Isolation, Identification and Evaluation of Potential Biological Control Agents for Sustainable Grapevine Pruning Wound Protection

Grapevine trunk diseases (GTDs), caused by many different fungal pathogens, threaten the economic sustainability of viticulture worldwide causing a significant reduction of both yields and quality of grapes. GTDs are referred to as a complex of diseases with Botryosphaeria dieback, Eutypa dieback and Esca being major contributors. Pruning wounds are the main point of entry for these fungal pathogens and majority of spore release occurs in the winter months when precipitation activates spore release. Traditionally, disease control is focused on cultural practices and preventative pruning wound protection by the application of chemical pastes and sprays. However, with an imperative need for sustainable agricultural practices there has been much interest in the use of biological control agents (BCAs) as pruning wound protectants. This study aimed to (i) identify naturally occurring potential BCAs from a variety of grapevine tissues, including sap, cane and pith and evaluate their antagonistic activity against selected fungal pathogens responsible for GTDs in vitro, (ii) take forward promising candidates to greenhouse and field trials to evaluate as pruning wound protectants alongside commercial chemical and biological protectants and (iii) determine the recovery rate of biologicals from treated canes at the end of greenhouse and field studies. Isolated bacterial and fungal isolates from grapevine structures were screened in vitro to determine their antifungal activity via a dual culture assay and volatile assay against Eutypa lata, Diplodia seriata, Diaporthe ampelina and Neofusicoccum parvum. Among the fungal isolates, Trichoderma spp. inhibited E. lata mycelial growth up to 64% and N. parvum mycelial growth up to 73% with overgrowth and stopped growth being the likely antagonistic mechanisms. Among the bacterial isolates, Bacillus spp. inhibited E. lata mycelial growth up to 20% and N. parvum mycelial growth up to 40%. Under greenhouse and field conditions, Trichoderma asperellum and Trichoderma gamsii consistently provided high pruning wound protection in greenhouse and field trials, with a mean percent disease control (MPDC) of 88% and 100% for E. lata and N. parvum respectively, when compared to the water treated-inoculated positive control. The chemical protectants, Thiophanate-methyl + Myclobutanil and Fluopyram + Trifloxystrobin were also able to effectively protect wounds with a MPDC of up to 86%. Lastly, when biological treatments were evaluated for recovery from treated canes at the end of the growing season, Trichoderma-based treatments had a rate of recovery between 0 and 100%