Detección de marcadores microsatélites asociados con la resistencia al añublo bacterial de la yuca (Manihot esculenta Crantz) en Colombia

Una de las principales estrategias para el manejo del Añublo Bacterial de la Yuca (CBB), causado por Xanthomonas axonopodis pv. manihotis es el uso de resistencia varietal, que implica desarrollar variedades de yuca con resistencia genética duradera. Para tal fin, es necesario conocer los genes que dominan la resistencia a la enfermedad, detectando inicialmente marcadores moleculares asociados con la respuesta fenotipica de la planta, siendo este el principal objetivo del presente estudio. Inicialmente, se evaluó la reacción a CBB de 4 familias de yuca BCI (retrocruce 1), se seleccionó la más segregante bajo presión natural de inóculo en Villavicencio (Meta, Colombia) y se confirmó la respuesta a CBB en condiciones de invernadero en CIAT (Palmira, Valle). La familia GM 315 presentó la mejor segregación, siendo la más adecuada para buscar asociación entre su reacción fenotípica y la presencia de un marcador molecular. Para esto, se evaluaron 486 cebadores microsatélites mediante análisis de grupos segregantes (BSA), encontrándose que 17 de ellos mostraron polimorfismo entre los grupos contrastantes y solo uno de ellos, el cebador SSRY 65, mostró diferencias significativas entre individuos resistentes y susceptibles. Al evaluar este cebador en toda la familia segregante se encontró asociación entre su presencia y los individuos evaluados fenotípicamente como resistentes en campo e invernadero, con una probabilidad mínima de P=0,OOl5 y P=0,OO7 respectivamente, en una prueba de Chicuadrado de independencia. Adicionalmente, a partir de los resultados obtenidos en el análisis estadístico, se calcularon los valores predictivos, especificidad y sensibilidad del marcador SSRY 65. Con base en los valores predictivos positivos generados, es posible sugerir la utilización de este marcador en pruebas diagnósticas para detectar la presencia de una banda específica en individuos resistentes de familias genéticamente relacionadas con la familia GM 315. = A major strategy for managing Cassava Bacterial blight (CBB), caused by Xanthomonas axonopodis pv. manihotis, is to use varietal resistance, that is, to develop cassava varieties with lasting genetic resistance. A search for the genes that dominate resistance to the disease was initially conducted by seeking the molecular markers associated with the plant s phenotypic response to CBB. The response in four BC1 (backcross 1) cassava families was accordingly evaluated under natural disease pressure at Villavicencio (Meta, Colombia). The most segregating family was then selected, and its response to CBB was verified under greenhouse conditions at CIAT (Palmira, Valle). Family GM 315 presented the best segregation, so, it was the most suitable for seeking association between its phenotypic reaction and the presence of a molecular marker. Of 486 microsatellite primers evaluated by bulked segregant analysis (BSA), 17 showed polymorphism among contrasting groups. Only one primer, SSRY 65, showed significant differences between resistant and susceptible individuals. On evaluating this primer for the entire segregating family, an association was found between its presence and individuals evaluated phenotypically as resistant in the field and greenhouse (minimum P = 0.0015 and P = 0.007, respectively, in a chi-square test of independence). With the results of the statistical analysis, the predictive values, specificity, and sensibility of marker SSRY 65 were calculated. The positive predictive values generated indicate that this marker can be used in diagnostic tests to detect the presence of a specific band in resistant individuals of families genetically related to the GM 315 family

Manejo de Moko de plátano en el Litoral Pacífico.

Desde 2001, el CIAT ha desarrollado estrategias de manejo de la enfermedad del Moko de plátano, causada por la bacteria Ralstonia solanacearum, como alternativa al uso de productos tóxicos, mediante la ejecución de diversas investigaciones en conjunto con entidades del estado y productores de plátano. Se encontró que aplicaciones al suelo de lixiviado de compost de raquis de plátano, extracto vegetal de flor de muerto (Tagetes patula), roca fosfórica o Calfos inhiben la bacteria entre 32% y 85%. Por otro lado, el fosfito de potasio, roca fosfórica, flor de muerto y lixiviado de compost de raquis de plátano, inhibieron completamente la bacteria in vitro. Además, con el uso de variedades tolerantes es posible reducir el impacto de la enfermedad. El presente artículo también describe los síntomas de la enfermedad y define estrategias para su prevención y manejo de focos de plantas afectadasEdicion N°

