Molecular Screening and Characterization of dsRNA from wild-type and Mutant Strains of Rhizoctonia solani Kühn Isolates
Doreen D. Domingo1, Paulina A. Bawingan2,
Seema Bharathan3 and Narayanaswamy Bharathan4
1Associate Professor, Mariano Marcos State University, City of Batac, I.N.
2Professor, Saint Louis University, Baguio City
1,4Professor, Indiana University of Pennsylvania, PA, USA
Rhizoctonia solani Kühn is a common soil-borne pathogen which infects a diversity of plant hosts. Previous studies have shown that the virulence of Rhizoctonia is affected by the presence of certain dsRNA segments which are of viral origin. This study attempted to screen dsRNA from wild type and mutant isolates of Rhizoctonia solani and characterize them as to size, concentration, and reaction to nucleases. The Rhizoctonia solani isolates were also compared as to color changes in the mycelia and in the culture media, in sclerotia formation and growth rate. There were twelve R. solani isolates utilized in the study: three wild type strains that are heterokaryons (RS29, RS 114 and TE2-4); and nine mutant strains that are homokaryons (RS29.5, RS 29.6. RS29.7, EGR4, EGR7, T2, Tom7, 123e and 115). Results of the study show that only five of the R. solani isolates had dsRNA namely: RS29, RS114, TE2-4, EGR4 and T2. The dsRNAs vary in their sizes and concentration and they showed resistance to nucleases. The R. solani isolates vary in the mycelia pigmentation: white (RS 29.6, RS 29.7, EGR4, EGR7 and 123E), light brown (RS29, RS114, RS 29.5 and 115), brown (TE2-4), and dark brown (T2 and Tom7). Color changes on the potato dextrose agar medium were light yellow (RS 29.5, RS 29.7, EGR4, EGR7), yellow brown (RS 29.6), light brown (RS 29, RS 114, TE2-4), brown (Tom 7, Strain 115) and dark brown (123E, T2).
Many plant pathogenic fungal isolates infected with dsRNA mycoviruses have shown increased virulence (hypervirulence) or reduced virulence (hypovirulence) with the presence of certain dsRNA segments (Tavantzis 2002; Lacksman et al. 1998). For instance, it was shown in previous studies that 3·6-kb dsRNA is associated with suppression of virulence, whereas a 6·4-kb dsRNA is involved with increased virulence in R. solani (Lakshman & Tavantzis 1994; Jian et al. 1997). These dsRNAs are common by-products of viral infection and may result directly from the virus or be produced through transcriptional processes (Karpala et al. 2005). . . .
ALIFERIS K, JABAJI S. 2010. H NMR and GC-MS metabolic fingerprinting of developmental stages of Rhizoctonia solani sclerotia. Metabolomics 6: 96-108.
ABO ELLIL AH. 2005. Biochemical changes and pathogenecity variations related to esterase polymorphism for morphological traits of Rhizoctonia solani. International Journal of Agriculture and Biology 7(4): 555-559.
ANDERSON NA. 1982. The genetics and pathology of Rhizoctonia solani. Annual Review of Phytopathology 20: 329-347.
ARORA P, DILBAGHI N, CHAUDHURY A. 2012. Detection of double stranded RNA in phytopathogenic Macrophormina phaseolina causing charcoal rot in Cyamopsis tetragonoloba. Molecular Biology Reports 39: 3047-54.
BHARATHAN N, TAVANTZIS SM. 1990. Genetic Diversity of double-stranded RNA in Rhizoctonia solani. Phytopathology 80: 631-635.
BHARATHAN N, SASO H, GUDIPATI L., BHARATHAN S, WHITED K, ANTHONY K. 2005. Double-stranded RNA: distribution and analysis among isolates of Rhizoctonia solani AG-2 to –13. Plant Pathology 54: 1196-2003.
BUCK KW. 1986. Fungal virology - an overview, in: Buck, KW (Ed), Fungal Virology. Boca Raton, Florida: CRC Press. p. 1-84.
CARLING DE, LEINER RH, KEBLER KM. 1987. Characterization of a new anastomosis group (AG-9) of Rhizoctonia solani. Phytopathology 77: 1609-12.
CASTANHO B, BUTLER EE. 1978. Rhizoctonia decline: Studies on hypovirulence and potential use in biological control. Phytopathology 68: 1511-14.
CHU YM, JEON JJ, YEA SJ, KIM YH, YUN SH, LEE YW, KIM KH. 2002. Double-stranded RNA Mycovirus from Fusarium graminearum. American Society for Microbiology Journal 68(5): 2529-34.
DUBEY SC, TRIPATHI A, UPADHYAY BK. 2012. Molecular diversity analysis of Rhizoctonia solani isolates infecting various pulse crops in different agroecological regions of India. PubMed 57(6): 513-24.
FRANKLIN R. 1966. Purification and properties of replicative intermediate of the RNA bacteriophage R17. Proceedings of National Academy of Science USA 55: 1504-11.
IHRMARK K. 2001. Double-stranded RNA elements in the Root Rot Fungus Heterobasidion annosum (Doctoral thesis). Swedish University of Agricultural Sciences, Uppsala. Available at http://pub.epsilon.slu. se/4/1/91-576-6094- 8.fulltext.pdf
JIAN J, LAKSHMAN DK, TAVANTZIS SM. 1997. Association of distinct double-stranded RNAs with enhanced or diminished virulence in Rhizoctonia solani infecting potato. Molecular Plant-Microbe Interactions 10: 1002-09.
