Identification and characterization of Candidatus Phytoplasma trifolii (16SrVI-D) inducing shoot proliferation disease of potato in India

Abstract

Potato (Solanum tuberosum) is a widely cultivated and commercially important vegetable crop widely grown in India. Profuse shoot proliferation symptoms were observed in potato plants of two varieties, Kufri Khyati and Kufri Surya at the Research plots of All India Co-ordinated Research Project on Potato, Orissa University of Agriculture and Technology, Bhubaneswar, Odisha, India during February, 2017. The symptomatic plants were tested for phytoplasma etiology. Nested and semi-nested PCR assays primed by primer pairs P1/P7 and 3Far/3Rev for 16S rRNA gene and secAF1/R3 for secA gene amplified DNA fragments of 1.3 kb (3Far/3Rev) and 840 bp (secAF1/R3), respectively from all the symptomatic potato samples but not from any asymptomatic plants. The 16S rDNA sequences of potato tuber (Acc. no. KY815100, Kufri Khyati; MG566064, Kufri Surya) and shoot phytoplasma strains (Acc. no. KY815099, Kufri Khyati; MG566063, Kufri Surya) shared 100% nucleotide sequence identity with 16S rDNA sequence of identified phytoplasma strains belonging to 16SrVI-D subgroup associated with Lens culinaris witches’ broom (Acc. no. KY439869), brinjal little leaf (Acc. no. KX284698) and Brassica oleracea var. capitata witches’ broom (Acc. no. KX671553) in pair wise sequence comparison. Sequence comparison of 840 bp of secA gene of potato phytoplasma strains from tuber of variety, Kufri Khyati (Acc. no. KY815101) and variety Kufri Surya (Acc. no. MG566065) showed 99% sequence identity to secA gene sequences of brinjal little leaf phytoplasma strains (Acc. no. KX610808, KX894792, KY064175), which belonged to 16SrVI-D subgroup. Phylogenetic analysis of 16S rRNA and secA gene sequences, and virtual RFLP analysis of 16S rDNA sequences of potato shoot proliferation phytoplasma strain confirmed their clustering and grouping with strains of clover proliferation subgroup D. Our results confirmed potato as a new host of 16SrVI-D subgroup of phytoplasmas in the world. The 16SrVI-D strain is wide spread in India and cause significant yield losses and hence poses serious socio-economic threat to commercially grown potato crops.