Residual analysis of chitosan-based agronanofungicides as a sustainable alternative in oil palm disease management.

The nanoformulations of pesticides have shown great interest from many parties due to their slow release capability and site-specific delivery. Hence, in this work, a new nanoformulation of a fungicide, namely chitosan-hexaconazole nanoparticles with a mean diameter size of 18 nm was subjected to the residual analysis on oil palm tissue, leaf and palm oil (crude palm oil and crude palm kernel oil) using a quick, easy, cheap, effective, rugged and safe (QuEChERS) method coupled with the gas chromatography-micro electron capture detector (GC-µECD). The chitosan-hexaconazole nanoparticles were applied using the trunk injection method at 4.5 g a.i./palm (standard single dose) and 9.0 g a.i./palm (double dose). The fungicide residue was analyzed at 0 (6 h after application), 1, 3, 7, 14, 30, 60, 90, and 120 days after treatment. The palm oil matrices; the crude palm oil (CPO) and crude palm kernel oil (CPKO) were found to be residue-free. However, it was observed that high accumulation of the fungicide in the stem tissue and leaf after the treatment using the chitosan-hexaconazole nanoparticles, which is good for better bioavailability for the treatment of the fungi, Ganoderma boninense. The dissipation kinetic at double dose treatment in the tissue and leaf was found to govern by the second-order kinetic with half-lives (t1/2) of 383 and 515 days, respectively.

Lanostane Triterpenoids from the Fruiting Bodies of Fomes officinalis and Their Anti-Inflammatory Activities.

In the current study, further chemical investigation of the fruiting bodies of Fomes officinalis led to isolate seven new 24-methyl-lanostane triterpenoids, named officimalonic acids I-O (1-7). Their structures were elucidated based on the analysis of spectroscopic data (HR-MS, 1D and 2D NMR, UV, IR). Compounds 1-3 possessed an unusual C-23 spirostructure moiety, while compounds 4-7 had 23,26-lactone unit. Anti-inflammatory assay revealed that compounds 3 and 5 exhibited significant inhibitory activities against NO production in LPS-induced RAW 264.7 cells and cyclooxygenase (COX-2).

Phenazine from Pseudomonas aeruginosa UPMP3 induced the host resistance in oil palm (Elaeis guineensis Jacq.)-Ganoderma boninense pathosystem.

Pseudomonas aeruginosa developed its biocontrol agent property through the production of antifungal derivatives, with the phenazine among them. In this study, the applications of crude phenazine synthesized by Pseudomonas aeruginosa UPMP3 and hexaconazole were comparatively evaluated for their effectiveness to suppress basal stem rot infection in artificially G. boninense-challenged oil palm seedlings. A glasshouse experiment under the randomized completely block design was set with the following treatments: non-inoculated seedlings, G. boninense inoculated seedlings, G. boninense inoculated seedlings with 1 mg/ml phenazine application, G. boninense inoculated seedlings with 2 mg/ml phenazine application and G. boninense inoculated seedlings with 0.048 mg/ml hexaconazole application. Seedlings were screened for disease parameters and plant vigour traits (plant height, plant fresh weight, root fresh, and dry weight, stem diameter, and total chlorophyll) at 1-to-4 month post-inoculation (mpi). The application of 2 mg/ml phenazine significantly reduced disease severity (DS) at 44% in comparison to fungicide application (DS = 67%). Plant vigour improved from 1 to 4 mpi and the rate of disease reduction in seedlings with phenazine application (2 mg/ml) was twofold greater than hexaconazole. At 4, 6 and 8 wpi, an up-regulation of chitinase and ß-1,3 glucanase genes in seedlings treated with phenazine suggests the involvement of induced resistance in G. boninense-oil palm pathosystem.

Application of Ground-Based LiDAR for Analysing Oil Palm Canopy Properties on the Occurrence of Basal Stem Rot (BSR) Disease.

