLoS ONE, 6(4), e19135. doi:10.1371/journal.pone.0019135Thompson, S. M., Johnson, C. P., Lu, A. Y., Frampton, R. A., Sullivan, K. L., Fiers, M. W. E. J., … Smith, G. R. (2015). Genomes of ‘Candidatus Liberibacter solanacearum’ Haplotype A from New Zealand and the United States Suggest Significant Genome Plasticity in the Species. Phytopathology®, 105(7), 863-871. doi:10.1094/phyto-12-14-0363-fiLin, H., Pietersen, G., Han, C., Read, D. A., Lou, B., Gupta, G., & Civerolo, E. L. (2015). Complete Genome Sequence of « Candidatus Liberibacter africanus,» a Bacterium Associated with Citrus Huanglongbing. Genome Announcements, 3(4). doi:10.1128/genomea.00733-15Wulff, N. A., Zhang, S., Setubal, J. C., Almeida, N. F., Martins, E. C., Harakava, R., … Gabriel, D. W. (2014). The Complete Genome Sequence of ‘Candidatus Liberibacter americanus’, Associated with Citrus Huanglongbing. Molecular Plant-Microbe Interactions®, 27(2), 163-176. doi:10.1094/mpmi-09-13-0292-rDuan, Y., Zhou, L., Hall, D. G., Li, W., Doddapaneni, H., Lin, H., … Gottwald, T. (2009). Complete Genome Sequence of Citrus Huanglongbing Bacterium, ‘CandidatusLiberibacter asiaticus’ Obtained Through Metagenomics. Molecular Plant-Microbe Interactions®, 22(8), 1011-1020. doi:10.1094/mpmi-22-8-1011Katoh, H., Miyata, S., Inoue, H., & Iwanami, T. (2014). Unique Features of a Japanese ‘Candidatus Liberibacter asiaticus’ Strain Revealed by Whole Genome Sequencing. PLoS ONE, 9(9), e106109. doi:10.1371/journal.pone.0106109Leonard, M. T., Fagen, J. R., Davis-Richardson, A. G., Davis, M. J., & Triplett, E. W. (2012). Complete genome sequence of Liberibacter crescens BT-1. Standards in Genomic Sciences, 7(2), 271-283. doi:10.4056/sigs.3326772Teresani, G. R., Bertolini, E., Alfaro-Fernández, A., Martínez, C., Tanaka, F. A. O., Kitajima, E. W., … Font, M. I. (2014). Association of ‘Candidatus Liberibacter solanacearum’ with a Vegetative Disorder of Celery in Spain and Development of a Real-Time PCR Method for Its Detection. Phytopathology®, 104(8), 804-811. doi:10.1094/phyto-07-13-0182-rLi, W., Hartung, J. S., & Levy, L. (2006). Quantitative real-time PCR for detection and identification of Candidatus Liberibacter species associated with citrus huanglongbing. Journal of Microbiological Methods, 66(1), 104-115. doi:10.1016/j.mimet.2005.10.018Munyaneza, J. E., Sengoda, V. G., Crosslin, J. M., De la Rosa-Lozano, G., & Sanchez, A. (2009). First Report of ‘Candidatus Liberibacter psyllaurous’ in Potato Tubers with Zebra Chip Disease in Mexico. Plant Disease, 93(5), 552-552. doi:10.1094/pdis-93-5-0552aPhillips, J. L., & Gnanakaran, S. (2014). A data-driven approach to modeling the tripartite structure of multidrug resistance efflux pumps. Proteins: Structure, Function, and Bioinformatics, 83(1), 46-65. doi:10.1002/prot.24632Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura, K. (2018). MEGA X: Molecular Evolutionary Genetics Analysis across Computing Platforms. Molecular Biology and Evolution, 35(6), 1547-1549. doi:10.1093/molbev/msy096Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. (1993). Molecular Biology and Evolution. doi:10.1093/oxfordjournals.molbev.a040023Rozas, J., Ferrer-Mata, A., Sánchez-DelBarrio, J. C., Guirao-Rico, S., Librado, P., Ramos-Onsins, S. E., & Sánchez-Gracia, A. (2017). DnaSP 6: DNA Sequence Polymorphism Analysis of Large Data Sets. 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Identificación y detección simultánea de aislados de Pepino mosaic virus (PepMV) mediante multiplex RT-PCR

A one-step reverse transcription-polymerase chain reaction (RT-PCR) followed by a restriction analysis has been developed for the simultaneous detection and identification of five different strains of Pepino mosaic virus (PepMV).Alfaro Fernández, AO. (2008). Simultaneous detection and identification of Pepino mosaic virus (PepMV) isolates by multiplex one-step RT-PCR. http://hdl.handle.net/10251/12149Archivo delegad

Detection, characterization and host range studies of Pepino mosaic virus in Cyprus

