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The inherent ability of seeds (orthodox, intermediate, and recalcitrant seeds and vegetative propagules) to serve as carriers of pests and pathogens (hereafter referred to as pests) and the risk of transboundary spread along with the seed movement present a high-risk factor for international germplasm distribution activities. Quarantine and phytosanitary procedures have been established by many countries around the world to minimize seed-borne pest spread by screening export and import consignments of germplasm. The effectiveness of these time-consuming and cost-intensive procedures depends on the knowledge of pest distribution, availability of diagnostic tools for seed health testing, qualified operators, procedures for inspection, and seed phytosanitation. This review describes a unique multidisciplinary approach used by the CGIAR Germplasm Health Units (GHUs) in ensuring phytosanitary protection for the safe conservation and global movement of germplasm from the 11 CGIAR genebanks and breeding programs that acquire and distribute germplasm to and from all parts of the world for agricultural research and food security. We also present the challenges, lessons learned, and recommendations stemming from the experience of GHUs, which collaborate with the national quarantine systems to export and distribute about 100,000 germplasm samples annually to partners located in about 90 to 100 countries. Furthermore, we describe how GHUs adjust their procedures to stay in alignment with evolving phytosanitary regulations and pest risk scenarios. In conclusion, we state the benefits of globally coordinated phytosanitary networks for the prevention of the intercontinental spread of pests that are transmissible through plant propagation materials.

En la distribución de germoplasma la calidad sanitaria juega un papel muy importante, debido a que en actividades de importación o exportación se puede correr el riesgo de propagar plagas y enfermedades a otros países. Por este motivo, en el proceso de certificación sanitaria deben utilizarse metodologías modernas, altamente sensibles y eficientes, que garanticen una transferencia segura. Para cumplir este objetivo existen legislaciones internacionales y nacionales que buscan prohibir la propagación de plagas cuarentenarias con las transferencias de germoplasma. Adicionalmente, las Naciones Unidas han declarado el 2020 como el “Año Internacional de la Sanidad Vegetal”, buscando sensibilizar a escala internacional sobre cómo la protección de la sanidad vegetal puede ayudar a acabar con el hambre, reducir la pobreza, proteger el medio ambiente y estimular el desarrollo económico.

Achira (Canna edulis Ker.) is a cultivated species for handcrafted food products and starch production. In Colombia is estimated an achira cultivated area of 800 ha; in the department of Nariño there has been a disturbance of viral etiology, known by farmers as Streak Virus, due to its symptoms in the leaves, but without previous records in the area. The disease causes losses in performance, although they have not been established precisely. In order to clarify the nature of this pathology and the identity of the pathogen associated with he problem, an investigation was carried out at the University of Nariño, by means of molecular tests of PCR and RT-PCR, sequencing, serology and electron microscopy, of foliar samples collected in the producing areas. The most outstanding symptoms in affected tissues were yellow mosaic, mottled, chlorotic streak and ribs discoloration, among others. There were no cytoplasmic inclusions similar to those produced by Potyvirus, nor viral particles were observed, nor serology positive results, but it was possible to achieve the amplifi cation of a cDNA fragment, with specifi c primers for Potyvirus and 98% of homology of the sequences with Sugarcane mosaic virus . This is the first S CMV report in achira in Nariño, Colombia La achira (Canna edulis Ker.) es una especie utilizada para la producción de almidón y alimentos artesanales. En Colombia, se estima un área cultivada de 800ha; en el departamento de Nariño, se viene presentando un disturbio de etiología viral, conocido por los agricultores como el rayado, por sus síntomas en las hojas, pero sin registros previos en esta zona. La enfermedad causa pérdidas en el rendimiento, aunque no se ha establecido con precisión. Con el objetivo de esclarecer la naturaleza de dicha patología y la identidad del patógeno asociado al problema, en la Universidad de Nariño, se realizó una investigación, mediante pruebas moleculares de PCR y RT-PCR, secuenciación, serología y microscopía electrónica, de muestras foliares colectadas en las zonas productoras. Los síntomas más sobresalientes en tejidos afectados fueron mosaico amarillo, moteado, rayado clorótico, aclaramiento de nervaduras entre otros. No se detectaron inclusiones citoplasmáticas similares a las producidas por Potyvirus, ni se observaron partículas virales, tampoco hubo resultados positivos con serología, pero sí se logró amplificación de un fragmento de cDNA, con cebadores específicos para Potyvirus y homología de 98% de las secuencias con el virus Sugarcane mosaic virus SCMV. Este es el primer reporte de SCMV en achira en Nariño, Colombia Incluye bibliografía

