Current Microbiology

, Volume 74, Issue 7, pp 877–884 | Cite as

Variation of rDNA Internal Transcribed Spacer Sequences in Rhizoctonia cerealis

  • Lei Ji
  • Chunju Liu
  • Li Zhang
  • Aixin Liu
  • Jinfeng Yu
Review Article


Fifty-four single protoplast isolates (SPIs) were regenerated from three Rhizoctonia cerealis strains. A total of 169 rDNA-ITS regions were cloned and sequenced from these 54 SPIs. Variations in the ITS1 and ITS2 regions that flank the 5.8S gene were found within clones from the same strain, as well as within clones from the same SPI. These include variations in GC content and ITS length, and single-nucleotide polymorphisms (SNPs). The different strains and SPIs GC contents range from 40.25 to 41.74% and from 42.40 to 45.02%, in the ITS1 and ITS2 regions, respectively. All SNPs occur in the ITS1 and ITS2 regions, with 3–6 and 4–6 polymorphic sites in each region, respectively, in the different strains. SNP variation is relatively stable within the same strain. For example, the 89 ITS sequences generated from isolate WK-207, regardless of SPI or clone, predominantly cluster into two separate clades on a phylogenetic tree, suggesting that nuclei genetic heterogeneity is related to ITS variation in R. cerealis. Although rDNA-ITS sequences from the three strains and different SPIs are somewhat variable, all of our ITS sequences cluster together in anastomosis subgroup AG-DI during phylogenetic analysis. The ITS variation we observed does not negatively influence R. cerealis anastomosis group or subgroup classification.


Rhizoctonia cerealis rDNA-ITS sequence Variation Protoplast preparation 



This study was supported by the National Natural Science Foundation, China, (Grant No. 30870007), and Shandong Province modern agricultural industry technology system innovation team building special funds (Grant No. SDAIT-01-09).


Note that the funders of this research had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

284_2017_1258_MOESM1_ESM.docx (391 kb)
Supplementary material 1 (DOCX 390 kb)