Ground-based LiDAR also known as Terrestrial Laser Scanning (TLS) technology is an active remote sensing imaging method said to be one of the latest advances and innovations for plant phenotyping. Basal Stem Rot (BSR) is the most destructive disease of oil palm in Malaysia that is caused by white-rot fungus Ganoderma boninense, the symptoms of which include flattening and hanging-down of the canopy, shorter leaves, wilting green fronds and smaller crown size. Therefore, until now there is no critical investigation on the characterisation of canopy architecture related to this disease using TLS method was carried out. This study proposed a novel technique of BSR classification at the oil palm canopy analysis using the point clouds data taken from the TLS. A total of 40 samples of oil palm trees at the age of nine-years-old were selected and 10 trees for each health level were randomly taken from the same plot. The trees were categorised into four health levels - T0, T1, T2 and T3, which represents the healthy, mildly infected, moderately infected and severely infected, respectively. The TLS scanner was mounted at a height of 1 m and each palm was scanned at four scan positions around the tree to get a full 3D image. Five parameters were analysed: S200 (canopy strata at 200 cm from the top), S850 (canopy strata at 850 cm from the top), crown pixel (number of pixels inside the crown), frond angle (degree of angle between fronds) and frond number. The results taken from statistical analysis revealed that frond number was the best single parameter to detect BSR disease as early as T1. In classification models, a linear model with a combination of parameters, ABD - A (frond number), B (frond angle) and D (S200), delivered the highest average accuracy for classification of healthy-unhealthy trees with an accuracy of 86.67 per cent. It also can classify the four severity levels of infection with an accuracy of 80 per cent. This model performed better when compared to the severity classification using frond number. The novelty of this research is therefore on the development of new approach to detect and classify BSR using point clouds data of TLS.

RNA-seq data of Ganoderma boninense at axenic culture condition and under in planta pathogen-oil palm (Elaeis guineensis Jacq.) interaction.

OBJECTIVE: Basal stem rot disease causes severe economic losses to oil palm production in South-east Asia and little is known on the pathogenicity of the pathogen, the basidiomyceteous Ganoderma boninense. Our data presented here aims to identify both the house-keeping and pathogenicity genes of G. boninense using Illumina sequencing reads. DESCRIPTION: The hemibiotroph G. boninense establishes via root contact during early stage of colonization and subsequently kills the host tissue as the disease progresses. Information on the pathogenicity factors/genes that causes BSR remain poorly understood. In addition, the molecular expressions corresponding to G. boninense growth and pathogenicity are not reported. Here, six transcriptome datasets of G. boninense from two contrasting conditions (three biological replicates per condition) are presented. The first datasets, collected from a 7-day-old axenic condition provide an insight onto genes responsible for sustenance, growth and development of G. boninense while datasets of the infecting G. boninense collected from oil palm-G. boninense pathosystem (in planta condition) at 1 month post-inoculation offer a comprehensive avenue to understand G. boninense pathogenesis and infection especially in regard to molecular mechanisms and pathways. Raw sequences deposited in Sequence Read Archive (SRA) are available at NCBI SRA portal with PRJNA514399, bioproject ID.

Pathogenicity of Ganoderma Species on Landscape Trees in the Southeastern United States.

The genus Ganoderma contains species that are associated with dead and declining host trees. Many species have been described as pathogens in literature, because anecdotally, the presence of fruiting bodies on living trees has been widely associated with a general decline in tree health. Few studies have investigated the pathogenicity of Ganoderma species on landscape trees in the southeastern U.S. Pathogenicity tests were used to determine the pathogenicity of G. curtisii, G. meredithiae, G. sessile, and G. zonatum on young, healthy landscape trees (Pinus elliottii var. elliottii, P. taeda, Quercus shumardii, Q. virginiana, and Butia odorata) common to the southeastern U.S. Inoculations were made by drilling into the sapwood of the lower bole and inserting wooden dowels that were infested with each Ganoderma species. In two field experiments, 11 to 12 months post inoculation, trees had no visual, external symptoms of disease. There were differences in the extent of internal xylem discoloration near the site of inoculation in comparison with the mock-inoculated control in experiment 1, but there were no differences relative to the control in experiment 2. In both experiments, G. sessile was the only species that was successfully reisolated from the pine and oak hosts. Although disease symptoms were not obvious, the reisolation of G. sessile outside the inoculation point was a significant finding, and suggests that this species was capable of infecting healthy sapwood. G. sessile constitutively produces chlamydospores within its vegetative mycelium, which may contribute to its persistence in the discolored sapwood. These data suggest that the Ganoderma species tested, following trunk wounding, are not pathogens of young, actively growing landscape trees that only possess sapwood. The establishment of these fungi using alternative infection courts (e.g., roots) and their interactions in older living trees (e.g., trees with heartwood) needs investigation to better understand their effects on tree health.

Evaluation of spatio-temporal Bayesian models for the spread of infectious diseases in oil palm.

