Activation of autophagy by Citri Reticulatae Semen extract ameliorates amyloid-beta-induced cell death and cognition deficits in Alzheimer’s disease

Amyloid-beta-induced neuronal cell death contributes to cognitive decline in Alzheimer’s disease.Citri Reticulatae Semen has diverse beneficial effects on neurodegenerative diseases,including Parkinson’s and Huntington’s diseases,however,the effect of Citri Reticulatae Semen on Alzheimer’s disease remains unelucidated.In the current study,the anti-apoptotic and autophagic roles of Citri Reticulatae Semen extract on amyloid-beta-induced apoptosis in PC12 cells were first investigated.Citri Reticulatae Semen extract protected PC12 cells from amyloid-beta-induced apoptosis by attenuating the Bax/Bcl-2 ratio via activation of autophagy.In addition,Citri Reticulatae Semen extract was confirmed to bind amyloid-beta as revealed by biolayer interferometry in vitro,and suppress amyloid-beta-induced pathology such as paralysis,in a transgenic Caenorhabditis elegans in vivo model.Moreover,genetically defective Caenorhabditis elegans further confirmed that the neuroprotective effect of Citri Reticulatae Semen extract was autophagy-dependent.Most importantly,Citri Reticulatae Semen extract was confirmed to improve cognitive impairment,neuronal injury and amyloid-beta burden in 3×Tg Alzheimer’s disease mice.As revealed by both in vitro and in vivo models,these results suggest that Citri Reticulatae Semen extract is a potential natural therapeutic agent for Alzheimer’s disease via its neuroprotective autophagic effects.

Resistance Monitoring of Field Populations of Panonychus citri to Three Pesticides

[Objectives]The paper was to investigate the development of Panonychus citri resistance to commonly used pesticides in major citrus producing areas in China,and to screen out new green and efficient agents for its control and prevention.[Methods]The resistance changes of field populations of P.citri to abamectin,pyridaben,and bifenazate in 12 locations across five provinces in China were compared using the leaf disc impregnation method.[Results]P.citri in the tested areas exhibited the most severe resistance to abamectin,with approximately 91.7%of the field populations showing high levels of resistance to abamectin(112.1-560.5 times);50%of P.citri populations exhibited high levels of resistance to pyridaben(123.0-202.7 times),while the remaining populations showed intermediate levels of resistance(25.6-80.3 times);except for the Zhejiang Xiangshan 2019 and Jiangxi Yudu 2019 populations,which exhibited a medium level of resistance to bifenazate,the remaining 10 monitored populations demonstrated a high level of resistance to bifenazate(140.4-686.1 times).[Conclusions]It is advisable to discontinue the use of abamectin and bifenazate due to significant resistance observed in populations of P.citri monitored in 12 locations across 5 provinces.It is recommended to reduce the frequency of pyridaben use and alternate with other acaricides that have different mechanisms of action due to the varying degrees of resistance developed.

A Network Pharmacology Study on Active Components and Targets of Citri Reticulatae Pericarpium for Treating Keloids

[Objectives]To investigate the mechanisms and pharmacologic effects of Citri Reticulatae Pericarpium against keloids by network pharmacology systematically.[Methods]TCMSP,Uniprot and BATMAN-TCM databases were used to obtain the active constituents and targets of Citri Reticulatae Pericarpium."Keloid"was used as key word to search for related therapeutic targets from Drug Bank,OMIM,TTD,and GEO databases.The Chinese medicine compound-target network was constructed by Cytoscape software.Besides,gene ontology(GO)and Kyoto Encyclopedia of genes and genome enrichment analysis were also performed.Afterward,Discovery Studio software was used to assess the interaction of key components and genes.[Results]Five active components of Citri Reticulatae Pericarpium,773 compound targets and 676 keloid treatment targets were obtained in the databases.After the intersection,there are 47 targets of Citri Reticulatae Pericarpium for treating keloids.Hub genes were identified such as MMP9,IL6,TNF,TP53,and VEGFA,which were enriched in tumor necrosis factor-α,nuclear factor kappa-B,and other signaling pathways.The molecular docking stimulation confirmed the interaction between the MMP9 and three components of Citri Reticulatae Pericarpium.[Conclusions]Citri Reticulatae Pericarpium may play an important role in treating keloids through modulating genes and signaling pathways.The present study sheds light on the mechanisms of active compounds of Citri Reticulatae Pericarpium for the treatment of keloids.

