Phenotypic Characterization and Molecular Phylogenetic Relationship of Erysiphe necator Infecting Grapes (Vitis vinifera)

Main Article Content

Marimuthu Karthick
Ayyanar Kamalakannan
Varagur Ganesan Malathi
Vaikuntavasan Paranidharan
Uthandi Sivakumar
Mathiyazhagan Kavino
Nagaraj Gowrisri

Abstract

Grapes powdery mildew is caused by the most destructive pathogen Erysiphe necator leading to severe yield losses around the world.  In order to study the phenotypic and molecular characters, the powdery mildew infected leaf samples were collected from eight different places in Coimbatore and Theni districts in the state of Tamil Nadu India. The identity of the pathogen as E. necator was established by microscopic studies and the isolates were further confirmed molecularly by amplification of Internal transcribed spacer (ITS) and Cytochrome b gene (Cyt b). Further molecular confirmation was obtained by characterizing Cytochrome b. An amplicon size of ~ 367 and ~ 470 bp were obtained from amplification with Uncin144 and Uncin511 and Cyt b F and Cyt b R gene primers respectively. The identity for cyt b gene segment was 96 to 98%, similarity with E. necator isolates deposited in NCBI genbank (KY418048.1, KY418049.1).

A phylogenetic tree was constructed on the basis of nucleotide sequence of cytochrome b gene of the study isolates as well as E. necator and other Erysiphe species in NCBI database. From the tree it was evident that the study isolates from Tamil Nadu, India were very distinct from other E. necator isolates deposited in NCBI genbank database.

Keywords:
Grapes, powdery mildew, E. necator, ITS, cytochrome b region, PCR, detection, phylogenetic analysis, sequence identity matrix.

Article Details

How to Cite
Karthick, M., Kamalakannan, A., Malathi, V., Paranidharan, V., Sivakumar, U., Kavino, M., & Gowrisri, N. (2019). Phenotypic Characterization and Molecular Phylogenetic Relationship of Erysiphe necator Infecting Grapes (Vitis vinifera). Current Journal of Applied Science and Technology, 37(3), 1-10. https://doi.org/10.9734/cjast/2019/v37i330291
Section
Original Research Article

References

Deliere L, Miclot AS, Sauris P, Rey P, Calonnec A. Efficacy of fungicides with various modes of action in controlling the early stages of an Erysiphe necator-induced epidemic. Pest Management Science. 2010; 66:1367-1373.

Dufour MC, Fontaine S, Montarry J, Corio-Costet MF. Assessment of fungicide resistance and pathogen diversity in Erysiphe necator using quantitative real-time PCR assays. Pest Management Science. 2011;67:60-69.

Yarwood CE. History and taxonomy. In The Powdery MíIdews. (Ed. D.M. Spencer), Academic press: London. 1978; 1-37.

Falacy JS, Gary G, Grove, Mahaffee WF, Galloway H, Dean A, Glawe, Larsen RC, Vandemark GJ. Detection of Erysiphe necator in Air Samples Using the Polymerase Chain Reaction and Species-Specific Primers. 2007;1290-1297.

Gadoury DM, Seem RC, Wilcox WF, Henick-Kling T, Conterno L, Day A, Ficke A. Effects of diffuse colonization of grape berries by Uncinula necator on bunch rots, berry microflora, and juice and wine quality. Phytopathology. 2007;97:1356-1365.

Venkateswaran K, Kamijoh Y, Ohashi E, Nakanishi H. A simple filtration technique to detect enterohemorrhagic Escherichia coli O157:H7 and its toxins in beef by multiplex PCR. Appl Environ Microbiol. 1997;63:4127–4131.

Kong P, Hong C, Jeffers SN, Richardson PA. A Species-specific polymerase chain reaction assay for rapid detection of Phytophthora nicotianae in irrigation water. Phytopathology. 2003;93:822–831. DOI: 10.1094/PHYTO.2003.93.7.822

Mior ZA, Tong PE, Mohammadpour lima M, Yun WM. Morphological and molecular characterizations of rice blast fungus, Magnaporthe oryzae. Pak. J. Agri. Sci. 2017;54(4):765-772.

Schena L, Cooke DEL. Assessing the potential of regions of the nuclear and mitochondrial genome to develop a ‘‘molecular tool box’’ for the detection and characterization of Phytophthora species. J Microbiol Methods. 2007;67:70– 85.

