Eficiencia de solubilización de fósforo de aislados nativos guatemaltecos de Pseudomonas fluorescens / Phosphorus solubilization efficiency of native Guatemalan isolates of Pseudomonas fluorescens

El fósforo (P) es un elemento esencial en la producción agrícola, pero debido a su compleja dinámica en el suelo, solo una pequeña cantidad es aprovechable para las plantas, ya que la mayoría del P se encuentra en formas insolubles, especialmente, en suelos Andisoles de origen volcánico. Los microorganismos con capacidad solubilizadora de fósforo (MSF) son una alternativa para transformar el P a formas solubles y aprovechables por las plantas; además de brindar múltiples beneficios ambientales. Este trabajo identificó y evaluó in vitro, aislados nativos de Pseudomonas fluorescens Mingula, obtenidos de regiones guatemaltecas con suelos Andisoles que limitan la producción agrícola por la alta fijación de P. Se realizaron cultivos in vitro de la bacteria en medio National Botanical Research Instituteís phosphate growth (NBRIP), con fosfato tricálcico Ca3(PO4)2 como fuente de P insoluble y se midió el índice de solubilización de fósforo (ISF). Un total de 35 aislados de P. fluorescensfueron identificados y confirmados por PCR específico. El análisis de relaciones genéticas con el marcador AFLP, mostró dos grupos: el grupo A incluyó a los aislados con ISF mayores a 1.75, mientras el grupo B incluyó a aquellos con ISF menor a 1.75. La comparación de ISF entre los aislados y departamentos, demostró diferencia estadísticamente significativa (p < .001), con el aislado Pf_33 como más eficiente. Debido al potencial de solubilización de los aislados nativos del grupo genético A (ISF > 1.75), estos se recomiendan para futuras investigaciones que determinen su respuesta a condiciones de campo y estrategias para el desarrollo de biofertilizantes.

Phosphorus (P) is an essential element in agricultural production, but due to its complex dynamics in the soil, only a tiny amount is usable by plants. This is because most P is in insoluble forms, especially in volcanic Andisol soils. Microorganisms with phosphorus solubilizing capacity (MSF) are an alternative for transforming P into soluble forms usable by plants and providing multiple environmental benefits. This research identified and evaluated in vitro native isolates of Pseudomonas fluorescens Mingula, obtained from Guatemalan regions with Andisol soils that limit agricultural production due to high P fixation. In vitro cultures of the bacteria were grown on the National Botanical Research Instituteís phosphate medium (NBRIP), with tricalcium phosphate Ca3(PO4)2 as a source of insoluble P, and We measured the phosphorus solubilization index (PSI). We identified and confirmed a total of 35 isolates of P. fluorescens by specific PCR. Using the AFLP marker, genetic relationship analysis showed two groups: group A included isolates with PSI greater than 1.75, while group B included those with FSI less than 1.75. Comparing of PSI between isolates and departments showed statistically significant dif-ferences (p < 0.001), respectively, with the Pf_33 isolate as the most efficient. Because of the high solubilization potential of the native isolates of genetic group A (FSI > 1.75), We recommend future research to determine their response to field conditions and strategies for biofertilizer development.

Diversidad genética de materiales nativos de aguacate guatemalteco a través del marcador molecular AFLP / Genetic diversity of native Guatemalan avocado through AFLP molecular marker

El aguacate es un cultivo de consumo a nivel mundial, y según teorías recientes, se sugiere a la región de la Sierra Nevada, en California, como centro de origen y, a Guatemala, como uno de los principales centros de domesticación. Mediante caracterizaciones morfológicas se ha reportado una alta diversidad genética en el país, pero debido al comportamiento de polinización cruzada e hibridaciones interraciales, no se ha podido detallar el estado genético actual de la especie. Sin embargo, los marcadores moleculares son útiles para este tipo de estudios al enfocarse en las diferencias a nivel del ADN. Este estudio analizó la diversidad genética del aguacate nativo guatemalteco de siete poblaciones geográficas con el marcador molecular AFLP. Los datos de estructura poblacional mostraron un alto grado de diversidad a nivel de individuos (Ht = 0.1933, Hw = 0.1872) y baja diferenciación entre poblaciones (Hb = 0.0061). Los resultados sugieren una alta tasa de migración que influye directamente en el grado de mezcla genética de los materiales analizados. El bajo índice de estructura poblacional apunta a un alto flujo genético entre las poblaciones, por lo que la especie no presenta mayor riesgo ante la deriva genética, minimizándose el riesgo de pérdida de alelos por fijación. Se sugiere el resguardado del recurso fitogénetico total y no únicamente de materiales promisorios, evitando así el riesgo de erosión genética de la especie y garantizando la permanencia de la diversidad genética, la cual será la base de futuros programas de mejoramiento.

