Whole-genome sequencing of Ganoderma boninense, the causal agent of basal stem rot disease in oil palm, via combined short- and long-read sequencing.

The hemibiotrophic Basidiomycete pathogen Ganoderma boninense (Gb) is the dominant causal agent of oil palm basal stem rot disease. Here, we report a complete chromosomal genome map of Gb using a combination of short-read Illumina and long-read Pacific Biosciences (PacBio) sequencing platforms combined with chromatin conformation capture data from the Chicago and Hi-C platforms. The genome was 55.87 Mb in length and assembled to a high contiguity (N50: 304.34 kb) of 12 chromosomes built from 112 scaffolds, with a total of only 4.34 Mb (~ 7.77%) remaining unplaced. The final assemblies were evaluated for completeness of the genome by using Benchmarking Universal Single Copy Orthologs (BUSCO) v4.1.4, and based on 4464 total BUSCO polyporales group searches, the assemblies yielded 4264 (95.52%) of the conserved orthologs as complete and only a few fragmented BUSCO of 42 (0.94%) as well as a missing BUSCO of 158 (3.53%). Genome annotation predicted a total of 21,074 coding genes, with a GC content ratio of 59.2%. The genome features were analyzed with different databases, which revealed 2471 Gene Ontology/GO (11.72%), 5418 KEGG (Kyoto Encyclopedia of Genes and Genomes) Orthologous/KO (25.71%), 13,913 Cluster of Orthologous Groups of proteins/COG (66.02%), 60 ABC transporter (0.28%), 1049 Carbohydrate-Active Enzymes/CAZy (4.98%), 4005 pathogen-host interactions/PHI (19%), and 515 fungal transcription factor/FTFD (2.44%) genes. The results obtained in this study provide deep insight for further studies in the future.

Population genetics provides insights into the important agronomic traits of Ganoderma cultivation varieties in China.

This study aimed to investigate the species diversity and genetic differentiation of the genome of the main cultivated strains of Ganoderma in China. Population genomics analysis was conducted based on 150 cultivated strains of Ganoderma collected nationwide. The results indicated that the main species currently cultivated in China were Ganoderma sichuanense and Ganoderma lucidum, with a minor proportion of Ganoderma sessile, Ganoderma weberianum, Ganoderma sinense, Ganoderma gibbosum and Ganoderma australe. A total of 336,506 high-quality single nucleotide polymorphism (SNP) loci were obtained through population evolution analysis. The Fst values were calculated using a 5-kb sliding window, which ranged from 0.11 to 0.74. This suggests varying degrees of genetic differentiation between populations and genetic exchange among varieties. On this basis, the genes related to the stipe length, cap color and branch phenotypes of Ganoderma were excavated, and the region with the top 1% ZFst value region was used as a candidate region. A total of 137, 270 and 222 candidate genes were identified in the aforementioned 3 phenotypes, respectively. Gene annotation revealed that genes associated with stipe length were mainly related to cell division and differentiation, including proteins such as Nse4 protein and DIM1 protein. The genes related to Ganoderma red color were mainly related to the metabolism of tryptophan and flavonoids. The genes related to the branch were mainly related to cytokinin synthesis, ABC transporter and cytochrome P450. This study provided 150 valuable genome resequencing data in assessing the diversity and genetic differentiation of Ganoderma and laid a foundation for agronomic trait analysis and the development of new varieties of Ganoderma.

Transcriptional effects of carbon and nitrogen starvation on Ganoderma boninense, an oil palm phytopathogen.

