Abu Seman Idris

Comparison of Different Protein Extraction Methods for Gel-Based Proteomic Analysis of Ganoderma spp.

Ganoderma species are a group of fungi that have the ability to degrade lignin polymers and cause severe diseases such as stem and root rot and can infect economically important plants and perennial crops such as oil palm, especially in tropical countries such as Malaysia. Unfortunately, very little is known about the complex interplay between oil palm and Ganoderma in the pathogenesis of the diseases. Proteomic technologies are simple yet powerful tools in comparing protein profile and have been widely used to study plant–fungus interaction. A critical step to perform a good proteome research is to establish a method that gives the best quality and a wide coverage of total proteins. Despite the availability of various protein extraction protocols from pathogenic fungi in the literature, no single extraction method was found suitable for all types of pathogenic fungi. To develop an optimized protein extraction protocol for 2-DE gel analysis of Ganoderma spp., three previously reported protein extraction protocols were compared: trichloroacetic acid, sucrose and phenol/ammonium acetate in methanol. The third method was found to give the most reproducible gels and highest protein concentration. Using the later method, a total of 10 protein spots (5 from each species) were successfully identified. Hence, the results from this study propose phenol/ammonium acetate in methanol as the most effective protein extraction method for 2-DE proteomic studies of Ganoderma spp.

Serine-rich protein is a novel positive regulator for silicon accumulation in mangrove

Silicon (Si) plays an important role in reducing plant susceptibility against a variety of different biotic and abiotic stresses; and also has an important regulatory role in soil to avoid heavy metal toxicity and providing suitable growing conditions for plants. A full-length cDNAs of 696bp of serine-rich protein was cloned from mangrove plant (Rhizophora apiculata) by amplification of cDNA ends from an expressed sequence tag homologous to groundnut (Arachis hypogaea), submitted to NCBI (KF211374). This serine-rich protein gene encodes a deduced protein of 223 amino acids. The transcript titre of the serine-rich protein was found to be strongly enriched in roots compared with the leaves of two month old mangrove plants and expression level of this serine-rich protein was found to be strongly induced when the mangrove seedlings were exposed to SiO2. Expression of the serine-rich protein transgenic was detected in transgenic Arabidopsis thaliana, where the amount of serine increased from 1.02 to 37.8mg/g. The same trend was also seen in Si content in the roots (14.3%) and leaves (7.4%) of the transgenic A. thaliana compared to the wild-type plants under Si treatment. The biological results demonstrated that the accumulation of the serine amino acid in the vegetative tissues of the transgenic plants enhanced their ability to absorb and accumulate more Si in the roots and leaves and suggests that the serine-rich protein gene has potential for use in genetic engineering of different stress tolerance characteristics.

Assessment of plant secondary metabolites in oil palm seedlings after being treated with calcium, copper ions and salicylic acid

The effect of calcium, copper ions and salicylic acid (SA) amendment on the incidence of basal stem rot and activity of secondary metabolites in oil palm seedlings were investigated in glasshouse study. Disease incidence (DI) in positive control (T8) was 75% at nine months after inoculation (9 MAI). However, weekly pre-immunisation with Ca2+ + Cu2+ + SA prior to inoculation significantly suppressed DI and delayed disease onset as noted in T7. In the present study, the lowest %DI was observed in T7 (15%) followed by T1, T5, T6, T3, T4 and T2. The Ca2+, Cu2+ and SA amendments were resulted in earlier and higher accumulation of plant secondary metabolites as noted in leaves, stems and root tissues in response to invasion by Ganoderma boninense. High total phenolic content concentration was detected in T7 (leaf: 233.38 ± 0.12 mg/g; stem: 132.78 ± 0.04 mg/g and root: 86.98 ± 0.28 mg/g). Similar trend was obtained in peroxidase activity, total lignin content and hydrogen peroxide scavenging activity. These results suggested that it could be due to the accumulation of phenolics, peroxidase activities, lignin content and hydrogen peroxide scavenging activities in oil palm seedling tissues which might have collectively contributed to induce resistance against G. boninense.