In the field of epidemiology, studies are often focused on mapping diseases in relation to time and space. Hierarchical modeling is a common flexible and effective tool for modeling problems related to disease spread. In the context of oil palm plantations infected by the fungal pathogen Ganoderma boninense, we propose and compare two spatio-temporal hierarchical Bayesian models addressing the lack of information on propagation modes and transmission vectors. We investigate two alternative process models to study the unobserved mechanism driving the infection process. The models help gain insight into the spatio-temporal dynamic of the infection by identifying a genetic component in the disease spread and by highlighting a spatial component acting at the end of the experiment. In this challenging context, we propose models that provide assumptions on the unobserved mechanism driving the infection process while making short-term predictions using ready-to-use software.

Characterization of Streptomyces spp. isolated from the rhizosphere of oil palm and evaluation of their ability to suppress basal stem rot disease in oil palm seedlings when applied as powder formulations in a glasshouse trial.

Ganoderma boninense, the main causal agent of oil palm (Elaeis guineensis) basal stem rot (BSR), severely reduces oil palm yields around the world. To reduce reliance on fungicide applications to control BSR, we are investigating the efficacy of alternative control methods, such as the application of biological control agents. In this study, we used four Streptomyces-like actinomycetes (isolates AGA43, AGA48, AGA347 and AGA506) that had been isolated from the oil palm rhizosphere and screened for antagonism towards G. boninense in a previous study. The aim of this study was to characterize these four isolates and then to assess their ability to suppress BSR in oil palm seedlings when applied individually to the soil in a vermiculite powder formulation. Analysis of partial 16S rRNA gene sequences (512 bp) revealed that the isolates exhibited a very high level of sequence similarity (> 98%) with GenBank reference sequences. Isolates AGA347 and AGA506 showed 99% similarity with Streptomyces hygroscopicus subsp. hygroscopicus and Streptomyces ahygroscopicus, respectively. Isolates AGA43 and AGA48 also belonged to the Streptomyces genus. The most effective formulation, AGA347, reduced BSR in seedlings by 73.1%. Formulations using the known antifungal producer Streptomyces noursei, AGA043, AGA048 or AGA506 reduced BSR by 47.4, 30.1, 54.8 and 44.1%, respectively. This glasshouse trial indicates that these Streptomyces spp. show promise as potential biological control agents against Ganoderma in oil palm. Further investigations are needed to determine the mechanism of antagonism and to increase the shelf life of Streptomyces formulations.

Comparative proteomic analysis of oil palm leaves infected with Ganoderma boninense revealed changes in proteins involved in photosynthesis, carbohydrate metabolism, and immunity and defense.

The basidiomycete fungal pathogen Ganoderma boninense is the causative agent for the incurable basal stem rot (BSR) disease in oil palm. This disease causes significant annual crop losses in the oil palm industry. Currently, there is no effective method for disease control and elimination, nor is any molecular marker for early detection of the disease available. An understanding of how BSR affects protein expression in plants may help identify and/or assist in the development of an early detection protocol. Although the mode of infection of BSR disease is primarily via the root system, defense-related genes have been shown to be expressed in both the root and leafs. Thus, to provide an insight into the changes in the global protein expression profile in infected plants, comparative 2DE was performed on leaf tissues sampled from palms with and without artificial inoculation of the Ganoderma fungus. Comparative 2DE revealed that 54 protein spots changed in abundance. A total of 51 protein spots were successfully identified by LC-QTOF MS/MS. The majority of these proteins were those involved in photosynthesis, carbohydrate metabolism as well as immunity and defense.

Cebadores para la amplificación del gen de la (1, 3)-Beta-glucano sintasa y caracterización parcial de la enzima en Ganoderma lucidum / Primers for (1, 3)-Beta-glucan synthase gene amplification and partial characterization of the enzyme in Ganoderma lucidum

Antecedentes. El β-(1,3)(1,6)-D-glucano es un compuesto de la pared celular de los hongos que presenta efectos inmunomoduladores y anticancerígenos. La (1,3)-β-glucano sintasa es una de las principales enzimas involucradas en su síntesis. Objetivos. Diseñar cebadores para amplificar y caracterizar parcialmente el gen correspondiente a la enzima (1,3)-β-glucano sintasa y probarlos en la cepa CP-132 de Ganoderma lucidum. Métodos. Los cebadores fueron diseñados realizando una búsqueda de la secuencia del gen en otros hongos. Después, con la técnica de PCR se probaron los cebadores utilizando ADN extraído de la cepa CP-382 de G. lucidum. Las secuencias obtenidas se compararon con aquellas de la base de datos del GenBank. Resultados. Se diseñaron 3 pares de cebadores. Todos los pares amplificaron productos de PCR de tamaño esperado. Las secuencias amplificadas con los pares BGS2113UmF y BGS3097UmR, y BGS547UmF y BGS2113UmR correspondieron a un par de secciones del gen de la (1,3)-β-glucano sintasa. Las secuencias deducidas de aminoácidos mostraron una similitud alta con genes homólogos de otros hongos, especialmente con aquellos de la clase Agaricomycetes. Conclusiones. El diseño de cebadores para amplificar parcialmente el gen de la (1,3)-β-glucano sintasa a partir de secuencias de genes homólogos fue exitoso. Estos cebadores permitirán en un futuro la caracterización de esta importante enzima en un amplio grupo de hongos (AU)

