Gamma-Aminobutyric Acid Accumulation Contributes to Citrus sinensis Response against ‘Candidatus Liberibacter Asiaticus’ via Modulation of Multiple Metabolic Pathways and Redox Status

Abstract

Huanglongbing (HLB; also known as citrus greening) is the most destructive bacterial disease of citrus worldwide with no known sustainable cure yet. Herein, we used non-targeted metabolomics and transcriptomics to prove that γ-aminobutyric acid (GABA) accumulation might influence the homeostasis of several metabolic pathways, as well as antioxidant defense machinery, and their metabolism-related genes. Overall, 41 metabolites were detected in ‘Valencia’ sweet orange (Citrus sinensis) leaf extract including 19 proteinogenic amino acids (PAA), 10 organic acids, 5 fatty acids, and 9 other amines (four phenolic amines and three non-PAA). Exogenous GABA application increased most PAA in healthy (except L-threonine, L-glutamine, L-glutamic acid, and L-methionine) and ‘Candidatus L. asiaticus’-infected citrus plants (with no exception). Moreover, GABA accumulation significantly induced L-tryptophan, L-phenylalanine, and α-linolenic acid, the main precursors of auxins, salicylic acid (SA), and jasmonic acid (JA), respectively. Furthermore, GABA supplementation upregulated most, if not all, of amino acids, phenolic amines, phytohormone metabolism-related, and GABA shunt-associated genes in both healthy and ‘Ca. L. asiaticus’-infected leaves. Moreover, although ‘Ca. L. asiaticus’ induced the accumulation of H2O2 and O2•− and generated strong oxidative stress in infected leaves, GABA possibly stimulates the activation of a multilayered antioxidative system to neutralize the deleterious effect of reactive oxygen species (ROS) and maintain redox status within infected leaves. This complex system comprises two major components: (i) the enzymatic antioxidant defense machinery (six POXs, four SODs, and CAT) that serves as the front line in antioxidant defenses, and (ii) the non-enzymatic antioxidant defense machinery (phenolic acids and phenolic amines) that works as a second defense line against ‘Ca. L. asiaticus’-induced ROS in citrus infected leaves. Collectively, our findings suggest that GABA might be a promising alternative eco-friendly strategy that helps citrus trees battle HLB particularly, and other diseases in general.

Non-proteinogenic amino acids mitigate oxidative stress and enhance the resistance of common bean plants against Sclerotinia sclerotiorum

“…Therefore, it is necessary to introduce some eco-friendly alternatives to control S. sclerotiorum particularly, and soil-borne phytopathogens in general. NPAAs have emerged as potent fungistatic agents that could be promising eco-friendly alternatives to control fungal phytopathogens ( Marcucci et al., 2010 ; Justyna and Ewa, 2013 ; de Oliveira et al., 2019 ; Guo et al., 2022 ), as well as bacterial ones ( Nehela and Killiny, 2019 ; Nehela and Killiny, 2023a , b ).…”

“…However, GABA was reported to be accumulated as a part of the metabolomic responses of soybean plants against S. sclerotiorum ( de Oliveira et al., 2019 ), as well as grapevine response against Neofusicoccum parvum . Additionally, GABA plays a positive role in the host response to bacterial phytopathogens such as Agrobacterium tumefaciens ( Chevrot et al., 2006 ), Pseudomonas syringae ( Park et al., 2010 ; Deng et al., 2020 ), and ‘ Candidatus Liberibacter Asiaticus’ ( Killiny and Nehela, 2017 ; Nehela and Killiny, 2019 , 2022 ; Nehela and Killiny, 2023a , b ). In other words, augmented GABA shunt activity was reported to be associated with plant defense against Agrobacterium in tobacco ( Chevrot et al., 2006 ), Botrytis cinerea in tomato ( Seifi et al., 2013 ), and ‘ Candidatus Liberibacter Asiaticus’ in citrus ( Killiny and Nehela, 2017 ; Nehela and Killiny, 2019 , 2022 ; Nehela and Killiny, 2023a , b ).…”

“…However, the biochemical and physiological mechanisms behind these roles are poorly understood. This might be due to the modulation of multiple metabolic pathways and redox status ( Nehela and Killiny, 2023a ). For instance, the tolerance of citrus plants against the bacterial pathogen ‘ Ca .…”

“…For instance, the tolerance of citrus plants against the bacterial pathogen ‘ Ca . Liberibacter asiaticus’ was enhanced by the positive regulation of the GABA shunt and associated pathways such as amino acids ( Killiny and Nehela, 2017 ; Nehela and Killiny, 2023a ), polyamines ( Nehela and Killiny, 2020 ; Nehela and Killiny, 2023a ), TCA cycle, phytohormones ( Nehela and Killiny, 2023b ), and other stress-associated metabolites ( Nehela and Killiny, 2020 ; Nehela and Killiny, 2023a ). The dual function of GABA as a metabolite and as a signaling component enables plants to cope with different conditions.…”

“…Histochemical localization of superoxide anion (O 2 •− ) was assessed as described by Adám 1989 ( Adám et al., 1989 ) with slight modifications ( Nehela et al., 2021 ; Nehela and Killiny, 2023a ). Briefly, 10 fresh leaf discs (~1 cm 2 ) were immersed and vacuum infiltrated with 0.1% nitro blue tetrazolium (NBT; Sigma–Aldrich, Darmstadt, Germany) until the development of a blue/purple color, then bleached, and intensity of developed blue/purple color was assessed the as described above.…”