Biocontrol of Orchid-pathogenic Mold, Phytophthora palmivora, by Antifungal Proteins from Pseudomonas aeruginosa RS1

Black rot disease in orchids is caused by the water mold Phytophthora palmivora. To gain better biocontrol performance, several factors affecting growth and antifungal substance production by Pseudomonas aeruginosa RS1 were verified. These factors include type and pH of media, temperature, and time for antifungal production. The results showed that the best conditions for P. aeruginosa RS1 to produce the active compounds was cultivating the bacteria in Luria-Bertani medium at pH 7.0 for 21 h at 37 °C. The culture filtrate was subjected to stepwise ammonium sulfate precipitation. The precipitated proteins from the 40% to 80% fraction showed antifungal activity and were further purified by column chromatography. The eluted proteins from fractions 9–10 and 33–34 had the highest antifungal activity at about 75% and 82% inhibition, respectively. SDS-PAGE revealed that the 9–10 fraction contained mixed proteins with molecular weights of 54 kDa, 32 kDa, and 20 kDa, while the 33–34 fraction contained mixed proteins with molecular weights of 40 kDa, 32 kDa, and 29 kDa. Each band of the proteins was analyzed by LC/MS to identify the protein. The result from Spectrum Modeler indicated that these proteins were closed similarly to three groups of the following proteins; catalase, chitin binding protein, and protease. Morphological study under scanning electron microscopy demonstrated that the partially purified proteins from P. aeruginosa RS1 caused abnormal growth and hypha elongation in P. palmivora. The bacteria and/or these proteins may be useful for controlling black rot disease caused by P. palmivora in orchid orchards.

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

Biological Control of Phytophthora palmivora Causing Root Rot of Pomelo Using Chaetomium spp

Phytophthora diseases have become a major impediment in the citrus production in Thailand. In this study, an isolate of Phytophthora denominated as PHY02 was proven to be causal pathogen of root rot of Pomelo (Citrus maxima) in Thailand. The isolate PHY02 was morphologically characterized and identified as Phytophthora palmivora based on molecular analysis of an internal transcribed spacer rDNA sequence. This work also presents in vitro evaluations of the capacities of Chaetomium spp. to control the P. palmivora PHY02. As antagonists, Chaetomium globosum CG05, Chaetomium cupreum CC3003, Chaetomium lucknowense CL01 inhibited 50~61% mycelial growth, degraded mycelia and reduced 92~99% sporangial production of P. palmivora PHY02 in bi-culture test after 30 days. Fungal metabolites from Chaetomium spp. were tested against PHY02. Results showed that, methanol extract of C. globosum CG05 expressed strongest inhibitory effects on mycelial growth and sporangium formation of P. palmivora PHY02 with effective dose ED50 values of 26.5 microg/mL and 2.3 microg/mL, respectively. It is interesting that C. lucknowense is reported for the first time as an effective antagonist against a species of Phytophthora.

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.

Electrophoretic protein patterns of three species of Phytophthora associated with black pod disease of cocoa (Theobroma cacao L.).

When electrophoretic profiles of native proteins from vegetative mycelia of Phytophthora palmivora, Phytophthora capsici and Phytophthora citrophthora causing black pod disease of cocoa in India were compared on a single Polyacrylamide gel, the isolates of same species were readily distinguished both qualitatively by visual similarity in banding patterns and quantitatively by calculating similarity coefficients. Similarity coefficients were generally much higher between isolates within a species than between isolates of different species. The dendrograms obtained after unweighted pair grouping with arithmetic averaging cluster analysis, revealed that all the isolates of Phytophthora capsici were highly homogenous and formed a single cluster. The isolates of Phytophthora citrophthora were resolved into two electrophoretic types which were clustered into two distinct sub groups. Phytophthora palmivora formed a separate group. Thus, the results reveal that polyacrylamide gel electrophoresis can be used successfully in distinguishing species and sub groups within a species of Phytophthora encountered on cocoa.