Threshold Dynamics of a Vector-Borne Epidemic Model for Huanglongbing with Impulsive Control

In this paper, the basic reproduction number is calculated for Huanglongbing (HLB) model with impulses which is a vector-borne epidemic model with impulses. For controlling HLB, farmers’ experience is replanting of healthy plants and removing infected plants. To reflect the real world, we construct an impulsive control model which considers replanting of healthy plants and removing infected plants at one fixed time. By analyzing the model, we conclude that the disease-free equilibrium is globally asymptotically stable if the basic reproduction number Ro 1, and we prove that the HLB is permanence if the basic reproduction number Ro > 1.

Digital Gene Expression Analysis of Huanglongbing Affected Mandarins(Citrus reticulata Blanco)In Response to Thermotherapy

Huanglongbing(HLB),which is mainly associated with“Candidatus Liberibacter asiaticus”(CLas),is currently threatening citrus production around the world.This destructive century-old disease results in callose deposition in phloem plasmodesmata and starch overaccumulation in leaf chloroplasts.Heat treatment with high humidity has proven to be effective in eliminating the CLas titers in potted citrus trees.This study explored the curative mechanisms and impact of heat treatment(from 32 to 48°C with 75%±5%relative humidity,14h in light and 10h in dark for four days)on mandarin trees(Citrus reticulata Blanco)that were severely infected by CLas using Solexa/Illumina’s digital gene expression(DGE)profiling.A total of 838 differentially expressed genes(DEGs)were identified,of which 510 were downregulated.Proteinprotein interaction analysis of the DEGs indicated that the thermotherapy of HLB trees downregulated the circadian clock related genes in chloroplasts and upregulated the RNA-editing process in the mitochondria,characterized by 141 genes encoding pentatricopeptide repeatcontaining proteins(PPR)being all upregulated.Furthermore,the expression of phenylpropanoid and flavonoid biosynthesis genes,and auxininduced genes were suppressed.The defense response pathway of the thermotherapy treated diseased trees was apparently disrupted,as indicated by the downregulation of LRR receptor-like serine/threonine-protein kinase FLS2 gene.This study improves our understanding of the curative mechanisms of heat treatment on HLB trees and CLas biological regulation.Importantly,this provides new insights on the involvement of RNA modification by PPR genes and changes in the circadian clock after the heat stress treatment of CLas-infected citrus.

Effects of Nutrients on Growth and Yield of Shatian Pomelo Trees with Huanglongbing

[Objectives]In order to effectively alleviate Huanglongbing of Shatian pomelo trees,a nutrient preparation was developed and its effect on the growth and yield of Shatian pomelo was explored.[Methods]A field experiment was carried out with four treatments:control group,nutrient solution group,agent group,and nutrient solution+agent group,and the liquids were infused into the fruit trees through the trunk using a hanging bag.[Results]Through field trials,it was found that the nutrient solution treatment of fruit trees significantly increased the leaf chlorophyll content and nitrogen content,which increased by 17%and 12%compared with the control group,respectively,reaching(70.80±1.57)and(5.53±0.1)mg/kg,respectively;and the combined application of the nutrient solution and powder agents greatly alleviated the symptoms of Huanglongbing in Shatian pomelo plants,and significantly improved the bacterial disease index,photosynthetic intensity and yield,which increased by 152%,158%and 213.53%respectively,reaching(371.00±7.22),(406.67±7.09)and 23.15 kg/hm^2.[Conclusions]The use of the nutrient solution and agents greatly alleviated the symptoms of Huanglongbing of Shatian pomelo plants,and significantly improved the chlorophyll content,nitrogen content,bacterial disease index and photosynthetic intensity.Meanwhile,the number of fruit per plant and the weight of single fruit were better than other treatment groups and the control group,the technical operation was simple and the effect was obvious.This provides a certain reference basis for the prevention and control measures of Huanglongbing in Shatian pomelo plants.

Field Identification and Prevention and Control Techniques of Citrus Huanglongbing of Orah Mandarin

In order to promote the development of Orah mandarin industry in Guangxi and reduce the harm of citrus huanglongbing to Orah mandarin,the pathogen,characteristics,occurrence regularity and field symptoms of citrus huanglongbing were summarized,and scientific prevention and control measures were put forward,such as strict implementation of quarantine system,cultivation and use of disease-free seedlings,strengthening field management,timely control of psyllid,timely removal of diseased trees,etc.