In the Americas, different disease symptoms have been reported in cassava including leaf mosaics, vein clearings, mottles, ring spots, leaf distortions and undeveloped and deformed storage roots. Some viruses have been identified and associated with these symptoms while others have been reported in symptomless plants or latent infections. We observed that reoviruses associated with severe root symptoms (RS) of Cassava Frogskin Disease (CFSD) are not associated with leaf symptoms (LS) observed in the cassava indicator plant ‘Secundina’. Neither were these LS associated with the previously characterized Cassava common mosaic virus, Cassava virus X, Cassava vein mosaic virus or phytoplasma, suggesting the presence of additional pathogens. In order to explain LS observed in cassava we used a combination of biological, serological and molecular tests. Here, we report three newly described viruses belonging to the families Secoviridae, Alphaflexiviridae and Luteoviridae found in cassava plants showing severe RS associated with CFSD. All tested plants were infected by a mix of viruses that induced distinct LS in ‘Secundina’. Out of the three newly described viruses, a member of family Secoviridae could experimentally induce LS in single infection. Our results confirm the common occurrence of complex viral infections in cassava field-collected since the 1980s.

Several potexviruses (Family Alphaflexiviridae) have been reported infecting cassava (Manihot esculenta Crantz) in the Americas. They were isolated from severely diseased plants during the last 30-40 years and include: Cassava common mosaic virus (CsCMV), Cassava Caribbean mosaic virus (CsCaMV), Cassava Colombian symptomless virus (CsCSV) and Cassava virus X (CsVX). However, their definitive classification as distinct species remains unresolved for several reasons, including the lack of sequence data and unavailability of samples from original isolates. This complicates disease diagnostics, cassava germplasm exchange certification, evaluation of virus cleaning protocols and epidemiological studies. Furthermore, a recently detected novel alphaflexivirus, indicates that cassava-infecting potexviruses may be more diverse. To solve the identity of these viruses, we started indexing samples from different parts of Colombia using different sets of PCR primers, antisera available and inoculation to indicator plants. Results show that there are three major phylogenetic groups of potexviruses infecting cassava, and they correspond to CsCMV, CsVX and the newly identified Cassava new alphaflexivirus (CsNAV). Bioassays and sequence analysis established that isolates of CsNAV and CsVX cause latent infections in different cassava landraces, they are not efficiently transmitted to the indicator plant Nicotiana benthamiana and they lack the gene 3 of the conserved potexviral 'triple gene block' (TGB). In contrast, all isolates of CsCMV (which have a characteristic potexvirus genome arrangement) caused Cassava Common Mosaic Disease (CCMD) in single infections and were efficiently transmitted to N. benthamiana. Although phylogenetic analysis of the replicase sequence placed CsNAV and CsVX as members of the Potexvirus genus, their distinct genome arrangement and biological characteristics suggest they can be considered as members of a separate taxonomic group.

Cassava Frogskin Disease (CFSD) can cause severe damage to cassava roots and is one of the most important diseases of cassava in Latin America. The principal objective of this study was to identify the causal agent of CFSD. Electron microscopy, viral purifications, double-stranded RNA (dsRNA) analysis, cloning, sequencing, rtPCR and hybridizations were carried out to characterize and associate a novel virus with the disease. Virus-like particles of 70 and 45 nm in diameter were found in affected cassava plants and partially purified preparations respectively. Nine species of dsRNA were associated with this disease and cDNA clones to six genomic segments were synthesized from the purified dsRNAs. The putative proteins predicted from the sequence of the cassava virus cDNA clones have similarity with the P1, P2, P3, P4, P5 and P10 proteins of Rice ragged stunt virus (RRSV). Phylogenic analysis confirmed that this virus is a member of the family Reoviridae and is most closed related to RRSV. Hybridization analyses of dsRNA identified S1, S2, S3, S4, S5 and S10 genomic segments in the CFSD-affected plants, but not in healthy controls. Additionally, 26 isolates were compared using a portion of the putative polymerase gene. The virus was detected in all 26 isolates, and they were classified into three distinct races. The association of this virus with CFSD was strengthened by the detection of this virus in diseased plants collected from different locations throughout Colombia.

