Phytochemical investigation and antioxidant activity of ganoderma boninense

Ganoderma lucidum is known as a functional mushroom and is traditionally being used as medicine but G. boninense is well known as the causal pathogen of basal stem rot disease of oil palm. Numerous secondary metabolites of mushrooms from this genus have been reported with various biological activities. However, there are not many report on the medicinal benefits of G. boninense. Hence, this study was designed to investigate the phytochemical constituents and antioxidant activities of the fruiting body G. boninense extracts. Various groups of phytochemicals were extracted using solvents with different polarities such as chloroform, ethyl acetate, acetone, methanol, ethanol and distilled water. Results confirmed the presence of numerous phytochemicals in G. boninense fruiting body. Ethanolic extract of G. boninense has the highest phenolic (33.05 ± 1.374 mg GAE/g DW) and total flavonoid (8.20 ± 0.059 mg QE/g DW) content. Furthermore, ethanolic extract of G. boninense also exhibited the greatest potency in antiradical activity with IC50 of 113.9 ± 2.78 µg/ml. Correlation coefficient, R2 = 0.97 suggested phenolic acids of ethanol extract was contributed to its radical scavenging activity

The role of phenolics in the interaction between oil palm and Ganoderma boninense the casual agent of basal stem rot

Basal Stem Rot (BSR) of oil palm (EIaeis guineensis) is caused by Ganoderma boninense, and is one of the most commercially devastating diseases in South East Asia. It can kill more than 80 percent of stands by the time they are halfway through their normal economic life. High incidence of BSR results in economic losses, due to zero yield from dead palms and significantly reduced weight and number of fruit bunches in infected but living palms. Many methods have been attempted to control BSR. These include clean clearing, fallow period before re-planting, burning of waste or dead material, windrowing, use of fungicides as soil drenching or by tree injection, surgical removal of infected material, biological control and development of varieties with enhanced resistance. To date no method gives good control of Ganoderma infection in established plantations and some have technical limitations in application. The aim of this study was to investigate a possible source of disease resistance associated with the accumulation of phenolics, after inoculation with G. boninense and elicitation with the biopolymer chitosan. The identity of isolates of Ganoderma from Langkon Oil Palm Estate in Sabah, Malaysia, which were used in this project, was confirmed using Ganoderma Selective Medium and DNA sequence analysis after PCR amplification. That latter method showed that the Sabah isolates were very similar to virulent G. boninense strains FA5017 or FA5035 from West Malaysia, with a maximum similarity of 98%. A method to inoculate oil palm seedlings with Ganoderma, based on root treatment with macerated mycelium, was developed which facilitated experiments with consistent and rapid disease development. This proved an improvement compared to the traditional technique of inoculation with colonised rubber wood blocks. Ekona and Calabar varieties of oil palm were found to be more susceptible to G. boninense infection compared to the variety AVROS, based on a higher accumulation of ergosterol (a fungal sterol) and infection scores based on the Modified Disease Severity Score. Results from both parameters showed a strong correlation between each other and provided a quick assessment on the progress of pathogen infection for use in this project. It should be noted, however, that none of the varieties tested exhibited a high level of disease resistance. Following preliminary studies to quantify the accumulation of total phenolics, using Folin-Ciocalteu reagent, High Performance Liquid Chromatography was used to monitor changes in the concentration of three specific phenolics, which were considered to be of primary importance. These were syringic acid, caffeic acid and 4-hydroxybenzoic acid (4-HBA). Their identification was confirmed using co-injection with pure standards and by LCMS/Q-tof. The variety AVROS showed a slightly higher content of all these three phenolics compared to Ekona and Calabar. Stimulation of their accumulation was promoted by the addition of chitosan to the plant-growing medium. By the end of the time course, the concentration of these phenolics decreased in the oil palm tissues inoculated with G. boninense suggesting possible metabolism by this pathogen. This loss was, however, not detected in tissues elicited with chitosan alone and was greatly reduced when G. boninense was combined with chitosan. In vitro studies on antifungal activity with phenolics incorporated either in 10% Potato Dextose Agar, Oil Palm Root Agar or Oil Palm Root Broth were done, using concentrations (50-110 pg mL 1) of phenolic acids typically detected in oil palm root tissues. Syringic acid was found to be the most antifungal of the phenolics tested. Caffeic acid had lower activity and 4-HBA was virtually inactive. Concentrations of syringic acid detected in root tissues, especially in the presence of chitosan, that might be expected to inhibit growth of G. boninense. The pathogen was shown to be able to degrade all of the phenolic acids tested. However, at the highest concentration tested, metabolism was greatly delayed, only occurring after a lag phase in the growth of pathogen. TLC bioassay showed the degradation products of all these phenolics were no longer antifungal. Thus, accumulation of phenolic acids, especially syringic acid, may provide a useful marker in future breeding of resistant varieties