A Rapid Identification Method for Germplasm Materials Resistant to Citrus Huanglongbing Disease

[Objectives]This study was conducted to develop a rapid identification method for citrus germline materials resistant to Huanglongbing disease and lay a basis for accelerating citrus breeding for resistance to Huanglongbing and increasing the breeding efficiency.[Methods]Thirty-six citrus germplasms suspected to be resistant to citrus Huanglongbing disease were collected.The method of direct high grafting to citrus trees infected with Huanglongbing pathogen was adopted.The resistance of the test materials was identified and evaluated by field symptoms combined with quantitative PCR.It was defined as the top grafting identification method.[Results]The test materials that were grafted in spring started to germinate after one month,and three months late(June 5,2018)typical mottled yellowing on leaves was observed on KH-14 for the first time.After four months(July 5,2018)of top grafting,typical mottled yellowing occurred on 23 materials,and 11 materials showed no such symptom.After six months(September 4,2018)of top grafting,although the growth of KH-18,KH-12,KHY-4,KHY-5 and KHY-6 were normal,yellowing was observed on their leaves.Only KH-21 grew well,and showed no yellow shoots and yellowing leaves.It was identified as the material with resistance to Huanglongbing disease.Quantitative PCR tests on the above six materials showed that KH-21 was negative and other five were positive.Real-time fluorescence quantitative PCR test indicated that the average Huanglongbing bacteria amount in KH-21 was 1870.0 cell/μg DNA,and the average Huanglongbing bacteria amount in the control material was 372285.5 cell/μg DNA,indicating KH-21 was resistant to Huanglongbing bacteria.[Conclusions]The method for infecting bacteria by top grafting takes six months,can detect large amount of seedlings,and is time-saving,efficient,cost-saving and accurate.This method can quickly identify the resistance of citrus varieties to citrus Huanglongbing disease,and can be popularized and used in the identification of citrus Huanglongbing disease resistance.

Prevention and Control Measures of Citrus Huanglongbing and Application Ideas of Repellent Plants

Citrus huanglongbing is the most infectious and destructive disease in citrus production in the world.The essential characteristics,transmission routes and severity,and diagnosis of citrus huanglongbing,as well as control methods and effects of Diaphorina citri are summarized,and the application ideas of repellent plants is put forward.

Global Dynamics of a New Huanglongbing Transmission Model with Quarantine Measures

An epidemic model which describes Huanglongbing transmission is proposed with the goal of investigating the effect of quarantine measures on the spread of diseases. First of all, the analytical formula for the basic reproduction number R0 is obtained by the means of next generation matrix, and the existence of disease-free equilibrium and endemic equilibrium is discussed. Then, the local stability and the global stability of equilibria are investigated by using Routh-Hurwitz criterion and Lyapunov function, respectively. Numerical simulations indicate that comprehensive quarantine measures can effectively control the spread of Huanglongbing. It provides a reliable tactic basis for preventing the epidemic outbreak.

Role of vector resistance and grafting infection in Huanglongbing control models

Citrus huanglongbing(HLB)is one of the most devastating diseases affecting citrus almost worldwide due to the lack of a cure.To better understand the impact of insecticide resistance and grafting infection on the spread of HLB disease,a vector-borne compartmental model is formulated to describe the transmission dynamics of HLB between citrus and Asian citrus psyllid(ACP).The basic reproduction number R0 is computed by using the next generation matrix approach,which is a threshold value of the uniform persistence and disappearance of HLB disease.By applying the sensitivity analysis of R0,we obtain some parameters with the most significant influence on the transmission dynamics of HLB.Moreover,we also obtain that grafting infection has the least influence on the transmission dynamics of HLB.Additionally,a time-dependent control model of HLB to minimize the cost of implementing control efforts and infected trees and ACPs is formulated.By using Pontryagin's Minimum Principle,we obtain the optimal integrated strategy and prove the uniqueness of optimal control solution.The simulation results illustrate that the strategy involving two time-dependent optimal controls is the most effective to suppress the spread of the disease.However,insecticide spraying is more effective measure compared with infected tree removing.

环境和气象因素对柑橘黄龙病重要传播媒介——柑橘木虱发生规律的影响

研究了柑橘木虱(Diaphorina citri)对环境和气象因素的敏感性,以及其与柑橘黄龙病(Huanglongbing,HLB)传播的关系。总结了柑橘木虱(Diaphorina citri)的基本情况,并探究了环境因素(如海拔、纬度、土壤、田间管理、寄主和生物因素)和气象因素(如温度、湿度、降雨、光照和气压)对其发生规律的影响。这些信息有助于预测柑橘木虱的发生情况和制定有效的防治策略,从而减少柑橘黄龙病的传播风险。

The Citrus Huanglongbing Crisis and Potential Solutions

Citrus is one of the top three fruit crops worldwide.Citrus production is facing an unprecedented crisis due to Huanglongbing(HLB,also known as citrus greening),which is the most devastating disease of citrus.HLB was first reported in Asia a century ago.HLB is associated with Candidatus Liberibacter asiaticus(Las),Ca.L.americanus(Lam),and Ca.L.africanus(Bove,2006),which belong to the gramegative a-proteobacteria.Among them,Las is the most widely spread.Las and Lam are vectored by Diaphorina citri Kuwayama(ACP),whereas Laf is transmitted by Trioza erytreae.

