Members of the form genus Rhizoctonia D.C. are considered as a complex mixture of filamentous fungi, having in common the possession of a non-spored imperfect state, usually referred to as the Rhizoctonia anamorph. The group includes several of the most devastating crop pathogens like Thanatephorus cucumeris (Frank) Donk (anamorph=Rhizoctonia solani Kuhn), the majority of orchid mycorrhizal symbionts (mainly belonging to genus Ceratobasidium D.P. Rogers) and a collection of saprotrophic organisms of different systematic placement. The Rhizoctonia anamorph is characterized by several common features present among members of the entire Rhizoctonia species complex. Taxa from the group have been rearranged into several groups of higher fungi, including both Ascomycota and Basidiomycota, and split into several genera, employing criteria such as the analysis and ultrastructural comparison of septal apparatus. Until very recently, classification for some of the groups within the complex has been exclusively based on criteria such as hyphal anastomosis, since other types of diagnostic features are usually scarce in these fungi. Phytopathological studies in the complex have represented the major contingent of contributions in the group, especially in the case of R. solani. Some members of the complex have been reported to be protective isolates against pathogenic members of Rhizoctonia and some other fungal pathogens. This review focuses on the knowledge of several aspects of the species of Rhizcotonia s. lato, such as its current taxonomic placement, the biology and systematics of some groups of the complex, and a revision of the methodologies employed in studying it.

http://digital.csic.es/bitstream/10261/15813/3/Rhizoctonia.pdf

Review. Biology and systematics of the form genus Rhizoctonia

Population Biology of the Rhizoctonia solani Complex.

A specific and sensitive PCR assay was developed for the detection and identification of Rhizoctonia solani AG-3, the main causal pathogen of stem canker and black scurf of potato. A conventional primer set (Rs1F2 and Rs2R1) was designed from the nuclear ribosomal internal transcribed spacer (ITS1 and ITS2) regions of R. solani. Following PCR amplification, a 0·5-kb product was amplified from DNA of all isolates of AG-3 using primers Rs1F2 and Rs2R1. No product was amplified when DNA from isolates belonging to a range of other R. solani anastomosis groups or from a selection of other potato pathogens was tested, confirming the specificity of the primers for AG-3 only. Rhizoctonia solani AG-3 was also detected in potato tissue with varying black scurf severity, and in soil inoculated with sclerotia of R. solani to a minimum detection level of 5 × 10−4 g sclerotia/g soil. In addition, specific primers RsTqF1 (based on the Rs1F2 sequence) and RsTqR1, and a TaqMan™ fluorogenic probe RQP1, were designed to perform real-time quantitative (TaqMan) PCR. The conventional PCR and real-time PCR assays were compared and combined with direct DNA extraction from soil and a seed-baiting method to determine the most reliable method for the detection and quantification of AG-3 in both artificially inoculated field soil and naturally infested soils. It was shown that direct DNA extractions from soil could be problematic, although AG-3 was detectable using this method combined with the real-time PCR assay. The amplification of Rhizoctonia solani by seed baiting increased the sensitivity of the assay compared with direct extraction of DNA from the soil, and AG-3 was detectable in artificially inoculated and naturally infested soils when seed baiting was combined with either the conventional PCR or the real-time PCR assay. The potential for using these rapid and quantitative AG-3-specific assays to address epidemiological questions and as tools for decision-making in disease management is discussed.

Development of conventional and quantitative real‐time PCR assays for the detection and identification of Rhizoctonia solani AG‐3 in potato and soil

Rhizoctonia solani is a soil-borne basidiomycete fungus with a necrotrophic lifestyle which is classified into fourteen reproductively incompatible anastomosis groups (AGs). One of these, AG8, is a devastating pathogen causing bare patch of cereals, brassicas and legumes. R. solani is a multinucleate heterokaryon containing significant heterozygosity within a single cell. This complexity posed significant challenges for the assembly of its genome. We present a high quality genome assembly of R. solani AG8 and a manually curated set of 13,964 genes supported by RNA-seq. The AG8 genome assembly used novel methods to produce a haploid representation of its heterokaryotic state. The whole-genomes of AG8, the rice pathogen AG1-IA and the potato pathogen AG3 were observed to be syntenic and co-linear. Genes and functions putatively relevant to pathogenicity were highlighted by comparing AG8 to known pathogenicity genes, orthology databases spanning 197 phytopathogenic taxa and AG1-IA. We also observed SNP-level “hypermutation” of CpG dinucleotides to TpG between AG8 nuclei, with similarities to repeat-induced point mutation (RIP). Interestingly, gene-coding regions were widely affected along with repetitive DNA, which has not been previously observed for RIP in mononuclear fungi of the Pezizomycotina. The rate of heterozygous SNP mutations within this single isolate of AG8 was observed to be higher than SNP mutation rates observed across populations of most fungal species compared. Comparative analyses were combined to predict biological processes relevant to AG8 and 308 proteins with effector-like characteristics, forming a valuable resource for further study of this pathosystem. Predicted effector-like proteins had elevated levels of non-synonymous point mutations relative to synonymous mutations (dN/dS), suggesting that they may be under diversifying selection pressures. In addition, the distant relationship to sequenced necrotrophs of the Ascomycota suggests the R. solani genome sequence may prove to be a useful resource in future comparative analysis of plant pathogens.

