Project description:BACKGROUND: The plant pathogenic basidiomycete Sclerotium rolfsii produces the industrially exploited exopolysaccharide scleroglucan, a polymer that consists of (1 --> 3)-beta-linked glucose with a (1 --> 6)-beta-glycosyl branch on every third unit. Although the physicochemical properties of scleroglucan are well understood, almost nothing is known about the genetics of scleroglucan biosynthesis. Similarly, the biosynthetic pathway of oxalate, the main by-product during scleroglucan production, has not been elucidated yet. In order to provide a basis for genetic and metabolic engineering approaches, we studied scleroglucan and oxalate biosynthesis in S. rolfsii using different transcriptomic approaches. RESULTS: Two S. rolfsii transcriptomes obtained from scleroglucan-producing and scleroglucan-nonproducing conditions were pooled and sequenced using the 454 pyrosequencing technique yielding approximately 350,000 reads. These could be assembled into 21,937 contigs and 171,833 singletons, for which 6,951 had significant matches in public protein data bases. Sequence data were used to obtain first insights into the genomics of scleroglucan and oxalate production and to predict putative proteins involved in the synthesis of both metabolites. Using comparative transcriptomics, namely Agilent microarray hybridization and suppression subtractive hybridization, we identified approximately 800 unigenes which are differently expressed under scleroglucan-producing and non-producing conditions. From these, candidate genes were identified which could represent potential leads for targeted modification of the S. rolfsii metabolism for increased scleroglucan yields. CONCLUSIONS: The results presented in this paper provide for the first time genomic and transcriptomic data about S. rolfsii and demonstrate the power and usefulness of combined transcriptome sequencing and comparative microarray analysis. The data obtained allowed us to predict the biosynthetic pathways of scleroglucan and oxalate synthesis and to identify important genes putatively involved in determining scleroglucan yields. Moreover, our data establish the first sequence database for S. rolfsii, which allows research into other biological processes of S. rolfsii, such as host-pathogen interaction.
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: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:Physalis minima is an herbaceous plant and inhabitant of the porous and organic matter containing soil of bunds in crop fields, wastelands, around the houses, and on the roadsides. S. rolfsii is soil borne and it can infect over 500 plant species of different families. It is of interest to study the pathogenesis of S. rolfsii on P. minima. The S. rolfsii isolated from P. minima (physr1) was characterized by morphology and sequence of Internal Transcribed Spacer (ITS) region. The population structure determination and phylogenetic analysis showed the isolate physr1 significantly differs from other isolates. The null hypothesis of equal evolutionary rate was rejected throughout the Maximum likelihood (ML) tree topology of different S. rolfsii ITS sequences. The site-specific mean (relative) evolutionary rate analysis showed that most of the sites (80.59 % sites) evolved at a slower rate than average. Finally, the result of Tajima's neutrality test indicated that the population of S. rolfsii has recently begun to expand and that's why the pathogen was infecting the new host P. minima and pose a serious threat of infecting several other cropped and non-cropped hosts.
Project description:Sclerotium rolfsii, which causes southern blight in a wide variety of crops, is a devastating plant pathogen worldwide. Mycoviruses that induce hypovirulence in phytopathogenic fungi are potential biological control resources against fungal plant diseases. However, in S. rolfsii, mycoviruses are rarely reported. In a previous study, we found a hypovirulent strain carrying a diverse pattern of dsRNAs. Here, we utilized the RNA_Seq technique to detect viral sequences. Deep sequencing, RT-PCR and Sanger sequencing validation analyses revealed that this strain harbors various new viral species that show affinity to the distinctly established and proposed families Benyviridae, Endornaviridae, Fusariviridae, Hypoviridae, and Fusagraviridae. Moreover, some viral sequences that could not be assigned to any of the existing families or orders were also identified and showed similarities to the Alphavirus, Ourmiavirus, phlegivirus-like and Curvularia thermal tolerance virus-like groups. In addition, we also conducted deep sequencing analysis of small RNAs in the virus-infecting fugal strain. The results indicated that the Dicer-mediated gene silencing mechanism was present in S. rolfsii. This is the first report of viral diversity in a single S. rolfsii fungal strain, and the results presented herein might provide insight into the taxonomy and evolution of mycoviruses and be useful for the exploration of mycoviruses as biocontrol agents.
Project description: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:Glyoxylate dehydrogenase (glyoxylate:NAD+ oxidoreductase) was purified 600-fold in three steps from crude extracts of the fungus Sclerotium rolfsii (Corticium rolfsii Curzi). Two of the purification steps involved dye-affinity chromatography. The enzyme is a tetramer of Mr 250 000, with identical subunits of Mr 57 000. Inhibition studies suggest that there is one essential thiol group per active site.