BACKGROUND: Ganoderma boninense is a phytopathogen of oil palm, causing basal and upper stem rot diseases. METHODS: The genome sequence was used as a reference to study gene expression during growth in a starved carbon (C) and nitrogen (N) environment with minimal sugar and sawdust as initial energy sources. This study was conducted to mimic possible limitations of the C-N nutrient sources during the growth of G. boninense in oil palm plantations. RESULTS: Genome sequencing of an isolate collected from a palm tree in West Malaysia generated an assembly of 67.12 Mb encoding 19,851 predicted genes. Transcriptomic analysis from a time course experiment during growth in this starvation media identified differentially expressed genes (DEGs) that were found to be associated with 29 metabolic pathways. During the active growth phase, 26 DEGs were related to four pathways, including secondary metabolite biosynthesis, carbohydrate metabolism, glycan metabolism and mycotoxin biosynthesis. G. boninense genes involved in the carbohydrate metabolism pathway that contribute to the degradation of plant cell walls were up-regulated. Interestingly, several genes associated with the mycotoxin biosynthesis pathway were identified as playing a possible role in pathogen-host interaction. In addition, metabolomics analysis revealed six metabolites, maltose, xylobiose, glucooligosaccharide, glycylproline, dimethylfumaric acid and arabitol that were up-regulated on Day2 of the time course experiment. CONCLUSIONS: This study provides information on genes expressed by G. boninense in metabolic pathways that may play a role in the initial infection of the host.

Identification a novel Ganoderma FIP gene from Ganoderma capense and its functional expression in Pichia pastoris.

Ganoderma capense is a precious medicinal fungus in China. In this study, a novel fungal immunomodulatory protein gene, named as FIP-gca, was cloned from G. capense by homologous cloning. Sequencing analysis indicated that FIP-gca was composed of 336 bp, which encoded a polypeptide of 110 amino acids. Protein sequence blasting and phylogenetic analysis showed that FIP-gca shared homology with other Ganoderma FIPs. FIP-gca was effectively expressed in Pichia pastoris GS115 at an expression level of 166.8 mg/L and purified using HisTrap™ fast-flow prepack columns. The immunomodulation capacity of rFIP-gca was demonstrated by that rFIP-gca could obviously stimulate cell proliferation and increase IL-2 secretion of murine spleen lymphocytes. Besides, antitumor activity of rFIP-gca towards human stomach cancer AGS cell line was evaluated in vitro. Cell wound scratch assay proved that rFIP-gca could inhibit migration of AGS cells. And flow cytometry assay revealed that rFIP-gca could significantly induce apoptosis of AGS cells. rFIP-gca was able to induce 18.12% and 22.29% cell apoptosis at 0.3 µM and 0.6 µM, respectively. Conclusively, the novel FIP-gca gene from G. capense has been functionally expressed in Pichia and rFIP-gca exhibited ideal immunomodulation and anti-tumour activities, which implies its potential application and study in future.

Increased production and anti-senescence activity of exopolysaccharides in Ganoderma lingzhi by co-overexpression of ß-1,3-glucan synthase and UDP-glucose pyrophosphorylase.

A ß-1,3-glucan synthase gene (gls) was cloned and overexpressed in Ganoderma lingzhi. The content of intracellular polysaccharides (IPS) in G. lingzhi overexpressing gls was 22.36 mg/100 mg dry weight (DW), 19 % higher than those in the wild-type (WT) strain. Overexpression of gls did not affect the expression of the phosphoglucomutase gene and the UDP-glucose pyrophosphorylase gene (ugp) in the polysaccharide biosynthesis. The gls and ugp were then simultaneously overexpressed in G. lingzhi for the first time. The combined overexpression of these two genes increased the IPS content and exopolysaccharides (EPS) production to a greater extent than the overexpression of gls independently. The maximum IPS content of the overexpressed strain was 24.61 mg/100 mg, and the maximum EPS production was 1.55 g/L, 1.31- and 1.50-fold higher than that in the WT strain, respectively. Moreover, the major EPS fractions from the overexpression strain contained more glucose (86.7 % and 72.5 %) than those from the WT strain (78.2 % and 62.9 %). Furthermore, the major fraction G+U-0.1 from the overexpression strain exhibited stronger antioxidant and anti-senescence activities than the WT-0.1 fraction from the WT strain. These findings will aid in the hyperproduction and application of Ganoderma polysaccharides and facilitate our understanding of mushroom polysaccharide biosynthesis.

Phylogenetic analysis and morphology of Ganoderma multipileum, a Ganoderma species associated with dieback of the metropolitan woody plant Delonix regia (Boj. ex Hook.) Raf. in Vietnam.