Enhancement of Thiamine Biosynthesis in Oil Palm Seedlings by Colonization of Endophytic Fungus Hendersonia toruloidea

Thiamine, or vitamin B1 plays an indispensable role as a cofactor in crucial metabolic reactions including glycolysis, pentose phosphate pathway and the tricarboxylic acid cycle in all living organisms. Thiamine has been shown to play a role in plant adaptation toward biotic and abiotic stresses. The modulation of thiamine biosynthetic genes in oil palm seedlings was evaluated in response to root colonization by endophytic Hendersonia toruloidea. Seven-month-old oil palm seedlings were inoculated with H. toruloidea and microscopic analyses were performed to visualize the localization of endophytic H. toruloidea in oil palm roots. Transmission electron microscopy confirmed that H. toruloidea colonized cortical cells. The expression of thiamine biosynthetic genes and accumulation of total thiamine in oil palm seedlings were also evaluated. Quantitative real-time PCR was performed to measure transcript abundances of four key thiamine biosynthesis genes (THI4, THIC, TH1, and TPK) on days 1, 7, 15, and 30 in response to H. toruloidea colonization. The results showed an increase of up to 12-fold in the expression of all gene transcripts on day 1 post-inoculation. On days 7, 15, and 30 post-inoculation, the relative expression levels of these genes were shown to be downregulated. Thiamine accumulation was observed on day 7 post-colonization and subsequently decreased until day 30. This work provides the first evidence for the enhancement of thiamine biosynthesis by endophytic colonization in oil palm seedlings.

Effect of formulated bioorganic containing Burkholderia GanoEB2 in suppressing Ganoderma disease in oil palm seedlings.

Ili Nadhrah N., Nulit R., Nurrashyeda R., Idris A.S. (2015): Effect of formulated bioorganic containing Burkholderia GanoEB2 in suppressing Ganoderma disease in oil palm seedlings. Plant Protect. Sci., 51: 80–87. The ability of Burkholderia GanoEB2 formulated in two different bioorganic powders as carriers – bioorganic empty fruit bunch (BEFB) and real strong bioorganic fertilizer (RSBF), and the efficacies of Burkholderia GanoEB2 in sup pressing Ganoderma boninense infection in oil palm seedlings were determined. Results showed that the highest reduction in disease incidence (DI, 18.2%), severity of foliar symptoms (SFS, 26.6%), and disease severity foliar index (DSFI, 27.7%) was achieved by the seedlings treated with RSBF containing Burkholderia GanoEB2 compared to the seedlings treated with BEFB containing Burkholderia GanoEB2 (36.4% DI, 33.4% SFS, and 31.8% DSFI). Untreated seedlings (as control) had 100.0% DI, 90.4% SFS, and 87.5% DSFI. The disease was reduced by as much as 85.3% for seedlings treated with RSBF and by 70.5% for seedlings treated with BEFB. The formulated bioorganics containing Burkholderia GanoEB2 were proven to suppress Ganoderma disease in oil palm.

DIFFERENTIAL PROTEOMIC STUDY OF OIL PALM LEAVES IN RESPONSE TO IN VITRO INOCULATION WITH PATHOGENIC AND NON-PATHOGENIC GANODERMA spp.

Basal stem rot is an aggressive disease in oil palm caused by Ganoderma species. The disease threatens the commercial oil palm plantations of South East Asia, especially in Malaysia. In order to understand the mechanism involved in the early stage of interaction between Ganoderma spp. and its host at systemic level, proteomic analysis of oil palm leaves was conducted on protein samples collected over 72 hours during inoculation with pathogenic Ganoderma boninense and non-pathogenic Ganoderma tornatum. A total of 82 proteins resolved during two-dimensional gel electrophoresis with significant differences in the spot abundance. However, only 24 differentially expressed proteins in response to Ganoderma spp. inoculations were successfully identified by mass spectrophotometry as compared to the non-inoculated control. These proteins are mainly involved in photosynthesis, signalling, stress/defense, energy and metabolism regulation. Changes in relative abundance of these proteins suggest an important role in disease susceptibility. Most proteins showed altered abundance in response to both G. boninense and G. tornatum, while some proteins were only affected by either G. boninense or G. tornatum. The putative role of the identified proteins in oil palm leaves during the interaction with both Ganoderma spp. is discussed.

Determination of optimum levels of nitrogen, phosphorus and potassium of oil palm seedlings in solution culture

Balanced nutrient elements in fertilizer play a critical role in oil palm seedling successful growth and development, and at the same time reduces of fertilizer losses in the environment. This study examines the effect of different levels of N, P2O5 and K2O for oil palm seedlings in solution culture on growth traits, nutrient uptake in plant tissues and biomass accumulation under nursery conditions. Five concentration levels of N (50, 100, 300, 600 and 900 mg L–1), P2O5 and K2O (15, 30, 60, 90 and 120 mg L–1) were used in a completely randomized design (CRD) with five replications for each. Parameters measured during the growing period include - plant height, leaf number/plant, stem diameter, SPAD chlorophyll value, and at harvest – total leaf area, root dry weight, shoot dry weight and total dry weight. Different levels of N, P2O5 and K2O showed significant effects on all the parameters studied. The highest values for diameter, plant height, leaf number/plant, total leaf area, root dry weight, shoot dry weight and total dry biomass were obtained using 100, 90 and 300 mg/L levels of N, P2O5 and K2O, respectively. Most of the growth parameters, declined with lower levels of N, P2O5 and K2O. The results of this study provide a new knowledge to produce oil palm plant with better nutrient management at the nursery under solution culture.

