Search

BACKGROUND: Asian citrus psyllid, Diaphorina citri, is a hemipteran that vectors the causal pathogen of citrus greening disease, or huanglongbing (HLB). HLB is a tree killing disease that has severely limited citrus production globally. Unfortunately, there is no cure for this disease, and mitigation depends on multiple insecticide applications to reduce vector populations. Silencing of cytochrome P450 expression associated with detoxification enzymes by RNA interference is known to increase susceptibility of D. citri to insecticides. However, dsRNA was previously introduced into psyllids by topical applications. The possible application of this technology for pest management will require effective field delivery of the dsRNA. Therefore, we evaluated a virus vector (Citrus tristeza virus; 'mild strain' T36) to deliver gene silencing directly to this sap‐sucking insect via plant phloem. Citrus macrophylla plants inoculated with CTV expressing a truncated consensus sequence of CYP450 (CTV‐tCYP450) constantly produced small interfering RNA in the plant phloem that targeted five cytochrome p540 (CYP450) genes in D. citri. RESULTS: Insecticide susceptible D. citri reared on citrus infected with CTV‐tCYP450 were subsequently more susceptible to imidacloprid, fenpropathrin, carbaryl, and chlorpyrifos than those reared on citrus infected with wildtype CTV or non‐infected negative controls. Additionally, nymph survival and adult lifespan were significantly reduced when psyllids were reared on CTV‐tCYP450 citrus plants compared with controls. Interestingly, similar results were obtained after one and two generations of rearing. Finally, field‐collected psyllids from areas with known broad‐spectrum insecticide resistance were rendered more susceptible to imidacloprid and fenpropathrin after feeding on CTV‐tCYP450 citrus trees as compared with those reared on controls. CONCLUSION: The integration of citrus‐mediated RNA inference targeting psyllid detoxification enzymes could function as a resistance management tool and reduce insecticide input in an integrated pest management program for HLB. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

BACKGROUND: Citrus huanglongbing (HLB) is a devastating disease caused by Candidatus Liberibacter asiaticus (CLas) that affects the citrus industry. In nature, CLas relies primarily on Diaphorina citri Kuwayama as its vector for dissemination. After D. citri ingests CLas‐infected citrus, the pathogen infiltrates the insect's body, where it thrives, reproduces, and exerts regulatory control over the growth and metabolism of D. citri. Previous studies have shown that CLas alters the composition of proteins in the saliva of D. citri, but the functions of these proteins remain largely unknown. RESULTS: In this study, we detected two proteins (DcitSGP1 and DcitSGP3) with high expression levels in CLas‐infected D. citri. Quantitative PCR and Western blotting analysis showed that the two proteins were highly expressed in the salivary glands and delivered into the host plant during feeding. Silencing the two genes significantly decreased the survival rate for D. citri, reduced phloem nutrition sucking and promoted jasmonic acid (JA) defenses in citrus. By contrast, after overexpressing the two genes in citrus, the expression levels of JA pathway‐associated genes decreased. CONCLUSION: Our results suggest that CLas can indirectly suppress the defenses of citrus and support feeding by D. citri via increasing the levels of effectors in the insect's saliva. This discovery facilitates further research into the interaction between insect vectors and pathogens. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Huanglongbing (HLB) is caused by “Candidatus Liberibacter asiaticus” and is spread by citrus psyllids. It is a highly damaging and infectious disease of citrus for which there is no reliable treatment. Timely detection and removal of diseased trees is an effective strategy to control HLB. Various citrus cultivars infected with HLB exhibit distinct symptoms, posing a challenge to generalize HLB detection methods across multiple cultivars. Volatile organic compounds (VOCs) in HLB‐infected leaves from cultivars grown in different regions were comprehensively analysed by gas chromatography–mass spectrometry (GC‐MS) and gas chromatography‐ion mobility spectrometry (GC‐IMS), aiming to establish a new generalized HLB detection method applicable to a range of citrus cultivars. The changes in VOCs were analysed in leaves of four citrus cultivars after infection with HLB. The results showed that there was a similar response to HLB infection in different citrus cultivars, and this response was reflected in both the concentration and type of VOCs. By combining GC‐MS and GC‐IMS with orthogonal partial least squares discriminant analysis (OPLS‐DA) model, the accurate identification of the HLB infection status of different citrus cultivars was achieved, with the prediction indices reaching 0.994 for GC‐MS and 0.972 for GC‐IMS. In addition, 27 compounds were identified that were significantly different between citrus cultivars affected by HLB and healthy plants. This study provides valuable insights into the changes in VOCs in citrus cultivars after HLB infection and lays the theoretical foundation for VOC‐based HLB detection strategies. [ABSTRACT FROM AUTHOR]

The Asian citrus psyllid, Diaphorina citri, is the primary vector of the HLB pathogen, Candidatus Liberibacter asiaticus (CLas). The acquisition of CLas shortens the developmental period of nymphs, accelerating the emergence into adulthood and thereby facilitating the spread of CLas. Cuticular proteins (CPs) are involved in insect emergence. In this study, we investigated the molecular mechanisms underlying CLas‐promoted emergence in D. citri via CP mediation. Here, a total of 159 CP genes were first identified in the D. citri genome. Chromosomal location analysis revealed an uneven distribution of these CP genes across the 13 D. citri chromosomes. Proteomic analysis identified 54 differentially expressed CPs during D. citri emergence, with 14 CPs exhibiting significant differential expression after CLas acquisition. Five key genes, Dc18aa‐1, Dc18aa‐2, DcCPR‐24, DcCPR‐38 and DcCPR‐58, were screened from the proteome and CLas acquisition. The silencing of these 5 genes through a modified feeding method significantly reduced the emergence rate and caused various abnormal phenotypes, indicating the crucial role that these genes play in D. citri emergence. This study provides a comprehensive overview of the role of CPs in D. citri and reveals that CLas can influence the emergence process of D. citri by regulating the expression of CPs. These key CPs may serve as potential targets for future research on controlling huanglongbing (HLB) transmission. [ABSTRACT FROM AUTHOR]