Detection of citrus Huanglongbing based on image feature extraction and two-stage BPNN modeling

Citrus Huanglongbing(HLB),which is spread by the citrus psyllid,is the most destructive disease of citrus industry.While no effective cure for the disease has been reported,detection and removal of infected trees can prevent spreading.Symptoms indicative of HLB can be present in both HLB-positive trees and HLB-negative trees,making identification of infected trees difficult.A detection method for citrus HLB based on image feature extraction and two-stage back propagation neural network(BPNN)modeling was investigated in this research.The identification method for eight different classes including healthy,HLB and non-HLB symptoms was studied.Thirty-four statistical features including color and texture were extracted for each leaf sample,following the two-stage BPNN to model and identify HLB-positive leaves from HLB-negative leaves.The discrimination accuracy can reach approximately 92%which shows that this method based on visual image processing can perform well in detecting citrus HLB.

Analysis and optimal control of a Huanglongbing mathematical model with resistant vector

Huanglongbing(HLB)is an incurable disease that affects citrus trees.To better understand the transmission of HLB,the mathematical model is developed to investigate the transmission dynamics of the disease between Asian citrus psyllid(ACP)and citrus trees.Through rigorous mathematical derivations,we derive the expression of the basic reproduction number(R_(0))of HLB.The findings show that the disease-free equilibrium is locally asymptotically stable if R_(0)<1,and if R_(0)>1 the system is uniformly persistent.By applying the global sensitivity analysis of R_(0),we can obtain some parameters that have the greatest influence on the HLB transmission dynamics.Additionally,the optimal control theory is used to explore the corresponding optimal control problem of the HLB model.Numerical simulations are conducted to reinforce the analytical results.These theoretical and numerical results provide useful insights for understanding the transmission dynamics of HLB and may help policy makers to develop intervention strategies for the disease.

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.

柑橘黄龙病高光谱早期鉴别及病情分级

为实现柑橘黄龙病的早期、快速确诊,有效阻止病害蔓延,达到早期防治、保障柑橘生产的目的,该文研究基于高光谱成像的柑橘黄龙病早期无损检测及病情分级,并对多种预处理方法的建模结果进行探讨。试验获取370～1 000 nm健康、不同染病程度及缺锌共5类柑橘叶片的高光谱图像,用遥感图像处理平台(environment for visualizing images,ENVI)得到各类样本感兴趣区域的光谱反射率平均值。运用一阶微分、移动窗口拟和多项式平滑(savitzky-golay,SG)进行数据处理,结合偏最小二乘判别分析(partial least squares-discriminate analysis,PLS-DA)构建黄龙病的早期鉴别及病情分级模型。结果表明:建立的3个判别模型,验证集相关系数均不低于0.9548。其中,经SG平滑及一阶微分预处理所建立的模型分类效果最佳,总体预测准确率达96.4%,预测均方根误差0.1344。该研究为柑橘病害早期诊断和预警提供了新方法,也为黄龙病病害程度遥感监测提供了基础。

黄龙病病菌侵染对茶枝柑果实类黄酮和挥发性物质的影响

为分析黄龙病(Citrus Huanglongbing,HLB)病菌侵染对茶枝柑(Citrus reticulata cv.chachiensis)果实品质的影响,采用高效液相色谱法和气相色谱-质谱联用技术,分析了花后180 d感病与正常茶枝柑果实中类黄酮物质和挥发性物质的差异。结果表明:茶枝柑感染黄龙病病菌后,果皮中橘皮素等4种聚甲氧基黄酮以及α-蒎烯、辛醛等39种挥发性物质含量显著升高,而氮甲基邻氨基苯甲酸甲酯、胡椒烯等16种挥发性物质显著降低;果肉中川陈皮素等2种聚甲氧基黄酮及D-柠檬烯、β-芳樟醇等27种挥发性物质含量显著降低。因此,黄龙病病菌侵染可能会显著影响茶枝柑果皮和果肉中的次生代谢,进而改变其苦味和香味品质,其改变可能与其感病防御应答机制有关。

Comparative analysis of Wanjincheng orange leaf and root responses to ‘Candidatus liberibacter asiaticus’ infection using leaf-disc grafting

