Population Structure of a Worldwide Phytophthora palmivora Collection Suggests Lack of Host Specificity and Reduced Genetic Diversity in South America and the Caribbean.

Phytophthora palmivora (Butler) is a highly destructive plant pathogen that infects tropical hosts worldwide, many of which are economically important crops. Despite the broad host range and wide distribution, the pathogen has displayed a considerable amount of variation in morphological characters, including virulence. However, the genetic variability at a global level, which is critical to understand the center of origin and the potential pathway(s) of introduction, was unclear. Here, we mapped the genetic variation of P. palmivora using isolates representing four regions, 15 countries, and 14 host species. We designed a large set of simple sequence repeat markers from the P. palmivora genome and picked 17 selectively neutral markers to screen 98 P. palmivora isolates. We found that P. palmivora populations from our collection generally did not cluster according to host; rather, some isolates from North America were generally distinct from all other populations. Isolates from South America and the Caribbean clustered and appeared to share ancestry with isolates from Asia. Populations from North America and Asia were the most genetically diverse, while the South American and Caribbean populations exhibited similar reduced genetic diversity. The isolates collected in various plantations in Colombia did not show host or geographic specificity. Our study brought a further understanding of this important plant pathogen, although the determination for hypothesized source of origin, spread, and evolution would need further sampling. The genomic resources developed in this study would facilitate further studies on P. palmivora diagnostics and management.

Novel Cell Death-Inducing Elicitors from Phytophthora palmivora Promote Infection on Hevea brasiliensis.

Elicitors play an important role in plant and pathogen interactions. The discovery of new elicitors and their effects on plant defense responses is significant and challenging. In this study, we investigated novel elicitors from Phytophthora palmivora and their effects on plant defenses. A crude elicitor isolated by ethanol precipitation from culture filtrates of P. palmivora induced cell death in tobacco leaves. When tobacco leaves were infiltrated with this cell death-inducing elicitor, the accumulations of H2O2, salicylic acid (SA), scopoletin (Scp), and abscisic acid (ABA) were detected. Accumulations of SA, Scp, and ABA were also induced in rubber tree leaves. P. palmivora infection significantly increased in rubber tree leaves pretreated with the elicitor and cotreated with the elicitor and zoospores of P. palmivora. This elicitor can be described as compound elicitor because Fourier-transform infrared spectroscopy revealed that it consisted of both polysaccharide and protein. We also found that the cell death effect caused by this compound elicitor was completely neutralized by Proteinase K. The compound elicitor was composed of four fractions which were beta-glucan, high-molecular-weight glycoprotein, broad-molecular-weight glycoprotein and 42-kDa protein. Interestingly, the broad-molecular-weight glycoprotein caused the highest level of cell death in tobacco leaves, while the beta-glucan had no effect. The high-molecular-weight glycoprotein, broad-molecular-weight glycoprotein and 42-kDa protein fractions not only caused cell death in tobacco leaves but also induced high levels of SA accumulation. Furthermore, these three fractions clearly promoted P. palmivora infection of rubber tree leaves.

Sulphated Polysaccharide from Acanthophora spicifera Induced Hevea brasiliensis Defense Responses Against Phytophthora palmivora Infection.

Elicitors from seaweeds are considered an alternative stimulant of plant defenses against pathogenic infection. Finding new sources of elicitors and exploring their effects on plant defenses is a significant undertaking. In this study, we extracted crude polysaccharide (CPS) from Acanthophora spicifera (a red alga) and tested the effects of the compound on rubber tree (Hevea brasiliensis) defense responses. Accumulations of salicylic acid (SA) and scopoletin (Scp) were measured by HPLC. The expression of SA- and Jasmonic acid (JA)-responsive genes was analyzed by semi-qRT-PCR. Strong anion exchange chromatography and Fourier-transform infrared (FTIR) spectroscopy were used for purification and functional characterization of CPS, respectively. The extracted CPS enhanced rubber tree defenses against Phytophthora palmivora infection. It induced SA and Scp accumulations and SA-responsive gene expression, but suppressed JA-responsive gene expression. We successfully separated the non-sulphated polysaccharide (F1) from the sulphated polysaccharides (SPS). Both peaks of SPS (F2 and F3) were identified as lambda (λ)-carrageenan. The F3 fraction showed greater elicitor activity on tobacco leaves. It induced SA and Scp accumulations and peroxidase activity but suppressed catalase activity. Furthermore, the purified λ-carrageenan did not cause cell death in tobacco or rubber tree leaves. Therefore, the elicitor from A. spicifera could be an alternative plant stimulant.