Plant pathogens can alter the behavior of their insect vectors as well as their survival and reproduction. The African psyllid, Trioza erytreae, is one of the vectors of Huanglongbing, a citrus disease caused mainly by "Candidatus Liberibacter asiaticus" (CLas). The purpose of this study was to characterize the effects of CLas on the psyllid, T. erytreae using Citrus volkamerina plants as the study system. The study focused more specifically on the CLas effects prior to and after its acquisition by the psyllid T. erytreae. Our results did not support the hypothesis that CLas effects psyllid probing behavior prior to acquisition; few differences were observed between uninfected T. erytrea feeding on CLas‐infected versus control plants. On the other hand, compared to psyllids that had completed their development on control plants, the ones that had completed their development on a CLas‐infected plant exhibited changes in their behavior (greater velocity), physiology (smaller mass) and biochemistry (lower water and lipid content). Altogether, our results confirm the existence of a marked postacquisition effect on the vector locomotor behavior and a minor preacquisition effect of CLas on the vector behavior, which can be partially explained by physiological and biochemical changes. [ABSTRACT FROM AUTHOR]

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 24 048 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. [ABSTRACT FROM AUTHOR]

The evolution of insect vector‐pathogen relationships has long been of interest in the field of molecular ecology. One system of special relevance, due to its economic impacts, is that between Diaphorina citri and 'Candidatus Liberibacter asiaticus' (CLas), the cause of the severe Asian form of huanglongbing. CLas‐positive D. citri are more fecund than their CLas‐negative counterparts, boosting opportunities for pathogens to acquire new vector hosts. The molecular mechanism behind this life‐history shift remains unclear. Here, we found that CLas promoted ovarian development and increased the expression of the vitellogenin receptor (DcVgR) in ovaries. DcVgR RNAi significantly decreased fecundity and CLas titer in ovaries, extended the preoviposition period, shortened the oviposition period and blocked ovarian development. Given their importance in gene regulation, we explored the role of miRNAs in shaping these phenotypes and their molecular triggers. Our results showed that one miRNA, miR‐275, suppressed DcVgR expression by binding to its 3' UTR. Overexpression of miR‐275 knocked down DcVgR expression and CLas titer in ovaries, causing reproductive defects that mimicked DcVgR knockdown phenotypes. We focused, further, on roles of the Juvenile Hormone (JH) pathway in shaping the observed fecundity phenotype, given its known impacts on ovarian development. After CLas infection, this pathway was upregulated, thereby increasing DcVgR expression. From these combined results, we conclude that CLas hijacks the JH signalling pathway and miR‐275, thereby targeting DcVgR to increase D. citri fecundity. These changes simultaneously increase CLas replication, suggesting a pathogen‐vector host mutualism, or a seemingly helpful, but cryptically costly life‐history manipulation. [ABSTRACT FROM AUTHOR]

The use of unmanned aerial vehicles (UAVs) combined with other methods to spray insecticides may be a useful technique for controlling Diaphorina citri Kuwayama (Hemiptera: Psyllidae) and consequently managing Huanglongbing. Therefore, this study was conducted to assess the efficacy of insecticide application using a UAV for D. citri control in citrus orchards of 1, 4 and 12 years old. In addition, spray deposition was assessed using copper as a marker. Seedlings of sweet orange were attached on the inner canopy and on the outer canopy of citrus trees before spraying insecticide or copper. An additional experiment was performed to assess the effect of 6 and 10 m spray swaths in the 4‐year‐old orchard. Ten adult psyllids were confined to each sprayed seedling. Untreated seedlings were used as a control. The first psyllid confinement was performed 4 h after spraying insecticide. The second, third and fourth confinements were performed 7, 14 and 21 days, respectively, after spraying. In general, insecticide application using a UAV resulted in psyllid mortality higher than 80% after the first and second confinements, regardless of the orchard age and seedling position on the citrus tree (inner or outer). There were no significant differences in D. citri mortality between the spray swaths assessed. Spray deposition on leaves was higher in the outer canopy than in the inner canopy of citrus trees. This study demonstrated, for the first time, the efficacy of insecticide application using a UAV for D. citri control in citrus orchards of different ages. These results broaden the prevailing perspectives on the use of UAVs as complements to ground spraying for psyllid management. [ABSTRACT FROM AUTHOR]

