Quantification of Flavonoids, Phenols and Antioxidant Potential from Dropped Citrus reticulata Blanco Fruits Influenced by Drying Techniques.

Physiologically dropped immature Citrus reticulata Blanco fruits are regarded as waste and discarded in the citrus orchard but are a good source of bioactive compounds including flavonoids, antioxidants and total phenols. A study was undertaken to identify and quantify these bioactive compounds and to investigate the influence of different drying techniques, namely freeze drying and hot air oven drying, on flavonoids namely flavanone glycosides, antioxidant potential and total phenol content in immature dropped fruits of Citrus reticulata Blanco. Flavonoids were quantified in high-performance liquid chromatography (HPLC). The antioxidant activity were investigated with three assays azino-bis [3-ethylbenzthiazoline-6-sulfonic acid]) (ABTS), 2,2-diphenyl-1-picrylhydrazyl radical (DPPH), Ferric Reducing Ability of Plasma (FRAP) and total phenol content was determined. Freeze dried samples of 12 and 14 mm size retained maximum hesperidin flavonoid content (27.03% and 27.20%) as compared to the hot air dried samples (17.99%) and retained higher phenolic content ranged from 50.54-54.19 mg GAEL-1. The antioxidant activity in freeze dried fruits was from 12.21-13.55 mM L-1 Trolox and 15.27-16.72 mM L-1 Trolox with ABTS, DPPH assay and FRAP values ranging from 7.31-9.07 mM L-1 Trolox. Significant positive correlation was found between the flavonoid hesperidin with antioxidant assays and total phenolic content (TPC). The results showed that waste citrus fruits can act as potential source of bioflavonoids, especially hesperidin, and antioxidants for pharmaceutical as well as nutraceutical industry.

Herbal treatments for migraine: A systematic review of randomised-controlled studies.

Herbal treatments are often used as a treatment for migraine. Therefore, an evaluation of their safety and efficacy is important. Based on the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, and Cochrane Collaboration's tool for assessing the risk of bias, a systematic literature review of randomised, controlled human trials assessing the effects of herbal treatments delivered as a single ingredient for the acute or prophylactic treatment of migraine were conducted. Studies were identified through electronic database searches on Medline (Pubmed), Cochrane Library, Scopus, and CINAHL. Nineteen studies were identified examining the effects on migraine of feverfew, butterbur, curcumin, menthol/peppermint oil, coriander, citron, Damask rose, chamomile, and lavender. Overall, findings on the efficacy of feverfew were mixed and there was positive, albeit limited evidence for butterbur. There were positive, preliminary findings on curcumin, citron, and coriander as a prophylactic treatment for migraine, and the use of menthol and chamomile as an acute treatment. However, the risk of bias was high for many studies. The results of this systematic review suggest that several herbal medicines, via their multifactorial physiological influences, present as potential options to enhance the treatment of migraine. However, further high-quality research is essential to examine their efficacy and safety as a treatment for migraine.

Mycorrhizal effectiveness in Citrus macrophylla at low phosphorus fertilization.