Identificación de genes análogos de resistencia a enfermedades en yuca, Manihot esculenta Crantz, y su relación con la resistencia a tres especies de Phytophthora

Los genes de resistencia se buscaron mediante dos estrategias. La primera, por medio de hibridización con sondas de maíz y arroz, utilizando RFLP. La segunda consistió en la amplificación de regiones conservadas de ADN, con cebadores degenerados NBS y Pto kinasa, en tres genotipos de yuca resistentes a Phytophthora tropicalis y P. palmivora, obteniendo clones que se secuenciaron y se homologaron con genes de resistencia conocidos. Con las secuencias se diseñaron cebadores específicos que permitieron amplificar regiones de ADN de los parentales e individuos resistentes y susceptibles. Las bandas se separaron mediante electroforesis en geles de poliacrilamida denaturantes y no denaturantes (SSCP - polimorfismo en la conformación de cadenas simples). Se identificaron cinco QTLs asociados con resistencia a Phytophthora. La yuca tuvo muy baja homología con los genes de maíz y arroz. Se obtuvieron 28 clones NBS y 2 Pto kinasa, de los cuales 5 mostraron secuencia homóloga con RGAs (genes análogos de resistencia) NBS-LRR y cuatro de ellos mostraron marco abierto de lectura con motivos conservados de la región NBS, y se consideraron como RGAs. Se identificaron tres clases de RGAs aunque no hubo evidencia de su asociación con resistencia a Phytophthora. = Identification of gene analogs for resistance to cassava (Manihot esculenta Crantz) Diseases, and their relationship to resistance to three Phytophthora species. Two strategies were used to find resistance genes in cassava. The first through hybridizing probes from maize and rice, using RFLP. The second strategy consisted of amplifying conserved regions of DNA, with degenerated NBS and Pto kinase primers, in three cassava genotypes resistant to Phytophthora tropicalis and P. palmivora, obtaining clones that were sequenced and compared with known resistance genes. Specific primers were designed from the sequences, allowing DNA regions of parental material and resistant and susceptible individuals, to be amplified. Bands were separated by denaturing polyacrylamide gel electrophoresis, and non-denaturing polyacrylamide gel (SSCP - single strand conformation polymorphism-). Five QTLs associated to Phytophthora spp resistance were identified. Cassava has a very low homology with the genes of the monocotyledons tested. A total of 28 NBS and 2 Pto kinase clones were obtained; of these, 5 showed homologous sequence with NBS-LRR RGAs (resistance gene analogs). Four of them had open reading frames with conserved motifs of the NBS region, and were considered as RGAs. Three different RGAs classes were identified. It remain to be shown if there are association to resistance to Phytophthora

Efecto de prácticas ecológicas sobre la población de Ralstonia solanacearum Smith, causante de moko de plátano