Huanglongbing (HLB) is highly contagious and cannot be cured, resulting in a decrease in the commercial value of citrus. Timely detection and removal of diseased trees is an effective way to reduce losses. Complex symptoms of HLB, such as nutrient deficiencies often accompany HLB; as a result effective and accurate identification of HLB remains a challenge. In this study, 175 volatile organic compounds (VOCs) were detected in three categories (healthy, HLB, and Zn‐deficiency) of samples using headspace solid‐phase microextraction gas chromatography–mass spectrometry (HS‐SPME‐GC/MS), highlighting the variability of VOCs present in different categories of samples. In order to simplify the testing steps and reduce the cost in practical agricultural production, a method based on electronic nose technology to collect VOCs from citrus leaves for HLB detection was proposed. Among them, limiting value features and linear discriminant analysis were identified as the best combination of feature extraction and pattern recognition methods. Multiple sets of comparison experiments were set up and the collection conditions of VOCs were optimized. The results showed that the best classification performance was achieved for a 0.2 g sample at a collection time of 20 min when the collection temperature was 40°C and the headspace volume was 200 mL. Four types of samples (healthy, HLB‐positive, Zn‐deficiency, Zn‐deficiency and HLB‐positive) were used for model reliability validation, with an accuracy of 97.79% for HLB samples for multiple symptoms (including HLB‐positive and Zn‐deficiency and HLB‐positive) identification. In addition, the accuracy of samples with a combined effect of Zn‐deficiency and HLB was 96.43%. The results show that the E‐nose‐based HLB detection method is conducive to suppressing the spread of HLB, which can ensure the quality of citrus products and reduce the economic loss to horticulturists, and has good practical value. [ABSTRACT FROM AUTHOR]

The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is the major citrus pest worldwide because it can transmit the bacteria associated with citrus greening disease (huanglongbing, HLB). The most common management strategy of this pest is chemical control, but this often results in pest resistance and threatens the sustainability of the citrus culture. There is therefore need for sustainable control strategies such as biological control. Previous experiments showed that the predatory mite Amblyseius herbicolus Chant (Acari: Phytoseiidae) is able to develop and reproduce on D. citri eggs. Here, we studied the population dynamics of this pest on plants with and without A. herbicolus on isolated orange jasmine plants, Murraya paniculata (L.) Jack (Rutaceae), an important host plant of the pest. Predatory mites were released 1 week before the release of D. citri and were provided with twines with pollen as oviposition sites and food source, respectively, to maintain populations of A. herbicolus in the absence of the pest. Densities of adults and nymphs of D. citri on plants with predators were about 85% lower than on plants without predators, showing that A. herbicolus is a potential biocontrol agent of D. citri. However, the stage structure of the D. citri populations at the end of the experiment did not differ significantly between control plants and plants with predators. This led us to investigate whether the predator attacked other stages besides eggs. Indeed, predators were able to feed and reproduce on first and second instar nymphs of D. citri, but not on instars 3–5. Concluding, the release of A. herbicolus with alternative food and oviposition sites can reduce infestations of D. citri. We suggest that field experiments should be done in areas where HLB occurs to test whether releases of this predator limit the spread of this devastating disease. [ABSTRACT FROM AUTHOR]

The development of sustainable methods to control Diaphorina citri Kuwayama (Hemiptera: Liviidae) is crucial to improve management of huanglongbing (HLB), the most important citrus disease worldwide. In a flushing Citrus × sinensis L. Osbeck (Rutaceae) orchard, this study assessed the combined effects of insecticide‐treated Bergera koenigii L. (Rutaceae) trees in the orchard border, as a lethal trap crop, and the application of processed kaolin to the citrus crop, as a repellent (push‐pull and kill strategy), in reducing D. citri infestation and dispersal. Additionally, the volatile organic compound (VOC) emission profile and attractiveness of B. koenigii shoots were investigated in indoor experiments. The push‐pull and kill strategy reduced the number of D. citri adults on citrus trees by 95, 84, and 63% compared to the untreated control, trap crop only, and kaolin only treatments, respectively. These high reductions can be attributed to the combination of attractive stimuli (VOCs) and repellent stimuli by the trap crop and kaolin, respectively. The number of psyllids observed on citrus trees subjected to treatments with trap crop and kaolin separately was 69 and 86% lower than on untreated control trees, respectively. In addition, all treatments reduced and delayed D. citri dispersal inside the orchard, especially the push‐pull and kill strategy. In olfactometric assays, D. citri females showed a significant preference for B. koenigii odors over C. × sinensis cv. 'Valencia' flush odors, likely related with the different VOC emission profiles of the two plants. This study shows the effectiveness of push‐pull and kill as a new management strategy to control D. citri and consequently HLB. [ABSTRACT FROM AUTHOR]

BACKGROUND: Host genetic resistance is a promising strategy for the management of Diaphorina citri Kuwayama (Hemiptera: Psyllidae), and consequently Huanglongbing (HLB). To date, no study has investigated the resistance to D. citri in the clonal and vegetatively propagated plants of the Microcitrus, Eremocitrus, and Atalantia genera. This study assesses Near and True Citrus genotype antixenosis and antibiosis against D. citri, with trichome density and volatile emission as possible mechanisms of resistance. RESULTS: All genotypes were oviposited by D. citri, however, 8 of 14 genotypes were less oviposited than Citrus × sinensis 'Valencia' (susceptible control). Diaphorina citri nymphs had lower nymphal viability in E. glauca (31%) and M. warburgiana (58%) than that in Citrus × sinensis (77%). The behavioral assay showed that 30% of D. citri nymphs in the last instars evaded E. glauca shoots, whereas no nymphs evaded Citrus × sinensis shoots. A higher trichome density was observed in E. glauca shoots compared to the other genotypes. Chemical analysis revealed differences in the volatile profiles of E. glauca and Citrus × sinensis. CONCLUSION: Eremocitrus glauca and M. warburgiana genotypes were more resistant to D. citri than Citrus × sinensis. Higher trichome density in the shoots may negatively influence the development of D. citri nymphs. Eremocitrus glauca volatiles may also be involved in their resistance to D. citri. © 2022 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