Avocado is one of the most widely consumed crops worldwide and according to new theories, the Sierra Nevada region in California is suggested as the center of origin and Guatemala as one of the main domestication cen-ters. Through morphological characterizations, a high genetic diversity has been reported in the country, but due to the behavior of cross pollination and interracial hybridizations, it has not been possible to detail the current genetic status of the species. Molecular markers are useful for this type of study by focusing on differences at DNA level. This study analyzed the genetic diversity of the native Guatemalan avocado from seven geographic populations with AFLP molecular marker. Population structure data showed a high degree of diversity at the individual level (Ht = 0.1933, Hw = 0.1872) and low differentiation between populations (Hb = 0.0061). The results suggest a high rate of migration that directly influences the degree of genetic mixing of the analyzed materials. The low index of population structure points to a high genetic flow between populations, so that the species does not present a greater risk due to genetic drift, minimizing the risk of loss of alleles due to fixation. The protection of the total genetic resource is suggested, and not only of promising materials, thus avoiding the risk of genetic erosion of the species and guaranteeing the permanence of genetic diversity, which will be the basis of future breeding programs.

Detección de patógenos asociados a la enfermedad punta morada en los cultivos de papa y tomate en Guatemala / Detection of pathogens associated to potato purple top disease in potato and tomato crops in Guatemala

La punta morada es una enfermedad que afecta la producción de algunas especies de solanáceas como la papa y el tomate, causando enrollamiento en las puntas de las hojas con una marcada coloración morada, decaimiento temprano de la planta y en la papa se observa tuberización aérea. Como patógenos asociados a la enfermedad se consideran al fitoplasma BLTVA y la bacteria Candidatus Liberibacter solanacearum. Dada la similitud en la sin-tomatología foliar que generan ambos patógenos, es difícil precisar cuál de ellos está implicado en la enfermedad. En Guatemala, existen reportes de la sintomatología típica de punta morada en las principales zonas productoras de papa y tomate, desconociéndose el agente asociado. La investigación determinó cuál de los dos patógenos reportados está asociados a la enfermedad en 12 municipios productores de papa y/o tomate en el país. Se realizaron ampli-ficaciones de ADN con cebadores específicos para cada patógeno asociado a la enfermedad. Por la alta incidencia del fitoplasma BLTVA en las muestras de papa (73.9%), en comparación a C. Liberibacter solanacearum (26%), este es considerado como el patógeno asociado más importante en papa. En las muestras de tomate, la incidencia del fitoplasma BLTVA (29.8%) y C. Liberibacter solanacearum del (27.6%) fue similar. Además, sobresale el primer reporte de la detección del fitoplasma BLTVA afectando el cultivo de tomate en Guatemala. Se sugiere un monitoreo constante, mediante métodos moleculares, para un diagnóstico certero y establecer medidas de manejo de la enfermedad para evitar su diseminación hacia zonas aún no afectadas.

The potato purple top is a disease that affects the production of some solanaceous species such as potatoes and tomatoes, causing curl at the tips of the leaves with a marked purple coloration, early decay of the plant, and aerial tuberization is observed in the potato. BLTVA phytoplasma and Candidatus Liberibacter solanacearum are considered as pathogens associated with the disease. Given the similarity in foliar symptoms generated by both pathogens, it is difficult to determine which one is involved in the disease. There are reports of the typical potato purple top symptoms in the main potato and tomato producing areas in Guatemala, being unknown the associated agent. The research determined which of the two reported pathogens is associated with the disease in 12 potatoes and/or tomato producing areas in the country. We performed DNA amplification with specific primers for each disease-associated pathogen. Due to the high incidence of BLTVA phytoplasma in potato samples (73.9%), com-pared to C. liberibacter solanacearum (26%), this is considered the most important associated pathogen in potatoes. In tomato samples, the incidence of BLTVA phytoplasma (29.8%) and C. liberibacter solanacearum (27.6%) was similar. Besides, the first report of the detection of the BLTVA phytoplasma affecting tomato cultivation in Gua-temala stands out. Using molecular methods, constant monitoring is suggested for an accurate diagnosis and to establish management measures for the disease to prevent its spread to areas not yet affected.