Evaluation on the efficacy of microbial approaches in the control of Ganoderma boninense and their effects to soil microbial diversity

Basal Stem Rot (BSR) is the most destructive disease of oil palm (Elaeis guineensis Jacq.) which caused by bracket fungus; Ganoderma boninense. With no remedy to date, this disease has jeopardized the industry of oil palm critically. Several approaches including cultural, chemical, mechanical and biological measures have been reported in attempt to control this disease, but unfortunately, none have given satisfactory result. Therefore, a more reliable approach that could manage this disease, besides a continuous and sustainable oil palm production is worth to be further investigated. Introduction of microbes in disease management have been used for decades. An astute observation of low incidence of disease due to Ganoderma pathogen in natural ecosystem suggests that a healthy natural ecosystem can hinder the incidence of Ganoderma pathogen outbreak. In this respect, study on the potential of microbes in suppressing the growth of Ganoderma boninense and its effect to the soil microbial diversity were investigated in this project. Three microbes-based products which are Living Soil Microbes (LSM), Agrinos (AGN) and Agriorganica (AO) were tested in this study. LSM contains multi-strain of Bacillus spp. and Trichoderma spp., whereas AGN consist of Bacillus spp., Pseudomonas spp. and Aspergillus sp. Meanwhile, AO comprises of three living food microbes which are Lactobacillus, Nattobacillus and Saccharomyces cerevisiae developed from Japanese fermentation and brewing technology. To examine the ability of the treatments in reducing BSR disease incidence, assessments in nursery and field trial were conducted. The results from both trials showed that LSM, AGN and AO were able to reduce the colonization of G. boninense based on the reduction of ergosterol content compared to untreated control

The role of phenolics in the interaction between oil palm and Ganoderma boninense the casual agent of basal stem rot