BACKGROUND: The citrus greening disease or Huanglongbing (HLB) is the most devastating disease of citrus crops. Trioza erytreae is a vector of HLB. Since its introduction in Europe, the insect reached the northern region of Spain and the southern region of Portugal, threatening relevant citrus production areas. Limiting the spread of HLB vectors is mandatory to prevent this disease. In this work, we assessed the effect of kaolin, a white mineral clay, on the landing, settling behavior and feeding behavior of Trioza erytreae on lemon plants. RESULTS: After kaolin application, the number of plants on which the insect was found was significantly lower than on untreated plants in the laboratory and in the field. Moreover, there were significantly fewer T. erytreae and a shorter duration of phloem‐related events on kaolin‐treated than untreated plants. CONCLUSION: The use of kaolin could be a suitable and efficient tool for inclusion into integrated pest management programs or organic production to reduce populations of T. erytreae and subsequently limit the spread of HLB in citrus crops. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The effects of five host plant species – tangerine, Citrus reticulata Blanco cv. Shatangju, C. reticulata cv. Ponkan, orange jasmine, Murraya paniculata (L.) Jack, lemon, Citrus limon (L.) Osbeck, and navel orange, Citrus sinensis (L.) Osbeck (all Rutaceae) – on the life‐table parameters and host preference of the citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), were evaluated under laboratory conditions. In the presence of the host plants tested, citrus psyllid showed higher preference for navel orange, followed by Shatangju and orange jasmine. Total nymph duration of D. citri was longest on lemon (14.1 days) and shortest on Shatangju (12.7 days). The survival rate (lx) of citrus psyllid on orange jasmine was significantly higher than on other tested host plants. The highest longevity of females was observed on lemon (47.6 days), whereas the shortest was on navel orange (40.5 days). The total number of eggs laid by females was not significantly different among host species. The net reproductive rate (R0) was significantly affected by host plants, being highest on Shatangju (162.1 females/female per generation) and lowest on navel orange (130.7). The intrinsic rates of natural increase (rm) was not significantly different among hosts, and ranged from 0.112 on lemon to 0.123 on orange jasmine. Cluster analysis of the life‐table parameters of D. citri on five host plant species indicated that Shatangju was the most suitable host to D. citri, followed by orange jasmine, and navel orange was the least suitable. [ABSTRACT FROM AUTHOR]

BACKGROUND: Processed kaolin is a promising tactic that can be utilized for managing Diaphorina citri Kuwayama infestation. However, the frequency of kaolin application to protect citrus trees during flushing remains undetermined. The objective of this study was to ascertain the frequency of kaolin application (2%) required to reduce D. citri infestation and dispersal in flushing citrus orchards and to measure the spectral reflectance of kaolin‐sprayed leaves. RESULTS: Trees sprayed with kaolin at 7‐ and 14‐day intervals showed an 80% psyllid reduction compared to untreated trees. In addition, there was lower psyllid dispersal in the kaolin‐sprayed plots. Spectral measurements (wavelengths between 320 and 700 nm) showed an increased reflectance of 35.4% (adaxial leaf side) and 21.9% (abaxial leaf side) in mature leaves, and 9% (adaxial leaf side) and 2.2% (abaxial leaf side) in kaolin‐sprayed immature leaves compared to untreated trees. CONCLUSION: Application of kaolin (2%) at 7‐ and 14‐day intervals can be an important and sustainable tool for reducing D. citri infestation and dispersal in flushing citrus orchards. The reduced D. citri population in plots with both kaolin application intervals may be related to the visual effect (whitish aspect) and increased light reflectance in citrus trees, which are probably the main mechanisms by which kaolin interferes on D. citri host finding behavior. © 2021 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Huanglongbing (HLB) causes extensive damage in citrus orchards worldwide. Symptoms include blotchy mottle leaf (BML) and little leaf chlorosis (LLC), and nutrient deficiency usually occurs concurrently. However, the relationship between plant mineral content and infection with Candidatus Liberibacter asiaticus (CLas) is not clearly established.We sampled 7‐month‐old autumn shoots with three characteristic phenotypes, asymptomatic leaf (AL), BML and LLC, representing HLB disease progression, and further divided samples into CLas‐infected and uninfected based on PCR analysis.HLB infection decreased transfer coefficients of Mg and K from leaf to phloem tissues through regulation of the transporter genes Cs3g03790.1 and PtrMGT5, increasing the content of leaf Mg and K. HLB infection also decreased leaf Zn, xylem Ca and phloem Ca and Zn content. Leaf Ca, Mg, Zn and B content decreased while leaf K content increased significantly as symptoms progressed from AL to LLC. The transport of P from leaf to phloem tissue, as evaluated by the transfer coefficient, was regulated by the transporter CsiPT2, resulting in irregular levels of leaf P.Our results provide insights into the nutrient dynamics in Citrus in response to CLas infection and the progression of HLB symptoms. [ABSTRACT FROM AUTHOR]

Asian Citrus Psyllid (ACP), Diaphorina citri, is a key vector transmitting the causative agent of Huanglongbing (HLB) disease. Population growth of ACP is evident after feeding on plant flush shoots, as they only oviposit here. However, the underlying mechanism as to why flush shoots govern oviposition is unclear. This study compares the fecundity and ovarian morphology of ACP between young flush and mature leaves. Furthermore, the transcriptome of mated females infesting Murraya paniculata was analysed. Finally, the gene of the key Target of Rapamycin (TOR) signalling pathway was silenced by RNAi. Results indicated that flush shoot feeding activated the development of the psyllids ovary and therefore induced oviposition. A total of 126 and 2794 differentially expressed genes were detected at 1 and 5 days, respectively, after pest infestation of flush shoots compared to mature leaves. Many genes are involved in protein metabolism, Mitogen‐Activated Protein Kinase (MAPK) signalling pathway, hormone synthesis, and TOR signalling pathway: all thought to activate reproduction. Silencing of the positive regulator gene DcRheb in the TOR pathway resulted in lower levels of ecdysone and juvenile hormone and decreased vitellogenin synthesis, further disrupting reproductive ability. This study enhances understanding of the molecular mechanism underlying ACP's reproductive strategy. [ABSTRACT FROM AUTHOR]