Molecular characterization of Phytophthora palmivora responsible for bud rot disease of oil palm in Colombia.

Bud rot disease is a damaging disease of oil palm in Colombia. The pathogen responsible for this disease is a species of oomyctes, Phytophthora palmivora which is also the causal pathogen of several tropical crop diseases such as fruit rot and stem canker of cocoa, rubber, durian and jackfruit. No outbreaks of bud rot have been reported in oil palm in Malaysia or other Southeast Asian countries, despite this particular species being present in the region. Analysis of the genomic sequences of several genetic markers; the internal transcribe spacer regions (ITS) of the ribosomal RNA gene cluster, beta-tubulin gene, translation elongation factor 1 alpha gene (EF-1α), cytochrome c oxidase subunit I & II (COXI and COXII) gene cluster along with amplified fragment length polymorphism (AFLP) analyses have been carried out to investigate the genetic diversity and variation of P. palmivora isolates from around the world and from different hosts in comparison to Colombian oil palm isolates, as one of the steps in understanding why this species of oomycetes causes devastating damage to oil palm in Latin America but not in other regions. Phylogenetic analyses of these regions showed that the Colombian oil palm isolates were not separated from Malaysian isolates. AFLP analysis and a new marker PPHPAV, targeting an unclassified hypothetical protein, was found to be able to differentiate Malaysian and Colombian isolates and showed a clear clade separations. Despite this, pathogenicity studies did not show any significant differences in the level of aggressiveness of different isolates against oil palm in glasshouse tests.

Identificación de estructuras de infección de Phytophthora palmivora en hojas de clones de palma de aceite Elaeis guinennsis Jacq / Identification of infection structures of Phytophthora palmivora in leaves of oil palm clones Elaeis guineensis Jacq

Este trabajo investigó el modelo de interacción planta-patógeno mediante discos de hojas de clones de palma de aceite inoculados en condiciones ex situ con un aislamiento de Phytophthora palmivora. Las inoculaciones se realizaron en condiciones controladas en cámara de crecimiento. En total, seis diferentes ortets fueron evaluados en seis tiempos de infección (2, 4, 6, 12, 24 y 48 horas postinfección, o hpi). Se determinó la presencia de estructuras de patogenicidad de P. palmivora como quiste, apresorio y tubos germinativos. Los quistes fueron identificados principalmente a las 2, 4 y 6 hpi. A partir de las 48 hpi no hubo presencia de zoosporas enquistadas para ningún ortet evaluado. En cuanto a los apresorios, estos se empezaron a desarrollar a las 4 horas de realizada la inoculación (siendo las 12 y 24 hpi los tiempos de mayor registro de estas estructuras). 121 Identificación de estructuras de infección de Phytophthora palmivora en hojas de clones de palma de aceite (Elaeis guineensis Jacq.) Méndez, K. et al. Introducción La Pudrición del cogollo (pc) es una de las principales enfermedades que afecta al cultivo de palma de aceite, destruyendo plantaciones desde 1964. En Colombia la enfermedad se presenta en las cuatro zonas palmeras y ha alcanzado proporciones epidémicas (Sarria et al., 2013a, 2016). A finales de 2004 en la Zona Suroccidental se registró un incremento en el número de casos, con un crecimiento exponencial de esta enfermedad, y a partir de 2007 se vieron afectadas más de 30.000 ha de cultivos de palma de aceite. En la Zona Central un comportamiento similar de la enfermedad fue registrado para la región de Puerto Wilches (Norte de Santander) entre 2009 y 2013, periodo en el cual se perdieron más de 40.000 ha como consecuencia de una epidemia de pc (Sanz, 2016). Por su parte, los productores de la Zona Norte actualmente hacen frente a la amenaza de una expansión epidémica de la pc con síntoma de hoja clorótica, la cual da cuenta de un estado avanzado de la enfermedad. Una amplia revisión del impacto de esta enfermedad desde sus inicios se encuentra en Benítez & García (2015), Sundram & Intan-Nur (2017) y Torres et al. (2016). La enfermedad de la Pudrición del cogollo es causada por el oomiceto hemibiótrofo P. palmivora (Sarria et al., 2008, 2013). Desde su identificación como agente causal de la pc en 2008, el Centro de Investigación en Palma de Aceite (Cenipalma) ha liderado diferentes investigaciones que han dado como resultado el desarrollo de estrategias de manejo integrado del cultivo de palma, así como la descripción del patógeno por medio de inoculación en condiciones in vitro en foliolos inmaduros de palma (Martínez et al., 2013, 2014a), entre otras. En cuanto a la presencia de esta enfermedad en otros cultivos, Mohamed-Azni et al. (2017) emplearon la técnica de foliolo inmaduro, logrando infectar foliolos de palma de aceite con P. palmivora aislada proveniente de cultivos de cacao y durián. Teniendo en cuenta que actualmente no se conocen fuentes probadas de resistencia de cultivares de Elaeis guineensis a la pc, y que un ciclo de mejoramiento genético de palma puede durar más de 25 años Por su parte, los tubos germinativos se encontraron a partir de las 48 hpi únicamente. Finalmente, se pudo establecer que el patógeno logra colonizar tejidos de foliolo no lignificados de clones de palma. Además, se encontró una relación entre el número de estructuras del patógeno con el comportamiento del cultivar de la palma donor (ramet)

