Identification and Expression Pattern of a Novel NAM,ATAF, and CUC-Like Gene from <Emphasis Type="Italic">Citrus sinensis</Emphasis> Osbeck

A citrus NAM, ATAF, and CUC (NAC)-like gene (CitNAC) was isolated from fruit tissues of Citrus sinensis Osbeck using complementary DNA (cDNA) amplified fragment length polymorphism and rapid amplification of cDNA ends techniques. Its full length was 988 bp in which 781 bp form the open reading frame, coding for a protein of 264 amino acids. Sequence comparison revealed that CitNAC possesses the general structural features at the N terminus of the NAC domains. Phylogenetic analysis results showed that CitNAC was closely related to AtNAP and PeNAP, which are involved in plant organ senescence. Gene expression analysis showed that the messenger RNA level of CitNAC was just detected in fruit peel and pulp during fruit ripening or senescence stage. The observed expression pattern of CitNAC along with the result of phylogenetic analysis suggested that CitNAC is related to fruit development and senescence.     

Suppressive effects of the peel of Citrus kawachiensis (Kawachi Bankan) on astroglial activation,tau phosphorylation,and inhibition of neurogenesis in the hippocampus of type 2 diabetic db/db mice

We previously reported that the dried peel powder of Citrus kawachiensis exerted anti-inflammatory effects in the brain in several animal models. Hyperglycemia induces inflammation and oxidative stress and causes massive damage in the brain; therefore, we herein examined the anti-inflammatory and other effects of the dried peel powder of C. kawachiensis in the streptozotocin-induced hyperglycemia mice model and in the type 2 diabetic db/db mice model. The C. kawachiensis administration inhibited microglial activation in the hippocampus in the streptozotocin-injected mice. Moreover, The C. kawachiensis treatment inhibited astroglial activation in the hippocampus and the hyperphosphorylation of tau at 231 of threonine and 396 of serine in hippocampal neurons, and also relieved the suppression of neurogenesis in the dentate gyrus of the hippocampus in the db/db mice. It was suggested that the dried peel powder of C. kawachiensis exerts anti-inflammatory and neuroprotective effects in the brain.     

Metabolomic analyses of plasma and liver of mice fed with immature Citrus tumida peel

ABSTRACT

In this study, we investigated the effects of dietary supplementation of Citrus tumida hort. ex Tanaka on food intake, body and fat tissue weights, and metabolic profiles of plasma and liver in mice. Supplementation with 5% (w/w) of peels of immature C. tumida (PIC) for 4 weeks significantly suppressed body weight gain and decreased adipose tissue weight in epididymal, perirenal, and subcutaneous fats. Metabolome analyses showed that 2-hydroxyvaleric acid levels were reduced in the blood plasma of mice fed with PIC. PIC supplementation significantly elevated dipeptide (Thr-Asp, Ser-Glu, and Ala-Ala), glucuronic acid, and S-methylglutathione levels, and significantly reduced betaine aldehyde levels in the liver. In conclusion, PIC supplementation affects the metabolism of fatty acids, pectin, glutathione, and choline, showing potential beneficial effects for metabolic syndrome and obesity. PIC may be developed as a functional food and used in the treatment of these diseases.     

Growth and root hydraulic conductivity of several citrus rootstocks under salt and polyethylene glycol stresses