An experiment was conducted with seedlings of Citrus macrophylla Wester to study the effects of P nutrition on plants inoculated with a mixture (Rhizophagus irregularis and Funneliformis mosseae) of arbuscular mycorrhizal (AM) fungi. The treatments consisted of factorial combinations of two factors: mycorrhization (-AM: non-inoculated plants, and +AM: inoculated plants) and P nutrition (0, 0.1, 1, and 5 mM P). After the P treatments had been applied for 165 days, the AM fungi showed an important effect on plant growth and P uptake, but this effect depended on the P fertilization. In the absence of P fertilization, inoculation with the AM fungi had little impact on P nutrition and plant growth. However, when 0.1 or 1 mM P was supplied, inoculation had a clear beneficial effect on plant growth, since P nutrition was significantly improved, the maximum growth of the +AM plants occurring at 1 mM P. The supply of 5 mM P did not increase plant growth with regard to 1 mM P due to a lack of improvement in leaf P nutrition and photosynthesis. The higher demand of the AM fungi in the roots of the +AM plants for sucrose reduced the concentration of sucrose in the leaves of plants receiving 5 mM P, and of fructose and glucose in the roots of plants supplied with 0.1 or 1 mM P, relative to the -AM plants. The inoculated plants grown with 5 mM P had a decreased starch concentration in their roots, in order to supply the high sugar demand of the AM fungi. The C drain towards the AM fungi in the +AM plants may have been compensated by a higher photosynthetic rate and improved mineral nutrition. Inoculation improved plant P nutrition in the 0.1 and 1 mM P treatments but had a lesser effect at 5 mM P. The tissue levels of certain nutrients, such as Mg, improved with inoculation regardless of the P treatment, but those of other nutrients - such as Zn or Fe - increased more in the +AM plants with lower P supply. So, in general, the +AM C. macrophylla plants receiving the highest P supply did not show improved mineral nutrition relative to the -AM plants. Overall, the results indicate that when the availability of P to C. macrophylla plants is high, the lower benefits received by the plants from the C-for-P trade can convert a mutualistic relationship between the host plant and AM fungi into a parasitic one since colonization can persist even when the availability of P in the soil is high.

Challenges for Managing Candidatus Liberibacter spp. (Huanglongbing Disease Pathogen): Current Control Measures and Future Directions.

Huanglongbing (HLB; "citrus greening" disease) has caused significant damages to the global citrus industry as it has become well established in leading citrus-producing regions and continues to spread worldwide. Insecticidal control has been a critical component of HLB disease management, as there is a direct relationship between vector control and Candidatus Liberibacter spp. (i.e., the HLB pathogen) titer in HLB-infected citrus trees. In recent years, there have been substantial efforts to develop practical strategies for specifically managing Ca. Liberibacter spp.; however, a literature review on the outcomes of such attempts is still lacking. This work summarizes the greenhouse and field studies that have documented the effects and implications of chemical-based treatments (i.e., applications of broad-spectrum antibiotics, small molecule compounds) and nonchemical measures (i.e., applications of plant-beneficial compounds, applications of inorganic fertilizers, biological control, thermotherapy) for phytopathogen control. The ongoing challenges associated with mitigating Ca. Liberibacter spp. populations at the field-scale, such as the seasonality of the phytopathogen and associated HLB disease symptoms, limitations for therapeutics to contact the phytopathogen in planta, adverse impacts of broad-spectrum treatments on plant-beneficial microbiota, and potential implications on public and ecosystem health, are also discussed.

Temperatures during flower bud development affect pollen germination, self-incompatibility reaction and early fruit development of clementine (Citrus clementina Hort. ex Tan.).

One of the key environmental factors affecting plant reproductive systems is temperature. Characterising such effects is especially relevant for some commercially important genera such as Citrus. In this genus, failure of fertilisation results in parthenocarpic fruit development and seedlessness, which is a much-prized character. Here, we characterise the effects of temperature on flower and ovary development, and on pollen-pistil interactions in 'Comune' clementine (Citrus clementina Hort. ex Tan.). We examine flower bud development, in vitro pollen germination and pollen-pistil interaction at different temperatures (15, 20, 25 or 30 °C). These temperatures span the range from 'cold' to 'hot' weather during the flowering season in many citrus-growing regions. Temperature had a strong effect on flower and ovary development, pollen germination, and pollen tube growth kinetics. In particular, parthenocarpic fruit development (indicated by juice vesicle growth) was initiated early if flowers were exposed to warmer temperatures during anthesis. Exposure to different temperatures during flower bud development also alters expression of the self-incompatibility reaction. This affects the point in the pistil at which pollen tube growth is arrested and confirms the role of sub- and supra-optimal temperatures in determining the numbers of pollen tubes reaching the ovary.

[SUBTROPICAL CULTURES FRUIT BIOCHIMICAL DEFIRNITION OF ITS CONSISTENCY CONSIDERING DIFFERENT METHODS OF SELECTION].

