Past studies have shown that the major source of infection of young grapevine plants by Botryosphaeriaceae fungi were through the use of infected rootstock and scion cuttings. To investigate the potential infection pathways of Botryosphaeriaceae species within a rootstock mother vine, three genotyping studies using universally-primed polymerase chain reaction (UP-PCR) were conducted with two Neofusicoccum species, namely N. luteum and N. parvum. The investigations identified genotypes of the fungal isolates in trunk and shoot infections of the same mother vine. Results showed that the trunk and shoot isolates from the same vine were of the same or different genotypes, suggesting multiple infections from different inoculum sources. This study further showed that the Neofusicoccum isolates recovered from the surfaces of the cuttings were of the same or different genotypes from those isolated from adjacent internal tissues, again suggesting multiple sources of external inoculum. Investigations into the spatial distribution of Botryosphaeriaceae fungi within an entire dormant cane also showed that multiple species and genotypes were distributed along the cane but most isolates were sited within the bark, being less frequently in the wood, which suggested that they were latent on surface tissues. Since some adjacent wood and bark infections were caused by the same genotypes, this indicated that wood infection may have originated from the bark. These fungi appear to cause latent infections in the bark of dormant cuttings which are used in plant propagation, thus providing an additional infection pathway for a disease that is known to show obvious symptoms only in older vineyards.