柑桔木虱(Diaphorina citri)抗药性及抗药性机理研究进展

柑桔木虱主要为害柑桔等芸香科植物,是传播柑桔黄龙病的主要自然媒介,严格防治柑桔木虱是当前防控柑桔黄龙病的一项关键性措施。目前防治柑桔木虱最重要的方式是化学防治,而大量化学农药的使用以及使用的不恰当,柑桔木虱对一些杀虫剂产生了不同程度的抗药性。研究柑桔木虱抗药性的发生情况以及抗药性机理对于抗药性治理具有重要意义。为全面了解和总结柑桔木虱抗药性相关认识成果,进而为相关研究的开展及柑桔木虱的科学防治提供参考,综述了近年特别是2018年以来国内外柑桔木虱抗药性测定方法、抗药性现状、交互抗性、抗药性机理以及抗性治理等方面的研究进展。

Transcriptomic analysis reveals the defense mechanisms of citrus infested with Diaphorina citri

Huanglongbing(HLB) is a devastating disease that has led to an acute crisis for growers of citrus, one of the world's most important fruit crops. The phloem-feeding Asian citrus psyllid(ACP), Diaphorina citri, is the main pest at the new shoot stage and is the only natural vector of HLB pathogenic bacteria. Little is known about how plants perceive and defend themselves from this destructive pest. Here, we characterized changes in the expression of various genes in citrus plants that were continuously infested by D. citri for different durations(12, 24, and 48 h). A total of 5 219 differentially expressed genes(DEGs) and 643 common DEGs were identified across all time points. Several pathways related to defense were activated, such as peroxisome, alpha-linolenic acid metabolism, and phenylpropanoid and terpenoid biosynthesis, and some pathways related to growth and signal transduction were suppressed in response to D. citri infestation. The expression of genes including kinases(CML44, CIPK6, and XTH6), phytohormones(SAMT, LOX6, and NPR3), transcription factors(bHLH162, WRKY70, and WRKY40), and secondary metabolite synthesis-related genes(PAL, 4CL2, UGT74B1 and CYP82G1) was significantly altered in response to D. citri infestation. The findings of this study greatly enhance our understanding of the mechanisms underlying the defense response of citrus plants to D. citri infestation at the molecular level. Functional characterization of the candidate defense-related genes identified in this study will aid the molecular breeding of insect-resistant citrus varieties.

Occurrence of Diaphorina citri in Guangxi and Integrated Control Technique

In order to promote the healthy and sustainable development of citrus industry in Guangxi and reduce the harm of Diaphorina citri and citrus Huanglongbing on citrus industry,the paper summarizes the occurrence of D.citri in Guangxi,and put forward comprehensive control technique,so as to provide the reference for guiding prevention and control of major pests in citrus industry.

Overview of Occurrence,Prevention and Control of Panonychus citri in China

In order to further promote the prevention and control research of Panonychus citri and improve its control effect,this paper summarizes the main influencing factors of the outbreak,main control technology and drug resistance of P.citri,and puts forward the research focus and control strategy.

The small and large subunits of carbamoyl-phosphate synthase exhibit diverse contributions to pathogenicity in Xanthomonas citri subsp.citri

Carbamoyl-phosphate synthase plays a vital role in the carbon and nitrogen metabolism cycles. In Xanthomonas citrisubsp. citri, carA and carB encode the small and large subunits of carbamoyl-phosphate synthase, respectively. The deletion mutation of the coding regions revealed that carA did not affect any of the phenotypes, while carB played multiple roles in pathogenicity. The deletion of carB rendered the loss of pathogenicity in host plants and the ability to induce a hyper- sensitive reaction in the non-hosts. Quantitative reverse transcription-PCR assays indicated that 11 hrp genes coding the type Ill secretion system were suppressed when interacting with citrus plants. The mutation in carB also affected bacterial utilization of several carbon and nitrogen resources in minimal medium MMX and extracellular enzyme activities. These data demonstrated that only the large subunit of carbamoyl-phosphate synthase was essential for canker development by X. citri subsp, citri.