McDermott, JM, Brandle U, Dutly F, Haemmerli UA. , Keller S, Muller KE, wolfe MS. Genetic variation in powdery mildew of barley: Development of RAPD, SCAR, and VNTR Markers. Phytopathology. 1994;84(11):1316-1321.
Available:https://doi.org/10.1094/Phyto-84-1316

Kumar S, Stecher G, Tamura K. MEGA7: Molecular evolutionary genetics analysis version 7.0. for bigger datasets. Mol. Biol. Evol. 2016:33(7):1870–1874.

Calonnec A, Cartolaro P, Poupot C, Dubourdieu D, Darriet P. Effects of Uncinula necator on the yield and quality of grapes (Vitis vinifera) and wine. Plant Pathol. 2004;53(4):434–445.

Stummer BE, Francis IL, Zanker T, Lattey KA, Scott ES. Effects of powdery mildew on the sensory properties and composition of Chardonnay juice and wine when grape sugar ripeness is standardised. Aust J Grape Wine Res. 2005;11(1):66–76.

Almeida AMR, Binneck E, Piuga FF, Marin SRR, Riberio do Valle PRZ, Silveira CA. Characterization of powdery mildew strains from soybean, bean, sunflower and weeds in Brazil using rDNA ITS sequences. Trop. Plant Pathol. 2008;33:20-26.

Braun U, Takamatsu S. Phylogeny of Erysiphe, Microsphaera, Uncinula (Erysipheae) and Cystotheca, Podosphaera, Sphaerotheca (Cystotheceae) inferred from rDNA ITS sequences – some taxonomic consequences. Schlechtendalia. 2000;4:1-33.

Cunnington JH, Takamatsu S, Lawrie, AC, Pascoe, IG. Molecular identification of anamorphic powdery mildews (Erysiphales). Australas. Plant Pathol. 2003;32:421-428.

Takamatsu S, Shin HD, Paksiri U, Limkaisang S, Taguchi Y, Nguyen TB, Sato Y. Two Erysiphe species associated with recent outbreak of soybean powdery mildew: Results of molecular phylogenetic analysis based on nuclear rDNA sequences. Mycoscience. 2002;43:333-341.

Zheng and Koller. Characterization of the mitochondrial cytochrome b gene from Venturia inaequalis. Current Genetics. 1997;32(5):361-366.

Lesemann SS, Schimpk S, Dunemann F, Deising, HB. Mitochondrial 464 heteroplasmy for the cytochrome b gene controls the level of strobilurin resistance in the 465 apple powdery mildew fungus Podosphaera leucotricha (Ell. & Ev.) E.S. Salmon. J. Plant Dis. Prot. 2006:113:259-266.

Villani SM, Cox KD. Heteroplasmy of the cytochrome b gene in Venturia inaequalis and its involvement in quantitative and practical resistance to trifloxistrobin. Phytopathology. 2014;104 (9):945-953.

Hashimoto M, Aoki Y, Saito S, Suzuki, S. Characterisation of heteroplasmic 429 status at codon 143 of the Botrytis cinerea cytochrome b gene in a semi-quantitative AS430 PCR assay. Pest Manag. Sci. 2015;71:467-477.

Miles LA, Miles TD, Kirk WW, Schilder, AMC. Schilder, strobilurin (QoI) Resistance in Populations of Erysiphe necator on Grapes in Michigan. Plant Disease. 2012;96 (11):1621-1628.

Fernandez AV, Gomez DB, Tores JA, Vicente AD, Garcia AP, Ortuno DF. Heteroplasmy for the Cytochrome b gene in Podosphaera xanthii and its role in resistance to QoI fungicides in spain. Plant Disease. 2018;1–29.

Kears M, Moir R, Wilso A, Stones-Havas S, Cheung M, Sturrock S, Buxton S, Cooper A, Markowitz S, Duran C, Thierer T, Ashton B, Mentjies P, Drummond A. Geneious Basic: An integrated and extendable desktop software 452 platform for the organization and analysis of sequence data; 2012.

Mosquera S, Chen L, Aegerter B, Miyao E, Salvucci A, Chang T, Epstein, L, Stergiopoulos I. Cloning of the Cytochrome b Gene from the tomato powdery mildew fungus Leveillula taurica. Reveals high levels of allelic variation and heteroplasmy for the G143A mutation. Front Microbiol. 2019;10:663.