Ganoderma multipileum, a wood decay mushroom, was initially discovered and classified in Taiwan through the analysis of its morphology and the internal transcribed spacer (ITS) region of nuclear ribosomal DNA. In this study, we identified a mushroom associated with the dieback of Delonix regia (Boj. ex Hook.) Raf., a woody ornamental street tree in Vietnam, as Ganoderma multipileum. This classification was based on phylogenetic analysis of ITS, RPB2, and TEF1 sequences, as well as morphology assessment and scanning electron microscope observation of basidiospores. The phylogenetic analysis revealed that the specimens collected in Vietnam formed a monophyletic group of Ganoderma multipileum with a high bootstrap value and posterior probability (100%/1.00). Furthermore, the morphological features consistent with laccate Ganoderma, including a thin pileipellis composed of enlarged and bulbous hyphae, and the basidiomes exhibited two different phenotypes. Notably, scanning electron microscopy of the basidiospores revealed ovoid spores with numerous echinules, providing the first documented evidence of this characteristic for Ganoderma multipileum. This research represents the first recorded instance of Ganoderma multipileum in Vietnam associated with the dieback of Delonix regia.

Exploring the potential of Bornean polypore fungi as biological control agents against pathogenic Ganoderma boninense causing basal stem rot in oil palm.

Basal stem rot due to a fungal pathogen, Ganoderma boninense, is one of the most devastating diseases in oil palm throughout the major palm oil producer countries. This study investigated the potential of polypore fungi as biological control agents against pathogenic G. boninense in oil palm. In vitro antagonistic screening of selected non-pathogenic polypore fungi was performed. Based on in planta fungi inoculation on oil palm seedlings, eight of the 21 fungi isolates tested (GL01, GL01, RDC06, RDC24, SRP11, SRP12, SRP17, and SRP18) were non-pathogenic. In vitro antagonistic assays against G. boninense revealed that the percentage inhibition of radial growth (PIRG) in dual culture assay for SRP11 (69.7%), SRP17 (67.3%), and SRP18 (72.7%) was relatively high. Percentage inhibition of diameter growth (PIDG) in volatile organic compounds (VOCs) in dual plate assay of SRP11, SRP17, and SRP18 isolates were 43.2%, 51.6%, and 52.1%, respectively. Molecular identification using the internal transcribed spacer gene sequences of SRP11, SRP17, and SRP18 isolates revealed that they were Fomes sp., Trametes elegans, and Trametes lactinea, respectively.

Comparative proteomic analysis of two divergent strains provides insights into thermotolerance mechanisms of Ganoderma lingzhi.

Heat stress (HS) is a major abiotic factor influencing fungal growth and metabolism. However, the genetic basis of thermotolerance in Ganoderma lingzhi (G. lingzhi) remains largely unknown. In this study, we investigated the thermotolerance capacities of 21 G. lingzhi strains and screened the thermo-tolerant (S566) and heat-sensitive (Z381) strains. The mycelia of S566 and Z381 were collected and subjected to a tandem mass tag (TMT)-based proteome assay. We identified 1493 differentially expressed proteins (DEPs), with 376 and 395 DEPs specific to the heat-tolerant and heat-susceptible genotypes, respectively. In the heat-tolerant genotype, upregulated proteins were linked to stimulus regulation and response. Proteins related to oxidative phosphorylation, glycosylphosphatidylinositol-anchor biosynthesis, and cell wall macromolecule metabolism were downregulated in susceptible genotypes. After HS, the mycelial growth of the heat-sensitive Z381 strain was inhibited, and mitochondrial cristae and cell wall integrity of this strain were severely impaired, suggesting that HS may inhibit mycelial growth of Z381 by damaging the cell wall and mitochondrial structure. Furthermore, thermotolerance-related regulatory pathways were explored by analyzing the protein-protein interaction network of DEPs considered to participate in the controlling the thermotolerance capacity. This study provides insights into G. lingzhi thermotolerance mechanisms and a basis for breeding a thermotolerant germplasm bank for G. lingzhi and other fungi.

The bHLH-zip transcription factor SREBP regulates triterpenoid and lipid metabolisms in the medicinal fungus Ganoderma lingzhi.