In conclusion, we firstly established a novel time-resolved fluorescent microsphere lateral flow biosensors (TRFM-LFBs) for the detection of I C i Las. Time-resolved fluorescent microsphere lateral flow biosensors for rapid detection of Candidatus Liberibacter asiaticus Keywords: Candidatus Liberibacter asiaticus; Citrus; detection; Huanglongbing; time-resolved fluorescent microsphere lateral flow biosensors EN Candidatus Liberibacter asiaticus Citrus detection Huanglongbing time-resolved fluorescent microsphere lateral flow biosensors 1235 1237 3 07/01/22 20220701 NES 220701 Citrus Huanglongbing (HLB) is one of the most devastating diseases causing unprecedented global economic losses and constituting a major hindrance to the development of the citrus industry. [Extracted from the article]

Whole‐genome sequencing of "Candidatus Liberibacter asiaticus" (Las) indicated some polymorphic gene regions enabling the molecular characterization of this bacterium. Although the population diversity of Las in China has been previously studied, no reports have used a combination of a prophage region and short tandem repeat (STR) loci for phylogenetic relationship characterization. In this study, we investigated the genetic diversity and structure of 667 Las strains from nine provinces in southern China using multiple genetic loci including three type‐specific prophage loci, two STR loci, and a miniature inverted‐repeat transposable element (MITE) region. The results indicated that the genetic diversity varied according to the gene loci used. The prophage regions, including the MITEs, revealed significant genetic differences of Las populations in Yunnan and Guizhou from other provinces, while the STR loci only indicated a difference of Las population in Guizhou from those of other provinces. In particular, the Las population shown to be diverse in Jiangxi at the CLIBASIA_01215_STR locus was not diverse when measured using the other loci. Considering all loci, the Las populations from Yunnan and Guizhou were different, those from Guangdong and Guangxi were complex, while the Las population in Jiangxi was comparatively simple. Las populations from five neighbouring provinces, including Guangdong, Guangxi, Hunan, Fujian, and Jiangxi were grouped in a big cluster. The Las population structure changed with age according to the prophage type. This study evaluated the method used for studying the molecular diversity of Las populations and provided more detailed information on the geographical origin and epidemic trend of Chinese Las populations. [ABSTRACT FROM AUTHOR]

Seasonality often influences multiple aspects of vector‐borne plant diseases. With respect to the management of plant disease vectors, an understanding of the factors driving seasonal changes in vector flight activity may improve management outcomes by facilitating more proactive application of control measures. One of the most challenging issues for citrus Huanglongbing management is to forecast when disease primary spread by bacteriliferous Diaphorina citri will occur. We monitored the temporal patterns of immigrating psyllids, using upwards of 1,200 traps spread among 9 citrus farms and checked on a weekly basis over 4 years. This dataset was analysed with a set of hierarchical models to estimate the effect of climatic variables on citrus foliage production (i.e. flush), and effects of climatic variables and citrus foliage dynamics on D. citri catches over the season. The results showed substantial seasonal variability in immigrating D. citri abundance, with the critical dispersal/migration period occurring between end of winter and spring. During this period, 65% of the total psyllids were collected on sticky traps. Seasonality of immigrating D. citri coincided with changes in certain climate variables, with negative effects of humidity and daily maximum temperatures, and a positive effect of rainfall amount in prior weeks. Maximum temperature and both daily minimum temperature and rainfall during prior weeks were also associated with new citrus flush production, which itself was positively related to immigrating D. citri abundance. Based on these results, citrus growers should be aware and intensify the frequency of psyllid control tactics (i.e. insecticide and/or kaolin sprays) during this period in order to prevent Ca. L. asiaticus transmission. These results are an important step towards developing the predictive framework needed to refine D. citri and huanglongbing management. [ABSTRACT FROM AUTHOR]

Summary: Huanglongbing (HLB) is the most devastating citrus disease in the world. Almost all commercial citrus varieties are susceptible to the causal bacterium, Candidatus Liberibacter asiaticus (CLas), which is transmitted by the Asian citrus psyllid (ACP). Currently, there are no effective management strategies to control HLB. HLB‐tolerant traits have been reported in some citrus relatives and citrus hybrids, which offer a direct pathway for discovering natural defence regulators to combat HLB. Through comparative analysis of small RNA profiles and target gene expression between an HLB‐tolerant citrus hybrid (Poncirus trifoliata × Citrus reticulata) and a susceptible citrus variety, we identified a panel of candidate defence regulators for HLB‐tolerance. These regulators display similar expression patterns in another HLB‐tolerant citrus relative, with a distinct genetic and geographic background, the Sydney hybrid (Microcitrus virgata). Because the functional validation of candidate regulators in tree crops is always challenging, we developed a novel rapid functional screening method, using a C. Liberibacter solanacearum (CLso)/potato psyllid/Nicotiana benthamiana interaction system to mimic the natural transmission and infection circuit of the HLB complex. When combined with efficient virus‐induced gene silencing in N. benthamiana, this innovative and cost‐effective screening method allows for rapid identification and functional characterization of regulators involved in plant immune responses against HLB, such as the positive regulator BRCA1‐Associated Protein, and the negative regulator Vascular Associated Death Protein. [ABSTRACT FROM AUTHOR]