Plant-pathogen model interaction was studied using leaf disks of oil palm clones inoculated ex situ with a Phytophthora palmivora isolate. The inoculation process was performed under growth chamber conditions. Six ortets were evaluated at six post-inoculation times (2, 4, 6, 12, 24 y 48 hours post infection, or hpi). Pathogen's infection structures as cyst, apressorium and germinative tubes were found. Cysts were identified mainly at 2, 4 and 6 hpi. After 48 hpi there were not any cysts for the evaluated ortets. The apresoria started to develop at 4 hpi, with the highest presence of these structures at 12 and 24 hpi. Germinative tubes were found only after 48 hpi. Therefore, it was established that the pathogen can colonize no-lignified tissue of oil palm clones. Finally, we found a relation between the pathogen's structures number and the response of the ortet related to its susceptibility and resistance response. Thus, it was found that the susceptible cultivar showed the highest number of germinative tubes

Molecular cloning and characterization of a novel bi-functional α-amylase/subtilisin inhibitor from Hevea brasiliensis.

A novel cDNA encoding a bi-functional α-amylase/subtilisin inhibitor (HbASI) was isolated from rubber (Hevea brasiliensis) leaves cultivar RRIM600. The HbASI had strong homology with the soybean trypsin inhibitor (Kunitz) family of protease inhibitors. Its putative amino acid sequence was similar to that of the α-amylase/subtilisin inhibitor from Ricinus communis (72% identity). Genomic sequencing indicated that the HbASI gene contained no introns. The messenger RNA of HbASI was detected in leaf, hypocotyl and root. The recombinant HbASI expressed extracellularly in Pichia pastoris exhibited inhibitory activity against α-amylase from Aspergillus oryzae, trypsin and subtilisin A. The HbASI gene was induced in the rubber leaves infected with a rubber tree pathogen, Phytophthora palmivora. It was also enhanced by salicylic acid (SA) treatment and mechanical wounding. In addition, the biological activity of the HbASI protein involving in the plant defence responses was also investigated. The HbASI at a concentration of 0.16 mg mL(-1) could inhibit the mycelium growth of P. palmivora. These data suggested that the HbASI protein might play a crucial role in defence against pathogen of rubber trees.

