Supercoiled filamentous virus particles with lengths of 400 to 700 nm and 3 nm wide were isolated... more Supercoiled filamentous virus particles with lengths of 400 to 700 nm and 3 nm wide were isolated from leaves of Carica papaya L. plants showing a mild yellowing between the veins. The morphological properties of this virus resemble those of tenuiviruses. However, it was serologically unrelated to three of the five definitive members of this group of plant viruses and had biochemical features quite different from tenuiviruses. Therefore, the virus described here is possible an unreported new virus infecting papaya for which the name of papaya mild yellow leaf virus (PMYLV) is proposed. PMYLV was mechanically transmitted to papaya and to several Cucurbitaceae species. Virus particles sedimented as one component in sucrose density gradients, containing one molecule of ssRNA with an apparent size of 6400 nucleotides which constitutes 5% of the particle weight. The buoyant density of PMYLV was 1.26 g/cm 3 in cesium chloride equilibrium gradients, and the virus coat protein consisted of a single polypeptide with mol. wt. of c. 39 kDa. Estimated virus yield in purified preparations was 2.6 g/kg leaf tissues. An antiserum was produced with a titer of 1 : 1500. Ultrastructural observations of PMYLV-infected leaf tissues showed crystalline aggregates of virus particles, closely associated with electron dense amorphous inclusion bodies only within xylem cells. 892 E. Marys et al. present investigation was undertaken to analyze the physico-chemical properties and host range of isolated virus particles.
SUMMARY The development of a DAC-ELISA assay for the diagnosis of Rice hoja blanca virus (RHBV) i... more SUMMARY The development of a DAC-ELISA assay for the diagnosis of Rice hoja blanca virus (RHBV) in Venezuela is reported. The virus was isolated and partially characterized from infected rice samples collected in the states of Portuguesa and Guarico. Highly specific immunoglobulin (IgGs) to virus particles re acted in 1:1000 dilution, when tested against viral particles by immunoassay. According to serological data, the incidence of RHBV in the main rice growing states was low (11%), possibly due to the use of resistant rice lines by local farmers and/or to the low incidence of the virus vector, the insect Sogatodes orizicola. The diagnostic will allow phytosanitary certification of RHBV free material.
La Dra. Frieda Herold, es considerada por muchos fitopatólogos como la "madre" de la vi... more La Dra. Frieda Herold, es considerada por muchos fitopatólogos como la "madre" de la virología vegetal en Venezuela. Viróloga eminente de origen alemán, la Dra. Herold llega al país durante la década de los 50, prestada por la Fundación Alemana Iberoamericana, y ejerce como Investigadora Invitada en el Instituto Nacional de Higiene (INH), a razón de su experiencia en enfermedades virales tropicales humanas. Sin embargo, su interés principal era la botánica, especialmente los virus de plantas. Por tal motivo, fue convocada en 1959 a organizar un laboratorio de virología vegetal en el Instituto Venezolano de Investigaciones Científicas (IVIC) por su compatriota, el Dr. Gernot Bergold, quien tenía entonces la tarea de crear el Departamento de Virología del IVIC. En 1964 se inaugura el "Centro de Virología Louis Daniel Beauperthuy", construido y dotado con ayuda del estado alemán.
Venezuela is currently experiencing the most severe humanitarian crisis in the Americas of this c... more Venezuela is currently experiencing the most severe humanitarian crisis in the Americas of this century. Little food is being produced locally, despite the population's right to food. Plant disease outbreaks are causing substantial declines in major staple food and cash crops, and this impacts on rural livelihoods, and poses a significant and growing threat to the already complex food insecurity crisis in the country. Nonetheless, phytosanitary services and hence the control of plant pests and diseases have been substantially weakened over the recent years as a consequence of the collapse of the economy and the substantial deterioration of government services. Therefore, most of the pathogens associated with symptoms-causing diseases remain unidentified or uncharacterized, and no surveillance or crop protection strategies have been implemented. In this review, we address the country's issues and challenges in diagnosing, monitoring and managing plant diseases to restore nati...
In surveys of common bean (Phaseolus vulgaris L.), 819 sam- ples from plants showing virus-like s... more In surveys of common bean (Phaseolus vulgaris L.), 819 sam- ples from plants showing virus-like symptoms were collected throughout the main growing fields in Aragua state, Venezue- la. The samples were tested for Southern bean mosaic virus (SBMV) using indirect enzyme linked immunosorbent assay (ELISA). More than 30% of the tested plants were infected with the virus. Sequence analyses targeting the 5’-terminal region of a virus isolate showed 90-97% similarity to corresponding sequences of other SBMV isolates deposited in the GenBank database. This is the first study on the incidence of SBMV in Venezuela and on the partial molecular characterization of a Venezuelan isolate of SBMV.
Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alim... more Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alimentación del venezolano. Recientemente, en una parcela experimental de fabáceas en la Facultad de Agronomía-UCV, Maracay, se detectó en vainita china (Vigna unguiculata subsp. sesquipedalis) un virus, cuya caracterización biológica y molecular lo tipificó como un miembro del género Carlavirus. El objetivo de esta investigación fue evaluar la transmisión de este virus por la mosca blanca B. tabaci. Se utilizaron individuos adultos provenientes de una cría sana, mantenida en capacho (Canna indica); como fuente de inóculo se utilizaron plantas de vainita china de 20 días de edad infectadas con el virus. A los insectos se les permitió un período de acceso a la adquisición de 24 h y un período de acceso a la inoculación de 48 h. Fueron utilizadas 21 plantas de frijol ‘Tuy’ y 71 plantas de vainita china de 8 días de edad y 10 individuos/planta. Después de la inoculación, los insectos fueron el...
ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subs... more ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subspecies of cowpea, with apparent origin in East Asia. Yardlong bean is characterized by extremely long and thin pods, and is considered one of the top ten Asian vegetables. It is now grown extensively in Asia, Europe, Oceania and Americas. In 2012, yardlong bean plants showing severe yellowing were found in an experimental plot at the Agronomy Faculty, UCV, in Aragua State, Venezuela. The observed incidence of foliar yellowing symptoms, resembling those associated with the infection of a bromovirus, was 30%. The field samples with virus-like symptoms of yellowing were collected for further studies. Previously, we reported the presence of Cowpea mild mottle virus (CPMMV), a carlavirus with filamentous particles, in yard long bean in the country (Brito et al., 2012), but electron microscope observations using the leaf-dip method excluded CPMMV, since symptomatic leaves contained icosahedral virus-like particles, ca. 26-28 nm in diameter. Leaf extracts from field samples were mechanically inoculated onto V. unguiculata cv. Tuy, Vigna. radiata, Phaseolus vulgaris L. cv. Tacarigua, Chenopodium quinoa Willd., C. amaranticolor Coste et A. Reyn, Gomphrena globosa L. and Glycine max (L.) Merr. All the seven mechanically inoculated plant species were found susceptible to the virus. Among these, C. amaranticolor, C. quinoa and G. globosa developed chlorotic local lesions 10-14 days post inoculation (dpi), while V. radiata developed local necrotic lesions on inoculated leaves. Seedlings of V. unguiculata and G. max were found systemically infected and showed mottle and mosaic symptoms on upper non-inoculated leaves at 7 dpi. Extensive severe systemic chlorosis was developed in cowpea after 15 dpi. A virus isolate was propagated in V. unguiculata cv. Tuy, grown under greenhouse conditions, purified (de Carvalho et al., 2013), and tested by Western blot using polyclonal antisera specific to Cowpea mosaic virus (CPMV), Southern bean mosaic virus (SBMV), Broad bean mottle virus (BBMV), Cucumber mosaic virus (CMV) and Cowpea chlorotic mottle virus (CCMV). A conspicuous immunoreactive band with an approximate size of 19 kDa was obtained in purified virus sample against BBMV and CCMV antisera. Total RNA was extracted from 20 symptomatic plants and from purified virions, and tested by RT-PCR using degenerate primers Ilar1F5 (GCNGGWTGYGGDAARWCNAC) and Ilar1R7 (AMDGGWAYYTGYTNYGTRTCACC), specific to the detection of members of the genus Ilarvirus and family Bromoviridae (Untiveros et al., 2010). A PCR fragment of the expected size (ca. 300 bp) was amplified for all samples. Two amplicons were sequenced and the identical (100%) consensus sequences submitted to the GenBank database (NCBI, USA), with the accession numbers KJ810515 (from field yardlong bean) and KJ810516 (from purified virions). The deduced amino acid sequence contained 97 residues encoding part of the viral methyltransferase which was 93% identical to that of a CCMV methyltransferase (AAN37635.1). A nucleotide BLAST analysis of the sequence revealed 85% identity with CCMV strain T (AF325739). The level of sequence similarity with CCMV suggested that the virus from yardlong bean may be a distinct strain of this species. Phylogenetic analysis of the putative methyltransferase gene demonstrated that CCMV yardlong bean clustered separately form the known strains of CCMV. Because chrysomelids are known vectors of bromoviruses, virus-free Andrector ruficornis Olivier, A. arcuatus Olivier and Ledesmodina auricollis Lèfevre adults were exposed to symptomatic cowpea leaves for a 48-hour acquisition access period and then cage-confined with 10 healthy V. unguiculata cv. Tuy and P.vulgaris cv. Tacarigua each for a 48-hour inoculation access period. Symptoms were reproduced in all tested plants after 21-day period and CCMV infection was confirmed by RT-PCR assay. To our knowledge, this is the first evidence of the presence of CCMV in yardlong bean, and the first report of CCMV transmission by chrysomelid beetle L. auricollis. This is an important information for the Venezuelan cowpea’s market as CCMV-infected yardlong bean plants could act as virus source for secondary spread by beetle vectors. References: de Carvalho et al. Tropical Plant Pathology 38(1): 49-54, 2013., Untiveros et al., J. Virol. Methods 165: 97-104, 2010., Brito et al., Viruses 4: 3804-3811, 2012.
Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alim... more Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alimentación del venezolano. Recientemente, en una parcela experimental de fabáceas en la Facultad de Agronomía-UCV, Maracay, se detectó en vainita china (Vigna unguiculata subsp. sesquipedalis) un virus, cuya caracterización biológica y molecular lo tipificó como un miembro del género Carlavirus. El objetivo de esta investigación fue evaluar la transmisión de este virus por la mosca blanca B. tabaci. Se utilizaron individuos adultos provenientes de una cría sana, mantenida en capacho (Canna indica); como fuente de inóculo se utilizaron plantas de vainita china de 20 días de edad infectadas con el virus. A los insectos se les permitió un período de acceso a la adquisición de 24 h y un período de acceso a la inoculación de 48 h. Fueron utilizadas 21 plantas de frijol ‘Tuy’ y 71 plantas de vainita china de 8 días de edad y 10 individuos/planta. Después de la inoculación, los insectos fueron el...
ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subs... more ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subspecies of cowpea, with apparent origin in East Asia. Yardlong bean is characterized by extremely long and thin pods, and is considered one of the top ten Asian vegetables. It is now grown extensively in Asia, Europe, Oceania and Americas. In 2012, yardlong bean plants showing severe yellowing were found in an experimental plot at the Agronomy Faculty, UCV, in Aragua State, Venezuela. The observed incidence of foliar yellowing symptoms, resembling those associated with the infection of a bromovirus, was 30%. The field samples with virus-like symptoms of yellowing were collected for further studies. Previously, we reported the presence of Cowpea mild mottle virus (CPMMV), a carlavirus with filamentous particles, in yard long bean in the country (Brito et al., 2012), but electron microscope observations using the leaf-dip method excluded CPMMV, since symptomatic leaves contained icosahedral virus-like particles, ca. 26-28 nm in diameter. Leaf extracts from field samples were mechanically inoculated onto V. unguiculata cv. Tuy, Vigna. radiata, Phaseolus vulgaris L. cv. Tacarigua, Chenopodium quinoa Willd., C. amaranticolor Coste et A. Reyn, Gomphrena globosa L. and Glycine max (L.) Merr. All the seven mechanically inoculated plant species were found susceptible to the virus. Among these, C. amaranticolor, C. quinoa and G. globosa developed chlorotic local lesions 10-14 days post inoculation (dpi), while V. radiata developed local necrotic lesions on inoculated leaves. Seedlings of V. unguiculata and G. max were found systemically infected and showed mottle and mosaic symptoms on upper non-inoculated leaves at 7 dpi. Extensive severe systemic chlorosis was developed in cowpea after 15 dpi. A virus isolate was propagated in V. unguiculata cv. Tuy, grown under greenhouse conditions, purified (de Carvalho et al., 2013), and tested by Western blot using polyclonal antisera specific to Cowpea mosaic virus (CPMV), Southern bean mosaic virus (SBMV), Broad bean mottle virus (BBMV), Cucumber mosaic virus (CMV) and Cowpea chlorotic mottle virus (CCMV). A conspicuous immunoreactive band with an approximate size of 19 kDa was obtained in purified virus sample against BBMV and CCMV antisera. Total RNA was extracted from 20 symptomatic plants and from purified virions, and tested by RT-PCR using degenerate primers Ilar1F5 (GCNGGWTGYGGDAARWCNAC) and Ilar1R7 (AMDGGWAYYTGYTNYGTRTCACC), specific to the detection of members of the genus Ilarvirus and family Bromoviridae (Untiveros et al., 2010). A PCR fragment of the expected size (ca. 300 bp) was amplified for all samples. Two amplicons were sequenced and the identical (100%) consensus sequences submitted to the GenBank database (NCBI, USA), with the accession numbers KJ810515 (from field yardlong bean) and KJ810516 (from purified virions). The deduced amino acid sequence contained 97 residues encoding part of the viral methyltransferase which was 93% identical to that of a CCMV methyltransferase (AAN37635.1). A nucleotide BLAST analysis of the sequence revealed 85% identity with CCMV strain T (AF325739). The level of sequence similarity with CCMV suggested that the virus from yardlong bean may be a distinct strain of this species. Phylogenetic analysis of the putative methyltransferase gene demonstrated that CCMV yardlong bean clustered separately form the known strains of CCMV. Because chrysomelids are known vectors of bromoviruses, virus-free Andrector ruficornis Olivier, A. arcuatus Olivier and Ledesmodina auricollis Lèfevre adults were exposed to symptomatic cowpea leaves for a 48-hour acquisition access period and then cage-confined with 10 healthy V. unguiculata cv. Tuy and P.vulgaris cv. Tacarigua each for a 48-hour inoculation access period. Symptoms were reproduced in all tested plants after 21-day period and CCMV infection was confirmed by RT-PCR assay. To our knowledge, this is the first evidence of the presence of CCMV in yardlong bean, and the first report of CCMV transmission by chrysomelid beetle L. auricollis. This is an important information for the Venezuelan cowpea’s market as CCMV-infected yardlong bean plants could act as virus source for secondary spread by beetle vectors. References: de Carvalho et al. Tropical Plant Pathology 38(1): 49-54, 2013., Untiveros et al., J. Virol. Methods 165: 97-104, 2010., Brito et al., Viruses 4: 3804-3811, 2012.
Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling a... more Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling and leaf distortion were observed in both experimental and commercial fields in Aragua State, Venezuela. Symptomatic leaves were shown to contain carlavirus-like particles. RT-PCR analysis with carlavirus-specific primers was positive in all tested samples. Nucleotide sequences of the obtained amplicons showed 84%-74% similarity to corresponding sequences of Cowpea mild mottle virus (CPMMV) isolates deposited in the GenBank database. This is the first report of CPMMV in Venezuela and is thought to be the first report of CPMMV infecting yardlong bean.
Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowe... more Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowers of the world, with great commercial value. Since 1998, Venezuela began a floral industry to produce and export fresh cut gerbera and chrysanthemum, with 40% of nurseries concentrated in Altos Mirandinos (Miranda State, north central region of the country). For the past 2 years, greenhouse-grown gerbera and chrysanthemum have been observed displaying symptoms resembling those associated with tospoviruses. Symptomatic plants showed concentric rings, irregular chlorotic blotches, and deformation on leaves. Disease incidence was estimated at 30%. Mechanical inoculation with extracts of symptomatic leaves reproduced the typical concentric ring symptoms on indicator plants Arachis hypogaea L. cv. San Martín, Capsicum chinense, and G. jamesonii 6 to 15 days after inoculation. In initial tests, leaves from each 30 symptomatic gerbera and chrysanthemum species from several greenhouse facilitie...
Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowe... more Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowers of the world, with great commercial value. Since 1998, Venezuela began a floral industry to produce and export fresh cut gerbera and chrysanthemum, with 40% of nurseries concentrated in Altos Mirandinos (Miranda State, north central region of the country). For the past 2 years, greenhouse-grown gerbera and chrysanthemum have been observed displaying symptoms resembling those associated with tospoviruses. Symptomatic plants showed concentric rings, irregular chlorotic blotches, and deformation on leaves. Disease incidence was estimated at 30%. Mechanical inoculation with extracts of symptomatic leaves reproduced the typical concentric ring symptoms on indicator plants Arachis hypogaea L. cv. San Martín, Capsicum chinense, and G. jamesonii 6 to 15 days after inoculation. In initial tests, leaves from each 30 symptomatic gerbera and chrysanthemum species from several greenhouse facilitie...
Huanglongbing (HLB), is the most devastating citrus disease in the world. Two of the three known ... more Huanglongbing (HLB), is the most devastating citrus disease in the world. Two of the three known HLB-associated (Candidatus Liberibacter spp.) species are present in the Americas. Among them, 'Ca. L. asiaticus' (CLas) was detected in Venezuela in 2017, although early characteristics HLB symptoms such as yellow shoots were first noticed in 2016. In this study, plant material from suspect citrus and from knowing HLB hosts, were collected in 17 municipalities in the states of Aragua, Carabobo, Yaracuy and Portuguesa (Venezuela) between August and November 2017, to determine CLas relative incidence and host range. Samples were then evaluated by PCR, using primers targeting the β-operon locus of ribosomal proteins. Out of 113 HLB-symptomatic citrus plants, 87 (76.9 %) were positive for CLas. Citrofortunella microcarpa, Swinglea glutinosa, and Murraya paniculata known host plants of the vector, the Asian citrus psyllid Diaphorina citri Kuwayama, were also found infected in all regions analyzed. Further spread of the pathogen is expected, given the nationwide distribution and abundance of the vector. Given the HLB-infection rates found in this study, additional surveys in eastern and western states are recommended to evaluate the spread and severity of the disease which will greatly assist the design of efficient management strategies based on research data.
The genetic variation of papaya ringspot virus (PRSV) in Venezuela was estimated by single strand... more The genetic variation of papaya ringspot virus (PRSV) in Venezuela was estimated by single strand conformation and nucleotide sequence analyses of two genomic regions of twenty-six isolates. These analyses showed that mutation, virus movement, selection, mixed infections and recombination contributed to shape the genetic variation observed. Phylogenetic analysis indicated that Venezuelan isolates were within a clade composed of isolates from the Americas and Australia. The genetic diversity of these isolates was sufficiently large that it must be taken into account when designing control strategies such as transgenic resistance and cross-protection.