  1. 1.
    Ahvenniemi P, Wolf M, Lehtonen MJ, Wilson P, German-Kinnari M, Valkonen JP (2009) Evolutionary diversification indicated by compensatory base changes in ITS2 secondary structures in a complex fungal species, Rhizoctonia solani. J Mol Evol 69:150–163CrossRefPubMedGoogle Scholar
  2. 2.
    Blair ID (1942) Studies on the growth in soil and the parasiticaction of certain Rhizoctonia solani isolates from wheat. Can J Res 20:174–185CrossRefGoogle Scholar
  3. 3.
    Boysen M, Borja M, del Moral C, Salazar O, Rubio V (1996) Identification at strain level of Rhizoctonia solani AG4 strains by direct sequence of asymmetric PCR products of the ITS regions. Curr Genet 29:174–181CrossRefPubMedGoogle Scholar
  4. 4.
    Carling DE, Kuniaga S, Brainard A et al (2002) Hyphal anastomosis reactions, rDNA-internal transcribed spacer sequences, and virulence levels among subsets of Rhizoctonia solani anastomosis group-2(AG-2) and AG-BI. Phytopathology 92:43–50CrossRefPubMedGoogle Scholar
  5. 5.
    Clarkson JDS, Cook RJ (1983) Effect of sharp eyespot (Rhizoctonia cerealis)on yield losses in winter wheat. Plant Pathol 32:421–428CrossRefGoogle Scholar
  6. 6.
    Shilin Chen, Xiaohui Pang, Kun Luo, Hui Yao, Jianping Han, Jingyuan Song (2013) DNA barcoding of biological resources. Chin Bull Life Sci 25:458–466Google Scholar
  7. 7.
    Cromey MG, Butler RC, Boddington HJ, Moorhead AR (2002) Effects of sharp eyespot on yield of wheat (Triticum aestivum). NZ Crop Hortic Sci 30:9–17CrossRefGoogle Scholar
  8. 8.
    Boerema GH, Verhoeven Adriana A (1977) Check-list for scientific names of common parasitic fungi. Series 2b: fungi on field crops: cereals and grasses. Neth J Plant Pathol 83:165–204CrossRefGoogle Scholar
  9. 9.
    Glynne MD (1950) Sharp eyespot as a severe disease of oats. Nature 166:232CrossRefPubMedGoogle Scholar
  10. 10.
    Glynne MD, Ritchie WM (1943) Sharp eyespot of wheat caused by Corticium (Rhizoctonia solani). Nature 152:160CrossRefGoogle Scholar
  11. 11.
    Gonzalez D, Carling DE, Kuninaga S, Vilgalys R et al (2001) Ribosomal DNA systematics of Ceratobasidium and Thanatephorus with Rhizoctonia anamorphs. Mycologia 93:1138–1150CrossRefGoogle Scholar
  12. 12.
    Gonzalez D, Cubeta A, Vilgalys R (2006) Phylogenetic utility of indels within ribosomal DNA and beta-tubulin sequences from fungi in the Rhizoctonia solani species complex. Mol Phylogent Evol 40:459–470CrossRefGoogle Scholar
  13. 13.
    Grosch R, Schneider JHM, Peth A, Waschke A, Franken P, Kofoet A, Jabaji-Hare SH (2007) Development of a specific PCR assay for the detection of Rhizoctonia solani AG 1-IB using SCAR primers. Appl Microbiol 102:806–819CrossRefGoogle Scholar
  14. 14.
    Hamada MS, Yin Y, Chen H, Ma Z (2011) The escalating threat of Rhizoctonia cerealis, the causal agent of sharp eyespot in wheat. Pest Manag Sci 67:1411–1419CrossRefPubMedGoogle Scholar
  15. 15.
    Yuepeng Han, Xiulan Chen, Zhentian He, Jinrong Wang, Hefeng Yang (2001) Wheat sharp eyespot research status problems and prospect. Tcrop 21:81–84Google Scholar
  16. 16.
    Hayakawa T, Toda T, Ping Q, Mghalu JM, Yaguchi S, Hyakumachi M (2006) A new subgroup of Rhizoctonia AG-D, AG-D III, obtained from Japanese zoysia grass exhibiting symptoms of a new disease. Plant Dis 90:1389–1394CrossRefGoogle Scholar
  17. 17.
    Hershkovitz MA, Lewis LA (1996) Deep-level diagnostic value of the rDNA-ITS region. Mol Biol Evol 13:1276–1295CrossRefPubMedGoogle Scholar
  18. 18.
    Hijri M, Hosny M, van Tuinen D, Dulieu H (1999) Intraspecific ITS polymorphism in Scutellospora castanea (Glomales, Zygomycota) is structured within multinucleate spores. Fungal Genet Biol 26:141–151CrossRefPubMedGoogle Scholar
  19. 19.
    Hillis DM, Dixon MT (1991) Ribosomal DNA: molecular evolution and phylogenetic inference. Q Rev Biol 66:411–453CrossRefPubMedGoogle Scholar
  20. 20.
    Joke P, Monica H (2009) Detection of rDNA ITS polymorphism in Rhizoctonia solani AG 2-1 isolates. Mycologia 101(1):26–33CrossRefGoogle Scholar
  21. 21.
    Julian MC, Hyakumachi M, Rubio V (2000) Phylogenetic grouping of cultural types of Rhizoctonia solani AG2-2 based on ribosomal ITS sequences. Mycologia 92:505–509CrossRefGoogle Scholar
  22. 22.
    Justesen AF, Yohalem D, Bay A, Nicolaisen M (2003) Genetic diversity in potato field populations of Thanatephorus cucumeris AG-3, revealed by ITS polymorphism and RAPD markers. Mycol Res 107:1323–1331CrossRefPubMedGoogle Scholar
  23. 23.
    Kataria MR, Hoffmann GM (1988) A critical review of plant pathogenic species of Ceratobasidium Rogers. Z Pflanzenk Pflanzen 95:81–107Google Scholar
  24. 24.