Cassava (Manihot esculenta Crantz) is the third most important source of calories for human nutrition in the world. In Argentina, cassava is largely produced in the northeastern region, with Misiones Province accounting for the majority of the production for industrial purposes while in Corrientes and Formosa provinces, cassava is primarily grown for direct human consumption. Since cassava is vegetatively propagated, it is prone to buildup of virus infections, which are associated with severe root and leaf symptoms (Carvajal-Yepes et al. 2014) and significant yield reductions. Recent field surveys in Argentina have identified the presence of severe leaf mosaic symptoms in local cassava varieties while historical virus indexing records of cassava plantlets maintained in vitro at the International Center for Tropical Agriculture (CIAT) indicate the presence of Cassava common mosaic virus (CsCMV; genus Potexvirus) in Argentinian accessions collected in 1993. To confirm the current presence of CsCMV in Argentina, a total of 19 samples were collected in 2012 and 2014 from the fields in Corrientes (Corrientes), El Colorado (Formosa), and Puerto Rico (Misiones), and assayed for CsCMV and other viruses reported in the Americas . These plants showed virus-like symptoms including leaf mosaic and leaf deformation. Plate trapped antigen (PTA)-ELISA tests readily detected CsCMV in 16 out of the 19 samples. Negative samples could be explained by low virus titers and/or the specificity of the antiserum used. Mechanical transmissions to experimental hosts induced the formation of characteristic symptoms previously described for CsCMV including systemic mild mosaic in Nicotiana benthamiana and N. occidentalis and local chlorotic lesions in Chenopodium quinoa and C. amaranticolor. RT-PCR confirmed the presence of CsCMV in the originally collected cassava samples and detected a mixed infection with Cassava frogskin-associated virus (CsFSaV; tentative genus Oryzavirus) in one plant. PCR products from three independent CsCMV-positive samples were cloned into plasmid vectors and sequenced using standard procedures. Sequence analysis of the replicase region of CsCMV obtained using universal potexvirus primers (GenBank Accession No. KP025969) showed a nucleotide identity of 87 and 92% with two Brazilian isolates sequences available in GenBank (U23414 and JF913280, respectively). For CsFSaV, sequence analysis of a conserved region (958 bp) of the segment 4 encoding the replicase gene (KJ742699) detected a nucleotide identity of 88 to 99% with Colombian and Brazilian isolates. Symptoms caused by CsCMV in single infection can reduce yields significantly and although no obvious difference in symptoms was observed in the mixed infected plant detected in this study, the inadvertent accumulation and propagation of additional virus infections could dramatically affect the growing cassava industry in Argentina as it has been occurring in other countries. Fil: Di Feo, Liliana del Valle. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Instituto de Patología Vegetal; Argentina Fil: Zanini, Andrea Alejandra. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Instituto de Patología Vegetal; Argentina Fil: Rodríguez Pardina, Patricia Elsa. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Instituto de Patología Vegetal; Argentina Fil: Cuervo, M.. International Center For Tropical Agriculture; Colombia Fil: Carvajal Yepesand, M.. Centro Internacional de Agricultura Tropical; Colombia Fil: Cuellar, W. J.. Centro Internacional de Agricultura Tropical; Colombia

Rice hoja blanca virus (RHBV) is a major virus disease of economic importance affecting rice in northern South America, Central America and the Caribbean. This is the first report of transgenic resistance to RHBV and the transformation of an indica rice variety from Latin America. Rice transformed with the RHBV nucleocapsid protein ( N) gene had a significant reduction in disease development. Several reactions were observed that ranged from susceptible to completely resistant plants (immunity). The resistant reactions were characterized by the production of local lesions like a hypersensitive reaction or a recovery phenotype with the emergence of symptom-less new leaves. These transgenic RHBV-resistant rice lines expressed the N gene RNA at low levels that were below the detection limit by Northern blots and only resolved by RT-PCR. The nucleocapsid protein could not be detected in any of the transgenic plants either by Western or ELISA tests. These results suggest that the resistance encoded by the N gene in these plants appears to be mediated by RNA. When challenged with RHBV, the resistant transgenic lines showed a significant increased performance for important agronomic traits including the number of tillers, the number of grains per plant and the yield as compared to the susceptible control. Furthermore, upon inoculation some of the most-resistant transgenic lines showed agronomic traits similar to the uninoculated non-transgenic Cica 8 control. Using both agronomic traits and disease severity as criteria, several of the most-resistant lines were followed through the R(4) generation and demonstrated that the N gene and RHBV resistance was inherited in a stable manner. These transgenic rice lines could become a new genetic resource in developing RHBV-resistant cultivars.

Morphology of the third antennal segment and compound eyes of adults of the whitefly species Bemisia tabaci (Gennadius), B. argentifolii Bellows & Perring, B. tuberculata (Bondar), Trialeurodes vaporariorum (Westwood), T. variablilis (Quaintance), and Aleurotrachelus socialis (Bondar) were studied using scanning electron microscopy to aid in identification of adult whiteflies in cassava and beans in Colombia. Random amplified polymorphic DNA polymerase chain reaction markers proved complementary to the morphological identification of whitefly species and the only rapid method to distinguish individuals in the Bemisia tabaci species complex. From each species of whitefly, a region of the mitochondrial 16S rDNA gene was amplified, cloned and the sequence determined. Parsimony and distance analyses were performed and the results were similar to those based on morphology. The distance between the two species of Trialeurodes was greater than expected for two species within the same genus. The combination of morphological and molecular traits is useful in understanding the diversity and evolution of these whitefly species.