Citrus Huanglongbing,associated with Candidatus Liberibacter asiaticus(Las),is the serious disease of citrus worldwide.Here,we compared differences between leaf and root responses in Wanjincheng Orange(Citrus sinensis Osbeck)to Las infection using leaf-disc grafting.Trees had no obvious symptoms in the first two months after grafting(MAG),but yellowing leaves,thickened midribs and decayed roots began to appear at 6 MAG.The Las growth rate in roots was greater than inmidribs from 2 to 6 MAG;however,by 10 MAG,it was significantly lower in roots than in midribs.Amicroscopic analysis revealed that starch accumulation,callose deposition,cellwall thickness and the number of cell layers increased in the phloem of infected trees.After Las infection,the starch content in the leaves was significantly higher than that in the roots,while the callose deposition in the midribs was 3–19 times that in the roots.A gene expression analysis showed that transcripts of callose-and starch metabolism-related genes were obviously affected by Las infection,and the pathogenesis-related genes PR1,PR2 and PR5 were significantly upregulated and downregulated in midrib and root,respectively.Our results indicated that the PR-mediated resistance response was repressed in root but activated in midrib,which may result in the more rapid Las growth in roots than in midribs during the early infection stage.

Limited infection by ‘Candidatus Liberibacter asiaticus’ in ‘Valencia’ sweet orange trees in the presence of Citrus tristeza virus

Huanglongbing (HLB) is the most destructive disease of citrus and is associated with ‘Candidatus Liberibacter asiaticus’(CLas), a member of the α-proteobacteria. Citrus tristeza virus (CTV) is another pathogen of citrus with very great historic as well as current importance. Both CLas and CTV are phloem-restricted pathogens. A severe CTV isolate, CTV-B6, and CLas-B232 induce a group of symptoms of phloem dysfunction that overlap, but the mild isolate CTV-B2 does not cause any loss to commercial trees. Prior inoculation and establishment of CLas-B232 did not affect subsequent establishment of either CTV-B2 or CTV-B6, while super infection by CLas-B232 was reduced by prior establishment of CTV-B2 and to a lesser extent by prior infection with CTV-B6. Trees co-infected with CTV-B6 and CLas-B232 developed more severe symptoms, typical of CTV-B6, than either of the two pathogens co-infected with CTV-B2. In this study, we confirmed that CLas established in the rootlets earlier and with higher concentration than in leaves. The distribution of CLas in the plant infected by CLas-B438 alone and with CTV-B2 fits a previously proposed model but CLas was more sporadically distributed in a plant co-infected by CLas and CTV-B2 than in a plant infected by CLas alone. These biological phenomena are aligned with previously analyzed transcriptome data and the study provides a novel idea that mild CTV strains may provide some protection against CLas by limiting its multiplication and spread. The protective effect may be due to opposite regulation of key host defense pathways in response to CTV-B2 and CLas-B438.

Decreasing detection frequency of MITE(MCLas-A)in the population of‘Candidatus Liberibacter asiaticus'recently collected in southern China

An active miniature inverted-repeat transposable element(MITE),MCL as-A,was previously identified from'Candidatus Liberibacter asiaticus'known to be associated with citrus Huanglongbing(HL B,yellow shoot disease).To explore the recent transposition status of MCLas-A,389'Ca.L.asiaticus'strains collected from nine regions in China were amplifed using a specific primer set and three representative'Ca.L.asiaticus'strains were analyzed by next-generation sequencing(NGS)approach.PCR and genomic analysis showed that the entire MCLas A was only present in 1.80%(7/389)and the jumping-out type of the MITE was predominant(8123%)in samples tested,suggesting high frequency transposition occurred in'Ca.L.asiaticus'strains recently collected from China.Biological roles of transposition of the active MITE remain to be determined.

Chromosome-level genome assembly of the Asian citrus psyllid,Diaphorina citri

Diaphorina citri is a global citrus pest.As a vector insect,it can transmit the causative agents of citrus huanglongbing,causing irreversible losses to the citrus industry.The acquisition of genomic information can provide a molecular genetic basis for effective control of D.citri.Here,the DNBSEQ™,Oxford Nanopore Technologies,and Hi-C technologies are applied to generate a high-quality chromosome-level genome of D.citri.The genome size of D.citri was 523.78 Mb with a scaffold N50 of 47.05 Mb distributed on 13 chromosomes.A total of 250.64 Mb(47.85%)repeat sequences and 24048 protein-coding genes were predicted.Genome resequencing of female and male individuals indicated that the sex chromosome system of D.citri is XO.Phylogenetic analysis demonstrated that D.citri and Pachypsylla venusta,which separated from their most recent common ancestor about 336.62 million years ago,were the most closely related.Additionally,we identified genes potentially involved in detoxification metabolism,pathogen transmission,and honeydew secretion for further investigation.The high-quality genome provides an important reference for developing effective management strategies of D.citri.