Ganoderic acids (GAs) are well recognized as important pharmacological components of the medicinal species belonging to the basidiomycete genus Ganoderma. However, transcription factors directly regulating the expression of GA biosynthesis genes remain poorly understood. Here, the genome of Ganoderma lingzhi is de novo sequenced. Using DNA affinity purification sequencing, we identify putative targets of the transcription factor sterol regulatory element-binding protein (SREBP), including the genes of triterpenoid synthesis and lipid metabolism. Interactions between SREBP and the targets are verified by electrophoretic mobility gel shift assay. RNA-seq shows that SREBP targets, mevalonate kinase and 3-hydroxy-3-methylglutaryl coenzyme A synthetase in mevalonate pathway, sterol isomerase and lanosterol 14-demethylase in ergosterol biosynthesis, are significantly upregulated in the SREBP overexpression (OE::SREBP) strain. In addition, 3 targets involved in glycerophospholipid/glycerolipid metabolism are upregulated. Then, the contents of mevalonic acid, lanosterol, ergosterol and 13 different GAs as well as a variety of lipids are significantly increased in this strain. Furthermore, the effects of SREBP overexpression on triterpenoid and lipid metabolisms are recovered when OE::SREBP strain are treated with exogenous fatostatin, a specific inhibitor of SREBP. Taken together, our genome-wide study clarify the role of SREBP in triterpenoid and lipid metabolisms of G. lingzhi.

Induced expression of Ganoderma boninense Lanosterol 14α-Demethylase (ERG11) during interaction with oil palm.

BACKGROUND: The basidiomycete fungus, Ganoderma boninense is the main contributor to oil palm Basal Stem Rot (BSR) in Malaysia and Indonesia. Lanosterol 14α-Demethylase (ERG11) is a key enzyme involved in biosynthesis of ergosterol, which is an important component in the fungal cell membrane. The Azole group fungicides are effective against pathogenic fungi including G. boninense by inhibiting the ERG11 activity. However, the work on molecular characterization of G. boninense ERG11 is still unavailable today. METHODS AND RESULTS: This study aimed to isolate and characterize the full-length cDNA encoding ERG11 from G. boninense. The G. boninense ERG11 gene expression during interaction with oil palm was also studied. A full-length 1860 bp cDNA encoding ERG11 was successfully isolated from G. boninense. The G. boninense ERG11 shared 91% similarity to ERG11 from other basidiomycete fungi. The protein structure homology modeling of GbERG11 was analyzed using the SWISS-MODEL workspace. Southern blot and genome data analyses showed that there is only a single copy of ERG11 gene in the G. boninense genome. Based on the in-vitro inoculation study, the ERG11 gene expression in G. boninense has shown almost 2-fold upregulation with the presence of oil palm. CONCLUSION: This study provided molecular information and characterization study on the G. boninense ERG11 and this knowledge could be used to design effective control measures to tackle the BSR disease of oil palm.

Multilocus, Multiplex Detection of Ganoderma zonatum from Environmental Samples.

Ganoderma butt rot of palms is caused by a white rot basidiomycete fungus, Ganoderma zonatum. Typical symptoms include wilting of fronds that starts in the lower canopy and moves to the top. As wilting symptoms are also associated with other diseases and disorders, appearance of basidiomata on the trunks is necessary to confirm this disease. Basidiomata develop late in the disease cycle, making early diagnostics challenging. Here, we describe a DNA-based molecular diagnostic assay that could be used to confirm the presence of G. zonatum in palm trunks before conks are observed. Primers tailored to end on single-nucleotide polymorphisms (SNPs), that differentiate G. zonatum from 14 other Ganoderma taxa, were designed from multiple regions in four genes: internal transcribed spacer (ITS), RNA polymerase 1 (rpb1), rpb2, and translation elongation factor 1-α (tef1-α). A set of three primer pairs could successfully determine the incidence of G. zonatum with high specificity and sensitivity in different environmental samples such as sawdust collected from naturally infected palm trunks and soil samples containing G. zonatum basidiospores. This rapid PCR-based assay could potentially be used to detect inoculum sources of the fungus and track its movement and survival in different palm tissues and environments. Early detection of G. zonatum is a crucial step toward building and implementing better disease management strategies and mitigating potential risks from palm failures due to decay.

