Huanglongbing (HLB) is a devastating citrus disease that is associated with bacteria of the genus 'Candidatus Liberibacter' (Ca. L.). Powerful diagnostic tools and management strategies are desired to control HLB. Host small RNAs (sRNA) play a vital role in regulating host responses to pathogen infection and are used as early diagnostic markers for many human diseases, including cancers. To determine whether citrus sRNAs regulate host responses to HLB, sRNAs were profiled from Citrus sinensis 10 and 14 weeks post grafting with Ca. L. asiaticus (Las)-positive or healthy tissue. Ten new microRNAs (miRNAs), 76 conserved miRNAs, and many small interfering RNAs (siRNAs) were discovered. Several miRNAs and siRNAs were highly induced by Las infection, and can be potentially developed into early diagnosis markers of HLB. miR399, which is induced by phosphorus starvation in other plant species, was induced specifically by infection of Las but not Spiroplasma citri that causes citrus stubborn-a disease with symptoms similar to HLB. We found a 35% reduction of phosphorus in Las-positive citrus trees compared to healthy trees. Applying phosphorus oxyanion solutions to HLB-positive sweet orange trees reduced HLB symptom severity and significantly improved fruit production during a 3-year field trial in south-west Florida. Our molecular, physiological, and field data suggest that phosphorus deficiency is linked to HLB disease symptomology.

Huanglongbing (HLB) causes citrus root systems to decline, which in turn contributes to deficiencies of essential nutrients followed by decline of the canopy and yield. This study was conducted on a 6-year-old ‘Valencia’ [Citrus sinensis (L.) Osb.] on Swingle rootstock (Citrus paradisi Macf. × Poncirus trifoliata (L.) Raf.) trees in a commercial grove near Immokalee, FL, to evaluate the effects of foliar applications of selected essential nutrients (N, K, Mn, Zn, B, and Mg) on growth and productivity of citrus trees infected with Candidatus Liberibacter asiaticus (CLas), the pathogen putatively associated with HLB in Florida. Mn, Zn, B, and Mg were applied in all experiments to drip at 0×, 0.5×, 1.0×, and 2.0×/spray of what has been traditionally recommended in Florida to correct deficiencies. Treatments were applied foliarly 3×/year with the sprays occurring during each growth flush for 5 years (2010–14). Thus, the 0×, 0.5×, 1.0×, and 2.0×/spray treatments resulted in 0×, 1.5×, 3.0×, and 6.0×/year to correct deficiencies. MnS04 and ZnSO4 were applied with or without KNO3 and in separate experiments were compared with Mn3(PO3)2 and Zn3(PO3)2, respectively. Disease incidence, foliar nutrient content, canopy volume, and yield were measured. At the beginning of the experiment, foliar N, P, Ca, Mg, Cu, and B were in the sufficient range and K, Mn, Zn, and Fe were slightly low. Disease incidence was very high with 83% and 98% of trees testing positive for CLas in 2010 and 2014, respectively. Nutrients that are not mobile or have limited mobility in plants, namely Mn, Zn, and B, demonstrated an increase in foliar concentration immediately after spray and in the annual averages. Foliar K increased from the deficient to the sufficient level by KNO3 sprays, but the mobile nutrients N and Mg did not show an increase in foliar levels, indicating that intraplant transport occurs in the presence of HLB. Foliar KNO3 application had a stronger effect on growth than yield. Yield was most strongly affected by application of MnSO4 where yield of the 3×/year treatment was 45% higher than that of the unsprayed control, but yield declined by 25% for the 6×/year treatment. Yield within 95% of the maximum occurred with foliar Mn concentrations of 70–100 µg·g−1 dry weight when Mn was applied as MnSO4, which is at the high end of the traditionally recommended 25–100 µg·g−1 dry weight range. The phosphite form of Mn [Mn3(PO3)2] depressed yield by an average of 25% across all application concentrations. Zn, B, and Mg did not significantly impact yield. Canopy volume demonstrated concave relationships across application concentrations for MnSO4 and ZnSO4 without KNO3 and Mn3(PO3)2, Zn3(PO3)2, Boron, and MgSO4 with KNO3, with the minimum occurring near the 3×/year application concentration. These data indicate a complex interaction in the amount of nutrients applied and their corresponding effects on foliar concentration, growth, and yield for HLB-affected trees. The results of this study at least partially explain the current confusion among scientists and the commercial industry in how to manage nutrition of HLB-affected citrus trees. The traditionally recommended approaches to correcting nutrient deficiencies need to be reconsidered for citrus with HLB.

Citrus trees affected by huanglongbing (HLB) become diminished, weak, and develop dieback resulting in reduced production. Decline in fruit yield ultimately prevents economically acceptable commercial citrus production. The objectives of this study were to evaluate the effects of severe pruning in combination with an enhanced foliar nutritional treatment on growth, yield, and juice quality of HLB-affected orange trees. The bacterial titer within the trees was monitored before and after treatments, and a cost–benefit analysis provided an economic evaluation of the treatments. Fifteen-year-old ‘Valencia’ orange (Citrus sinensis Macf.) trees on Swingle citrumelo rootstocks [C. paradisi × Poncirus trifoliata (L.) Raf.] with 100% incidence of HLB, confirmed by real-time polymerase chain reaction (PCR), were severely pruned back to the main scaffold branches. Between 2010 and 2015, foliar nutrients were sprayed on both pruned and nonpruned trees to target new flush growth. Three enhanced nutritional foliar treatments were evaluated and compared with a control foliar nutritional treatment that was considered to be a standard practice before endemic HLB. The enhanced nutritional treatments included a mixture of micro- and macronutrients commonly known as the “Boyd cocktail,” a micronutrient package labeled Fortress © (Florida Phosphorus LLC, Key Largo, FL) sprayed with potassium nitrate (KNO3), and the Fortress © micronutrient package sprayed with urea. The experiment was a split-plot with seven replications, with pruning as the main plots, and a foliar nutritional treatment as subplots. Tree pruning was performed in Feb. 2010 before the spring flush. Pruned trees grew longer shoots than the controls the year after pruning. Canopy volume and leaf area were greater with nonpruned trees, but the chlorophyll content per cm2 leaf area was higher in the pruned trees compared with nonpruned trees in 3 years of the 5-year experiment. Pruned and nonpruned trees bloomed and set fruit the first year of the experiment in the spring of 2010–11. The fruit crop for the 2010–11 and 2014–15 seasons, and the overall total fruit crop for the 2010–15 season on pruned trees were significantly lower than those on nonpruned trees. However, no significant yield differences were found between pruned and nonpruned trees in the 2011–12, 2012–13, and 2013–14 growing seasons. Fruit yields from pruned trees never surpassed the yields from nonpruned trees, and this was possibly due to the severe-pruning treatment. Thus, severe pruning, as used in this trial, was not cost effective through the first 5 years after pruning. The rapid regrowth response of the pruned trees, however, may indicate that a reduced pruning approach could be effective at rejuvenating the HLB-affected trees, and an alternative to tree removal and replanting. Enhanced foliar nutrition treatments provided some yield benefits, especially in the early years of the trial. However, the enhanced foliar nutrition treatments did not prove to be cost effective.