Transcriptional response of susceptible and tolerant citrus to infection with Candidatus Liberibacter asiaticus

Candidatus Liberibacter asiaticus (Las), a non-culturable phloem-limited bacterium, is the suspected causal agent of Huanglongbing (HLB) in Florida. HLB is one of the most devastating diseases of citrus and no resistant cultivars have been identified to date, though tolerance has been suggested in the genus Poncirus. A recent study conducted in our laboratory demonstrated tolerance of US-897, a hybrid of Poncirus trifoliata (L. Raf.) and the susceptible ‘Cleopatra’ mandarin (Citrus reticulata Blanco), to Ca. L. asiaticus, the presumed causal agent of HLB in Florida (Albrecht & Bowman, HortScience 46 (2011) 16-22). This study compares transcriptional changes in tolerant US-897 and susceptible ‘Cleopatra’ mandarin seedlings in response to infection with Las using the Affymetrix GeneChip citrus array with the main objective of identifying genes associated with tolerance to HLB. Such genes may be suitable as potential targets for biotechnology approaches, providing one strategy to possibly control this destructive disease of citrus. Overall design: Fifteen months-old seedlings of the genotypes 'Cleopatra' mandarin and US-897 were graft-inoculated with non-infected (control) or Las-infected tissue from greenhouse-grown 'Valencia' (C. sinensis L.) plants. Six non-infected control plants each from 'Cleopatra' (MC) and US-897 (TC) and six infected plants each from Cleopatra (MI) and US-897 (TI) were used. Four to six leaves per plant were excised at 32 weeks after inoculation (wai) and immediately frozen in liquid nitrogen. Total RNA was extracted from all 24 plants. Equal amounts of RNA from two samples were pooled to obtain three biological replicates per genotype and treatment and used for hybridization on Affymetrix citrus microarrays.

Candidatus Liberibacter asiaticus (Las)，一种非可培养的韧皮部局限细菌，被认为是佛罗里达州黄龙病的疑似病原体。黄龙病是柑橘类植物中最具破坏性的疾病之一，迄今为止尚未发现具有抗性的栽培品种，尽管在 Poncirus 属中已提出耐受性。我们实验室近期进行的一项研究证明了 US-897 株（Poncirus trifoliata (L. Raf.) 与易感‘克洛巴特拉’橘 (Citrus reticulata Blanco) 的杂交种）对 Ca. L. asiaticus（佛罗里达州黄龙病的疑似病原体）的耐受性（Albrecht & Bowman, HortScience 46 (2011) 16-22）。本研究通过 Affymetrix 柑橘基因芯片，对比了耐受性 US-897 株和易感‘克洛巴特拉’橘幼苗在感染 Las 时的转录组变化，主要目的是鉴定与黄龙病耐受性相关的基因。这些基因可能成为生物技术方法中的潜在靶点，为控制柑橘类植物这种毁灭性疾病的策略之一提供可能。总体设计：将15个月大的 'Cleopatra' 橘和 US-897 等基因型幼苗通过嫁接接种来自温室种植的 'Valencia'（C. sinensis L.）植物的未感染（对照组）或 Las 感染组织。每组分别使用来自 'Cleopatra'（MC）和 US-897（TC）的六株非感染对照组植物以及来自 Cleopatra（MI）和 US-897（TI）的六株感染植物。在接种后32周，每株植物取4至6片叶子，并立即用液氮冷冻。从所有24株植物中提取总RNA。将两个样本的等量RNA混合，以获得每个基因型和处理的三个生物重复，并用于 Affymetrix 柑橘微阵列的杂交。