Aplicação de fosfito de potássio, cálcio ou magnésio para a redução da podridão-do-pé do mamoeiro em casa de vegetação / Papaya foot rot reduction under nursery conditions with application of potassium, calcium, or magnesium phosphate

O controle da podridão-do-pé (Phytophthora palmivora) do mamoeiro (Carica papaya) é essencial para reduzir perdas e aumentar a produtividade dos plantios. Sete fosfitos foram avaliados em plântulas (~10cm de altura) de mamoeiro da cultivar 'Tailândia Roxão'. Experimentos foram conduzidos com os seguintes objetivos: determinar a melhor forma de aplicação do fosfito (pulverização ou rega); determinar o melhor período prévio à inoculação do patógeno para aplicação do fosfito; determinar a quantidade de aplicações de fosfito; determinar o efeito de diferentes fosfitos sobre a doença. Os resultados dos testes mostraram que: o método mais adequado para aplicação dos fosfitos foi a pulverização da parte aérea; a aplicação dos fosfitos deve ocorrer com pelo menos 48h antes da inoculação do patógeno; duas pulverizações semanais por três semanas consecutivas antes da inoculação do patógeno reduziram a doença; e fosetyl-Al e todos os fosfitos testados [fosfito A (40 por cento P2O5 + 20 por cento K2O, 150mL p.c. 100L-1), B (40 por cento P2O5 + 20 por cento K2O, 250mL), C (10 por cento P2O5 + 6 por cento Ca, 400mL), D (20 por cento P2O5 + 20 por cento K2O, 200mL), E (40 por cento P2O5 + 6 por cento Mg, 150mL), F (30 por cento P2O5 + 20 por cento K2O, 175mL), G (20 por cento P2O5 + 20 por cento K2O, 200mL)] reduziram a doença.

Foot rot, caused by Phytophthora palmivora, is an important disease of papaya (Carica papaya). Disease control is of extreme importance to reduce losses and to increase productivity. Seven phosphites were tested in greenhouse using papaya seedlings (~ 10cm height) of the cultivar 'Tailândia Roxão'. Experiments were conducted with the following objectives: to determine the most effective method for phosphite application (spray or drench) to reduce disease; to determine the period for phosphite application before pathogen inoculation on plants; to determine the number of phosphite applications; to determine the effects of different phosphites on disease severity. Data from experiments showed that: spraying of aerial plant parts was the most adequate method for phosphite application; phosphites should be applied at least 48h before the inoculation of the pathogen on plants; two weekly phosphite sprays for three consecutive weeks before the pathogen inoculation on plants reduced disease, and; fosetyl-Al and all tested phosphites [phosphite A (40 percent P2O5 + 20 percent K2O, 150mL c.p. 100L-1), B (40 percent P2O5 + 20 percent K2O, 250mL), C (10 percent P2O5 + 6 percent Ca, 400mL), D (20 percent P2O5 + 20 percent K2O, 200mL), E (40 percent P2O5 + 6 percent Mg, 150mL), F (30 percent P2O5 + 20 percent K2O, 175mL), G (20 percent P2O5 + 20 percent K2O, 200mL)] reduced disease severity.

Aplicação de fosfito de potássio, cálcio ou magnésio para a redução da podridão-do-pé do mamoeiro em casa de vegetação

Foot rot, caused by Phytophthora palmivora, is an important disease of papaya (Carica papaya). Disease control is of extreme importance to reduce losses and to increase productivity. Seven phosphites were tested in greenhouse using papaya seedlings (~ 10cm height) of the cultivar 'Tailândia Roxão'. Experiments were conducted with the following objectives: to determine the most effective method for phosphite application (spray or drench) to reduce disease; to determine the period for phosphite application before pathogen inoculation on plants; to determine the number of phosphite applications; to determine the effects of different phosphites on disease severity. Data from experiments showed that: spraying of aerial plant parts was the most adequate method for phosphite application; phosphites should be applied at least 48h before the inoculation of the pathogen on plants; two weekly phosphite sprays for three consecutive weeks before the pathogen inoculation on plants reduced disease, and; fosetyl-Al and all tested phosphites [phosphite A (40% P2O5 + 20% K2O, 150mL c.p. 100L-1), B (40% P2O5 + 20% K2O, 250mL), C (10% P2O5 + 6% Ca, 400mL), D (20% P2O5 + 20% K2O, 200mL), E (40% P2O5 + 6% Mg, 150mL), F (30% P2O5 + 20% K2O, 175mL), G (20% P2O5 + 20% K2O, 200mL)] reduced disease severity.