[EN] Pepino mosaic virus (PepMV, Genus Potexvirus, Family Flexiviridae) is a mechanically transmitted viral disease that has emerged as a significant problem of greenhouse tomato crops in Europe and around the world. Although previous studies in Cyprus suggested that the virus was not present on the island, in 2009 tomato fruits from two major tomato production areas exhibited symptoms of yellow mosaic and discolouration, similar to those induced by PepMV. Consequently, an extensive survey was conducted in all tomato producing areas of the country to identify the incidence and prevalence of PepMV in protected and open field tomato crops. Analysis of 3500 leaf samples from tomato plants and weeds with DAS-ELISA and real-time RT-PCR showed that PepMV was present in all tomato growing areas of the island. The virus was detected in both protected and open field tomato plants, as well as in 20 weed species in the families of Amaranthaceae, Chenopodiaceae, Compositae, Convolvulaceae, Malvaceae, Plantaginaceae and Solanaceae. All Cypriot isolates assayed belonged to the CH2 genotype. Biological assays with two Cypriot isolates showed that they could infect cultivated and weed species including Vigna unguiculata, Solanum melongena, Nicotiana tabacum, Malva parviflora, Sonchus oleraceus, Solanum nigrum, Convolvulus arvensis, Chrysanthemum segetum and Calendula arvensis. To our knowledge, this is the first study to report Chrysanthemum segetum and Calendula arvensis as hosts of PepMV.This work was funded by a grant from the Cyprus Research Promotion Foundation and supported by the European Commission in the 6th Framework Programme (PEPEIRA EC contract No 044189). The authors thank Dr. R. van der Vlugt (Plant Research International, The Netherlands), Prof. M. Ravnikar and Dr. I. Gutierrez-Aguirre (National Institute of Biology, Slovenia), Prof. C. Jorda (Instituto Agroforestal Mediterraneao, Valencia, Spain), for providing Pepino mosaic virus isolates used as positive controls. The authors would also like to thank Drs. G. Neophytou, A. Melifronidou, D. Koudounas (Cyprus Department of Agriculture) and I. Harkou and Y. Markou for their valuable assistance in surveys and laboratory experiments and G. Economides for identification of weed species. Dr. M. Stavrinides and Dr. A. Kyriakou are acknowledged for critically reviewing this manuscript.Papayiannis, L.; Kokkinos, C.; Alfaro Fernández, AO. (2012). Detection, characterization and host range studies of Pepino mosaic virus in Cyprus. European Journal of Plant Pathology. 132(1):1-7. https://doi.org/10.1007/s10658-011-9854-7S171321Alfaro-Fernández, A., Cebrián, M. C., Córdoba-Sellés, M. C., Herrera-Vásquez, J. A., & Jordá, C. (2008). First report of the US1 strain of Pepino mosaic virus in tomato in the Canary Isands, Spain. Plant Disease, 92, 1590.Alfaro-Fernández, A., Sánchez-Navarro, J. A., Cebrián, M. C., Córdoba-Sellés, M. C., Pallás, V., & Jordá, C. (2009). Simultaneous detection and identification of Pepino mosaic virus (PepMV) isolates by multiplex one-step RT-PCR. European Journal of Plant Pathology, 125, 143–158.Campanella, J. J., Bitincka, L., & Smalley, J. (2003). MatGAT: an application that generates similarity/identity matrices using protein or DNA sequences. BMC Bioinformatics, 4, 29–33.Córdoba, M. C., Martínez-Priego, L., & Jordá, C. (2004). New natural hosts of Pepino mosaic virus in Spain. Plant Disease, 88, 906.Córdoba-Sellés, M. C., García-Rández, A., Alfaro-Fernández, A., & Jordá, C. (2007). Seed transmission of Pepino mosaic virus and efficacy of tomato seed disinfection treatments. Plant Disease, 91, 1250–1254.Gómez, P., Sempere, R. N., Elena, S. F., & Aranda, M. (2009). Mixed infections of Pepino mosaic virus strains modulate the evolutionary dinamics of this emerging virus. Journal of Virology, 83, 12378–12387.Gutiérrez-Aguirre, I., Mehle, N., Delić, D., Gruden, K., Mumford, R., & Ravnikar, M. (2009). Real-time quantitative PCR based sensitive detection and genotype discrimination of Pepino mosaic virus. Journal of Virological Methods, 162, 46–55.Hanssen, I. M., & Thomma, B. P. H. J. (2010). Pepino mosaic virus: a successful pathogen that rapidly evolved from emerging to endemic in tomato crops. Molecular Plant Pathology, 11, 179–189.Hanssen, I. M., Mumford, R., Blystad, D.