JUAREZ M, JUAREZ D, RUSSO VM. 2000. Growth and protein content in Colletotrichum circinas, Fusarium solani and Rhizoctonia solani in liquid culture. Biologia Plantarum 43(4): 577-582.
KARPALA AJ, DORSAN TJ, BEAN A. 2005. Immune responses to dsRNA: Implications for gene silencing technologies. Immunology and Cell Biology 83: 211- 216.
KIM HT, CHUNG YR, CHO KY. 2001. Mycelial Melanization of Rhizoctonia solani AG1 Affecting Pathogenicity in Rice. Plant Pathol J 17(4): 210-215.
KOUSIK CS, SNOW JP, VALVERDE RA. 1993. Comparison of double-stranded RNA components and virulence among isolates of Rhizoctonia solani AG1-1A and AG1-1B. Genetics 84(1): 44-49.
LAKSHMAN DK, JIAN J, TAVANTZIS SM. 1998. A double-stranded RNA element from a hypovirulent strain of Rhizoctonia solani occurs in DNA form and is genetically related to the pentafunctional AROM protein of the shikimate pathway. Microbiology 95: 6425-29.
LEHTONEN MJ. 2009. Rhizoctonia solani as a potato pathogen – variation of isolates in Finland and host response [Doctoral thesis in Plant Pathology]. Viikki Graduate School in Molecular Biosciences, University of Helsinki, Finland.
LIU C, LAKSHMAN DK, TAVANTZIS SM. 2003. Quinic acid induces hypovirulence and expression of a hypovirulenceassociated double-stranded RNA in Rhizoctonia solani. Current Genetics 43: 103-11.
MCCABE PM, PFELFFER P, VANALFEN NK. 1999. The influence of dsRNA viruses on the biology of plant pathogenic fungi. Trends Microbiol 7: 377-381.
MORRIS TJ, DODDS JA. 1979. Isolation and analysis of double-stranded RNA from virus-infected plant and fungal tissue. Phytopathology 69: 854-858.
MUTUKU JM, NOSE A. 2012. Changes in the contents of metabolites and enzyme activities in rice plants responding to Rhizoctonia solani Kuhn infection: activation of glycolysis and connection to phenylpropanoid pathway. PubMed, Plant Cell Physiology 53(6): 1017-32.
NUSS D, KOLTIN Y. 1990. Significance of dsRNA Genetic Elements in Plant Pathogenic Fungi. Annual Review of Phytopathology 28: 38-57.
OGOSHI A. 1987. Ecology and pathogenicity of anastomosis and intraspecific groups of Rhizoctonia solani Kühn. Annual Review of Phytopathology 25: 125-143.
ORELLANA RG, MANDAVA NV. 2008. m-Hydroxyphenylacetic and m-Methoxyphenylacetic Acids of Rhizoctonia solani: Their Effect on Specific Root-Nodule Activity and Histopathology in Soybean. Journal of Phytopathology 107(2): 159-167.
RITCHIE F, BAIN RA, MCQUILKEN MP. 2009. Effects of nutrient status, temperature, and pH on mycelia growth, sclerotial production and germination of Rhizoctonia solani from potato. Journal of Plant Pathology 91(3): 589-596.
SAMBROOK J, FRITSCH EF, MANIATIS T. 1989. Molecular cloning: a laboratory manual. Cold Spring Harbor, NY: Cold Spring Harbor Press. 253p.
SCULLY E. 2007. In Vitro Assays to Genetically Cure Rhizoctonia solani of Mycoviral Double-Stranded RNA. [MS Thesis] Indiana, PA, USA: Indiana University of Pennsylvania.
SHOAEI NS, HASHEMI M, ETRBARIAN HR, MAHMOUDI SB. 2012. Correlation between specific double-stranded (ds) RNA elements and up- or down-regulation of virulence, laccase activity and mycelial growth of Rhizoctonia solani (AG2) isolates, International Journal of AgriScience 2(3): 204-21.
SNEH B, JABAJI-HARE S, NEATE S, DIJST G. 1996. Rhizoctonia Species: Taxonomy, Molecular Biology, Ecology, Pathology, and Disease Control. Doordrecht, The Netherlands: Kluwer Academic Publishers.
SOLEIMANI MJ, KASHI L. 2005. A New Disease of Gladiolus Caused by Binucleate Rhizoctonia sp. Plant Pathology Journal 4(2): 138-142.
TAVANTZIS SM. 1995. In: Advances in Potato Pest Biology and Management. Zehnder GW, Powelson ML, Jansson RK, Raman KV (eds). St. Paul, MN: Am. Phytopathology Society Press. p. 565-579.
TAVANTZIS SM. 2002. DsRNA genetic elements: Concepts and applications in agriculture. In: Forestry and Medicine. Boca Raton, FL: CRC Press LLC. p. 304.
VILGALYS R, CUBETA MA. 1994. Molecular Systematics and Population Biology of Rhizoctonia. Annual Review of Phytopathology 32: 135-155.
ZANZINGER DH, BANDY BP, TAVANTZIS SM. 1984. High-frequency of finding double-stranded RNA in naturally occurring isolates of Rhizoctonia solani. Journal of General Virology 65: 1601-05.