Evaluación de la resistencia de genotipos de tomate frente a aislados de Phytophthora infestans provenientes de Guatemala / Evaluation of the resistance of tomato genotypes against isolates of Phytophthora infestans from Guatemala

El oomicete Phytophthora infestans (Mont) DeBary es el patógeno responsable de causar la enfermedad denominada comúnmente como tizón tardío. Dicho patógeno afecta cultivos de importancia económica para el país, entre ellos el tomate, cultivo en el cual puede generar pérdidas totales en la producción. Dada la agresividad del patógeno, los programas de mejoramiento desarrollan nuevos genotipos de tomate resistentes a esta enfermedad mediante la incorporación de genes de resistencia, como los genes Ph. Sin embargo, las nuevas cepas de P. infestans, producto de la recombinación genética, pueden sobrepasar la resistencia de los genotipos mejorados. En este estudio se evaluó la resistencia genética de 13 genotipos de tomate mejorados, ante cinco aislados de P. infestans obtenidos de un estudio previo. Mediante inoculaciones in vitro y con la variable de respuesta de área bajo la curva del progreso de la enfermedad (AUDPC), se determinó que existió diferencia estadísticamente significativa (p < 0.05) entre el comportamiento de la resistencia, la agresividad de los aislados del patógeno y la interacción entre ambos factores. Además, se sugiere el uso de algunas líneas para ser incorporadas en programas de mejoramiento genético y así desarrollar nuevos genotipos aptos para el país y, por último, la profundización para establecer las bases genéticas de la resistencia de los dos cultivares evaluados en este estudio.

The oomycete Phytophthora infestans (Mont) DeBary causes late blight disease. This pathogen affects economically important crops for the country, including tomato, a crop in which it can causes total losses in production. Given the aggressiveness of the pathogen, tomato breeding programs develop new genotypes with resistance to P. infestans, being a source of resistance Ph genes. However, the new strains of P. infestans, product of the genetic variability of their populations, can overcome the resistance of the genotypes. In this study, the genetic resistance of 13 tomato materials was evaluated against 5 isolates of P. infestans obtained from a previous study. Through in vitro inoculations and with the area under the disease progress curve (AUDPC), it was determined that there was a statistically significant difference (p < 0.05) between the behavior of the genetic resistance, the aggressiveness of the isolates of the pathogen and the interaction between both factors. It is also suggested the use of some lines to be incorporated into tomato breeding programs and develop new genotypes suitable for the country and, finally, to establish the genetic basis of the resistance of the two cultivars evaluated in this study.

Diversidad genética de aislados de Phytophthora infestans colectados en zonas productoras de papa y tomate de Guatemala / Genetic diversity of Phytophthora infestans isolates collected from tomato and potato producing areas in Guatemala

Phytophthora infestans (Mont) DeBary es el agente causal de la enfermedad conocida como tizón tardío, la cual ha sido catalogada como la enfermedad de plantas más devastadora reportada en la historia de la humanidad. Este patógeno afecta plantas de importancia económica de la familia solanaceae, como el tomate y la papa. P. infestans es un oomicete heterotálico y necesita de dos tipos de apareamiento, A1 y A2, para presentar reproducción sexual, la cual es la vía por la que este patógeno incrementa su grado de diversidad, a través de una recombinación de su material genético, que representa el mayor desafío para el manejo de la enfermedad. Este estudio determinó el nivel de variabilidad genética, a través del marcador molecular amplified fragment length polymorphism (AFLP), de 22 aislados de P. infestans colectados en diferentes zonas productoras de papa y tomate. Con el perfil de bandas generado por el marcador molecular, se realizó un análisis cluster y se elaboró un dendograma de tipo unweighted pair group method with arithmetic mean (UPGMA), con el índice de Dice, mediante una matriz de distancias genéticas. Los aislados fueron situados en tres grupos principales, los cuales responden al lugar de procedencia y al tipo de planta hospedera. Se encontró un valor de similitud de 0.49 entre los aislados analizados, por lo que se concluyó que la variabilidad genética de P. infestans en Guatemala es alta.

