Abstract:
Horse hair blight (HHB) prevails in tea plantations of Sri Lanka below 1200
m altitude, and it has been reported from a wide range of host plants
including tea. HHB forms black, horse hair-like network on the canopy of
the affected tea bush. Although there are no records supporting HHB as a
pathogen in tea plants, its fungal network directly interferes plucking,
pruning and other agronomic practices during tea cultivation. The causal
agent of HHB in tea had been previously identified as Marasmius crinisequi
(Basidiomycetes) by using morphological characters. However,
identification of HHB fungus under in-vitro conditions is always difficult as
the fungus does not produce conidia or any other distinctive structures in
culture. Therefore, the aim of the present study was to confirm the identity
of HHB fungus by DNA sequencing. DNA was extracted both from pure
cultures of HHB established from rhizomorphs (strands) as well as fruit
bodies collected from the field. Polymerase chain reaction was conducted
for four samples using universal primers, ITS1 and ITS4-B which amplifies
internal transcribed spacer (ITS) region of the 18S-26S nuclear ribosomal
DNA. During the BLAST search, DNA sequences of the resultant ITS
amplicons (approximately 850 bp) showed 96-97% identity to Marasmius
crinis-equi at nucleotide level. Thus, the ITS sequence-based identification
confirmed the phenotypic identity of HHB causal agent as Marasmius
crinis-equi. Further, the identity of pure cultures of HHB was possible only
by adopting sequence based identification approach.