BACKGROUND: Citrus greening disease (Huanglongbing, HLB) has infected >90% of Florida's oranges and thus growers are desperate to improve pest management strategies. In this field study, insecticide application efficacy was investigated with liquid chromatography mass spectrometry to determine if insecticide concentration and distribution were effective at killing the target pest Asian citrus psyllids (ACP). Sample discs attached to leaves were sprayed with imidacloprid and malathion at a field site in Florida. Application method, canopy height and depth, cardinal side of tree, and leaf side were considered to assess the spatial distribution of insecticides throughout citrus trees. Furthermore, ACP were inspected before and after insecticide applications to quantify psyllid population response. RESULTS: Our findings show that although insecticide concentrations were high enough to kill ACP, the spatial distribution of insecticides throughout individual trees was highly variable and live ACP were detected after insecticide application. The top side of leaves received significantly more insecticide than the underside of leaves. Additionally, inadequate distribution to different areas of the tree canopy was observed for all application methods tested (aerial, ground speed-sprayer, and ground sidesprayer). Inspections of ACP populations before and after insecticide applications resulted in reductions of 85% (malathion) and 48–80% (imidacloprid). CONCLUSIONS: The variability in insecticide spatial distribution due to application method allows remaining ACP to continue spreading citrus greening disease to unprotected trees. Further research is needed to improve insecticide application methods and technology for citrus trees in order to implement effective pest management strategies and fully target ACP to eliminate HLB. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

SUMMARY: Trifoliate orange (Poncirus trifoliata), a deciduous close relative of evergreen Citrus, has important traits for citrus production, including tolerance/resistance to citrus greening disease (Huanglongbing, HLB) and other major diseases, and cold tolerance. It has been one of the most important rootstocks, and one of the most valuable sources of resistance and tolerance genes for citrus. Here we present a high‐quality, chromosome‐scale genome assembly of P. trifoliata. The 264.9‐Mb assembly contains nine chromosomal pseudomolecules with 25 538 protein‐coding genes, covering 97.2% of the estimated gene space. Comparative analyses of P. trifoliata and nine Citrus genomes revealed 605 species‐specific genes and six rapidly evolving gene families in the P. trifoliata genome. Poncirus trifoliata has evolved specific adaptation in the C‐repeat/DREB binding factor (CBF)‐dependent and CBF‐independent cold signaling pathways to tolerate cold. We identified candidate genes within quantitative trait loci for HLB tolerance, and at the loci for resistance to citrus tristeza virus and citrus nematode. Genetic diversity analysis of Poncirus accessions and Poncirus/Citrus hybrids shows a narrow genetic base in the US germplasm collection, and points to the importance of collecting and preserving more natural genetic variation. Two phenotypically divergent Poncirus accessions are found to be clonally related, supporting a previous conjecture that dwarf Flying Dragon originated as a mutant of a non‐dwarfing type. The high‐quality genome reveals features and evolutionary insights of Poncirus, and it will serve as a valuable resource for genetic, genomic and molecular research and manipulation in citrus. Significance Statement: A high‐quality, chromosome‐scale reference genome of trifoliate orange has enabled a comprehensive comparative genomics analysis of Citrus‐related genomes, discovery of gene families under expansion or positive selection, identification of gene families associated with important disease resistance and cold tolerance, and localization of candidate genes for tolerance to Huanglongbing, the most destructive disease in citrus. This genome will be a valuable resource for genomic research and manipulation in citrus and related plants. [ABSTRACT FROM AUTHOR]

Keywords: citrus; Huanglongbing; viable CLas; pollen; distribution; cross-pollination; dissemination EN citrus Huanglongbing viable CLas pollen distribution cross-pollination dissemination 247 249 3 01/17/22 20220201 NES 220201 Huanglongbing (HLB), also known as citrus greening, is the most severe pandemics in citrus in more than 50 countries in Asia, Africa, and America, causing serious economic losses worldwide (Gottwald,2020 2020; Wang, 2019). The distribution of I C i Las in pollen grains and pollen tubes opened a new possible dissemination pathway through pollination which should be also taken into account for the integrated control of HLB. After cross-pollination, I C i Las were firstly detected on the stigma in 4 hpp, and then moved from stigma to stylet via pollen tubes. [Extracted from the article]

Citrus greening (Huanglongbing, HLB) is a destructive disease associated with the uncultivable, phloem‐limited, gram‐negative bacteria Candidatus Liberibacter spp., which affects citrus crops and other Rutaceae species. HLB is rapidly transmitted by the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae). We developed an artificial medium on which D. citri eggs can hatch and first‐instar nymphs can feed. The medium could be used to study insect physiology and screen molecules that may interfere with egg hatching and nymph development. [ABSTRACT FROM AUTHOR]