-R., Cortez, I., Hasiów-Jaroszewska, B., Hristova, D., et al. (2009). Seed transmission of Pepino mosaic virus in tomato. European Journal of Plant Pathology, 126, 145–152.Jones, R. A. C., Koening, R., & Lesemann, D. E. (1980). Pepino mosaic virus, a new Potexvirus from pepino (Solanum muricatum). Annals of Applied Biology, 94, 61–68.Jordá, C., Lázaro-Pérez, A., Martínez-Culebras, P., & Lacasa, A. (2001). First report of Pepino mosaic virus on natural hosts. Plant Disease, 85, 1292.Jordá, C., Lázaro-Pérez, A., Martínez-Culebras, P., & Abad, P. (2001). First report of Pepino mosaic virus on tomato in Spain. Plant Disease, 85, 1292.Kumar, S., Tamura, K., & Nei, M. (2004). MEGA3: integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinformatic, 5, 150–163.Ling, K. S., Wechter, W. P., & Jordan, R. (2007). Development of a one-step immunocapture real-time TaqMan RT-PCR assay for the broad spectrum detection of Pepino mosaic virus. Journal of Virological Methods, 144, 65–72.Mumford, R. A., & Metcalfe, E. J. (2001). The partial sequencing of the genomic RNA of a UK isolate of Pepino mosaic virus and the comparison of the coat protein sequence with othes isolates from Europe and Peru. Archives of Virology, 146, 2455–2460.Pagán, I., Córdoba-Sellés, M. C., Martínez-Priego, L., Fraile, A., Malpica, J. M., Jordá, C., et al. (2006). Genetic structure of the population of Pepino mosaic virus infecting tomato crops in Spain. Phytopathology, 96, 274–279.Papayiannis, L. C., Savvides, A., Chatziafksentis, K., Kapari-Isaia, T., Dovas, C., Katis, N. I., et al. (2008). Incidence of viruses infecting tomato crops in Cyprus. Phytopathologia Mediterranea, 47, 159.Pospieszny, H., Hásiow, B., & Borodynko, N. (2008). Characterization of two distinct Polish isolates of Pepino mosaic virus. European Journal of Pathology, 122, 443–445.Spence, N. J., Basham, J., Mumford, R. A., Hayman, G., Edmondson, R., & Jones, D. R. (2006). Effect of Pepino mosaic virus on the yield and quality of glasshouse-grown tomatoes in the UK. Plant Pathology, 55, 595–606.Soler, S., Prohens, J., Diez, M. J., & Nuez, E. (2002). Natural occurrence of Pepino mosaic virus in Lycopersicon species in Central and Southern Peru. Journal of Phytopathology, 150, 45–53.Van der Vlugt, R. A. A. (2009). Pepino mosaic virus. Hellenic Plant Protection Journal, 2, 47–56.Van der Vlugt, R. A. A., Stijger, C. C. M. M., Verhoeven, J. Th. J., & Leserman, D. E. (2000). First report of Pepino mosaic virus on tomato. Plant Disease, 84, 103.Van der Vlugt, R. A. A., Cuperus, C., Vink, J., Stijger, C. C. M. M., Lesemann, D.-E., Verhoeven, J. Th. J., et al. (2002). Identification and characterisation of Pepino mosaic potex virus in tomato. Bulletin OEPP/EPPO Bulletin, 32, 503–508.Verhoeven, J. T. J., Van der Vlugt, R. A. A., & Roenhorst, J. W. (2003). High similarity between tomato isolates of Pepino mosaic virus suggest a common origin. European Journal of Plant Pathology, 109, 419–425.Wright, D., & Mumford, R. (1999). Pepino mosaic potexvirus (PepMV). First records in tomato in United Kingdom. Central Science Laboratory, York, UK. Plant Disease Notice, 89

Detection of stolbur group (16SrXII) phytoplasma in willow (Salix babylonica).

Copyright of the bulletin (©Bulletin of Insectology). Full text articles are available at the publisher site freely two years after publication.[EN] Preliminary results of nested-PCR indicated that phytoplasmas were detected in willow (Salix babylonica Linn) showing yellows, ball-like structures and small leaves symptoms collected in Valencia Province (Eastern Spain). RFLP analyses showed that the phytoplasmas belonged to the stolbur group (16SrXII).Alfaro Fernández, AO.; Abad Campos, P.; Hernández Llopis, D.; Serrano Fernández, A.; Font San Ambrosio, MI. (2011). Detection of stolbur phytoplasma in willow in Spain. Bulletin of insectology. 64(Supplement):111-112. http://hdl.handle.net/10251/63861S11111264Supplemen

First report of Eggplant mottled dwarf virus in Pittosporum tobira in Spain

Alfaro Fernández, AO.; Córdoba-Sellés, MDC.; Tornos, T.; Cebrián, M.; Font San Ambrosio, MI. (2011). First report of Eggplant mottled dwarf virus in Pittosporum tobira in Spain. Plant Disease. 95(1):75-75. doi:10.1094/PDIS-07-10-0491S757595