Supercoiled filamentous virus particles with lengths of 400 to 700 nm and 3 nm wide were isolated... more Supercoiled filamentous virus particles with lengths of 400 to 700 nm and 3 nm wide were isolated from leaves of Carica papaya L. plants showing a mild yellowing between the veins. The morphological properties of this virus resemble those of tenuiviruses. However, it was serologically unrelated to three of the five definitive members of this group of plant viruses and had biochemical features quite different from tenuiviruses. Therefore, the virus described here is possible an unreported new virus infecting papaya for which the name of papaya mild yellow leaf virus (PMYLV) is proposed. PMYLV was mechanically transmitted to papaya and to several Cucurbitaceae species. Virus particles sedimented as one component in sucrose density gradients, containing one molecule of ssRNA with an apparent size of 6400 nucleotides which constitutes 5% of the particle weight. The buoyant density of PMYLV was 1.26 g/cm 3 in cesium chloride equilibrium gradients, and the virus coat protein consisted of a single polypeptide with mol. wt. of c. 39 kDa. Estimated virus yield in purified preparations was 2.6 g/kg leaf tissues. An antiserum was produced with a titer of 1 : 1500. Ultrastructural observations of PMYLV-infected leaf tissues showed crystalline aggregates of virus particles, closely associated with electron dense amorphous inclusion bodies only within xylem cells. 892 E. Marys et al. present investigation was undertaken to analyze the physico-chemical properties and host range of isolated virus particles.
SUMMARY The development of a DAC-ELISA assay for the diagnosis of Rice hoja blanca virus (RHBV) i... more SUMMARY The development of a DAC-ELISA assay for the diagnosis of Rice hoja blanca virus (RHBV) in Venezuela is reported. The virus was isolated and partially characterized from infected rice samples collected in the states of Portuguesa and Guarico. Highly specific immunoglobulin (IgGs) to virus particles re acted in 1:1000 dilution, when tested against viral particles by immunoassay. According to serological data, the incidence of RHBV in the main rice growing states was low (11%), possibly due to the use of resistant rice lines by local farmers and/or to the low incidence of the virus vector, the insect Sogatodes orizicola. The diagnostic will allow phytosanitary certification of RHBV free material.
La Dra. Frieda Herold, es considerada por muchos fitopatólogos como la "madre" de la vi... more La Dra. Frieda Herold, es considerada por muchos fitopatólogos como la "madre" de la virología vegetal en Venezuela. Viróloga eminente de origen alemán, la Dra. Herold llega al país durante la década de los 50, prestada por la Fundación Alemana Iberoamericana, y ejerce como Investigadora Invitada en el Instituto Nacional de Higiene (INH), a razón de su experiencia en enfermedades virales tropicales humanas. Sin embargo, su interés principal era la botánica, especialmente los virus de plantas. Por tal motivo, fue convocada en 1959 a organizar un laboratorio de virología vegetal en el Instituto Venezolano de Investigaciones Científicas (IVIC) por su compatriota, el Dr. Gernot Bergold, quien tenía entonces la tarea de crear el Departamento de Virología del IVIC. En 1964 se inaugura el "Centro de Virología Louis Daniel Beauperthuy", construido y dotado con ayuda del estado alemán.
Venezuela is currently experiencing the most severe humanitarian crisis in the Americas of this c... more Venezuela is currently experiencing the most severe humanitarian crisis in the Americas of this century. Little food is being produced locally, despite the population's right to food. Plant disease outbreaks are causing substantial declines in major staple food and cash crops, and this impacts on rural livelihoods, and poses a significant and growing threat to the already complex food insecurity crisis in the country. Nonetheless, phytosanitary services and hence the control of plant pests and diseases have been substantially weakened over the recent years as a consequence of the collapse of the economy and the substantial deterioration of government services. Therefore, most of the pathogens associated with symptoms-causing diseases remain unidentified or uncharacterized, and no surveillance or crop protection strategies have been implemented. In this review, we address the country's issues and challenges in diagnosing, monitoring and managing plant diseases to restore nati...
In surveys of common bean (Phaseolus vulgaris L.), 819 sam- ples from plants showing virus-like s... more In surveys of common bean (Phaseolus vulgaris L.), 819 sam- ples from plants showing virus-like symptoms were collected throughout the main growing fields in Aragua state, Venezue- la. The samples were tested for Southern bean mosaic virus (SBMV) using indirect enzyme linked immunosorbent assay (ELISA). More than 30% of the tested plants were infected with the virus. Sequence analyses targeting the 5’-terminal region of a virus isolate showed 90-97% similarity to corresponding sequences of other SBMV isolates deposited in the GenBank database. This is the first study on the incidence of SBMV in Venezuela and on the partial molecular characterization of a Venezuelan isolate of SBMV.
Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alim... more Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alimentación del venezolano. Recientemente, en una parcela experimental de fabáceas en la Facultad de Agronomía-UCV, Maracay, se detectó en vainita china (Vigna unguiculata subsp. sesquipedalis) un virus, cuya caracterización biológica y molecular lo tipificó como un miembro del género Carlavirus. El objetivo de esta investigación fue evaluar la transmisión de este virus por la mosca blanca B. tabaci. Se utilizaron individuos adultos provenientes de una cría sana, mantenida en capacho (Canna indica); como fuente de inóculo se utilizaron plantas de vainita china de 20 días de edad infectadas con el virus. A los insectos se les permitió un período de acceso a la adquisición de 24 h y un período de acceso a la inoculación de 48 h. Fueron utilizadas 21 plantas de frijol ‘Tuy’ y 71 plantas de vainita china de 8 días de edad y 10 individuos/planta. Después de la inoculación, los insectos fueron el...
ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subs... more ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subspecies of cowpea, with apparent origin in East Asia. Yardlong bean is characterized by extremely long and thin pods, and is considered one of the top ten Asian vegetables. It is now grown extensively in Asia, Europe, Oceania and Americas. In 2012, yardlong bean plants showing severe yellowing were found in an experimental plot at the Agronomy Faculty, UCV, in Aragua State, Venezuela. The observed incidence of foliar yellowing symptoms, resembling those associated with the infection of a bromovirus, was 30%. The field samples with virus-like symptoms of yellowing were collected for further studies. Previously, we reported the presence of Cowpea mild mottle virus (CPMMV), a carlavirus with filamentous particles, in yard long bean in the country (Brito et al., 2012), but electron microscope observations using the leaf-dip method excluded CPMMV, since symptomatic leaves contained icosahedral virus-like particles, ca. 26-28 nm in diameter. Leaf extracts from field samples were mechanically inoculated onto V. unguiculata cv. Tuy, Vigna. radiata, Phaseolus vulgaris L. cv. Tacarigua, Chenopodium quinoa Willd., C. amaranticolor Coste et A. Reyn, Gomphrena globosa L. and Glycine max (L.) Merr. All the seven mechanically inoculated plant species were found susceptible to the virus. Among these, C. amaranticolor, C. quinoa and G. globosa developed chlorotic local lesions 10-14 days post inoculation (dpi), while V. radiata developed local necrotic lesions on inoculated leaves. Seedlings of V. unguiculata and G. max were found systemically infected and showed mottle and mosaic symptoms on upper non-inoculated leaves at 7 dpi. Extensive severe systemic chlorosis was developed in cowpea after 15 dpi. A virus isolate was propagated in V. unguiculata cv. Tuy, grown under greenhouse conditions, purified (de Carvalho et al., 2013), and tested by Western blot using polyclonal antisera specific to Cowpea mosaic virus (CPMV), Southern bean mosaic virus (SBMV), Broad bean mottle virus (BBMV), Cucumber mosaic virus (CMV) and Cowpea chlorotic mottle virus (CCMV). A conspicuous immunoreactive band with an approximate size of 19 kDa was obtained in purified virus sample against BBMV and CCMV antisera. Total RNA was extracted from 20 symptomatic plants and from purified virions, and tested by RT-PCR using degenerate primers Ilar1F5 (GCNGGWTGYGGDAARWCNAC) and Ilar1R7 (AMDGGWAYYTGYTNYGTRTCACC), specific to the detection of members of the genus Ilarvirus and family Bromoviridae (Untiveros et al., 2010). A PCR fragment of the expected size (ca. 300 bp) was amplified for all samples. Two amplicons were sequenced and the identical (100%) consensus sequences submitted to the GenBank database (NCBI, USA), with the accession numbers KJ810515 (from field yardlong bean) and KJ810516 (from purified virions). The deduced amino acid sequence contained 97 residues encoding part of the viral methyltransferase which was 93% identical to that of a CCMV methyltransferase (AAN37635.1). A nucleotide BLAST analysis of the sequence revealed 85% identity with CCMV strain T (AF325739). The level of sequence similarity with CCMV suggested that the virus from yardlong bean may be a distinct strain of this species. Phylogenetic analysis of the putative methyltransferase gene demonstrated that CCMV yardlong bean clustered separately form the known strains of CCMV. Because chrysomelids are known vectors of bromoviruses, virus-free Andrector ruficornis Olivier, A. arcuatus Olivier and Ledesmodina auricollis Lèfevre adults were exposed to symptomatic cowpea leaves for a 48-hour acquisition access period and then cage-confined with 10 healthy V. unguiculata cv. Tuy and P.vulgaris cv. Tacarigua each for a 48-hour inoculation access period. Symptoms were reproduced in all tested plants after 21-day period and CCMV infection was confirmed by RT-PCR assay. To our knowledge, this is the first evidence of the presence of CCMV in yardlong bean, and the first report of CCMV transmission by chrysomelid beetle L. auricollis. This is an important information for the Venezuelan cowpea’s market as CCMV-infected yardlong bean plants could act as virus source for secondary spread by beetle vectors. References: de Carvalho et al. Tropical Plant Pathology 38(1): 49-54, 2013., Untiveros et al., J. Virol. Methods 165: 97-104, 2010., Brito et al., Viruses 4: 3804-3811, 2012.
Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alim... more Las fabáceas constituyen una fuente de proteínas, carbohidratos y minerales importante en la alimentación del venezolano. Recientemente, en una parcela experimental de fabáceas en la Facultad de Agronomía-UCV, Maracay, se detectó en vainita china (Vigna unguiculata subsp. sesquipedalis) un virus, cuya caracterización biológica y molecular lo tipificó como un miembro del género Carlavirus. El objetivo de esta investigación fue evaluar la transmisión de este virus por la mosca blanca B. tabaci. Se utilizaron individuos adultos provenientes de una cría sana, mantenida en capacho (Canna indica); como fuente de inóculo se utilizaron plantas de vainita china de 20 días de edad infectadas con el virus. A los insectos se les permitió un período de acceso a la adquisición de 24 h y un período de acceso a la inoculación de 48 h. Fueron utilizadas 21 plantas de frijol ‘Tuy’ y 71 plantas de vainita china de 8 días de edad y 10 individuos/planta. Después de la inoculación, los insectos fueron el...
ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subs... more ABSTRACT Yardlong bean [Vigna unguiculata subsp. sesquipedalis (L.) Verdc.] is a distinctive subspecies of cowpea, with apparent origin in East Asia. Yardlong bean is characterized by extremely long and thin pods, and is considered one of the top ten Asian vegetables. It is now grown extensively in Asia, Europe, Oceania and Americas. In 2012, yardlong bean plants showing severe yellowing were found in an experimental plot at the Agronomy Faculty, UCV, in Aragua State, Venezuela. The observed incidence of foliar yellowing symptoms, resembling those associated with the infection of a bromovirus, was 30%. The field samples with virus-like symptoms of yellowing were collected for further studies. Previously, we reported the presence of Cowpea mild mottle virus (CPMMV), a carlavirus with filamentous particles, in yard long bean in the country (Brito et al., 2012), but electron microscope observations using the leaf-dip method excluded CPMMV, since symptomatic leaves contained icosahedral virus-like particles, ca. 26-28 nm in diameter. Leaf extracts from field samples were mechanically inoculated onto V. unguiculata cv. Tuy, Vigna. radiata, Phaseolus vulgaris L. cv. Tacarigua, Chenopodium quinoa Willd., C. amaranticolor Coste et A. Reyn, Gomphrena globosa L. and Glycine max (L.) Merr. All the seven mechanically inoculated plant species were found susceptible to the virus. Among these, C. amaranticolor, C. quinoa and G. globosa developed chlorotic local lesions 10-14 days post inoculation (dpi), while V. radiata developed local necrotic lesions on inoculated leaves. Seedlings of V. unguiculata and G. max were found systemically infected and showed mottle and mosaic symptoms on upper non-inoculated leaves at 7 dpi. Extensive severe systemic chlorosis was developed in cowpea after 15 dpi. A virus isolate was propagated in V. unguiculata cv. Tuy, grown under greenhouse conditions, purified (de Carvalho et al., 2013), and tested by Western blot using polyclonal antisera specific to Cowpea mosaic virus (CPMV), Southern bean mosaic virus (SBMV), Broad bean mottle virus (BBMV), Cucumber mosaic virus (CMV) and Cowpea chlorotic mottle virus (CCMV). A conspicuous immunoreactive band with an approximate size of 19 kDa was obtained in purified virus sample against BBMV and CCMV antisera. Total RNA was extracted from 20 symptomatic plants and from purified virions, and tested by RT-PCR using degenerate primers Ilar1F5 (GCNGGWTGYGGDAARWCNAC) and Ilar1R7 (AMDGGWAYYTGYTNYGTRTCACC), specific to the detection of members of the genus Ilarvirus and family Bromoviridae (Untiveros et al., 2010). A PCR fragment of the expected size (ca. 300 bp) was amplified for all samples. Two amplicons were sequenced and the identical (100%) consensus sequences submitted to the GenBank database (NCBI, USA), with the accession numbers KJ810515 (from field yardlong bean) and KJ810516 (from purified virions). The deduced amino acid sequence contained 97 residues encoding part of the viral methyltransferase which was 93% identical to that of a CCMV methyltransferase (AAN37635.1). A nucleotide BLAST analysis of the sequence revealed 85% identity with CCMV strain T (AF325739). The level of sequence similarity with CCMV suggested that the virus from yardlong bean may be a distinct strain of this species. Phylogenetic analysis of the putative methyltransferase gene demonstrated that CCMV yardlong bean clustered separately form the known strains of CCMV. Because chrysomelids are known vectors of bromoviruses, virus-free Andrector ruficornis Olivier, A. arcuatus Olivier and Ledesmodina auricollis Lèfevre adults were exposed to symptomatic cowpea leaves for a 48-hour acquisition access period and then cage-confined with 10 healthy V. unguiculata cv. Tuy and P.vulgaris cv. Tacarigua each for a 48-hour inoculation access period. Symptoms were reproduced in all tested plants after 21-day period and CCMV infection was confirmed by RT-PCR assay. To our knowledge, this is the first evidence of the presence of CCMV in yardlong bean, and the first report of CCMV transmission by chrysomelid beetle L. auricollis. This is an important information for the Venezuelan cowpea’s market as CCMV-infected yardlong bean plants could act as virus source for secondary spread by beetle vectors. References: de Carvalho et al. Tropical Plant Pathology 38(1): 49-54, 2013., Untiveros et al., J. Virol. Methods 165: 97-104, 2010., Brito et al., Viruses 4: 3804-3811, 2012.
Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling a... more Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling and leaf distortion were observed in both experimental and commercial fields in Aragua State, Venezuela. Symptomatic leaves were shown to contain carlavirus-like particles. RT-PCR analysis with carlavirus-specific primers was positive in all tested samples. Nucleotide sequences of the obtained amplicons showed 84%-74% similarity to corresponding sequences of Cowpea mild mottle virus (CPMMV) isolates deposited in the GenBank database. This is the first report of CPMMV in Venezuela and is thought to be the first report of CPMMV infecting yardlong bean.
Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowe... more Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowers of the world, with great commercial value. Since 1998, Venezuela began a floral industry to produce and export fresh cut gerbera and chrysanthemum, with 40% of nurseries concentrated in Altos Mirandinos (Miranda State, north central region of the country). For the past 2 years, greenhouse-grown gerbera and chrysanthemum have been observed displaying symptoms resembling those associated with tospoviruses. Symptomatic plants showed concentric rings, irregular chlorotic blotches, and deformation on leaves. Disease incidence was estimated at 30%. Mechanical inoculation with extracts of symptomatic leaves reproduced the typical concentric ring symptoms on indicator plants Arachis hypogaea L. cv. San Martín, Capsicum chinense, and G. jamesonii 6 to 15 days after inoculation. In initial tests, leaves from each 30 symptomatic gerbera and chrysanthemum species from several greenhouse facilitie...
Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowe... more Gerbera (Gerbera jamesonii) and Chrysanthemum (family Asteraceae) are two of the top 10 cut flowers of the world, with great commercial value. Since 1998, Venezuela began a floral industry to produce and export fresh cut gerbera and chrysanthemum, with 40% of nurseries concentrated in Altos Mirandinos (Miranda State, north central region of the country). For the past 2 years, greenhouse-grown gerbera and chrysanthemum have been observed displaying symptoms resembling those associated with tospoviruses. Symptomatic plants showed concentric rings, irregular chlorotic blotches, and deformation on leaves. Disease incidence was estimated at 30%. Mechanical inoculation with extracts of symptomatic leaves reproduced the typical concentric ring symptoms on indicator plants Arachis hypogaea L. cv. San Martín, Capsicum chinense, and G. jamesonii 6 to 15 days after inoculation. In initial tests, leaves from each 30 symptomatic gerbera and chrysanthemum species from several greenhouse facilitie...
Huanglongbing (HLB), is the most devastating citrus disease in the world. Two of the three known ... more Huanglongbing (HLB), is the most devastating citrus disease in the world. Two of the three known HLB-associated (Candidatus Liberibacter spp.) species are present in the Americas. Among them, 'Ca. L. asiaticus' (CLas) was detected in Venezuela in 2017, although early characteristics HLB symptoms such as yellow shoots were first noticed in 2016. In this study, plant material from suspect citrus and from knowing HLB hosts, were collected in 17 municipalities in the states of Aragua, Carabobo, Yaracuy and Portuguesa (Venezuela) between August and November 2017, to determine CLas relative incidence and host range. Samples were then evaluated by PCR, using primers targeting the β-operon locus of ribosomal proteins. Out of 113 HLB-symptomatic citrus plants, 87 (76.9 %) were positive for CLas. Citrofortunella microcarpa, Swinglea glutinosa, and Murraya paniculata known host plants of the vector, the Asian citrus psyllid Diaphorina citri Kuwayama, were also found infected in all regions analyzed. Further spread of the pathogen is expected, given the nationwide distribution and abundance of the vector. Given the HLB-infection rates found in this study, additional surveys in eastern and western states are recommended to evaluate the spread and severity of the disease which will greatly assist the design of efficient management strategies based on research data.
The genetic variation of papaya ringspot virus (PRSV) in Venezuela was estimated by single strand... more The genetic variation of papaya ringspot virus (PRSV) in Venezuela was estimated by single strand conformation and nucleotide sequence analyses of two genomic regions of twenty-six isolates. These analyses showed that mutation, virus movement, selection, mixed infections and recombination contributed to shape the genetic variation observed. Phylogenetic analysis indicated that Venezuelan isolates were within a clade composed of isolates from the Americas and Australia. The genetic diversity of these isolates was sufficiently large that it must be taken into account when designing control strategies such as transgenic resistance and cross-protection.
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