Draft whole genome sequence of groundnut stem rot fungus Athelia rolfsii revealing genetic architect of its pathogenicity and virulence.
ABSTRACT: Groundnut (Arachis hypogaea L.) is an important oil seed crop having major biotic constraint in production due to stem rot disease caused by fungus, Athelia rolfsii causing 25-80% loss in productivity. As chemical and biological combating strategies of this fungus are not very effective, thus genome sequencing can reveal virulence and pathogenicity related genes for better understanding of the host-parasite interaction. We report draft assembly of Athelia rolfsii genome of ~73?Mb having 8919 contigs. Annotation analysis revealed 16830 genes which are involved in fungicide resistance, virulence and pathogenicity along with putative effector and lethal genes. Secretome analysis revealed CAZY genes representing 1085 enzymatic genes, glycoside hydrolases, carbohydrate esterases, carbohydrate-binding modules, auxillary activities, glycosyl transferases and polysaccharide lyases. Repeat analysis revealed 11171 SSRs, LTR, GYPSY and COPIA elements. Comparative analysis with other existing ascomycotina genome predicted conserved domain family of WD40, CYP450, Pkinase and ABC transporter revealing insight of evolution of pathogenicity and virulence. This study would help in understanding pathogenicity and virulence at molecular level and development of new combating strategies. Such approach is imperative in endeavour of genome based solution in stem rot disease management leading to better productivity of groundnut crop in tropical region of world.
Project description:Aconitum carmichaelii, commonly known as Fuzi, is a typical traditional Chinese medicine (TCM) herb that has been grown for more than one thousand years in China. Although root rot disease has been seriously threatening this crop in recent years, few studies have investigated root rot disease in Fuzi, and no pathogens have been identified. In this study, fungal libraries from rhizosphere soils were constructed by internal transcribed spacer (ITS) sequencing using the HiSeq 2500 high-throughput platform. A total of 948,843 tags were obtained from 17 soil samples, and these corresponded to 195,583,495 nt. At 97% identity, the libraries yielded 12,266 operational taxonomic units (OTUs), of which 97.5% could be annotated. In sick soils, Athelia, Mucor and Mortierella were the dominant fungi, comprising 10.3%, 10.1% and 7.7% of the fungal community, respectively. These fungi showed 2.6-, 1.53- to 6.31- and 1.38- to 2.65-fold higher enrichment in sick soils compared with healthy soils, and their high densities reduced the fungal richness in the areas surrounding the rotted Fuzi roots. An abundance analysis suggested that A. rolfsii and Mucor racemosus, as the dominant pathogens, might play important roles in the invading Fuzi tissue, and Phoma adonidicola could be another pathogenic fungus of root rot. In contrast, Mortierella chlamydospora, Penicillium simplicissimum, Epicoccum nigrum, Cyberlindnera saturnus and Rhodotorula ingeniosa might antagonize root rot pathogens in sick soils. In addition, A. rolfsii was further verified as a main pathogen of Fuzi root rot disease through hypha purification, morphological observation, molecular identification and an infection test. These results provide theoretical guidance for the prevention and treatment of Fuzi root rot disease.
Project description:Sclerotium rot was found on Cymbidium orchids at Seosan-si, Chungcheongnam-do, Korea, in July, 2010. Symptoms occurred on low leaves, which turned yellowish, after which the entire plant wilted. Severely infected plants were blighted and eventually died. White mycelial mats and sclerotia appeared on pseudobulbs. Based on the mycological characteristics and pathogenicity, the causal fungus was identified as Sclerotium rolfsii. This is the first report of new Sclerotium rot on Cymbidium spp. caused by S. rolfsii in Korea.
Project description:In this study, we characterized sporadically occurring sclerotium rot caused by Sclerotium rolfsii in Chinese chive (Allium tuberosum Roth.) in farm fields in Sacheon, Korea. The initial symptom of the disease was water-soaked, which progressed to rotting, wilting, blighting, and eventually death. Further, mycelial mats spread over the lesions near the soil line, and sclerotia formed on the scaly stem and leaves. The sclerotia were globoid, 1~3 mm, and white to brown. The optimum temperature for growth and sclerotia formation on potato dextrose agar (PDA) was 30℃. The diameter of the hypae ranged from 4 to 8 µm. Clamp connection was observed on PDA medium after 5 days of incubation. Based on the mycological characteristics, internal transcribed spacer sequence analysis, and pathogenicity test, the causal agent was identified as Sclerotium rolfsii Saccardo. This is the first report of sclerotium rot in Chinese chive caused by S. rolfsii in Korea.