The Viable but Non-culturable State in Xanthomonas citri subsp, citri is a Reversible State Induced by Low Nutrient Availability and Copper Stress Conditions

Xcc （Xanthomonas citri subsp, citri） causes citrus bacterial canker, a leaf, stem and fruit spotting disease that affects most commercial citrus species and cultivars. Copper compounds, widely used for management of this pathogen, have been reported as inducers of a VBNC （viable but non-culturable state） in plant pathogenic bacteria. VBNC may be considered as a state preceding bacterial death or as a survival mechanism under adverse conditions. Several experiments were performed to characterize the reversibility and persistence of the VBNC state in Xcc. VBNC was induced in low nutrient medium or with amendment of copper at concentrations used for field disease control. The VBNC condition was demonstrated to persist up to 150 days after copper treatment and was reversed after the addition of culture media without copper or amendment with citrus leaf extract. Xcc viability was evaluated by recovery of colonies on culture media, confirmed by membrane integrity, respiratory activity and by real-time RT-PCR targeting a sequence from the gumD gene. Besides, the colonies recovered were pathogenic on citrus leaves. These results confirm that the VBNC state in Xcc is inducible and reversible and therefore may occur in the phyllosphere when Xcc is under copper stress or starvation.

^(60)Coγ射线对柑桔大实蝇Dacus citri雄性生殖细胞的影响

本文研究了^(60)Coγ射线对柑桔大实蝇(Dacus citri)精子发生的影响。用9krad辐照羽化前2天的蛹,成虫羽化后解剖观察,雄虫已有大量成熟精子,精子形态、数量和活力与未经辐照处理的雄虫无显著差异,但其超微结构则发生了某些异常变化,如营养细胞出现空泡、精子排列零乱、线粒体衍生体和轴丝增多或减少、构型被破坏、精子细胞分裂异常等。

^(60)Coγ射线对柑桔大实蝇Dacus citri雌虫生殖细胞的影响

本文研究了柑桔大实蝇的卵子发生及(60)~Coγ射线对卵子发生的影响。作者将大实蝇的卵子发生划分为8个阶段。经观察,蛹期的卵子发生处于原囊时期,此期对辐射较敏感。用6、9、12、15krad辐照处理羽化前2天的蛹,都能导致雌虫不育,使大部份卵母细胞的发育停于第4—5阶段,不能发育为正常的卵。

Establishment of HPLC Fingerprint, Cluster Analysis and Principle Component Analysis of Citri Reticulatae Pericarpium Viride

[Objectives] This study aimed to establish HPLC fingerprint and conduct cluster analysis and principle component analysis for Citri Reticulatae Pericarpium Viride. [Methods] Using the HPLC method, the determination was performed on XSelect~® HSS T3-C_(18) column with mobile phase of acetonitrile-0.5% acetic acid solution(gradient elution) at the flow rate of 1.0 mL/min. The detection wavelength was 360 nm. The column temperature was 25℃. The sample size was 10 μL. With peak of hesperidin as the reference, HPLC fingerprints of 10 batches of Citri Reticulatae Pericarpium Viride were determined. The similarity of the 10 batches of samples was evaluated by Similarity Evaluation System for Chromatographic Fingerprint of TCM(2012 edition) to determine the common peaks. Cluster analysis and principal component analysis were performed by using SPSS 17.0 statistical software. [Results] The HPLC fingerprints of the 10 batches of medicinal materials had total 11 common peaks, and the similarity was 0.919-1.000, indicating that the chemical composition of the 10 batches of medicinal materials was consistent. There were 11 common components in the 10 batches of medicinal materials, but their contents were different. When the Euclidean distance was 20, the 10 batches of samples were divided into two categories, S4 in the first category, and the others in the second one. When the Euclidean distance was 5, the second category could be further divided into two sub-categories, S1 and S10 in one sub-category, and S2, S3, S5, S6, S7, S8 and S9 in the other one. The principle component analysis showed that cumulative contribution rate of the two main component factors was 92.797%, and the comprehensive score of S7 was the highest with the best quality. [Conclusions] The results of HPLC fingerprinting, cluster analysis and principle component analysis can provide reference for the quality control of Citri Reticulatae Pericarpium Viride.

芒果柑桔花蕾蛆（Contarinia citri Barnes）的生物学及其防治研究

柑桔花蕾蛆是云南芒果的主要害虫之一。其幼虫对芒果花蕊的危害率为６０％～７０％，最高达１００％，严重影响芒果开花、座果。该虫在云南景谷湿热气候区，１年发生５～６代，完成１代需１６～３２日，世代重叠。采取综合防治措施，危害率可控制在０．２９％左右。

A survey of FLS2 genes from multiple citrus species identifies candidates for enhancing disease resistance to Xanthomonas citri ssp. citri.