Many plant mixtures play a key role in human health. Bioactive mixtures synthesis ability gives an unchangeable role to plants. Polyphenols included in some food products and xenogenic kinds of antioxidants play an important role to neutralize free radicals. From this point of you is very interesting a lot of subtropical cultures. By the help of this cultures methodic selection it's possible to regulate in them consistence of useful mixtures for medicine and for health. The facts given in this work represent the results of research on subtropical cultures (raw and ready materials). The ways are found to regulate bioactive mixtures consistence of researching plants. The plant is introduced as one of the strongest guaranty of preventing a lot of diseases and successful treatment as it's less characterized for medicines produced by raw materials to have side affect. It's natural that during methodological selection it's possible to achieve results with the knowledge of classical methods and producing level. To achieve the desirable result is depended on selectionist's qualification level.

A L-type lectin gene is involved in the response to hormonal treatment and water deficit in Volkamer lemon.

Combination of biotic and abiotic stress is a major challenge for crop and fruit production. Thus, identification of genes involved in cross-response to abiotic and biotic stress is of great importance for breeding superior genotypes. Lectins are glycan-binding proteins with a functions in the developmental processes as well as in the response to biotic and abiotic stress. In this work, a lectin like gene, namely ClLectin1, was characterized in Volkamer lemon and its expression was studied in plants exposed to either water stress, hormonal elicitors (JA, SA, ABA) or wounding to understand whether this gene may have a function in the response to multiple stress combination. Results showed that ClLectin1 has 100% homology with a L-type lectin gene from C. sinensis and the in silico study of the 5'UTR region showed the presence of cis-responsive elements to SA, DRE2 and ABA. ClLectin1 was rapidly induced by hormonal treatments and wounding, at local and systemic levels, suggesting an involvement in defence signalling pathways and a possible role as fast detection biomarker of biotic stress. On the other hand, the induction of ClLectin1 by water stress pointed out a role of the gene in the response to drought. The simultaneous response of ClLectin1 expression to water stress and SA treatment could be further investigated to assess whether a moderate drought stress may be useful to improve citrus performance by stimulating the SA-dependent response to biotic stress.

Comparative transcriptional survey between self-incompatibility and self-compatibility in Citrus reticulata Blanco.

Seedlessness is an excellent economical trait, and self-incompatibility (SI) is one of important factors resulting in seedless fruit in Citrus. However, SI molecular mechanism in Citrus is still unclear. In this study, RNA-Seq technology was used to identify differentially expressed genes related to SI reaction of 'Wuzishatangju' (Citrus reticulata Blanco). A total of 35.67GB raw RNA-Seq data was generated and was de novo assembled into 50,364 unigenes with an average length of 897bp and N50 value of 1549. Twenty-three candidate unigenes related to SI were analyzed using qPCR at different tissues and stages after self- and cross-pollination. Seven pollen S genes (Unigene0050323, Unigene0001060, Unigene0004230, Unigene0004222, Unigene0012037, Unigene0048889 and Unigene0004272), three pistil S genes (Unigene0019191, Unigene0040115, Unigene0036542) and three genes (Unigene0038751, Unigene0031435 and Unigene0029897) associated with the pathway of ubiquitin-mediated proteolysis were identified. Unigene0031435, Unigene0038751 and Unigene0029897 are probably involved in SI reaction of 'Wuzishatangju' based on expression analyses. The present study provides a new insight into the molecular mechanism of SI in Citrus at the transcriptional level.

An Additional Phytosanitary Cold Treatment Against Ceratitis capitata (Diptera: Tephritidae) in 'Oroblanco' Citrus Fruit.