Project description:Plants being sessile are under constant threat of multiple abiotic and biotic stresses within its natural habitat. A combined stress involving an abiotic and a biotic factor reportedly increases susceptibility of the plants to pathogens. The emerging threat, collar rot disease of chickpea (caused by Sclerotium rolfsii Sacc.) is reported to be influenced by soil moisture condition (SMC). Hence, we studied the influence of differential SMC viz. upper optimum (100%), optimum (80%), lower optimum (60%), and limiting (40%) soil moisture conditions on colonization and collar rot development over the course of infection in two chickpea cultivars, Annigeri (susceptible to collar rot) and ICCV 05530 (moderately resistant to collar rot). Disease incidence was found to be directly proportional to increase in soil moisture (R2 = 0.794). Maximum incidence was observed at 80% SMC, followed by 100 and 60% SMC. Expression of genes (qPCR analysis) associated with host cell wall binding (lectin) and degradation viz. endopolygalacturonase-2, endoglucosidase, and cellobiohydrolase during collar rot development in chickpea were relatively less at limiting soil moisture condition (40%) as compared to optimum soil moisture condition (80%). As compared to individual stress, the expression of defense response genes in chickpea seedlings were highly up-regulated in seedlings challenged with combined stress. Our qPCR results indicated that the expression of defense-related genes in chickpea during interaction with S. rolfsii at low SMC was primarily responsible for delayed disease reaction. Involvement of moisture and biotic stress-related genes in combined stress showed a tailored defense mechanism.
Project description:The application of antagonistic fungi for plant protection has attracted considerable interest because they may potentially replace the use of chemical pesticides. Antipathogenic activities confirmed in volatile organic compounds (VOCs) from microorganisms have potential to serve as biocontrol agents against pre- and post-harvest diseases. In the present study, we investigated <i>Galactomyces</i> fungi isolated from rotten leaves and the rhizosphere of cherry tomato (<i>Lycopersicon esculentum</i> var. <i>cerasiforme</i>). VOCs produced by <i>Galactomyces</i> fungi negatively affected the growth of phytopathogenic fungi and the survival of nematodes. Mycelial growths of all nine examined phytopathogenic fungi were inhibited on agar plate, although the inhibition was more intense in <i>Athelia rolfsii</i> JYC2163 and <i>Cladosporium cladosporioides</i> JYC2144 and relatively moderate in <i>Fusarium</i> sp. JYC2145. VOCs also efficiently suppressed the spore germination and mycelial growth of <i>A. rolfsii</i> JYC2163 on tomatoes. The soil nematode <i>Caenorhabditis elegans</i> exhibited higher mortality in 24?h in the presence of VOCs. These results suggest the broad-spectrum activity of <i>Galactomyces</i> fungi against various plant pathogens and the potential to use VOCs from <i>Galactomyces</i> as biocontrol agents.
Project description:Thirty-four endophytic fungal isolates were obtained from the leaves of the medicinal plant Polyscias fruticosa, and their antagonistic activities against the growth of the common tomatoes plant pathogenic fungus Athelia rolfsii were initially screened using a dual culture assay. The endophytic fungus MFLUCC 17-0313, which was later molecularly identified as Diatrype palmicola, displayed the highest inhibition percentage (49.98%) in comparison to the others. This fungus was then chosen for further evaluation. Its culture broth and mycelia from a 10 L scale were separated and extracted using ethyl acetate, methanol, and hexane. Each extract was tested for antifungal activity against the same pathogen using a disc diffusion assay. Only the crude hexane extract of fungal mycelium showed antifungal activity. The hexane extract was fractioned using sephadex gel filtration chromatography and each fraction was tested for antifungal activity until the one with the highest inhibition percentage was obtained. The bioactive compound was identified as 8-methoxynaphthalen-1-ol using nuclear magnetic resonance spectroscopy and mass spectrometry. The minimum inhibition concentration of 8-methoxynaphthalen-1-ol was demonstrated at 250 µg/mL against the selected pathogen. Using the leaf assay, the solution of 8-methoxynapthalen-1-ol was tested for phytotoxic activity against A. rolfsii and was found to have no phytotoxic effects. These results showed that 8-methoxynaphthalen-1-ol has the potential for controlling the growth of A. rolfsii, the cause of Southern blight disease on tomatoes. This study may provide the foundation for future use of this compound as a biofungicide.