Pathogen-associated molecular patterns(PAMPs)-triggered immunity(PTI)is an important component of plant innate immunity.In a previous study,we showed that the PAMP flg22 from Xanthomonas citri ssp.citri(Xflg22),the causal agent of citrus canker,induced PTI in citrus,which correlated with the observed levels of canker resistance.Here,we identified and sequenced two bacterial flagellin/flg22 receptors(FLS2-1 and FLS2-2)from‘Duncan’grapefruit(Citrus paradisi,CpFLS2-1 and CpFLS2-2)and‘Sun Chu Sha’mandarin(C.reticulata,CrFLS2-1 and CrFLS2-2).We were able to isolate only one FLS2 from‘Nagami’kumquat(Fortunella margarita,FmFLS2-1)and gene flanking sequences suggest a rearrangement event that resulted in the deletion of FLS2-2 from the genome.Phylogenetic analysis,gene structure and presence of critical amino acid domains all indicate we identified the true FLS2 genes in citrus.FLS2-2 was more transcriptionally responsive to Xflg22 than FLS2-1,with induced expression levels higher in canker-resistant citrus than in susceptible ones.Interestingly,‘Nagami’kumquat showed the highest FLS2-1 steady-state expression levels,although it was not induced by Xflg22.We selected FmFLS2-1,CrFLS2-2 and CpFLS2-2 to further evaluate their capacity to enhance bacterial resistance using Agrobacterium-mediated transient expression assays.Both FmFLS2-1 and CrFLS2-2,the two proteins from canker-resistant species,conferred stronger Xflg22 responses and reduced canker symptoms in leaves of the susceptible grapefruit genotype.These two citrus genes will be useful resources to enhance PTI and achieve resistance against canker and possibly other bacterial pathogens in susceptible citrus types.

Acaricidal Activity of Boenninghausenia sessilicarpa Against Panonychus citri

Acaricidal activity of Boenninghausenia sessilicarpa against Panonychus cirri was tested in the laboratory. Four solvents were used to prepare crude extracts, petroleum ether, chloroform, ethyl acetate, and ethanol, among which ethanol was the most effective one. The results suggested that ethanol extracts of B. sessilicarpa had eminent acaricidal and ovicidal activities. Concentrated extracts were prepared using petroleum ether, chloroform, ethyl acetate, or distilled water as solvent. Mite mortality rates in the concentrated extracts by petroleum ether, chloroform, or distilled water were significantly lower than those by ethyl acetate. The LC50 values of eggs and female mites were 0.7639 and 1.1033 mg mL^-1, respectively. After liquid chromatography and thin layer chromatography, the concentrated extracts were separated into 14 groups of fractions and further tests for their acaricidal and ovicidal activities were conducted. Fraction 2 was found to possess higher acaricidal and ovicidal activities. The mortality of eggs and adult mites were 85.83 and 63.07%, respectively. Moreover, fraction 2 showed moderate oviposition inhibition effect （0.8795） against P. citri when the used dose was higher than 2.5 mg mL^-1

Comparison of PCR, DIA and Pathogenicity Assay for Detection of Xanthomonas axonopodis pv.citri,the Causal Agent of Citrus Bacterial Canker Disease

Polymerase chain reaction (PCR) approach based on newly designed primers, JYF5/JYR5, wasapplied for specific detection of Xanthomonas axonopodis pv.citri(Xac). The efficiencyand reliability of PCR method were compared with dot immunobinding assay (DIA) andclassical pathogenicity test techniques for detecting suspensions of pure cells of Xacand soaking sap of citrus tissues. Detection sensitivity of PCR was about 4.5 cells or1.56 pg target DNA per reaction which was higher than that of DIA (ca. 450 cells per dot).These three techniques (PCR assay, DIA and Pathogenecity test) could always detect Xacfrom symptomatic citrus samples. Different performances were obtained from citrusmaterials without symptoms, and the positive detection frequency was PCR, DIA andpathogenicity test.

Evaluation of bioactive flavonoids in Citri Reticulatae Pericarpium from different regions and its association with antioxidant andα-glucosidase inhibitory activities