O controle da podridão-do-pé (Phytophthora palmivora) do mamoeiro (Carica papaya) é essencial para reduzir perdas e aumentar a produtividade dos plantios. Sete fosfitos foram avaliados em plântulas (~10cm de altura) de mamoeiro da cultivar 'Tailândia Roxão'. Experimentos foram conduzidos com os seguintes objetivos: determinar a melhor forma de aplicação do fosfito (pulverização ou rega); determinar o melhor período prévio à inoculação do patógeno para aplicação do fosfito; determinar a quantidade de aplicações de fosfito; determinar o efeito de diferentes fosfitos sobre a doença. Os resultados dos testes mostraram que: o método mais adequado para aplicação dos fosfitos foi a pulverização da parte aérea; a aplicação dos fosfitos deve ocorrer com pelo menos 48h antes da inoculação do patógeno; duas pulverizações semanais por três semanas consecutivas antes da inoculação do patógeno reduziram a doença; e fosetyl-Al e todos os fosfitos testados [fosfito A (40% P2O5 + 20% K2O, 150mL p.c. 100L-1), B (40% P2O5 + 20% K2O, 250mL), C (10% P2O5 + 6% Ca, 400mL), D (20% P2O5 + 20% K2O, 200mL), E (40% P2O5 + 6% Mg, 150mL), F (30% P2O5 + 20% K2O, 175mL), G (20% P2O5 + 20% K2O, 200mL)] reduziram a doença.

Aplicação de fosfito de potássio, cálcio ou magnésio para a redução da podridão-do-pé do mamoeiro em casa de vegetação

Foot rot, caused by Phytophthora palmivora, is an important disease of papaya (Carica papaya). Disease control is of extreme importance to reduce losses and to increase productivity. Seven phosphites were tested in greenhouse using papaya seedlings (~ 10cm height) of the cultivar 'Tailândia Roxão'. Experiments were conducted with the following objectives: to determine the most effective method for phosphite application (spray or drench) to reduce disease; to determine the period for phosphite application before pathogen inoculation on plants; to determine the number of phosphite applications; to determine the effects of different phosphites on disease severity. Data from experiments showed that: spraying of aerial plant parts was the most adequate method for phosphite application; phosphites should be applied at least 48h before the inoculation of the pathogen on plants; two weekly phosphite sprays for three consecutive weeks before the pathogen inoculation on plants reduced disease, and; fosetyl-Al and all tested phosphites [phosphite A (40% P2O5 + 20% K2O, 150mL c.p. 100L-1), B (40% P2O5 + 20% K2O, 250mL), C (10% P2O5 + 6% Ca, 400mL), D (20% P2O5 + 20% K2O, 200mL), E (40% P2O5 + 6% Mg, 150mL), F (30% P2O5 + 20% K2O, 175mL), G (20% P2O5 + 20% K2O, 200mL)] reduced disease severity.

O controle da podridão-do-pé (Phytophthora palmivora) do mamoeiro (Carica papaya) é essencial para reduzir perdas e aumentar a produtividade dos plantios. Sete fosfitos foram avaliados em plântulas (~10cm de altura) de mamoeiro da cultivar 'Tailândia Roxão'. Experimentos foram conduzidos com os seguintes objetivos: determinar a melhor forma de aplicação do fosfito (pulverização ou rega); determinar o melhor período prévio à inoculação do patógeno para aplicação do fosfito; determinar a quantidade de aplicações de fosfito; determinar o efeito de diferentes fosfitos sobre a doença. Os resultados dos testes mostraram que: o método mais adequado para aplicação dos fosfitos foi a pulverização da parte aérea; a aplicação dos fosfitos deve ocorrer com pelo menos 48h antes da inoculação do patógeno; duas pulverizações semanais por três semanas consecutivas antes da inoculação do patógeno reduziram a doença; e fosetyl-Al e todos os fosfitos testados [fosfito A (40% P2O5 + 20% K2O, 150mL p.c. 100L-1), B (40% P2O5 + 20% K2O, 250mL), C (10% P2O5 + 6% Ca, 400mL), D (20% P2O5 + 20% K2O, 200mL), E (40% P2O5 + 6% Mg, 150mL), F (30% P2O5 + 20% K2O, 175mL), G (20% P2O5 + 20% K2O, 200mL)] reduziram a doença.