The African citrus triozid (ACT) Trioza erytreae (Del Guercio) (Hemiptera: Triozidae) and the Asian citrus psyllid (ACP) Diaphorina citri (Kuwayama) (Hemiptera: Liviidae) are primary vectors of the 'Candidatus' Liberibacter spp. ACT is associated with 'Candidatus Liberibacter africanus' (CLaf) causal agent of the African citrus greening disease (ACGD), whereas ACP vectors 'Candidatus Liberibacter asiaticus' (CLas) and 'Candidatus Liberibacter americanus' (CLam), the Asian and the American strains, respectively, associated with huanglongbing.A preliminary survey in Kenyan citrus groves after the invasion of ACP had revealed that lime‐green sticky traps (Asacp) were not effective in detecting and monitoring ACT and ACP. Therefore, this study compared eight differently coloured double‐sided sticky traps to evaluate, which colour was most effective for detecting ACT and ACP, particularly at low densities. The traps were coded Red, Blue, Asgreen1, GLMgreen, Asacp, Asyellow, White, and Black.Asyellow and GLMgreen traps captured more ACTs and ACPs than any other trap type. However, since there was no clear difference in the efficiency of Asyellow and GLMgreen in trapping ACP, any of the two trap types would be useful in the presence of both pests.Among the eight traps, Asyellow and GLMgreen traps are potentially promising options for monitoring and detecting the pests in areas where they coexist.Our findings will guide biosecurity agencies in decision‐making and designing ecologically friendly integrated pest management (IPM) strategies for citrus greening vectors in Africa as well as serve as an early warning to safeguard against pests' invasion into unaffected areas. [ABSTRACT FROM AUTHOR]

Bacillus thuringiensis (Bt) toxins are effective in controlling insect pests either through the spraying of products or when expressed in transgenic crops. The discovery of endophytic Bt strains opened new perspectives for studies aimed at the control of sap‐sucking insects, such as the Asian citrus psyllid Diaphorina citri Kuwayama (Hemiptera: Liviidae), a vector of "Candidatus Liberibacter spp.," associated with citrus huanglongbing (HLB). In this study, translocation of endophytic Bt strains in citrus seedlings inoculated with Bt suspension delivered by soil‐drench, and their systemic pathogenicity to D. citri nymphs were investigated. The pathogenicity of three wild‐type Bt strains against D. citri third‐instar nymphs was demonstrated. Among the 10 recombinant strains tested (each of them harboring a single cry or cyt gene), 3 can be highlighted, causing 42%–77% and 66%–90% nymphal mortality at 2 and 5 d after inoculation, respectively. The isolation of Bt cells from young citrus shoots and dead nymphs, and PCR performed with specific primers, confirmed the involvement of the Bt strains in the psyllid mortality. This is the first report showing the translocation of Bt strains from citrus seedling roots to shoots and their potential to control D. citri nymphs that fed on these soil‐drench inoculated seedlings. The Bt strains that caused the highest mortality rates have the potential to be used as bioinsecticides to control D. citri and the identified genes can be used for the production of transgenic Bt citrus. [ABSTRACT FROM AUTHOR]

'Candidatus Liberibacter' species are insect‐transmitted, phloem‐limited α‐Proteobacteria in the order of Rhizobiales. The citrus industry is facing significant challenges due to huanglongbing, associated with infection from 'Candidatus Liberibacter asiaticus' (Las). In order to gain greater insight into 'Ca. Liberibacter' biology and genetic diversity, we have performed genome sequencing and comparative analyses of diverse 'Ca. Liberibacter' species, including those that can infect citrus. Our phylogenetic analysis differentiates 'Ca. Liberibacter' species and Rhizobiales in separate clades and suggests stepwise evolution from a common ancestor splitting first into nonpathogenic Liberibacter crescens followed by diversification of pathogenic 'Ca. Liberibacter' species. Further analysis of Las genomes from different geographical locations revealed diversity among isolates from the United States. Our phylogenetic study also indicates multiple Las introduction events in California and spread of the pathogen from Florida to Texas. Texan Las isolates were closely related, while Florida and Asian isolates exhibited the most genetic variation. We have identified conserved Sec translocon (SEC)‐dependent effectors likely involved in bacterial survival and virulence of Las and analysed their expression in their plant host (citrus) and insect vector (Diaphorina citri). Individual SEC‐dependent effectors exhibited differential expression patterns between host and vector, indicating that Las uses its effector repertoire to differentially modulate diverse organisms. Collectively, this work provides insights into the evolution of 'Ca. Liberibacter' species, the introduction of Las in the United States and identifies promising Las targets for disease management. [ABSTRACT FROM AUTHOR]

Huanglongbing (HLB) is a devastating disease that affects the entire citrus plant, including the root system. Previous studies have shown that sulphonamide antibiotics can suppress titres of the pathogen that causes HLB, 'Candidatus Liberibacter asiaticus' (CLas), and affect root morphology in the plant through unknown mechanisms. To better understand the response of CLas‐infected roots to sulphonamide antibiotics, hydroponic cultures of CLas‐infected citrus roots were treated with sulfadimethoxine sodium (SDX), indole butyric acid (IBA), or water for 60 days to evaluate root metabolism and resistance against CLas via transcriptomic analysis. This study indicated that SDX and IBA treatments increased active root surface area in HLB‐affected citrus, which may be due to root hair and lateral root growth. CLas titres in HLB‐affected citrus roots treated with SDX were lower than those of IBA and control treatments. Function categorization indicated that plant hormone signal transduction and plant–pathogen interaction were the most markedly enriched pathways in the HLB‐affected citrus root after SDX treatment. The expression of genes involved in biosynthesis of auxin and ethylene, which are related to root hair and lateral root growth, were up‐regulated by SDX. Moreover, SDX also induced genes related to the metabolism of jasmonates, brassinosteroids, reactive oxygen species, and secondary metabolites, which are beneficial for resistance against HLB. In conclusion, we propose a model for SDX in regulating metabolic pathways in the root and resistance against CLas in HLB‐affected citrus root, which is beneficial for recovering an HLB‐affected citrus root system and combating CLas. [ABSTRACT FROM AUTHOR]