For 'Oroblanco' ('Sweetie'), the sweet seedless pummelo-grapefruit hybrid, when exported from Israel to Japan, the standard cold treatment against Ceratitis capitata (Wied.) (Diptera: Tephritidae) is conducted at ≤ 1.5 °C, for 16 d. In recent years, the transportation means of exported citrus was changed from reefer vessels to individual refrigerated containers, where the fruit bulk is relatively small and may be exposed to temperature fluctuations and to the risk of chilling injuries. To reduce this risk, Israel proposed to Japan to increase the treatment temperature and extend its duration to 2.2 °C and 18 d, respectively. This study shows that the proposed treatment effectively kills the third instar larva of C. capitata, in Oroblanco.

Partial Cold Treatment of Citrus Fruit for Export Risk Mitigation for Thaumatotibia leucotreta (Lepidoptera: Tortricidae) as Part of a Systems Approach.

Some of South Africa's citrus export markets require mandatory postharvest cold treatment of citrus fruit as a phytosanitary risk mitigation treatment for Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae). An alternative to this may be partial cold treatment as one of the final steps in a systems approach to mitigate phytosanitary risk. Consequently, the efficacy of such partial cold treatments was evaluated. It was first determined that a 2°C cold treatment was significantly more effective against fourth and fifth instars (the most cold-tolerant instars) than treatments at 3°C and 4°C for a duration of 18 d. Secondly, it was determined that 2°C for 18 d and 1°C for 16 d were similarly effective, but both treatments were significantly more effective than 1°C for 14 d. Mean mortality of fourth and fifth instars treated with 2°C for 18 d in seven replicates from four trials was 99.94%. Finally, it was determined that the inability of the majority of surviving larvae to develop to adulthood would further increase the efficacy of a 2°C for 18 d treatment to 99.96%. Inclusion of reproductive nonviability of survivors increased mortality to 99.99%.

Alterations of physiology and gene expression due to long-term magnesium-deficiency differ between leaves and roots of Citrus reticulata.

Seedlings of Ponkan (Citrus reticulata) were irrigated with nutrient solution containing 0 (Mg-deficiency) or 1mM MgSO4 (control) every two day for 16 weeks. Thereafter, we examined magnesium (Mg)-deficiency-induced changes in leaf and root gas exchange, total soluble proteins and gene expression. Mg-deficiency lowered leaf CO2 assimilation, and increased leaf dark respiration. However, Mg-deficient roots had lower respiration. Total soluble protein level was not significantly altered by Mg-deficiency in roots, but was lower in Mg-deficient leaves than in controls. Using cDNA-AFLP, we obtained 70 and 71 differentially expressed genes from leaves and roots. These genes mainly functioned in signal transduction, stress response, carbohydrate and energy metabolism, cell transport, cell wall and cytoskeleton metabolism, nucleic acid, and protein metabolisms. Lipid metabolism (Ca(2+) signals)-related Mg-deficiency-responsive genes were isolated only from roots (leaves). Although little difference existed in the number of Mg-deficiency-responsive genes between them both, most of these genes only presented in Mg-deficient leaves or roots, and only four genes were shared by them both. Our data clearly demonstrated that Mg-deficiency-induced alterations of physiology and gene expression greatly differed between leaves and roots. In addition, we focused our discussion on the causes for photosynthetic decline in Mg-deficient leaves and the responses of roots to Mg-deficiency.

cDNA-AFLP analysis reveals the adaptive responses of citrus to long-term boron-toxicity.