    Ikuko O, Masao A, Naoyuki M (2001) ITS polymorphism within a single strain of Sclerotium rolfsii. Mycoscience 42:107–113CrossRefGoogle Scholar
  25. 25.
    Kuninaga S, Natsuaki T, Takeuchi T, Yokosawa R (1997) Sequence variation of the rDNA ITS regions within and between anastomosis groups in Rhizoctonia solani. Curr Genet 32:237–243CrossRefPubMedGoogle Scholar
  26. 26.
    Kuninaga S, Natsuaki T, Takeuchi T (1997) Sequence Variation of the rDNA-ITS Regions within and between Anastomosis Groups in Rhizoctonia solani. Curr Genet 32:237–243CrossRefPubMedGoogle Scholar
  27. 27.
    Lipps PE, Herr LJ (1982) Etiology of Rhizoctonia cerealis in sharp eyespot of wheat. Phytopathology 72:1574–1577CrossRefGoogle Scholar
  28. 28.
    Wei Li, Haiyan Sun, Yuanyu Deng, Aixiang Zhang, Huaigu Chen (2014) The heterogeneity of the rDNA-ITS sequence and its phylogeny in Rhizoctonia cerealis, the cause of sharp eyespot in wheat. Curr Genet 60:1–9CrossRefGoogle Scholar
  29. 29.
    Hyakumachi Mitsuro, Priyatmojo Achmadi, Kubota Mayumi, Fukui Hirokazu (2005) New Anastomosis Groups, AG-T and AG-U, of binucleate Rhizoctonia spp. causing root and stem rot of cut-flower and miniature Roses. Phytopathol 95:784–792CrossRefGoogle Scholar
  30. 30.
    Nicholson P, Parry DW (1996) Development and use of a PCR assay to detect Rhizoctonia cerealis, the cause of sharp eyespot in wheat. Plant Pathol 45:872–883CrossRefGoogle Scholar
  31. 31.
    Okabe I, Matsumoto N (2003) Phylogenetic relationship of Sclerotium rolfsii (teleomorph Athelia rolfsii) and S. delphinii based on ITS sequences. Mycol Res 107:164–168CrossRefPubMedGoogle Scholar
  32. 32.
    Pannecoucque J, Hofte M (2009) Detection of rDNA ITS polymorphism in Rhizoctonia solani AG 2-1 isolates. Mycologia 101:26–33CrossRefPubMedGoogle Scholar
  33. 33.
    Pitt D (1964) Studies on sharp eyespot disease of cereals, Disease symptoms and pathogenicity of isolates of Rhizoctonia solani Kuhn and the influence of soil factors and temperature on disease development. Ann Appl Biol 54:77–89CrossRefGoogle Scholar
  34. 34.
    Reinecke P, Fehrmann H (1979) Infection experiments with Rhizoctonia cerealis Van der Hoeven on cereals. J Plant Dis Prot 86:241–246Google Scholar
  35. 35.
    Salazar O, Julian M et al (2000) Primers based on specific rDNA-ITS sequences for PCR detection of Rhizoctonia solani, AG-2 subgroups and ecological types, and binucleate Rhizoctonia. Mycol Res 104(3):281–285CrossRefGoogle Scholar
  36. 36.
    Scott D, Visser C, Ruzenacht E (1979) Crater disease of summerwheat in African dry lands. Plant Dis Rep 63:836–840Google Scholar
  37. 37.
    Seifert K (2009) Progress towards DNA barcoding of fungi. Mol Ecol Resour 9:83–89CrossRefPubMedGoogle Scholar
  38. 38.
    Sharon M, Kuninaga S, Hyakumachi M, Naito S, Sneh B (2008) Classification of Rhizoctonia spp. using rDNA-ITS sequence analysis supports the genetic basis of the classical anastomosis grouping. Mycoscience 49:93–114CrossRefGoogle Scholar
  39. 39.
    Strausbaugh CA, Eujayl IA, Panella LW, Hanson LE (2011) Virulence, distribution, and diversity of Rhizoctonia solani from sugar beet in Idaho and Oregon. Can J Plant Pathol 33:210–226CrossRefGoogle Scholar
  40. 40.
    Toda Takeshi, Hyakumachi Mitsuro, Suga Haruhisa, Kageyama Koji, Tanaka Akemi, Tani Toshikazu (1999) Differentiation of Rhizoctonia AG-D isolates from turfgrass into subgroups I and II based on rDNA and RAPD analyses. Eur J Plant Patho 105:835–846CrossRefGoogle Scholar
  41. 41.
    Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony method. Mol Biol Evol 28:2731–2739CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    TomasonPM and Trevathan LE (2007) Characterization of Rhizoctonia-like fungi isolated from agronomic crops and turfgrasses in Mississippi. Plant Dis 91:260–265Google Scholar
  43. 43.
    Havakawa Toshihiro, Takeshi Toda Qu, Ping Misturo Hvakumachi (2006) A New subgroup of Rhizoctonia AG-D, AG-D III, obtained from Japanese Zoysia grass exhibiting symptoms of a new disease. Plant Dis 90:1389–1394CrossRefGoogle Scholar
  44. 44.
    Yan SQ, Wu BC, Tang XF, Liu ZJ, Jiang L (1984) Sheath blight of cereal crops. I. Studies on the relation between sheath blight of rice, maize and wheat as well as soreshin of cotton (Chinese with English abstract). Chin J Plant Pathol 14:25–32Google Scholar
  45. 45.
    Dezhen Zhang, Xiaoxia Chen, Xianyue Gao, Jinfeng Yu (2015) Protoplast preparation and regeneration of Rhizoctonia cerealis. Plant Prot 41:79–85Google Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  1. 1.Department of Plant PathologyShandong Agriculture UniversityTaianChina
  2. 2.Shandong Gaomi Tobacco Monopoly BureauGaomiChina

Personalised recommendations