The water relations responses to salt of several important citrus rootstocks such as Swingle citrumelo, sour orange, and Milam lemon have not been studied in detail before. Studies were set up to compare growth and root hydraulic properties of these rootstocks to other citrus rootstocks by exposing them to NaCl and polyethylene glycol (PEG) stresses. Seedlings of 7 citrus rootstocks were irrigated for 5 months with nutrient solutions containing NaCl or PEG that had been adjusted to osmotic potentials of -0.10, -0.20 or -0.35 MPa. The 7 rootstocks studied were sour orange (Citrus aurantium), Cleopatra mandarin (Citrus reticulata Blanco), Swingle citrumelo (C. paradisi x P. trifoliata), Carrizo citrange (C. sinensis x P. trifoliata), rough lemon (Citrus jambhiri Lush), Milam lemon (C. jambhiri hybrid), and trifoliate orange (Poncirus trifoliata [L.] Raf.). In both shoot and root growth, Cleopatra mandarin and sour orange were the least sensitive to salt, Milam and trifoliate orange were the most sensitive, and rough lemon, Swingle, and Carrizo were intermediate in sensitivity. Even though the roots were exposed to solutions of equal osmotic potentials, plant growth and root conductivity were reduced more by the PEG treatments than the corresponding NaCl treatments. At -0.10 and -0.20 MPa, shoot and root dry weights were reduced 16 to 55% by NaCl and 24 to 68% by PEG. Shoot root ratio was lowered at the higher concentrations, particularly by PEG. There was a major decrease in root conductivity caused by NaCl at -0.10 MPa (19 to 30% in sour orange and Cleopatra mandarin and 78 to 85% in trifoliate orange and Milam). Conductivity decreased more at -0.20 and -0.35 MPa, but not proportionally as much as at -0.10 MPa. Root weight per unit length increased at the higher salt levels, particularly in trifoliate orange. Water flow rate through root systems followed the same trend as root conductivity; salt affected sour orange and Cleopatra mandarin the least and trifoliate orange and Milam the most. However, reductions in fibrous root length by salt treatment differed. Root lengths of Swingle and Carrizo were least affected by salt while sour orange. Milam, and rough lemon were the most affected. Hence, even though sour orange and Cleopatra mandarin were more tolerant than the other rootstocks in terms of water flow rate or root conductivity, these 2 rootstocks showed a proportionally greater decrease in root length than Carrizo, Swingle, or trifoliate orange.     

绿肥种植配施减量氮肥对柑橘园杂草群落组成及多样性的影响

以湖北当阳柑橘园杂草群落为对象,研究绿肥种植(自然生草和间种光叶苕子)下配施减量氮肥处理(当地推荐施氮肥量的0%、70%、85%、100%)对橘园杂草群落组成和多样性的影响,并运用Pearson相关分析和典范对应分析探究土壤因子对不同处理下杂草多样性和物种组成的影响。结果显示:(1)经样方调查,试验区共记录杂草植物21科29属30种,以禾本科和菊科为主,稗(Echinochloa crusgalli(L.)Beauv.)、酢浆草(Oxalis corniculata L.)、野老鹳草(Geranium carolinianum L.)等是所有处理中的共有种,自然生草下以匙叶鼠麴草(Gnaphalium pensylvanicum Willd.)和稗为优势种,间种光叶苕子(Vicia villosa Roth.var.glabrescens Koch.)下以酢浆草为优势种;(2)不同氮肥施入量下柑橘园草本物种Margalef指数、Patrick指数、Shannon-Wiener多样性指数和Pielou均匀度指数均无显著差异;相同施肥量下,间种光叶苕子能显著降低柑橘园杂草群落的物种多样性指数;(3)速效磷是显著影响柑橘园草本植物多样性指数的土壤因子;(4)不同处理下影响杂草物种分布的主要土壤因子是速效磷、pH值、含水率和温度。本研究认为无论减施氮肥与否,间种光叶苕子比自然生草更能显著降低杂草群落的物种多样性,从而避免恶性杂草与果树争肥。     

Evaluation of Aspergillus aculeatus GC-09 for the biological control of citrus blue mold caused by Penicillium italicum