Intraspecific comparison of mitochondrial genomes reveals the evolution in medicinal fungus Ganoderma lingzhi.

Several mitogenomes of the genus Ganoderma have been assembled, but intraspecific comparisons of mitogenomes in Ganoderma lingzhi have not been reported. In this study, 19 G. lingzhi mitogenomes were assembled and analyzed combined with three mitogenomes of G. lingzhi from GenBank in term of the characteristics, evolution, and phylogeny. The results showed that the mitogenomes of the G. lingzhi strains are closed circular ranging from 49.23 kb to 68.37 kb. The genetic distance, selective pressure, and base variation indicate that the 14 common protein coding genes were highly conserved. The differences in introns, open reading frames, and repetitive sequences in the mitogenome were the main factors leaded to the variations in mitogenome. The introns were horizontally transferred in mitogenomes, and the differences between introns in the same insertion, which were primarily caused by the repetitive sequence, showed that the introns may be under degeneration. Besides, the frequent insertion and deletion of introns showed an evolutionary rate faster than protein coding genes. Phylogenetic analysis showed that the G. lingzhi strains gathered with high support, and those with the same intron distribution law had closer clustering relationships.

Intraspecific Identification of Lingzhi Medicinal Mushroom, Ganoderma lingzhi (Agaricomycetes), Using ISSR Markers.

This study aimed to obtain a set of specific inter simple sequence repeat (ISSR) primers and establish a stable and accurate intraspecific identification method for Ganoderma lingzhi. A total of 117 G. lingzhi strains were identified using internal transcribed spacer sequences from 147 strains determined as G. lingzhi via simple morphological identification. Based on the sequences obtained, specific ISSR primers for G. lingzhi were screened and validated, and 15 specific ISSR primers showed polymorphic banding pattern with clear band resolution. Subsequently, ISSR PCRs of the 15 specific primers were performed for the 117 G. lingzhi strains. As expected, DNA analysis of the ISSR markers could distinguish G. lingzhi strains, with similarity coefficients ranging from 0.11 to 0.89. Thus, the 15 specific ISSR primers can be used for intraspecific identification and polymorphism analysis of G. lingzhi.

Optimization, purification and characterization of laccase from Ganoderma leucocontextum along with its phylogenetic relationship.

The aim of this work to study an efficient laccase producing fungus Ganoderma leucocontextum, which was identified by ITS regions of DNA and phylogenetic tree was constructed. This study showed the laccase first-time from G. leucocontextum by using medium containing guaiacol. The growth cultural (pH, temperature, incubation days, rpm) and nutritional (carbon and nitrogen sources) conditions were optimized, which enhanced the enzyme production up to 4.5-folds. Laccase production increased 855 U/L at 40 °C. The pH 5.0 was suitable for laccase secretion (2517 U/L) on the 7th day of incubation at 100 rpm (698.3 U/L). Glucose and sucrose were good carbon source to enhance the laccase synthesis. The 10 g/L beef (4671 U/L) and yeast extract (5776 U/L) were the best nitrogen source for laccase secretion from G. leucocontextum. The laccase was purified from the 80% ammonium sulphate precipitations of protein identified by nucleotides sequence. The molecular weight (65.0 kDa) of purified laccase was identified through SDS and native PAGE entitled as Glacc110. The Glacc110 was characterized under different parameters. It retained > 90% of its activity for 16 min incubation at 60 °C in acidic medium (pH 4.0). This enzyme exerted its optimal activity at pH 3.0 and temperature 70 °C with guaiacol substrate. The catalytic parameters Km and Vmax was 1.658 (mM) and 2.452 (mM/min), respectively. The thermo stability of the laccase produced by submerged fermentation of G. leucocontextum has potential for industrial and biotechnology applications. The results remarked the G. leucocontextum is a good source for laccase production.

Revision of Ganoderma species associated with stem rot of coconut palm.

