Aureobasidium pullulans and Epicoccum nigrum are frequently reported as endophytes of various
crops, including grapevine (Vitis vinifera). Because of their potential role as biological control
agents against grapevine pathogens, we examined the occurrence of A. pullulans and E.
nigrum in two grapevine varieties (Merlot and Prosecco) in Italian vineyards where spontaneous
recovery from phytoplasma disease is recurrent. Species-specific primers for A. pullulans and
two genetically distinct strains of E. nigrum were designed in variable regions of ITS1 and ITS2.
Primer specificity was confirmed by polymerase chain reaction using purified DNA from other
fungal endophytes that are usually encountered during isolation attempts from grapevine tissues
and from several other strains of A. pullulans and E. nigrum isolated from other sources. In order
to determine the occurrence of the two endophytes in grapevine plants, DNA was extracted from
shoots of 44 grapevines collected in six vineyards from different localities of northeast Italy.
Both endophytes were detected and their identity was confirmed by restriction fragment length
polymorphism (RFLP) patterns obtained from reference strains. RFLP analyses confirmed the
presence of two E. nigrum strains belonging to different RFLP groups in grapevine. The molecular
methods described allowed a sensitive, specific, and reliable identification of the two endophytes
in grapevine.