Blue mold caused by Penicillium italicum is a severe postharvest disease in citrus fruits. In this study, the fermentation product (FP-E) of Aspergillus aculeatus GC-09, an endophytic fungus isolated from a citrus plant, was found to exhibit antifungal activity against P. italicum with a MIC of 0.3125 mg/mL. The fungus A. aculeatus GC-09 was identified based on the studies of morphology and ITS nucleotide sequence. FP-E significantly inhibited the spore germination and mycelial growth of P. italicum. Scanning electron microscopy (SEM) results of P. italicum treated with FP-E showed shrunken, distorted and collapsed hyphae and conidiospores, indicative of the cell membrane damage, which was further confirmed by the propidium iodide (PI) fluorescent staining analysis. Consistent with the microscopy observation, FP-E led to the leakage of cellular constituents from P. italicum, which is evident from the increase in electrical conductivity and nucleic acid contents in the mycelial solution incubated with FP-E. In addition, FP-E treatment considerably increased the intracellular reactive oxygen species (ROS) content, and reduced the enzyme activities of both catalase (CAT) and peroxidase (POD) in P. italicum cells. Furthermore, orange fruits treated with FP-E showed fewer disease symptoms compared to the untreated fruits. These results suggested that the antifungal activity of FP-E might be associated with the disruption of cell membrane integrity, the accumulation of ROS level, and the reduction of the antioxidant enzymes activity of P. italicum. Therefore, A. aculeatus GC-09 might be a potential microbial resource for the biocontrol of citrus postharvest blue mold.     

PRpnp,a novel dual activity PNP family protein improves plant vigour and confers multiple stress tolerance in Citrus aurantifolia

Under field conditions, plants are often simultaneously exposed to several abiotic and biotic stresses resulting in significant reductions in growth and yield; thus, developing a multi-stress tolerant variety is imperative. Previously, we reported the neofunctionalization of a novel PNP family protein, Putranjiva roxburghii purine nucleoside phosphorylase (PRpnp) to trypsin inhibitor to cater to the needs of plant defence. However, to date, no study has revealed the potential role and mechanism of either member of this protein group in plant defence. Here, we overexpressed PRpnp in Citrus aurantifolia which showed nuclear-cytoplasmic localization, where it functions in maintaining the intracellular purine reservoir. Overexpression of PRpnp significantly enhanced tolerance to salt, oxidative stress, alkaline pH, drought and two pests, Papilio demoleus and Scirtothrips citri in transgenic plants. Global gene expression studies revealed that PRpnp overexpression up-regulated differentially expressed genes (DEGs) related to ABA- and JA-biosynthesis and signalling, plant defence, growth and development. LC–MS/MS analysis validated higher endogenous ABA and JA accumulation in transgenic plants. Taken together, our results suggest that PRpnp functions by enhancing the endogenous ABA and JA, which interact synergistically and it also inhibits trypsin proteases in the insect gut. Also, like other purine salvage genes, PRpnp also regulates CK metabolism and increases the levels of CK-free bases in transgenic Mexican lime. We also suggest that PRpnp can be used as a potential candidate to develop new varieties with improved plant vigour and enhanced multiple stress resistance.     

Proteins associated with salt adaptation in citrus and tomato cells: Involvement of 26 kDa polypeptides

Stress-induced changes in proteins of molecular weights of 25 and 27 kDa are reported for NaCl-adapted cells of Shamouti orange ( Citrus sinensis L. Osbeck) and the cultivated VFNT cherry tomato ( Lysopersicon esculentum Mill.), respectively. Salt-adapted cell lines of both species grew much faster in the presence of salt than non-adapted cells and their growth rate was similar to that of non-adapted cells in the absence of salt. Sodium dodecyl sulfate-extractable citrus and tomato polypeptides were analyzed by one- and two-dimensional polyacrylamide gel electrophoresis (PAGE). The enhancement of a citrus 25 kDa protein associated with salt tolerance was actually resolved only on two-dimensional PAGE. It is a constitutive protein, i.e. it was present in salt-tolerant ceils whether grown in the presence or absence of salt. The NaCl-induced changes of the tomato 27 kDa protein could be resolved on one-dimensional PAGE. This protein was first markedly suppressed by a salt shock and the time required for its recovery was dependent on the degree of salt tolerance. Prolonged adaptation resulted in an increase of this protein. Protein profiles of citrus and tomato cells grown in the absence and presence of salt revealed that while in citrus the level of most proteins was suppressed in the presence of salt, the opposite was true for tomato proteins under similar conditions. Comparison between the changes in citrus and tomato and results already published for tobacco show that many salt-induced changes in proteins are species-specific and that no obvious similarities exist.