Stem rot by Ganoderma is a disease of major concern for coconut farmers. Many species (G. applanatum, G. boninense, G. lucidum, G. zonatum) have been implicated as the causal agents of the disease. Despite its importance, systematics of the rot-associated Ganoderma remains uncertain and unresolved. Morphologically heterogeneous basidiomata of the putative pathogen(s) from infected palms in multiple disease sites were collected during an outbreak in the coastal state of India, Kerala. Morphological and molecular investigations revealed that these specimens were distinct from all the Ganoderma species so far identified and reported from coconut. Although with a close morphological resemblance to many Ganoderma species collected from palms, they exhibited unique genetics and geographic distribution patterns. We present a taxonomic reassessment of the species collected from infected coconut palms in India and also propose two new species, Ganoderma keralense and G. pseudoapplanatum. A taxonomic key to Ganoderma species on palms is given.

Analysis of rhizobacterial community associated with the occurrence of Ganoderma basal stem rot disease in oil palm by Illumina next-generation sequencing.

The fungus Ganoderma boninense is a causal pathogen of basal stem rot, a serious disease of oil palm plantation systems. As previously observed, some oil palm trees show no appearance of disease symptoms (asymptomatic oil palm), although they have grown close to a tree that showed severe symptoms of basal stem rot disease (symptomatic oil palm). The microbial community difference between asymptomatic and symptomatic oil palm will help understand disease suppression. Thus, in this study, rhizosphere soil was sampled around asymptomatic (OP - G) and symptomatic (OP + G) oil palm trees in Ganoderma-infected oil palm orchards. Illumina next-generation sequencing (NGS), bioinformatics analysis, bacterial diversity, and soil physicochemical properties were evaluated. The results demonstrated that soil physicochemical properties and species richness around rhizosphere soil of OP - G and OP + G samples were not significantly different. The age of the oil palm trees and oil palm variety showed negligible correlation and were not significant with bacterial diversity. However, the top ten most abundant analysis of the bacterial communities showed that phyla Actinobacteria and Firmicutes were significantly increased in rhizosphere soil around OP - G samples relative to the OP+ G samples. The unique operational taxonomic units (OTUs) of OP - G (2137) were higher than in the OP+ G samples (1747 OTUs). These bacterial communities have been reported as biological control agents and/or plant growth-promoting rhizosphere bacteria that are related to disease suppression. Thus, the data provided are useful for developing suppressive soil to biologically control G. boninense.

Whole-genome assembly of Ganoderma leucocontextum (Ganodermataceae, Fungi) discovered from the Tibetan Plateau of China.

Ganoderma leucocontextum, a newly discovered species of Ganodermataceae in China, has diverse pharmacological activities. Ganoderma leucocontextum was widely cultivated in southwest China, but the systematic genetic study has been impeded by the lack of a reference genome. Herein, we present the first whole-genome assembly of G. leucocontextum based on the Illumina and Nanopore platform from high-quality DNA extracted from a monokaryon strain (DH-8). The generated genome was 50.05 Mb in size with an N50 scaffold size of 3.06 Mb, 78,206 coding sequences, and 13,390 putative genes. Genome completeness was assessed using the Benchmarking Universal Single-Copy Orthologs (BUSCO) tool, which identified 96.55% of the 280 Fungi BUSCO genes. Furthermore, differences in functional genes of secondary metabolites (terpenoids) were analyzed between G. leucocontextum and Ganoderma lucidum. Ganoderma leucocontextum has more genes related to terpenoids synthesis compared to G. lucidum, which may be one of the reasons why they exhibit different biological activities. This is the first genome assembly and annotation for G. leucocontextum, which would enrich the toolbox for biological and genetic studies in G. leucocontextum.

Identification of non-ribosomal peptide synthetase in Ganoderma boninense Pat. that was expressed during the interaction with oil palm.

Basal stem rot (BSR) of oil palm is a disastrous disease caused by a white-rot fungus Ganoderma boninense Pat. Non-ribosomal peptides (NRPs) synthesized by non-ribosomal peptide synthetases (NRPSs) are a group of secondary metabolites that act as fungal virulent factors during pathogenesis in the host. In this study, we aimed to isolate NRPS gene of G. boninense strain UPMGB001 and investigate the role of this gene during G. boninense-oil palm interaction. The isolated NRPS DNA fragment of 8322 bp was used to predict the putative peptide sequence of different domains and showed similarity with G. sinense (85%) at conserved motifs of three main NRPS domains. Phylogenetic analysis of NRPS peptide sequences demonstrated that NRPS of G. boninense belongs to the type VI siderophore family. The roots of 6-month-old oil palm seedlings were artificially inoculated for studying NRPS gene expression and disease severity in the greenhouse. The correlation between high disease severity (50%) and high expression (67-fold) of G. boninense NRPS gene at 4 months after inoculation and above indicated that this gene played a significant role in the advancement of BSR disease. Overall, these findings increase our knowledge on the gene structure of NRPS in G. boninense and its involvement in BSR pathogenesis as an effector gene.