BACKGROUND: Boron (B)-toxicity is an important disorder in agricultural regions across the world. Seedlings of 'Sour pummelo' (Citrus grandis) and 'Xuegan' (Citrus sinensis) were fertigated every other day until drip with 10 µM (control) or 400 µM (B-toxic) H3BO3 in a complete nutrient solution for 15 weeks. The aims of this study were to elucidate the adaptive mechanisms of citrus plants to B-toxicity and to identify B-tolerant genes. RESULTS: B-toxicity-induced changes in seedlings growth, leaf CO2 assimilation, pigments, total soluble protein, malondialdehyde (MDA) and phosphorus were less pronounced in C. sinensis than in C. grandis. B concentration was higher in B-toxic C. sinensis leaves than in B-toxic C. grandis ones. Here we successfully used cDNA-AFLP to isolate 67 up-regulated and 65 down-regulated transcript-derived fragments (TDFs) from B-toxic C. grandis leaves, whilst only 31 up-regulated and 37 down-regulated TDFs from B-toxic C. sinensis ones, demonstrating that gene expression is less affected in B-toxic C. sinensis leaves than in B-toxic C. grandis ones. These differentially expressed TDFs were related to signal transduction, carbohydrate and energy metabolism, nucleic acid metabolism, protein and amino acid metabolism, lipid metabolism, cell wall and cytoskeleton modification, stress responses and cell transport. The higher B-tolerance of C. sinensis might be related to the findings that B-toxic C. sinensis leaves had higher expression levels of genes involved in photosynthesis, which might contribute to the higher photosyntheis and light utilization and less excess light energy, and in reactive oxygen species (ROS) scavenging compared to B-toxic C. grandis leaves, thus preventing them from photo-oxidative damage. In addition, B-toxicity-induced alteration in the expression levels of genes encoding inorganic pyrophosphatase 1, AT4G01850 and methionine synthase differed between the two species, which might play a role in the B-tolerance of C. sinensis. CONCLUSIONS: C. sinensis leaves could tolerate higher level of B than C. grandis ones, thus improving the B-tolerance of C. sinensis plants. Our findings reveal some novel mechanisms on the tolerance of plants to B-toxicity at the gene expression level.

Efficacy of indigenous predatory mites (Acari: Phytoseiidae) against the citrus rust mite Phyllocoptruta oleivora (Acari: Eriophyidae): augmentation and conservation biological control in Israeli citrus orchards.

The citrus rust mite (CRM), Phyllocoptruta oleivora (Acari: Eriophyidae) is a cosmopolitan key pest of citrus, inflicting severe economic damage if not controlled. In Israel, CRM damages all citrus cultivars. International regulation and increasing control failures of CRM led growers to seek sustainable biological control solutions such as acarine biological control agents. Laboratory studies conducted in Israel have indicated that the indigenous predator species Amblyseius swirskii, Iphiseius degenerans, Typhlodromus athiasae and Euseius scutalis (all Acari: Phytoseiidae) can potentially control CRM. Our general objective in the present study was to bridge the gap of knowledge between laboratory studies and the lack of control efficacy of these species in commercial orchards. Predator augmentation in the field showed that although predator populations increased immediately following releases they later decreased and did not affect CRM populations. When A. swirskii augmentation was combined with a series of maize pollen applications, A. swirskii populations were enhanced substantially and continuously but again CRM populations were not affected. Growth chamber studies with CRM-infested seedlings, with or without a maize pollen supplement, indicated that pollen provisioning led to population increase of E. scutalis and A. swirskii but only E. scutalis significantly lowered CRM populations. Control with E. scutalis was confirmed in the field on CRM infested seedlings with pollen provisioned by adjacent flowering Rhodes grass. While experiments in mature citrus orchard showed that pollen supplement usually increased predator populations they also indicated that other factors such as intraguild interactions and pesticide treatments should be taken into account when devising CRM biological control programs.

Leaf cDNA-AFLP analysis of two citrus species differing in manganese tolerance in response to long-term manganese-toxicity.