Control of Basal Stem Rot Disease in Oil Palm by Supplementation of Calcium, Copper, and Salicylic Acid.

Continuous supplementation of mineral nutrients and salicylic acid (SA) as foliar application could improve efficacy in controlling basal stem rot (BSR) disease in oil palm seedling. It is revealed from the results that the highest disease severity index (58.3%) was recorded in T8 treatments at 9 months after inoculation. The best disease control was achieved by T7 treatments (calcium/copper/SA [Ca/Cu/SA]) (5.0%) followed by T1 (5.5%), T5 (5.8%), T3 (8.3%), T6 (8.3%), T4 (13.3%), and T2 (15.8%) treatments. Continuous supplementation of Ca/Cu/SA was found to be the most effective in controlling the disease and the high performance liquid chromatography results showed the detection of ergosterol at very low concentration in the treated samples. Moreover, the transmission electron microscopy analysis results clearly indicated that T7 treatment was also enhancing lignification, which was responsible for the thickness of the secondary cell walls and middle lamella compared to untreated samples. It was therefore, concluded that continuous supplementation of minerals nutrients and SA could effectively suppress disease severity by reducing ergosterol activity and also improve the process of lignification in the treated plants. Furthermore, this treatment also managed to delay the onset of BSR symptoms and promote the growth of the seedlings and eventually suppress the BSR disease.

Antioxidant Enzyme Activities and Secondary Metabolite Profiling of Oil Palm Seedlings Treated with Combination of NPK Fertilizers Infected with Ganoderma boninense

Oil palm (Elaeis guineensis Jacq) is one of the major sources of edible oil. Reducing the effect of Ganoderma, main cause of basal stem rot (BSR) on oil palm, is the main propose of this study. Understanding the oil palm defense mechanism against Ganoderma infection through monitoring changes in the secondary metabolite compounds levels before/after infection by Ganoderma under different fertilizing treatment is required. Oil palm requires macro- and microelements for growth and yield. Manipulating the nutrient for oil palm is a method to control the disease. The 3-4-month-old oil palm seedlings were given different macronutrient treatments to evaluate induction of defense related enzymes and production of secondary metabolite compounds in response to G. boninense inoculation. The observed trend of changes in the infected and uninfected seedlings was a slightly higher activity for β-1,3-glucanases, chitinase, peroxidase, and phenylalanine ammonia-lyase during the process of pathogenesis. It was found that PR proteins gave positive response to the interaction between oil palm seedlings and Ganoderma infection. Although the responses were activated systematically, they were short-lasting as the changes in enzymes activities appeared before the occurrence of visible symptoms. Effect of different nutrients doses was obviously observed among the results of the secondary metabolite compounds. Many identified/unidentified metabolite compounds were presented, of which some were involved in plant cell defense mechanism against pathogens, mostly belonging to alkaloids with bitter-tasting nitrogenous-compounds, and some had the potential to be used as new markers to detect basal stem rot at the initial step of disease.

The development of spectral indices for early detection of Ganoderma disease in oil palm seedlings

ABSTRACT Field spectroscopy is a rapid and non-destructive analytical technique that may be used for assessing plant stress and disease. The objective of this study was to develop spectral indices for detection of Ganoderma disease in oil palm seedlings. The reflectance spectra of oil palm seedlings from three levels of Ganoderma disease severity were acquired using a spectroradiometer. Denoizing and data transformation using first derivative analysis was conducted on the original reflectance spectra. Then, comparative statistical analysis was used to select significant wavelength from transformed data. Wavelength pairs of spectral indices were selected using optimum index factor. The spectral indices were produced using the wavelength ratios and a modified simple ratio method. The relationship analysis between spectral indices and total leaf chlorophyll (TLC) was conducted using regression technique. The results suggested that six spectral indices are suitable for the early detection of Ganoderma disease in oil palm seedlings. Final results after regression with TLC showed that Ratio 3 is the best spectral index for the early detection of Ganoderma infection in oil palm seedlings. For future works, this can be used for the development of robust spectral indices for Ganoderma disease detection in young and mature oil palm using airborne hyperspectral imaging.