Genetic diversity and gene flow amongst admixed populations of Ganoderma boninense, causal agent of basal stem rot in African oil palm (Elaeis guineensis Jacq.) in Sarawak (Malaysia), Peninsular Malaysia, and Sumatra (Indonesia).

In 1911 and 1917, the first commercial plantings of African oil palm (Elaeis guineensis Jacq.) were made in Indonesia and Malaysia in Southeast Asia. In less than 15 years, basal stem rot (BSR) was reported in Malaysia. It took nearly another seven decades to identify the main causal agent of BSR as the fungus, Ganoderma boninense. Since then, research efforts have focused on understanding G. boninense disease epidemiology, biology, and etiology, but limited progress was made to characterize pathogen genetic diversity, spatial structure, pathogenicity, and virulence. This study describes pathogen variability, gene flow, population differentiation, and genetic structure of G. boninense in Sarawak (Malaysia), Peninsular Malaysia, and Sumatra (Indonesia) inferred by 16 highly polymorphic cDNA-SSR (simple sequence repeat) markers. Marker-inferred genotypic diversity indicated a high level of pathogen variability among individuals within a population and among different populations. This genetic variability is clearly the result of outcrossing between basidiospores to produce recombinant genotypes. Although our results indicated high gene flow among the populations, there was no significant genetic differentiation among G. boninense populations on a regional scale. It suggested that G. boninense genetic makeup is similar across a wide region. Furthermore, our results revealed the existence of three admixed genetic clusters of G. boninense associated with BSR-diseased oil palms sampled throughout Sarawak, Peninsular Malaysia, and Sumatra. We postulate that the population structure is likely a reflection of the high genetic variability of G. boninense populations. This, in turn, could be explained by highly successful outcrossing between basidiospores of G. boninense from Southeast Asia and introduced genetic sources from various regions of the world, as well as regional adaptation of various pathogen genotypes to different palm hosts. Pathogen variability and population structure could be employed to deduce the epidemiology of G. boninense, as well as the implications of plantation cultural practices on BSR disease control in different regions.

Transcriptional profile of oil palm pathogen, Ganoderma boninense, reveals activation of lignin degradation machinery and possible evasion of host immune response.

BACKGROUND: The white-rot fungi in the genus Ganoderma interact with both living and dead angiosperm tree hosts. Two Ganoderma species, a North American taxon, G. zonatum and an Asian taxon, G. boninense, have primarily been found associated with live palm hosts. During the host plant colonization process, a massive transcriptional reorganization helps the fungus evade the host immune response and utilize plant cell wall polysaccharides. RESULTS: A publicly available transcriptome of G. boninense - oil palm interaction was surveyed to profile transcripts that were differentially expressed in planta. Ten percent of the G. boninense transcript loci had altered expression as it colonized oil palm plants one-month post inoculation. Carbohydrate active enzymes (CAZymes), particularly those with a role in lignin degradation, and auxiliary enzymes that facilitate lignin modification, like cytochrome P450s and haloacid dehalogenases, were up-regulated in planta. Several lineage specific proteins and secreted proteins that lack known functional domains were also up-regulated in planta, but their role in the interaction could not be established. A slowdown in G. boninense respiration during the interaction can be inferred from the down-regulation of proteins involved in electron transport chain and mitochondrial biogenesis. Additionally, pathogenicity related genes and chitin degradation machinery were down-regulated during the interaction indicating G. boninense may be evading detection by the host immune system. CONCLUSIONS: This analysis offers an overview of the dynamic processes at play in G. boninense - oil palm interaction and provides a framework to investigate biology of Ganoderma fungi across plantations and landscape.