BACKGROUND: Very little is known about manganese (Mn)-toxicity-responsive genes in citrus plants. Seedlings of 'Xuegan' (Citrus sinensis) and 'Sour pummelo' (Citrus grandis) were irrigated for 17 weeks with nutrient solution containing 2 µM (control) or 600 µM (Mn-toxicity) MnSO4. The objectives of this study were to understand the mechanisms of citrus Mn-tolerance and to identify differentially expressed genes, which might be involved in Mn-tolerance. RESULTS: Under Mn-toxicity, the majority of Mn in seedlings was retained in the roots; C. sinensis seedlings accumulated more Mn in roots and less Mn in shoots (leaves) than C. grandis ones and Mn concentration was lower in Mn-toxicity C. sinensis leaves compared to Mn-toxicity C. grandis ones. Mn-toxicity affected C. grandis seedling growth, leaf CO2 assimilation, total soluble concentration, phosphorus (P) and magenisum (Mg) more than C. sinensis. Using cDNA-AFLP, we isolated 42 up-regulated and 80 down-regulated genes in Mn-toxicity C. grandis leaves. They were grouped into the following functional categories: biological regulation and signal transduction, carbohydrate and energy metabolism, nucleic acid metabolism, protein metabolism, lipid metabolism, cell wall metabolism, stress responses and cell transport. However, only 7 up-regulated and 8 down-regulated genes were identified in Mn-toxicity C. sinensis ones. The responses of C. grandis leaves to Mn-toxicity might include following several aspects: (1) accelerating leaf senescence; (2) activating the metabolic pathway related to ATPase synthesis and reducing power production; (3) decreasing cell transport; (4) inhibiting protein and nucleic acid metabolisms; (5) impairing the formation of cell wall; and (6) triggering multiple signal transduction pathways. We also identified many new Mn-toxicity-responsive genes involved in biological and signal transduction, carbohydrate and protein metabolisms, stress responses and cell transport. CONCLUSIONS: Our results demonstrated that C. sinensis was more tolerant to Mn-toxicity than C. grandis, and that Mn-toxicity affected gene expression far less in C. sinensis leaves. This might be associated with more Mn accumulation in roots and less Mn accumulation in leaves of Mn-toxicity C. sinensis seedlings than those of C. grandis seedlings. Our findings increase our understanding of the molecular mechanisms involved in the responses of plants to Mn-toxicity.

Identifying differentially expressed genes in pollen from self-incompatible "Wuzishatangju" and self-compatible "Shatangju" mandarins.

Self-incompatibility (SI) is one of the important factors that can result in seedless fruit in Citrus. However, the molecular mechanism of SI in Citrus is not yet clear. In this study, two suppression subtractive hybridization (SSH) libraries (forward, F and reverse, R) were constructed to isolate differentially expressed genes in pollen from "Wuzishatangju" (SI) and "Shatangju" (self-compatibility, SC) mandarins. Four hundred and sixty-eight differentially expressed cDNA clones from 2077 positive clones were sequenced and identified. Differentially expressed ESTs are possibly involved in the SI reaction of "Wuzishatangju" by regulating pollen development, kinase activity, ubiquitin pathway, pollen-pistil interaction, and calcium ion binding. Twenty five SI candidate genes were obtained, six of which displayed specific expression patterns in various organs and stages after self- and cross-pollination. The expression level of the F-box gene (H304) and S1 (F78) in the pollen of "Wuzishatangju" was 5-fold higher than that in "Shatangju" pollen. The F-box gene, S1, UBE2, UBE3, RNaseHII, and PCP were obviously up-regulated in pistils at 3 d after self-pollination of "Wuzishatangju", approximately 3-, 2-, 10-, 5-, 5-, and 2-fold higher, respectively than that at the same stage after cross-pollination of "Wuzishatangju" × "Shatangju" pistils. The potential involvement of these genes in the pollen SI reaction of "Wuzishatangju" is discussed.

Bicarbonate blocks iron translocation from cotyledons inducing iron stress responses in Citrus roots.