Se inoculó suelo estéril de Montenegro (Quindío, Colombia), con Ralstonia solanacearum Raza 2, bacteria causante de Moko de plátano, bajo condiciones de invernadero, para evaluar el efecto de prácticas no contaminantes alternativas al formol, sobre la población de esta bacteria. En un arreglo de bloques completos al azar, con cinco repeticiones, se probaron los siguientes tratamientos: incorporación de marigold o flor de muerto, Tagetes patula (1 Kg/m2), incorporación de calfos (0,5 Kg/m2), fertilizante Fulvan® líquido (20 L/m2) y lixiviado de compostaje de plátano (2,7 L/m2), los cuales se compararon con formol 20% (9,3 L/m2). Como controles se utilizó suelo inoculado y estéril, sin tratamiento. La unidad experimental consistió en dos materos con 250 mL de suelo. La población bacteriana se evaluó semanalmente durante 51 días después del tratamiento. Mediante la incorporación de marigold se logró reducir en 84,7 % la población de bacteria, mientras que formol la redujo 100 %, siendo esta diferencia no significativa (DMS ?=5 %). Además se lograron reducciones de 58,2 %, 50,80 % y 31,6 % con Fulvan®, calfos y lixiviado, respectivamente. Veinte días después de aplicados los tratamientos, marigold y Fulvan® fueron los más efectivos reduciendo la población hasta 80 % y 90 %, respectivamente. El calfos, por su baja solubilidad, tomó 20 días para actuar sobre la bacteria. Los resultados permitieron concluir que hay alternativas ecológicamente seguras y eficientes, para reducir la población del patógeno en el suelo. = Sterile soil of Montenegro (Quindío, Colombia) was inoculated with the bacterium Ralstonia solanacearum Race 2, causal agent of Moko or banana bacterial wilt in plantain, under greenhouse conditions to evaluate the effect of alternative non-polluting products other than formol on the population of this bacterium. The following treatments were evaluated in a random complete block design, with five replications, and results were compared with those obtained with the formol 20% treatment (9.3 L/m2): incorporation of marigold (Tagetes patula) at 1 kg/m2; incorporation of calfos (0.5 kg/m2); Fulvan® liquid fertilizer (20 L/m2); and plantain compost lixiviate (2.7 L/m2). Non-treated inoculated and sterile soil was used as controls. The experimental unit consisted of two pots with 250 mL of soil. After treatment, the bacterial population was evaluated weekly over a period of 51 days. The population of the bacterium was reduced by 84,7 % by incorporating marigold as compared with 100 % with the application of formol; this difference, however, was non-significant (DMS ?=5 %). Reductions of 58.2 %, 50.8 %, and 31.6 % were also achieved with the application of Fulvan®, calfos, and lixiviate, respectively. Marigold and Fulvan® were the most effective treatments, 20 days after application, reducing the bacterial population 80 % and 90%, respectively. Because its low solubility, calfos took 20 days to act on the bacterium. Based on the results, it concludes that there are safe and efficient ecological alternatives to reduce the population of this pathogen in the soil

Detection and molecular characterization of a phytoplasma associated with frogskin disease in Cassava.

none7Cassava frogskin disease (CFSD) is an economically important root disease of cassava (Manihot esculenta) in Colombia and other South American countries, including Brazil, Venezuela, Peru, Costa Rica, and Panama. The roots of severely affected plants are thin, making them unsuitable for consumption. In Colombia, phytoplasma infections were confirmed in 35 of 39 genotypes exhibiting mild or severe CFSD symptoms either by direct or nested polymerase chain reaction (PCR) assays employing ribosomal (r)RNA operon primer pairs. The CFSD-associated phytoplasmas were identified as group 16SrIII strains by restriction fragment length polymorphism (RFLP) and sequence analyses of amplified rDNA products, and results were corroborated by PCRs employing group 16SrIII-specific rRNA gene or ribosomal protein (rp) gene primers. Collectively, RFLP analyses indicated that CFSD strains differed from all phytoplasmas described previously in group 16SrIII and, on this basis, the strains were tentatively assigned to new ribosomal and ribosomal protein subgroups 16SrIII-L and rpIII-H, respectively. This is the first molecular identification of a phytoplasma associated with CFSD in cassava in Colombia.mixedAlvarez E.; J.F. Mejia; G.A. Llano; J.B. Loke; A. Calari; B. Duduk; A. BertacciniAlvarez E.; J.F. Mejia; G.A. Llano; J.B. Loke; A. Calari; B. Duduk; A. Bertaccin

Characterization of a phytoplasma associated with frogskin disease in cassava

Cassava frogskin disease (CFSD) is an economically important root disease of cassava (Manihot esculenta) in Colombia and other South American countries, including Brazil, Venezuela, Peru, Costa Rica, and Panama. The roots of severely affected plants are thin, making them unsuitable for consumption. In Colombia, phytoplasma infections were confirmed in 35 of 39 genotypes exhibiting mild or severe CFSD symptoms either by direct or nested polymerase chain reaction (PCR) assays employing ribosomal (r)RNA operon primer pairs. The CFSD-associated phytoplasmas were identified as group 16SrIII strains by restriction fragment length polymorphism (RFLP) and sequence analyses of amplified rDNA products, and results were corroborated by PCRs employing group 16SrIII-specific rRNA gene or ribosomal protein (rp) gene primers. Collectively, RFLP analyses indicated that CFSD strains differed from all phytoplasmas described previously in group 16SrIII and, on this basis, the strains were tentatively assigned to new ribosomal and ribosomal protein subgroups 16SrIII-L and rpIII-H, respectively. This is the first molecular identification of a phytoplasma associated with CFSD in cassava in Colombia