Basal Stem Rot (BSR) of oil palm (EIaeis guineensis) is caused by Ganoderma boninense, and is one of the most commercially devastating diseases in South East Asia. It can kill more than 80 percent of stands by the time they are halfway through their normal economic life. High incidence of BSR results in economic losses, due to zero yield from dead palms and significantly reduced weight and number of fruit bunches in infected but living palms. Many methods have been attempted to control BSR. These include clean clearing, fallow period before re-planting, burning of waste or dead material, windrowing, use of fungicides as soil drenching or by tree injection, surgical removal of infected material, biological control and development of varieties with enhanced resistance. To date no method gives good control of Ganoderma infection in established plantations and some have technical limitations in application. The aim of this study was to investigate a possible source of disease resistance associated with the accumulation of phenolics, after inoculation with G. boninense and elicitation with the biopolymer chitosan. The identity of isolates of Ganoderma from Langkon Oil Palm Estate in Sabah, Malaysia, which were used in this project, was confirmed using Ganoderma Selective Medium and DNA sequence analysis after PCR amplification. That latter method showed that the Sabah isolates were very similar to virulent G. boninense strains FA5017 or FA5035 from West Malaysia, with a maximum similarity of 98%. A method to inoculate oil palm seedlings with Ganoderma, based on root treatment with macerated mycelium, was developed which facilitated experiments with consistent and rapid disease development. This proved an improvement compared to the traditional technique of inoculation with colonised rubber wood blocks. Ekona and Calabar varieties of oil palm were found to be more susceptible to G. boninense infection compared to the variety AVROS, based on a higher accumulation of ergosterol (a fungal sterol) and infection scores based on the Modified Disease Severity Score. Results from both parameters showed a strong correlation between each other and provided a quick assessment on the progress of pathogen infection for use in this project. It should be noted, however, that none of the varieties tested exhibited a high level of disease resistance. Following preliminary studies to quantify the accumulation of total phenolics, using Folin-Ciocalteu reagent, High Performance Liquid Chromatography was used to monitor changes in the concentration of three specific phenolics, which were considered to be of primary importance. These were syringic acid, caffeic acid and 4-hydroxybenzoic acid (4-HBA). Their identification was confirmed using co-injection with pure standards and by LCMS/Q-tof. The variety AVROS showed a slightly higher content of all these three phenolics compared to Ekona and Calabar. Stimulation of their accumulation was promoted by the addition of chitosan to the plant-growing medium. By the end of the time course, the concentration of these phenolics decreased in the oil palm tissues inoculated with G. boninense suggesting possible metabolism by this pathogen. This loss was, however, not detected in tissues elicited with chitosan alone and was greatly reduced when G. boninense was combined with chitosan. In vitro studies on antifungal activity with phenolics incorporated either in 10% Potato Dextose Agar, Oil Palm Root Agar or Oil Palm Root Broth were done, using concentrations (50-110 pg mL 1) of phenolic acids typically detected in oil palm root tissues. Syringic acid was found to be the most antifungal of the phenolics tested. Caffeic acid had lower activity and 4-HBA was virtually inactive. Concentrations of syringic acid detected in root tissues, especially in the presence of chitosan, that might be expected to inhibit growth of G. boninense. The pathogen was shown to be able to degrade all of the phenolic acids tested. However, at the highest concentration tested, metabolism was greatly delayed, only occurring after a lag phase in the growth of pathogen. TLC bioassay showed the degradation products of all these phenolics were no longer antifungal. Thus, accumulation of phenolic acids, especially syringic acid, may provide a useful marker in future breeding of resistant varieties

In vitro antimicrobial activity of Cynodon dactylon (L.) pers. (bermudas) against selected pathogens

Cynodon dactylon (L.) Pers. is a type of perennial grass that possesses great medicinal values. In this study, the antimicrobial activity of the plant crude extract from seven different solvents (acetone, chloroform, diethyl ether, ethanol, ethyl acetate, methanol, and n-pentane) was investigated against some pathogens (Bacillus cereus, Bacillus subtilis, Escherichia coli, Klebsiella spp, Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus pyogenes, and Streptococcus pneumonia) using disc diffusion method and thin-layer chromatographic (TLC) bioassay for plant-SPE extracts against Aspergillus niger. Crude extraction showed that ethanolic extraction produced highest yield (7.065 %) followed by methanolic (5.420 %) and chloroform (3.550 %) extraction. The lowest yield was obtained from n-pentane extraction (0.500 %). The antimicrobial study revealed broad spectrum of antimicrobial activity from ethanol (7.0–10.0 ± 0.0–1.0 mm) and ethyl acetate (7.0–12.0 ± 0.0–1.0 mm) extracts against all of the bacterial pathogens. Both methanol and acetone extracts showed activity to B. cereus (8.0 ± 0.0 mm) and B. subtilis (7.0 ± 0.0 mm), while chloroform extract showed activity to B. subtilis (7.0 ± 0.0 mm) and S.pyogenes (8.3 ± 0.6 mm), respectively. Diethyl ether extraction showed activity only to S. pyogenes (7.3 ± 0.6 mm), while no activity was observed from n-pentane extraction. Great antimicrobial activity were observed for both ethyl acetate and ethanol SPE-based extracts (SBE) with size of inhibition ranging from 8.0 ± 0.0 mm to 15.7 ± 0.6 mm for ethyl acetate SBE and 8.0 ± 0.0 mm to 13.0 ± 0.0 mm for ethanol SBE. No significant antimicrobial activity was observed from thin-layer chromatographic bioassay against A. niger

Selected biomarkers from oil palm-ganoderma infected tissues for detection of basal stem rot disease

