Key role of heat shock protein expression induced by ampicillin in citrus defense against huanglongbing: a transcriptomics study

Authors: YANG, CHUANYU - UNIVERSITY OF FLORIDA; POWELL, CHARLES - UNIVERSITY OF FLORIDA; Duan, Ping; LIN, XIONGJIE - FUJIAN ACADEMY; FAN, GOUCHENG - FUJIAN ACADEMY; HU, HANQING - FUJIAN ACADEMY; ZHANG, MUQING - UNIVERSITY OF FLORIDA

Submitted to: Agronomy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/29/2022
Publication Date: 6/2/2022
Citation: Yang, C., Powell, C., Duan, Y., Lin, X., Fan, G., Hu, H., Zhang, M. 2022. Key role of heat shock protein expression induced by ampicillin in citrus defense against huanglongbing: a transcriptomics study. Agronomy. 12:1356. https://doi.org/10.3390/agronomy12061356.
DOI: https://doi.org/10.3390/agronomy12061356

Interpretive Summary: Citrus huanglongbing (HLB) is a catastrophic disease that threatens the citrus industry worldwide. HLB is caused by the psyllid-transmitted, phloem-limited bacteria Candidatus Liberibacter asiaticus (CLas). Our previous studies demonstrated that Ampicillin (Amp) can reduce CLas titer and mitigate HLB symptoms in HLB-affected citrus. In addition, Amp treatment of HLB-affected citrus promotes plant growth and enriches the population of beneficial bacteria that can induce pathogen defense pathways. However, due to public concerns about the emergence of antibiotic-resistant bacteria and their potential effects on humans, application of Amp to citrus crops in commercial groves has not yet been approved by the Environmental Protection Agency or other regulatory agencies. In this study, we applied nano-emulsions coupled with Amp that had different droplet sizes to HLB-affected citrus. We then examined changes in gene expression to identify which pathways are altered by different concentrations of Amp using RNA-seq. Our results indicated that the heat shock proteins (Hsps) were related to a number of defense related such as Gibberellic acid methyltransferase 2 and L-ascorbate peroxidase 2. The expression of these defense related genes was positively or negatively related to Hsps in response to treatment with Amp at different concentrations. On the other hand, the Amp did not directly kill or suppress CLas in our co-culture system of CLas in vitro, indicating effects of Amp on CLas may rely on the up-regulated Hsp and related defense genes of host plants. These findings are valuable for developing novel strategies to combat citrus HLB.

Technical Abstract: Citrus huanglongbing (HLB) is a devastating disease for the citrus industry. Earlier studies showed that ampicillin (Amp) can suppress titers of the pathogen that causes HLB, the bacteria Candidatus Liberibacter asiaticus (CLas), in HLB-affected citrus. CLas has not yet been cultured, so the mechanisms of Amp against CLas are unclear. Some chemicals were demonstrated to trigger citrus defense systems against CLas. Therefore, we hypothesize that Amp may induce citrus defenses against CLas. Here we applied three nano-formulations of varying droplet size to HLB-affected citrus to achieve different accumulated concentrations of Amp (high, medium, low) in the plants. We then used RNA-seq to analyze induction of gene expression of citrus defense systems against CLas in response to different concentrations of Amp. The results indicated that at all accumulated concentrations of Amp can significantly suppress CLas titer and mitigate HLB symptoms. Transcriptomic analyses showed that Amp treatment induced expression of heat shock proteins (Hsps) in HLB-affected citrus, and these Hsps were significantly related to several defense genes encoding R proteins, transcription factors, splicing factors, RNA-binding proteins, RNA-dependent RNA polymerase, Gibberellic acid methyltransferase 2, L-ascorbate peroxidase 2, and ferruginol synthase that confer resistance to CLas in citrus plants. Taken together, these results suggest that Amp treatment of citrus plants can trigger expression of Hsps and related defense genes to respond to CLas infection. These findings are valuable for developing novel strategies to combat citrus HLB.