Engineering d‐limonene synthase down‐regulation in orange fruit induces resistance against the fungus Phyllosticta citricarpa through enhanced accumulation of monoterpene alcohols and activation of defence

Abstract

Terpene volatiles play an important role in the interactions between specialized pathogens and fruits. Citrus black spot (CBS), caused by the fungus Phyllosticta citricarpa, is associated with crop losses in different citrus-growing areas worldwide. The pathogen may infect the fruit for 20-24 weeks after petal fall, but the typical hard spot symptoms appear when the fruit have almost reached maturity, caused by fungal colonization and the induction of cell lysis around essential oil cavities. d-Limonene represents approximately 95% of the total oil gland content in mature orange fruit. Herein, we investigated whether orange fruit with reduced d-limonene content in peel oil glands via an antisense (AS) approach may affect fruit interaction with P. citricarpa relative to empty vector (EV) controls. AS fruit showed enhanced resistance to the fungus relative to EV fruit. Because of the reduced d-limonene content, an over-accumulation of linalool and other monoterpene alcohols was found in AS relative to EV fruit. A global gene expression analysis at 2 h and 8 days after inoculation with P. citricarpa revealed the activation of defence responses in AS fruit via the up-regulation of different pathogenesis-related (PR) protein genes, probably as a result of enhanced constitutive accumulation of linalool and other alcohols. When assayed in vitro and in vivo, monoterpene alcohols at the concentrations present in AS fruit showed strong antifungal activity. We show here that terpene engineering in fruit peels could be a promising method for the development of new strategies to obtain resistance to fruit diseases.

Addition of Orange Peel in Orange Jam: Evaluation of Sensory, Physicochemical, and Nutritional Characteristics

“…The peel has lower amounts of TS as it protects the fruit from physical, chemical, and biological damage. Thus, it contains higher concentrations of essential oils [37]. TS include RS (glucose and fructose), NRS (sucrose), starch, and cellulose.…”

“…The lipophilic antioxidant capacity of orange peel was higher than that in the pulp (p < 0.05), contrary to the evaluation of hydrophilic antioxidant capacity. This effect occurs because the peel has essential oil vacuoles and a greater amount of fat-soluble bioactive compounds, such as carotenoids [37]. Pulp, on the other hand, contains a higher amount of ascorbic acid and water-soluble bioactive compounds as phenols [61,62].…”

Phyllosticta citricarpa and sister species of global importance to Citrus

“…Genetic methods may improve the CBS resistance of citrus. Rodriguez et al (2018) produced transgenic sweet orange trees with reduced production of d-limonene in the fruit flavedo and enhanced resistance against P. citricarpa, but these materials need to be assessed under field conditions.…”

Aspectos físico-químicos, nutricionais e sensoriais da adição de subprodutos de frutos do gênero Citrus em produtos de gelificação: uma revisão sistemática

“…A quantidade de lipídio da polpa dos frutos foi inferior àquelas das cascas e sementes, já que as glândulas de óleos essenciais presentes na casca (Raeissi et al, 2008) produzem o D-limoneno (casca da laranja) e o L-limoneno (casca do limão). Essas são as principais substâncias que atuam na proteção dos frutos contra o crescimento microbiano e na retenção da umidade na casca (Rodríguez et al, 2018). Maior teor de carboidrato foi observado na casca da Pera (66,05 g 100 g -1 ) e menor na polpa da Duncan (6,53 g 100 g -1 ).…”

“…Alterações sensoriais em alimentos adicionados de subprodutos de frutos do gênero Citrus ocorrem devido aos compostos fenólicos, fitatos e oxalatos. Exemplos dessas substâncias são a neosperidina (Pozzo et al, 2018), o D-limoneno e L-limoneno (Rodríguez et al, 2018) e as agliconas (Zoccali et al, 2018), que promovem sabores adoce, azedo e amargo (Ghasemi et al, 2018) e sensações de adstringência, pungência e refrescância (Glabasnia et al, 2018). Além disso, compostos solúveis e voláteis da casca, como os monoterpenos (limoneno, mirceno), sesquiterpenoides (α e β-sinensal), sesquiterpeno (valenceno) (Xie et al, 2018), monoterpenoides (decanal, linalol, neral, geranial, citronelal) dentre outros (Gonçalves et al, 2018), podem alterar o sabor e o aroma em produtos alimentícios.…”