A PCR-based method for the detection of Phaeomoniella chlamydospora in grapevines

The oligonucleotide primers, PCL1 and PCL2, were synthesized for Phaeomonie lla chlamydospora from the variable internal transcribed spacers ITS1 and I TS2 of the ribosomal DNA, respectively. Polymerase chain reaction (PCR ampl ification) with PCL1 and PCL2 produced a 325-bp fragment from isolates of P a. chlamydospora. Amplification of this fragment was achieved from as littl e as 16 pg of fungal DNA. The specific primers also amplified a 325-bp frag ment from infected grapevine tissue. Fungal DNA from closely related genera , Phaeoacremonium and Phialophora, as well as several other fungi commonly occurring in grapevine stems, showed no amplification with the species-spec ific primers. These primers can now be used to detect the presence of Pa. c hlamydospora in infected nursery material, and thus form the basis for a ph ytosanitary certification scheme.