Genetic diversity of 50 isolates of Neofusicoccum parvum, the predominant species of the Botryosphaeriaceae recovered from grapevines displaying symptoms of dieback and decline in New Zealand, was compared to that of isolates from Australia, South Africa, and California. The eight universally primed polymerase chain reaction (UP-PCR) primers distinguished 56 genotypes, with only four clonal pairs found. Seven main groups were identified in a neighbour-joining (NJ) tree with isolates from different regions and vineyards of New Zealand, Australia, and California distributed in different groups, indicating a high level of intra and intervineyard genetic variation. All of the South African isolates were positioned in a separate UP-PCR group, indicating that these isolates were less related to the other N. parvum isolates. When compared to fungi that reproduce sexually the genetic diversity and Shannon diversity indices were low (0.076-0.249; 0.109-0.367, respectively), genetic identity levels were high (0.76-0.95), and genetic distance levels were low (0.04-0.27). The large number of genotypes and the low number of clones in the New Zealand N. parvum populations may be explained by parasexual recombination as anastomosis was observed between nonself pairings. Pathogenicity tests using isolates from different UP-PCR groups inoculated onto either green shoots or 1-y-old grapevines detected virulence diversity, indicating intra and intervineyard variation between isolates, however, no correlation was detected between UP-PCR group and virulence.