Detection and molecular identification of a 16SrI group phytoplasma associated with sisal purple leafroll disease

Abstract

Sisal purple leafroll disease (SPLD) is a destructive disease affecting sisal in China, however, its etiology remains unknown. This disease is characterized by purple margins and rolling and yellowing of the leaves. Recently, phytoplasma was found to be present in SPLD-affected sisal according to PCR assay. To further verify the presence of phytoplasma, determine its association with SPLD and identify the possible pathogenic phytoplasma infecting sisal, 80 symptomatic and 65 asymptomatic sisal samples were collected from the main sisal farms in three Chinese provinces, including Hainan, Guangdong and Guangxi provinces, from 2018 to 2021. PCR assays and transmission electron microscopy further revealed the presence of phytoplasma. Nested PCR using the universal primer R16mF2/R16mR1 followed by R16F2n/R16R2 was confirmed as the most effective molecular method for the detection of phytoplasma in sisal. A comparison of the detection rate of phytoplasma in symptomatic (83%) versus asymptomatic (17%) sisal populations showed some association between phytoplasma and SPLD. Sequencing analyses, phylogenetic analyses, and virtual restriction fragment length polymorphism analyses confirmed the presence of phytoplasma in sisal belonging to 16SrI-B. This is the first report of 16SrI-B phytoplasma infecting sisal in China.

Keywords

• pcr assays
• 16s rrna
• universal primers
• transmission electron microscopy
• sequence analysis