Background:To explore the differences of 14 batches of Citri Reticulatae Pericarpium from different regions.Methods:The main aim of this study was to develop a high-performance liquid chromatography method-photodiode array detection for determining the contents of five flavonoids and the chromatographic fingerprints of Citri Reticulatae Pericarpium from different regions.Theα-glucosidase inhibitory activities and antioxidant properties of Citri Reticulatae Pericarpium,based on free-radical scavenging assays against 1,1-diphenyl-2-picrylhydrazyl,2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid),and hydroxyl radicals,were estimated and compared.Results:Among the tested compounds,the content of hesperidin(13.386-68.235 mg/g)was the highest and that of hesperitin(0.045-0.277 mg/g)was the lowest.In comparing different sources of Citri Reticulatae Pericarpium,the contents of narirutin in Citri Reticulatae Pericarpium from Guangdong(0.824-0.851 mg/g)and Sichuan(1.069-1.204 mg/g)provinces of China were lower than in other provinces.In contrast,nobiletin(8.429-12.237 mg/g)and tangeretin(3.947-4.613 mg/g)were most abundant in Guangdong sources of Citri Reticulatae Pericarpium,followed by samples from Sichuan Province(nobiletin:6.761-7.658 mg/g;tangeretin:3.422-3.933 mg/g).Correlation analysis showed that nobiletin and tangeretin were the main contributors to the antioxidant capacity,and narirutin was the main active component inhibiting theα-glucosidase activity of Citri Reticulatae Pericarpium.Conclusion:This work revealed that the intrinsic quality of Citri Reticulatae Pericarpium was affected by different growing regions,which provides a scientific basis for controlling the quality of Citri Reticulatae Pericarpium and rationally developing and utilizing Citri Reticulatae Pericarpium.

DR5 is a Suitable System for Studying the Auxin Response in the Poncirus trifoliata-Xanthomonas axonopodis pv.citri Interaction

The local auxin distribution characteristics in the roots,stems,and leaves of stably transformed plantlets of trifoliate orange(Poncirus trifoliata)with auxin reporter system DR5::GUS-YFP were elucidated in this research.The auxin response maxima could be observed in the apex of the root tip,primary phloem of the tender stem,and the margin of the young leaves according to the activity of theβ-glucuronidase(GUS)reporter gene triggered by the auxin responsive DR5 promoter.Auxin responses in the apex of the root tips increased when treated with synthetic auxin 1-naphthylacetic acid(NAA),but decreased when treated with the auxin polar transportation inhibitor 2,3,5-triiodobenzoic acid(TIBA).These results indicated that the DR5 reporter system worked in P.trifoliata for auxin distribution and response observation.Trifoliate orange is highly susceptible to citrus canker disease.Auxin accumulation was observed visually in the invasion sites of the detached leaves inoculated with Xanthomonas axonopodis pv.citri(Xac)by GUS staining;the upregulated expression of the YFP,GH3.1,GH3.9,and SAUR genes assessed by quantitative real-time PCR(qRT-PCR)also identified auxin accumulation in the inoculated tissues following Xac infection.Overall,these findings indicated that the plantlets of P.trifoliata engineered with the auxin reporter gene provided a promising system for studying auxin responses during Xac infection.

构建Citrix XenApp平台实现移动办公

移动办公作为信息时代的办公方式之一,越来越被广泛使用,特别是在企业管理中提高办公效率方面受到前所未有的重视。本文从背景、技术和内容等方面阐述一个在Windows2003环境下,以Citrix XenApp为技术核心,以AD用户控制为基础办公平台的设计和实现,并利用该平台方便地为终端用户发布应用程序及权限分配,不仅可以解决移动办公的问题,也为企业的部门在软件部署、运维和管理上节约了大量的成本和时间,其应用前景相当广阔。

Characterization of Xanthomonas citri pv.citri from China based on spoligotyping

Xanthomonas citri pv.citri(Xcc),a gram-negative bacterium,is the causal agent of citrus canker,one of the most devastating diseases threatening the citrus industry worldwide.Understanding the diversity and population structure of Xcc is a prerequisite for disease epidemiological monitoring and effective disease management.Recent characterization of the clustered regularly interspaced short palindromic repeats(CRISPR)/cas(CRISPR-associated proteins genes)system with a highly variable repeat number among species provides a new molecular typing method for bacterial genetic analysis.In this study,we performed systematic in silico analyses of 28 Xcc genomes and identified a credible CRISPR/cas in Xcc strains.Further analysis of CRISPR polymorphisms(repeat number and spacer types)in 129 Xcc A strains collected from six provinces in China identified 15 types of CRISPR arrays with 25 spacers.Phylogenetic analysis of Xcc strains based on the CRISPR locus produced a more reliable and accurate typing result compared to the commonly used loci.In addition,seven associated cas genes—cas1,cas2,cas3,cas4,cas5,cas7(csd2),and cas8(csd1)—were found located adjacent to the CRISPR array.BLAST results showed>99%similarity of seven cas genes among Xcc strains.Homology analysis of spacer sequences showed that six spacers had possible phage/prophage origin.The characterization of the CRISPR/cas system among Xcc strains provided an updated strain typing method for Xcc diversity analysis and yielded a panoramic view of CRISPR evolution for further studies of Xcc-phage interactions.