The oil palm industry has become a substantial contributor to the international market for vegetable oils, with Malaysia and Indonesia as the major producers of palm oil. However, basal stem rot disease caused by a certain fungi Ganoderma posed a major problem that affects the oil palm tree and eventually kills it. The aim of this research project is to investigate the specific organic compounds being released during the Ganoderma infection with different stages of severity among oil palm. Fresh trunk tissues were collected from oil palm trees at Langkon, Sawit Kinabalu Plantation, Sabah, Malaysia. The oil palm trees were selected based on their disease severity indices which was categorized to Index 0 (healthy), Index 1 (moderate healthy), Index 2 (moderate severe) and Index 3 (severe). The oil colour of palm trunk tissue was distinguished into white-yellowish (index 0), light yellow (index 1), dark yellow (index 2) and yellowish brown (index 3). The identity of the causal pathogen was confirmed as Ganoderma sp. BRIUMSa using Ganoderma Selective Medium, molecular identification and observation under the light microscope. Differences of the organic compounds released from oil palm tissues among different indices were studied using Gas Chromatography Mass Spectrometry (GC-MS). The GC-MS analysis showed the presence of eleven compounds: mainly, fatty acid methyl esters, and fatty acids, amino acids, esters, alcohols, alkaloids. Among them, dodecanoic acid, methyl ester, 9-Octadecenoic acid (Z)-, methyl ester and methyl stearate may have the potential as the biomarkers for detection of infection in oil palm caused by Ganoderma sp

Formulation of biofertilizers from oil palm empty fruit bunches and plant growth-promoting microbes: A comprehensive and novel approach towards plant health

Our heavy reliance on chemical fertilizers for agricultural practices has fostered the development of a vast industry that is producing chemicals that are toxic not only for humans but also for the environment. Biofertilizers are microbial formulations containing native plant growth-promoting microorganisms (PGPM) which have the potential to enhance plant growth either directly or indirectly by producing different types of phytohormones, iron-binding metabolites (siderophores), and solubilizing soil nutrients and minerals. The positive impacts on crop growth and development were documented by many researchers while using biofertilizers. Thus, biofertilizers offer enormous promise for sustainable agriculture, particularly in the face of climate change. Despite the growing interest in this technology, its entire potential remains untapped. This review collectively describes the potential use of empty fruit bunches (EFB) biomass as a biofertilizer for sustainable agricultural practices and the roles of plant growth-promoting microbes (PGPM) in plant growth and development. Attempts were also made to give insights into the oil palm industry in Malaysia and the nutrient profile of EFB biomass. We concluded that more research, fund and development activities are needed to improve traits of beneficial microbes that will potentially enhance the biological pathway of different biocompounds production and find solutions for the current issues related to converting EFB biomass into biofertilizers

Effects of Liming on Soil Properties and Its Roles in Increasing the Productivity and Profitability of the Oil Palm Industry in Malaysia

As global demand for edible oil increases, palm oil-producing countries in Southeast Asia are experiencing a rapid expansion of agricultural land for industrial oil palm cultivation by converting existing agricultural lands and some tropical rainforests; however, soil acidity and nutrient depletion are two major constraints in oil palm cultivation in the tropics. Several factors may cause soil acidification, including natural processes, industrial pollutants and extensive agricultural production. Soil acidity increases the leaching of many essential plant nutrient elements and the availability of toxic elements by modifying various geochemical and biological reactions in the soil. Even though acidic soil is less fertile, the productivity of tropical soil is among the highest in the world once the chemical constraints are removed by applying a sufficient quantity of lime and fertilizers. Lime is a widely used alkali to improve soil fertility by retaining nutrients, increasing soil biota, decreasing heavy-metal availability and potentially achieving resistance against Ganoderma disease at oil palm estates. Liming materials are not simple compounds with consistent chemical properties; thus, selecting the appropriate lime must be based on soil type and price compared to the products neutralizing value, composition, and fineness. Since the primary aim of liming is to improve soil pH, numerous reviews have been reported on the impacts of soil acidification, nutrient deficiencies and heavy-metal toxicity; however, no extensive review has been published that discusses the effects of liming on oil palm growth and yield. It is not enough to emphasize just soil impacts alone, and a thorough assessment must also be given on crops (oil palm) and soil biodiversity. This review synthesizes current understanding and introduces a holistic approach to provide insights into the far-reaching effects liming has on the biogeochemical properties of tropical soil and oil palm crops