The effect of bicarbonate ion (HCO3(-)) on the mobilization of iron (Fe) reserves from cotyledons to roots during early growth of citrus seedlings and its influence on the components of the iron acquisition system were studied. Monoembryonic seeds of Citrus limon (L.) were germinated "in vitro" on two iron-deprived media, supplemented or not with 10mM HCO3(-) (-Fe+Bic and -Fe, respectively). After 21d of culture, Fe concentration in seedling organs was measured, as well as gene expression and enzymatic activities. Finally, the effect of Fe resupply on the above responses was tested in the presence and absence of HCO3(-) (+Fe+Bic or +Fe, respectively). -Fe+Bic seedlings exhibited lower Fe concentration in shoots and roots than -Fe ones but higher in cotyledons, associated to a significative inhibition of NRAMP3 expression. HCO3(-) upregulated Strategy I related genes (FRO1, FRO2, HA1 and IRT1) and FC-R and H(+)-ATPase activities in roots of Fe-starved seedlings. PEPC1 expression and PEPCase activity were also increased. When -Fe+Bic pre-treated seedlings were transferred to Fe-containing media for 15d, Fe content in shoots and roots increased, although to a lower extent in the +Fe+Bic medium. Consequently, the above-described root responses became markedly repressed, however, this effect was less pronounced in +Fe+Bic seedlings. In conclusion, it appears that HCO3(-) prevents Fe translocation from cotyledons to shoot and root, therefore reducing their Fe levels. This triggers Fe-stress responses in the root, enhancing the expression of genes related with Fe uptake and the corresponding enzymatic activities.

Molecular characterization and expression analysis of S1 self-incompatibility locus-linked pollen 3.15 gene in Citrus reticulata.

Gametophytic self-incompatibility (GSI) is controlled by a highly polymorphic locus called the S-locus, which is an important factor that can result in seedless fruit in Citrus. The S1 self-incompatibility locus-linked pollen 3.15 gene (S1-3.15 ) belongs to a type of S locus gene. The role of S1-3.15 in the SI reaction of Citrus has not yet been reported. In this study, full-length sequences of cDNA and DNA encoding the S1-3.15 gene, referred to as CrS1-3.15 , were isolated from 'Wuzishatangju' (Self-incompatibility, SI) and 'Shatangju' (Self-compatibility, SC). The predicted amino acid sequences of CrS1-3.15 between 'Wuzishatangju' and 'Shatangju' differ by only three amino acids. Compared to 'Wuzishatangju', three bases were substituted in the genomic DNA of CrS1-3.15 from 'Shatangju'. Southern blot results showed that one copy of CrS1-3.15 existed in the genomic DNA of both 'Wuzishatangju' and 'Shatangju'. The expression level of the CrS1-3.15 gene in the ovaries of 'Shatangju' was approximately 60-fold higher than that in the ovaries of 'Wuzishatangju'. When 'Wuzishatangju' was cross-pollinated, the expression of CrS1-3.15 was upregulated in the ovaries at 3 d, and the highest expression levels were detected in the ovaries at 6 d after cross-pollination of 'Wuzishatangju' × 'Shatangju'. To obtain the CrS1-3.15 protein, the full-length cDNA of CrS1-3.15 genes from 'Wuzishatangju' and 'Shatangju' was successfully expressed in Pichia pastoris. Pollen germination frequency of 'Wuzishatangju' was inhibited significantly with increasing CrS1-3.15 protein concentrations from SI 'Wuzishatangju'.

Mechanism of seedlessness in a new lemon cultivar 'Xiangshui' [Citrus limon (L.) Burm. F].

Seedlessness is an important economic trait of lemon. Understanding the cellular and molecular mechanisms of seedlessness in 'Xiangshui' lemon requires detailed data on pollen and embryo sac fertility, embryo development and compatibility mechanisms governing self- and cross-pollination. The results of the current study indicate that the fertility of pollen and mature embryo sac remains normal. When flowers were self- or cross-pollinated, pollen grains of 'Xiangshui' were able to germinate on the stigma. In the case of self-pollination, pollen tubes became twisted, tube tips enlarged and tubes ruptured in the bottom of stigma. Following cross-pollination, tubes were able to grow normally in the style and ovary and enter the embryo sac, where double fertilization took place. Embryonic development resulting from cross-pollination was normal. After cross-pollination, the zygote began to divide at 2 weeks post-pollination, with early globular embryos observed after 3 weeks, globular and heart-shaped embryos at 4 weeks, torpedo-shaped embryos at 5 weeks, cotyledonary embryos at 6 weeks and thereafter germinable seeds. After self-pollination, however, ovules began to abort at 2 weeks post-pollination, with ovules disappearing at 5 weeks, ultimately producing seedless fruits. Emasculated unpollinated flowers also developed into seedless fruits, indicating that seedlessness contributes to parthenocarpy. However, gametophytic self-incompatibility has a major role in seedlessness in 'Xiangshui' lemon by blocking fertilization at the bottom of the stigma.