/pdf/genome-sequencing-and-comparative-genomics-of-the-broad-host-3qn7qdymxg.pdf

Genome Sequencing and Comparative Genomics of the Broad Host-Range Pathogen Rhizoctonia solani AG8

The advancing identification and classification of Rhizoctonia spp. using molecular and biotechnological methods compared with the classical anastomosis grouping

The pectic enzymes of 140 isolates of Rhizoctonia-1ike fungi from the Western Australian grainbelt were examined by electrophoresis and found to fall into 11 distinct zymogram groups (ZG). Isolates within a ZG had a similar cultural and morphological appearance and were either all multinucleate or all binucleate. Some isolates from most ZGs sporulated when transferred from potato-dextrose-marmite agar to water agar. Isolates from within a ZG had the same teleomorph. Pathogenicity of the isolates was tested against wheat and lupins. All isolates from within a ZG were consistent in the type of lesions they produced and in their virulence towards these hosts. Rhizoctonia patch disease of cereals and lupins appears to be caused by isolates from ZG 1 and ZG 2. Severe reddish-brown and brown hypocotyl rots of lupins were caused by ZG 3 and ZG 4 isolates respectively. Five Ceratobasidium groups (CZG), one Waitea group (WZG) and ZG 5 had weak to nil pathogenicity towards wheat and lupins.

Pectic zymograms and taxonomy and pathogenicity of the Ceratobasidiaceae

Zymograms of the extracellular pectic enzymes of Sclerotina sclerotiorum, S. minor and S. trifoliorum were prepared by electrophoresis of untreated fluid from liquid cultures. Each species gave a distinctive zymogram which allowed a rapid and non-subjective identification.

Distinction between Sclerotinia species by their pectic zymograms

Pectic enzymes produced by isolates of Rhizoctonia solani from many hosts and localities within South Australia were subjected to electrophoresis in pectin acrylamide gels. Seven different zymogram groups (ZG) were detected with the majority of isolates being ZG1-1, ZG1–2, ZG1–3 or ZG2. Representatives of these zymogram groups were found in all localities sampled. Of isolates from cereals and legumes, 92% were ZG1-1, ZG1–2, ZG1–3 or ZG2; in contrast, only 23% of isolates from soil were from these zymogram groups. Isolates in ZG1-1, ZG1–2, ZG1–3 and ZG2 were all anastomosis group AG-8, ZG5 were AG-2-1, ZG7 were AG-3 and ZG8 were AG-4. Pectic zymogram patterns are proposed as a fast and reliable method of determining anastomosis group.

Pectic enzyme patterns of Rhizoctonia solani isolates from agricultural soils in South Australia

As a means of identifying groups within binucleate Rhizoctonias isolated from soil and cereal plants, extracellular pectic enzymes produced by the isolates were subjected to electrophoresis in pectin acrylamide gels. Five binucleate zymogram groups CZG1 to CZG5 were isolated from both soil and cereals. Several binucleate zymogram groups and zymogram groups within Thanatephorus cucumeris were isolated from sharp eyespot-like lesions on cereals but Rhizoctonia cerealis was not found. Pathogenicity testing showed that isolates of Ceratobasidium cornigerum and T. cucumeris, particularly AG-4 the stem strain, produced sharp eyespot-like lesions on cereal stems but they were not as pathogenic as authentic isolates of R. cerealis from Europe.

R.H. Cruickshank

Papers

Pectic zymograms and taxonomy and pathogenicity of the Ceratobasidiaceae

Distinction between Sclerotinia species by their pectic zymograms

Pectic enzyme patterns of Rhizoctonia solani isolates from agricultural soils in South Australia

Pectic enzyme patterns of Ceratobasidium and Rhizoctonia spp. associated with sharp eyespot-like lesions on cereals in South Australia