Phytophthora infestans (Mont) DeBary is the causal agent of late blight disease, which has been cataloged as the most devasting plant disease in the history of humankind. This pathogen is capable of affecting economically important plants of the solanaceae family, such as tomato and potato. P. infestans is a heterothallic oomycete for which it needs two types of mating known as A1 and A2 to present a sexual reproduction, which is the main way by this pathogen increases its degree of genetic diversity through a recombination of its genetic material; this condition represents the major defiance to control this disease. This study determined the level of genetic variability, through the molecular marker amplified fragment length polymorphism (AFLP), of 22 P. infestans isolates collected in different potato and tomato producing areas in Guatemala. With the band profile generated by the molecular marker AFLP, a cluster analysis was performed creating a UPGMA dendrogram with Dice´s index through a genetic distances matrix. The isolates were located in three main groups, which respond to the place of origin and the type of host plant. A similarity value of 0.49 was found among the analyzed isolates. It is concluded that genetic variability of the isolates analyzed is high.

Competition between Phytophthora infestans effectors leads to increased aggressiveness on plants containing broad-spectrum late blight resistance.

BACKGROUND: The destructive plant disease potato late blight is caused by the oomycete pathogen Phytophthora infestans (Mont.) de Bary. This disease has remained particularly problematic despite intensive breeding efforts to integrate resistance into cultivated potato, largely because of the pathogen's ability to quickly evolve to overcome major resistance genes. The RB gene, identified in the wild potato species S. bulbocastanum, encodes a protein that confers broad-spectrum resistance to most P. infestans isolates through its recognition of highly conserved members of the corresponding pathogen effector family IPI-O. IpiO is a multigene family of effectors and while the majority of IPI-O proteins are recognized by RB to elicit host resistance, some variants exist that are able to elude detection (e.g. IPI-O4). METHODS AND FINDINGS: In the present study, analysis of ipiO variants among 40 different P. infestans isolates collected from Guatemala, Thailand, and the United States revealed a high degree of complexity within this gene family. Isolate aggressiveness was correlated with increased ipiO diversity and especially the presence of the ipiO4 variant. Furthermore, isolates expressing IPI-O4 overcame RB-mediated resistance in transgenic potato plants even when the resistance-eliciting IPI-O1 variant was present. In support of this finding, we observed that expression of IPI-O4 via Agrobacterium blocked recognition of IPI-O1, leading to inactivation of RB-mediated programmed cell death in Nicotiana benthamiana. CONCLUSIONS: In this study we definitively demonstrate and provide the first evidence that P. infestans can defeat an R protein through inhibition of recognition of the corresponding effector protein.

New Diversity of Ralstonia solanacearum Strains Associated with Vegetable and Ornamental Crops in Florida.

In 2003 and 2004, 15 isolates of Ralstonia solanacearum were obtained from wilting plants of field-grown pepper (Capsicum annuum) in south Florida and from pot-grown hydrangea (Hydrangea paniculata and H. macrophylla) and geranium (Pelargonium × hortorum) in commercial nurseries and retention ponds in north Florida. Diagnostic immunoassays and polymerase chain reaction (PCR) analyses identified all the isolates as R. solanacearum but not race 3 biovar 2. Pathogenicity studies on tomato, pepper, and tobacco revealed that all 15 strains had similar high virulence on tomato and all caused wilting of tobacco, although there were significant differences among the strains in aggressiveness on tobacco. An indigenous Florida tomato strain, race 1 biovar 1 (Rs5), caused no disease on tobacco and little or none on pepper. The three pepper strains were more aggressive than Rs5 or two hydrangea strains on all three pepper cultivars studied. Phylogenetic analysis based on an endoglucanase gene sequence indicated that these strains had three distinct origins. The three pepper strains belonged to phylotype I biovar 3 and clustered with strains from diverse hosts in Asia belonging to sequevar 13. The six geranium strains and four of the hydrangea strains were closely related to strains in sequevar 5, a distinct subcluster of phylotype II biovar 1 strains isolated from the French West Indies and Brazil. Two other biovar 1 strains from hydrangea and strains K60, AW, and Rs5 belonged to sequevar 7 in phylotype II and probably are native to North America. None of the Florida isolates belong to the highly regulated Select Agent race 3 biovar 2 subgroup, according to both the DNA sequence analysis and the biovar phenotypic test results. However, the race 3 biovar 2-specific B2 primers weakly amplified a product from some race 1 biovar 1 strains in real-time PCR, indicating that this assay may give false positives under some conditions. Given the high cost of a misdiagnosis, it seems advisable to use at least two independent diagnostic methods to confirm that a suspect isolate is R. solanacearum R3B2. This is the first report of the presence of R. solanacearum race 1 biovar 3 or phylotype I strains in North America, and the first report confirming R. solanacearum causing natural infection of hydrangea in Florida. Thus, R. solanacearum strains that are quite distinct from presumably indigenous strains are present and can infect diverse hosts in Florida.