Wang, Yuanzhi Zimmy

The global citrus industry faces a great threat from Huanglongbing (HLB), a destructive disease caused by ‘Candidatus Liberibacter asiaticus’ (CLas) that induces significant economic losses without any known cure. Understanding how citrus plants defend against HLB, particularly at the early stages of infection, is crucial for developing long-term solutions. This study investigated the earliest metabolic responses of fresh citrus leaves to CLas infection using untargeted metabolomics and machine learning models. HLB-tolerant and HLB-sensitive cultivars were compared to analyze their biochemical reactions within 48 hours post-infection. HESI/Q-Orbitrap MS analysis identified temporal differential metabolites, revealing distinct metabolic pathways activated in response to CLas infection. Both cultivars responded by increasing specific metabolite concentrations, such as flavonoids, within 2 hours post-infection, but the HLB-tolerant cultivar maintained higher levels throughout the 48-hour period. This early metabolic activity could influence long-term plant health by enhancing disease resistance and reducing pathogen impact. These findings provide potential biomarkers for breeding HLB-resistant cultivars and offer valuable insights for developing sustainable management strategies to mitigate the impact of HLB on the citrus industry, ensuring its long-term productivity and economic viability.

Long non-coding RNAs (lncRNAs) serve as crucial regulators in plant response to various diseases, while none have been systematically identified and characterized in response to citrus Huanglongbing (HLB) caused by Candidatus Liberibacter asiaticus (CLas) bacteria. Here, we comprehensively investigated the transcriptional and regulatory dynamics of the lncRNAs in response to CLas. Samples were collected from leaf midribs of CLas- and mock-inoculated HLB-tolerant rough lemon (Citrus jambhiri) and HLB-sensitive sweet orange (C. sinensis) at week 0, 7, 17, and 34 following inoculation using CLas+ budwood of three biological replicates in the greenhouse. A total of 8,742 lncRNAs, including 2,529 novel lncRNAs, were identified from RNA-seq data with rRNA-removed from strand-specific libraries. Genomic variation analyses of conserved lncRNAs from 38 citrus accessions showed that 26 single nucleotide polymorphisms (SNPs) were significantly correlated with HLB. In addition, lncRNA-mRNA weighted gene co-expression network analysis (WGCNA) showed a significant module correlated with CLas-inoculation in rough lemon. Notably, the most significant LNC_28805 and multiple co-expressed genes related to plant defense in the module were targeted by miRNA5021, suggesting that LNC28805 might compete with endogenous miR5021 to maintain the homeostasis of immune gene expression levels. Candidate WRKY33 and SYP121 genes targeted by miRNA5021 were identified as two key hub genes interacting with bacteria pathogen response genes based on the prediction of protein-protein interaction (PPI) network. These two genes were also found within HLB-associated QTL in linkage group 6. Overall, our findings provide a reference for a better understanding of the role of lncRNAs involved in citrus HLB regulation.