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The production of phenolic metabolites has been optimized in parallel to sugarcane micropropagation in temporary immersion bioreactors (TIBs). Culturing micropropagated plants in 0.4% CO 2 -rich air induced their photosynthetic activity by enhancing the change from a heterotrophic to a photomixotrophic metabolic stage. Under 0.4% CO 2 enrichment, the transcript levels of both phenylalanine ammonia-lyase (PAL EF189195) and ribulose-1,5-biphosphate carboxylase/oxygenase (Rubisco CF576750) increased and were correlated to a 4.6-and 6.3-fold increase in the phenolic levels when plants were multiplied in 20 or 30 g/l sucrose, respectively. A novel application of plant phenolic metabolites as elicitors of resistance to tomato bacterial wilt in the Solanum lycopersicum-Ralstonia solanacearum pathosystem has been identified. The culture media was collected, and the phenolics were sprayed onto tomato plants infected with R. solanacearum, eliciting and/or maintaining an early defense signaling mechanism that resulted in the protection of the plant against the tomato bacterial wilt disease. RT-PCR analyses confirmed that selected genes from defense-related pathways were differentially expressed between plants treated with sugarcane metabolites, non-treated pathogenfree plants, and non-treated plants inoculated with R. solanacearum.

For the first time both priming and biopriming approaches have been integrated into the sugarcane micropropagation technology by temporary immersion bioreactors (TIBs). Sugarcane micropropagation in CO 2 -rich TIBs induces a mixotrophic condition adequate for the production of natural phenolic metabolites. Scaling up has been conducted in the C86-56 and C90-317 commercial genotypes. While phenolics demonstrate to act as priming molecules during the in vitro culture, vitroplantlets growing and shooting in the presence of phenolic metabolites display an enhanced vigour (measure as plant size), emitted functional roots and increase adaptability to the natural environment. Additionally, when combined with the inoculation of the endophytic Gluconacetobacter diazotrophicus during transplanting, a significant improvement of the percentage of survival has been attached through this critical step. Altogether, results indicate a promising potential for diversification of the sugarcane micropropagation industry by the production of useful metabolites as byproducts.