Investigation of phenolic and polyphenolic responses in phloem sap of citrus cultivars infected with Candidatus Liberibacter asiaticus

Abstract

Huanglongbing (HLB), caused by Candidatus liberibacter asiaticus, is a destructive citrus disease that disrupts plant physiology and is transmitted by Diaphorina citri. It is hypothesized that higher degree phenolic contents showed higher tolerance to many plant pathogens. In this regard, present study was designed to aimed to characterize phenolic and polyphenolic content in the phloem sap of healthy and HLB-infected citrus varieties, including Citrus reticulata (Kinnow, Daisy and Nagpur), Citrus sinensis (Saddis and Valencia Late), and Citrus limon (Lezben). Infection was confirmed through RT-PCR, and phenolic content was analyzed using Gas Chromatography–Mass Spectrometry (GC–MS). A total of 79–137 distinct compounds were identified per cultivar, revealing striking differences in metabolite accumulation. Several phenolics, such as hydroxyhydrocodone, cyclotrisiloxane derivatives, and 2-ferrocenyl-3-(m-tolyl)thiazolidin-4-one, were exclusively abundant in healthy peels but depleted in infected tissues, suggesting their protective roles in defense mechanism. Conversely, stress-associated metabolites, including oxycodone, 2-methoxy-4-vinylphenol, indole derivatives, and sesquiterpenes like longifolene and aromandendrene, were predominantly induced in CLas-infected samples, reflecting pathogen-triggered defense activation. Some compounds (4-Pyridinol, 2,6-bis[p-chlorophenyl], Oxycodone, 2-Methoxy-4-vinylphenol, 3-Methoxyacetophenone, Acethydrazide derivatives) displayed significant fold-changes between healthy and diseased states, with p-values <0.05, underscoring their potential as metabolic biomarkers for early HLB detection. Importantly, cultivars with higher concentration and diversity of phenolic compounds, such as in Kinnow and Valencia Late, demonstrated broader chemical defense responses, supporting the hypothesis that elevated phenolic abundance enhances tolerance against pathogen stress. By employing a multi-varietal comparative approach, this study provides novel evidence of infection-induced metabolic markers that may facilitate the development of early diagnostic tools resistance breeding strategies. Deciphering the mechanisms underlying citrus tolerance to HLB will aid in the development of commercially viable tolerant cultivars.