No abstract

Boron (B) toxicity in Citrus is a common physiological disorder leading to reductions in both productivity and quality. Studies on how Citrus roots evade B toxicity may provide new insight into plant tolerance to B toxicity. Here, using Illumina sequencing, differentially expressed microRNAs (miRNAs) were identified in B toxicity-treated Citrus sinensis (tolerant) and C. grandis (intolerant) roots. The results showed that 37 miRNAs in C. grandis and 11 miRNAs in C. sinensis were differentially expressed when exposed to B toxicity. Among them, miR319, miR171, and miR396g-5p were confirmed via 5′-RACE and qRT-PCR to target a myeloblastosis (MYB) transcription factor gene, a SCARECROW-like protein gene, and a cation transporting ATPase gene, respectively. Maintenance of SCARECROW expression in B treated Citrus roots might fulfill stem cell maintenance, quiescent center, and endodermis specification, thus allowing regular root elongation under B-toxic stress. Down-regulation of MYB due to up-regulation of miR319 in B toxicity-treated C. grandis roots might decrease the number of root tips, thereby dramatically changing root system architecture. Our findings suggested that miR319 and miR171 play a pivotal role in Citrus adaptation to long-term B toxicity by targeting MYB and SCARECROW, respectively, both of which are responsible for root growth and development.

Citrus huanglongbing (HLB) is a devastating disease for the citrus industry. The previous studies demonstrated that oxytetracycline and penicillin are effective antibiotics against Candidatus Liberibacter asiaticus (CLas). However, since CLas is uncultured, the mechanisms of action of antibiotics against CLas are still unclear. It was recently reported that the endophytic microbial communities are associated with the progression of citrus HLB after oxytetracycline and penicillin treatment. Therefore, we hypothesize that penicillin has greater antibacterial activity against CLas than oxytetracycline, which may be associated with the alteration of the structure and function of endophytic microbial communities in HLB-affected citrus in response to these antibiotics. To test this hypothesis, the microbiome of HLB-affected citrus leaves treated with these two antibiotics was analyzed using a metagenomic method. Our results indicate that the microbial structure and function in HLB-affected citrus were altered by these two antibiotics. The relative abundance of beneficial bacterial species, including Streptomyces avermitilis and Bradyrhizobium, was higher in penicillin-treated plants compared to those treated with oxytetracycline, and the relative abundance of the bacterial species (such as Propionibacterium acnes and Synechocystis sp PCC 6803) associated with CLas survival was lower for penicillin-treated plants compared to oxytetracycline-treated plants. These results indicate that penicillin has greater antibacterial activity against CLas. Based on the metagenomic analysis, this study elucidated the mechanism for the observed increase in antibacterial activity of penicillin against CLas. The data presented here are not only invaluable for developing eco-friendly and effective biocontrol strategies to combat citrus HLB, but also provide a method for revealing mechanism of antimicrobial against uncultured bacteria in host.

The mechanisms underlying plant tolerance to boron (B) excess are far from fully understood. Here we characterized the role of the miR397-CsiLAC4/CsiLAC17 (from Citrus sinensis) module in regulation of B flow.Live-cell imaging techniques were used in localization studies. A tobacco transient expression system tested modulations of CsiLAC4 and CsiLAC17 by miR397. Transgenic Arabidopsis were generated to analyze the biological functions of CsiLAC4 and CsiLAC17. CsiLAC4's role in xylem lignification was determined by mRNA hybridization and cytochemistry. In situ B distribution was analyzed by laser ablation inductively coupled plasma mass spectrometry.CsiLAC4 and CsiLAC17 are predominantly localized in the apoplast of tobacco epidermal cells. Overexpression of CsiLAC4 in Arabidopsis improves the plants' tolerance to boric acid excess by triggering high-B-dependent lignification of the vascular system's cell wall and reducing free B content in roots and shoots. In Citrus, CsiLAC4 is expressed explicitly in the xylem parenchyma and is modulated by B-responsive miR397. Upregulation of CsiLAC4 in Citrus results in lignification of the xylem cell walls, restricting B flow from xylem vessels to the phloem.CsiLAC4 contributes to plant tolerance to boric acid excess via high-B-dependent lignification of cell walls, which set up a 'physical barrier' preventing B flow.