Project description:The genome of Fusarium oxysporum (Fo) consists of a set of eleven 'core' chromosomes, shared by most strains and responsible for housekeeping, and one or several accessory chromosomes. We sequenced a strain of Fo f.sp. radicis-cucumerinum (Forc) using PacBio SMRT sequencing. All but one of the core chromosomes were assembled into single contigs, and a chromosome that shows all the hallmarks of a pathogenicity chromosome comprised two contigs. A central part of this chromosome contains all identified candidate effector genes, including homologs of SIX6, SIX9, SIX11 and SIX 13. We show that SIX6 contributes to virulence of Forc. Through horizontal chromosome transfer (HCT) to a non-pathogenic strain, we also show that the accessory chromosome containing the SIX gene homologs is indeed a pathogenicity chromosome for cucurbit infection. Conversely, complete loss of virulence was observed in Forc016 strains that lost this chromosome. We conclude that also a non-wilt-inducing Fo pathogen relies on effector proteins for successful infection and that the Forc pathogenicity chromosome contains all the information necessary for causing root rot of cucurbits. Three out of nine HCT strains investigated have undergone large-scale chromosome alterations, reflecting the remarkable plasticity of Fo genomes.
Project description:BACKGROUND: Xanthomonas campestris pathovar campestris (Xcc) is the causal agent of black rot disease of crucifers worldwide. The molecular genetic diversity and host specificity of Xcc are poorly understood. RESULTS: We constructed a microarray based on the complete genome sequence of Xcc strain 8004 and investigated the genetic diversity and host specificity of Xcc by array-based comparative genome hybridization analyses of 18 virulent strains. The results demonstrate that a genetic core comprising 3,405 of the 4,186 coding sequences (CDSs) spotted on the array are conserved and a flexible gene pool with 730 CDSs is absent/highly divergent (AHD). The results also revealed that 258 of the 304 proved/presumed pathogenicity genes are conserved and 46 are AHD. The conserved pathogenicity genes include mainly the genes involved in type I, II and III secretion systems, the quorum sensing system, extracellular enzymes and polysaccharide production, as well as many other proved pathogenicity genes, while the AHD CDSs contain the genes encoding type IV secretion system (T4SS) and type III-effectors. A Xcc T4SS-deletion mutant displayed the same virulence as wild type. Furthermore, three avirulence genes (avrXccC, avrXccE1 and avrBs1) were identified. avrXccC and avrXccE1 conferred avirulence on the hosts mustard cultivar Guangtou and Chinese cabbage cultivar Zhongbai-83, respectively, and avrBs1 conferred hypersensitive response on the nonhost pepper ECW10R. CONCLUSION: About 80% of the Xcc CDSs, including 258 proved/presumed pathogenicity genes, is conserved in different strains. Xcc T4SS is not involved in pathogenicity. An efficient strategy to identify avr genes determining host specificity from the AHD genes was developed.
Project description:The bacterial family Enterobacteriaceae is notable for its well studied human pathogens, including Salmonella, Yersinia, Shigella, and Escherichia spp. However, it also contains several plant pathogens. We report the genome sequence of a plant pathogenic enterobacterium, Erwinia carotovora subsp. atroseptica (Eca) strain SCRI1043, the causative agent of soft rot and blackleg potato diseases. Approximately 33% of Eca genes are not shared with sequenced enterobacterial human pathogens, including some predicted to facilitate unexpected metabolic traits, such as nitrogen fixation and opine catabolism. This proportion of genes also contains an overrepresentation of pathogenicity determinants, including possible horizontally acquired gene clusters for putative type IV secretion and polyketide phytotoxin synthesis. To investigate whether these gene clusters play a role in the disease process, an arrayed set of insertional mutants was generated, and mutations were identified. Plant bioassays showed that these mutants were significantly reduced in virulence, demonstrating both the presence of novel pathogenicity determinants in Eca, and the impact of functional genomics in expanding our understanding of phytopathogenicity in the Enterobacteriaceae.