Protective effects of an extract from Citrus bergamia against inflammatory injury in interferon-γ and histamine exposed human keratinocytes.

AIMS: The present work evaluated the anti-inflammatory/antioxidant activity of a well characterized extract from Citrus bergamia Risso and Poiteau (CBE), containing neoeriocitrin, naringin, neohesperidin and other flavonoids, on human NCTC 2544 keratinocytes treated with interferon-gamma (IFN-γ) and histamine (H). MAIN METHODS: High performance liquid chromatography (HPLC) coupled with diode array detectors was used to characterize and quantify phenolic compounds in CBE. Anti-inflammatory/antioxidant ability on keratinocytes was determined through evaluation of inter-cellular adhesion molecule-1 (ICAM-1) and inducible nitric oxide synthase (iNOS) expression by Western blot, production of nitric oxide (NO) with Griess reagent and concentration of reactive oxygen species (ROS) by fluorescent quantitative analysis with 2',7'-dichlorfluorescein-diacetate (DCFH-DA). Cell viability was assessed using 3-(4,5-dimethyl-2 thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Antioxidant activity was also measured by oxygen radical absorbance capacity (ORAC) assay. Glycosaminoglycans (GAGs) were quantified using 1.9-dimethyl methylene blue (DMB). KEY FINDINGS: CBE exhibited high antioxidant activity confirmed by elevated ORAC values related to high capacity in oxygen radical scavenging. The assays on keratinocytes demonstrated that CBE does not inhibit cell proliferation and is shown to significantly reduce dose-dependently ICAM-1, iNOS, NO, ROS and GAG production in cells exposed to IFN-γ and H. SIGNIFICANCE: Our study demonstrates that the pools of compounds of an extract from C. bergamia efficiently block the proinflammatory actions induced by IFN-γ and H on human keratinocytes. CBE may be used for topic employment in some inflammatory diseases of the skin and to represent an important opportunity for the essential oil processing industries.

Extensive citrus triploid hybrid production by 2x×4x sexual hybridizations and parent-effect on the length of the juvenile phase.

UNLABELLED: The citrus fresh market demands the production of seedless citrus fruits, as seedy fruits are not accepted by consumers. The recovery of triploid plants has proven to be the most promising approach to achieve this goal, since triploids have very low fertility, are generally seedless and do not induce seeds in other cultivars by cross pollination. Triploid plants can be recovered by 2x×4x sexual hybridization. In this work, we present an effective methodology to recover triploid plants from 2x×4x hybridizations based on in vitro embryo rescue, ploidy level analysis by flow cytometry and genetic origin of triploid plants. The pollen viability of diploid and tetraploid citrus genotypes was analyzed by comparing the pollen germination rate in vitro. The pollen viability of tetraploid (doubled-diploid) genotypes is generally reduced but sufficient for successful pollination. Triploid embryos were identified in normal and undeveloped seeds that did not germinate under greenhouse conditions. The influence of parents and environmental conditions on obtaining triploid plants was analyzed and a strong interaction was noted between the parents and environmental conditions. The parental effect on the length of the juvenile phase was also demonstrated through observations of a large number of progeny over the last 15 years. The juvenile phase length of the triploid hybrids obtained with 'Fortune' mandarin as female parent and tetraploid 'Orlando' tangelo as male parent was shorter than the juvenile phase obtained with a clementine as female parent and tetraploids of 'Nova', 'W. Leaf' and 'Pineapple' male parents. KEY MESSAGE: Effective methodology to recover citrus triploid plants from 2x×4x sexual hybridizations and the parental effect on the length of the juvenile phase.