Sequence analysis and gene expression of putative exo- and endo-glucanases from oil palm (Elaeis guineensis) during fungal infection.

Glucanases are enzymes that hydrolyze a variety ß-d-glucosidic linkages. Plant ß-1,3-glucanases are able to degrade fungal cell walls; and promote the release of cell-wall derived fungal elicitors. In this study, three full-length cDNA sequences encoding oil palm (Elaeis guineensis) glucanases were analyzed. Sequence analyses of the cDNA sequences suggested that EgGlc1-1 is a putative ß-d-glucan exohydolase belonging to glycosyl hydrolase (GH) family 3 while EgGlc5-1 and EgGlc5-2 are putative glucan endo-1,3-ß-glucosidases belonging to GH family 17. The transcript abundance of these genes in the roots and leaves of oil palm seedlings treated with Ganoderma boninense and Trichoderma harzianum was profiled to investigate the involvement of these glucanases in oil palm during fungal infection. The gene expression of EgGlc1-1 in the root of oil palm seedlings was increased by T. harzianum but suppressed by G. boninense; while the gene expression of both EgGlc5-1 and EgGlc5-2 in the roots of oil palm seedlings was suppressed by G. boninense or/and T. harzianum.

Molecular biology of Ganoderma pathogenicity and diagnosis in coconut seedlings.

The pathogenicity of Ganoderma boninense was tested on coconut seedlings under greenhouse conditions and infection confirmed by using immunological and molecular diagnostic tools. Desiccation of older leaves and the emergence of sporophores were observed from pathogen-inoculated seedlings, whereas a control seedling does not show any pathogenic symptoms. Mature sporophores were formed within 10-13 weeks after inoculation. Polyclonal antibodies raised against mycelial proteins of Ganoderma were used for detection of Ganoderma in infected field palm and seedlings through indirect enzyme-linked immunosorbent assay technique. We adopted dot-immunobinding assay for the detection of Ganoderma from greenhouse and field samples. Under nucleic-acid-based diagnosis, G. boninense (167 bp) was detected from artificially inoculated seedlings and infected field palms by polymerase chain reaction. Apart from these, histopathological studies also support the Ganoderma pathogenicity in coconut seedlings. The pathogenicity test and combination of all the three diagnostic methods for Ganoderma could be highly reliable, rapid, sensitive and effective screening of resistance in planting material in the future.

Research on basal stem rot (BSR) of ornamental palms caused by basidiospores from Ganoderma boninense.

Basidiospores were isolated from the fruiting bodies of Ganoderma infecting oil palms from an estate in Johor and from ornamental palms (including oil palms) from Singapore. The spores were then germinated to obtain homokaryotic mycelia. Based on clamp connection formation in paired hyphal fusions, tester strains were identified from the homokaryons isolated. Compatibility tests were then carried out using these testers to determine the relatedness of the homokaryotic Ganoderma isolates, both from Johor and from Singapore. Results from the compatibility tests showed that Ganoderma from both locations belong to the same species, while the Ganoderma isolates from Singapore share some common alleles. The pathogenicity tests carried out on Chrysalidocarpus lutescens seedlings using inoculum growing on rubber wood blocks showed that dikaryotic mycelia can cause basal stem rot infection.

Morphological variation and host range of two Ganoderma species from Papua New Guinea.

Two species of Ganoderma belonging to different subgenera which cause disease on oil palms in PNG are identified by basidiome morphology and the morphology of their basidiospores. The names G. boninense and G. tornatum have been applied. Significant pleiomorphy was observed in basidiome characters amongst the specimens examined. This variation in most instances did not correlate well with host or host status. Spore morphology appeared uniform within a species and spore indices varied only slightly. G. tornatum was found to have a broad host range whereas G. boninense appears to be restricted to palms in Papua New Guinea.