Spatial recovery and persistence of Pseudomonas chlororaphis M71 and Trichoderma atroviride SC1 in grapevine following stem injection

ABSTRACT Establishing biocontrol agents (BCAs) within grapevine tissues could provide a means of protecting internal plant compartments, but little is known about their spatial and temporal fate after vascular delivery or whether the graft union constrains their distribution. We compared the internal recovery of two biologically contrasting BCAs, the bacterium Pseudomonas chlororaphis M71 and the filamentous fungus Trichoderma atroviride SC1, following controlled stem injection into rooted and grafted grapevine plants. A xylem-mobile dye was used as a reference for immediate passive fluid transport, while viable microbial recovery was assessed over time and complemented by quantitative isolation and microscopy. At 48 h, the dye generally showed broader longitudinal distribution than either of the microorganisms. The two microorganisms displayed markedly different patterns. M71 persisted for up to 56 days, but remained predominantly associated with tissues close to the injection site. In contrast, SC1 was recovered from a broader range of tissues, including roots, and remained detectable for up to 56 days. In grafted plants, both microorganisms were recovered on both sides of the graft union, indicating that the graft interface was not an absolute barrier, although recovery depended on microbial identity and inoculation position. Green fluorescent protein-labeled M71 cells and SC1 hyphae were observed within xylem vessels, confirming internal localization and, for SC1, vegetative development after injection. These results show that, and how, both microbial biology and grafted-plant architecture influence the internal fate of injected BCAs in grapevine. IMPORTANCE Beneficial microorganisms established inside grapevine tissues could potentially protect from infections regarding internal tissues, but little is known about how such microorganisms distribute and persist inside grapevine after injection into the plant tissues. This study shows that two biocontrol agents (BCAs) with contrasting biological characteristics follow markedly different spatial and temporal patterns. Pseudomonas chlororaphis M71 persisted mainly near the site of introduction, whereas Trichoderma atroviride SC1 was recovered from a broader range of tissues and developed hyphae within xylem vessels. Importantly, the graft union was not an absolute barrier to microbial recovery. These findings provide a framework for selecting where and when BCAs should be introduced during grapevine propagation and highlight that vascular access, persistence, and broad within-plant establishment are distinct properties that should be considered when developing endophytic biocontrol strategies.