Abstract:
Blister blight caused by biotrophic fungus Exobasidium vexans Massee is the most problematic leaf disease in tea
(Camellia sinensis (L.) 0. Kuntze) However, there is little information available on the genetic variation of E. vexansin
Sri Lanka. Understanding the molecular make-up of the pathogen and pathogen population will be helpful for
identifying virulent strains, developing and deploying cultivars with durable resistance and development of
appropriate disease control strategies. This study was carried out to develop a method to collect pathogen materials
for DNA analysis and to test the feasibility of using Basidiomycetes specific ITS-1F and 1TS-4B primers for studying
molecular diversity of E. vexans. DNA was extracted using DNeasy® mini extraction kit from Basidiospores of E.
vexanscollected by spore fall technique and pure cultures of Pestolotia theae, Cladosporium sp. (Ascomycetes) and
Poria hypolateritia (Basidiomycetes) for comparisons. PCR assays were carried out using ITS-F and ITS-4B primer
pairs. The primer pair preferentially amplified a prominent and reproducible region (700 bp) for both of E. vexansand
P. hypolateritiabut not the two ascomycetes fungi tested. The excised bands of E. vexans collected from Talawakelle
and P. hypolateritiawere then used as templates in a second PCR, using the same primers and the re-PCRed product
was sequenced" The DNA sequences of both species showed higher similarity (>80% ) with the DNA sequences of
Basidiomycetes fungi in the Genbank. ITS sequences of E. vexansshowed 77-89% homology with other Exobasidium
spp. when subjected to BLAST analysis confirming the accuracy of the amplified region. The results of this study
suggest that ITS-1F and ITS-4B primer pair can be successfully used to discriminate E. vexansfrom mixed templates
and to study the genetic diversity of this pathogen in Sri Lanka.
