Molecular characterization of 'Candidatus Phytoplasma australasia' 16SrII subgroups associated with eggplant, cabbage, beetroot, and celery in Saudi Arabia

Abstract

From 2015 to 2017 symptoms associated with phytoplasmas were observed in vegetable crop farms located in Uyun AlJiwa and Al-Dawadmi, cities located in the Al-Qassim Region and the Riyadh Province, respectively. Affected eggplants exhibited symptoms of phyllody, little leaves and witches broom, symptomatic cabbage showed multiple heads, beetroot showed reddening, stunting and leaves proliferation symptoms, while celery plants showed stunting and yellowing symptoms. To determine if the symptomatic samples were affected by phytoplasmas, PCR was performed with phytoplasma universal primers P1/P7 followed by a nested PCR with primer pair R16mF2/R16mR2, which target the 16S rRNA-encoding gene locus. PCR revealed that 20/20 symptomatic eggplants, 15/20 symptomatic cabbage, 26/30 symptomatic beetroot, and 9/11 symptomatic celery plants were infected with phytoplasma, and sequencing of the amplified genes revealed that the phytoplasmas belonged to the peanut witches’-broom group (16SrII). Sequence analysis using in silico restriction fragment length polymorphism (RFLP) and phylogenetic analysis revealed the presence of two closely related but distinct sequences from the 16SrII-D and the recently described 16SrII-X subgroups. The two sequences differed by a single base pair and both subgroups were detected in cabbage, beetroot, and celery, while eggplant was only affected by subgroup 16SrII-X. Sequence analysis of the chaperonin-60 universal target confirmed the classification of these phytoplasma strains in the 16SrII group and their close phylogenetic relationship. This is the first report of eggplant, cabbage, beetroot, and celery affected by phytoplasma in Saudi Arabia, and the first report of these plant hosts affected by the 16SrII-X subgroup.