Aim: To quantify current and predict future distribution of the citrus greening pathogens "Candidatus Liberibacter asiaticus" (Las) in Africa and "Candidatus Liberibacter africanus" (Laf) globally. Location: Africa. Methods: Three species distribution models (MaxEnt, BIOCLIM and Boosted Regression Trees) were used to predict the current and future potential distribution of Las in Africa, and the potential global distribution of Laf, using long‐term bioclimatic variables. Two climate change scenarios (moderate and extreme) were employed to determine how future climate alterations may affect the potential distribution of Las in Africa. Presence data from global reports of Las, as well as the new positional points obtained in this survey, were used to predict the habitat suitability of the pathogen in Africa, while the presence data points of Laf were used to predict the global habitat suitability. Testing data comprised 25% of the presence only points. Results: Consensus of the three models predicted a potential distribution of Las in large areas of Western, Eastern and sub‐Saharan Africa. North Africa was mostly unsuitable for Las, except for the northern fringes. The potential distribution of Laf included South and Central America, Asia and Australia. In Europe, the United Kingdom and the Iberian Peninsula showed marginal suitability for Laf. The projections under the future climate change scenarios showed an increase in the Laf habitat suitability hotspots under the extreme scenario. Main conclusions: This study highlights the potential establishment and distribution in Africa of Las‐associated Huanglongbing and globally for Laf‐associated with African citrus greening disease. The ensemble modelling approach for the distribution of plant pathogens is a valuable tool for the development of strategies for crop protection. These results constitute an early alert for citrus‐producing regions that should inform strategies for surveillance and preventive management against the invasion and spread of this destructive disease. [ABSTRACT FROM AUTHOR]

Summary: Citrus huanglongbing (HLB) is the most devastating citrus disease worldwide. 'Candidatus Liberibacter asiaticus' (Las) is the most prevalent HLB causal agent that is yet to be cultured. Here, we analysed the flagellar genes of Las and Rhizobiaceae and observed two characteristics unique to the flagellar proteins of Las: (i) a shorter primary structure of the rod capping protein FlgJ than other Rhizobiaceae bacteria and (ii) Las contains only one flagellin‐encoding gene flaA (CLIBASIA_02090), whereas other Rhizobiaceae species carry at least three flagellin‐encoding genes. Only flgJAtu but not flgJLas restored the swimming motility of Agrobacterium tumefaciens flgJ mutant. Pull‐down assays demonstrated that FlgJLas interacts with FlgB but not with FliE. Ectopic expression of flaALas in A. tumefaciens mutants restored the swimming motility of ∆flaA mutant and ∆flaAD mutant, but not that of the null mutant ∆flaABCD. No flagellum was observed for Las in citrus and dodder. The expression of flagellar genes was higher in psyllids than in planta. In addition, western blotting using flagellin‐specific antibody indicates that Las expresses flagellin protein in psyllids, but not in planta. The flagellar features of Las in planta suggest that Las movement in the phloem is not mediated by flagella. We also characterized the movement of Las after psyllid transmission into young flush. Our data support a model that Las remains inside young flush after psyllid transmission and before the flush matures. The delayed movement of Las out of young flush after psyllid transmission provides opportunities for targeted treatment of young flush for HLB control. [ABSTRACT FROM AUTHOR]

Wolbachia can profoundly influence the survival, reproduction, and defenses of insect hosts. These interactions could potentially be harnessed for managing pests or insect‐transmitted diseases. Diaphorina citri Kuwayama is a phloem‐feeding pest capable of transmitting the putative causal agent of citrus greening, Candidatus Liberibacter asiaticus (CLas). Like many insects, D. citri is also infected with Wolbachia (wDi). Recent studies indicate that the relative abundance of wDi could be associated with the abundance of CLas, and that wDi may contribute to regulating expression of phage lytic cycle genes in CLas, suggesting the need for better understanding of wDi biology in general. This study investigated the genetic diversity of wDi among D. citri in populations spanning eleven countries and two U.S. territories. Six Wolbachia genes, wsp, coxA, fbpA, ftsZ, gatB, and hcpA, were sequenced and compared across samples. Two prevalent wDi strains were identified across the samples, and screening of clone libraries revealed possible coinfection of wDi strains in specific populations. D. citri mitochondrial cytochrome oxidase subunit I gene (mtCOI) were more divergent between D. citri populations that were infected with different wDi strains or had different infection statuses (single infection vs. coinfection). While we could not eliminate the possibility that maternal transmission may contribute to such patterns, it is also possible that wDi may induce cytoplasmic incompatibility in their host. These findings should contribute to the understanding of wDi population ecology, which may facilitate manipulation of this endosymbiont for management of citrus greening disease worldwide. [ABSTRACT FROM AUTHOR]

Predicting where crop pests and diseases can occur, both now and in the future under different climate change scenarios, is a major challenge for crop management. One solution is to estimate the fundamental thermal niche of the pest/disease to indicate where establishment is possible. Here, we develop methods for estimating and displaying the fundamental thermal niche of pests and pathogens and apply these methods to Huanglongbing (HLB), a vector‐borne disease that is currently threatening the citrus industry worldwide.We derive a suitability metric based on a mathematical model of HLB transmission between tree hosts and its vector Diaphorina citri, and incorporate the effect of temperature on vector traits using data from laboratory experiments performed at different temperatures. We validate the model using data on the historical range of HLB.Our model predicts that transmission of HLB is possible between 16 and 33°C with peak transmission at ~25°C. The greatest uncertainty in our suitability metric is associated with the mortality of the vectors at peak transmission, and fecundity at the edges of the thermal range, indicating that these parameters need further experimental work.We produce global thermal niche maps by plotting how many months each location is suitable for establishment of the pest/disease. This analysis reveals that the highest suitability for HLB occurs near the equator in large citrus‐producing regions, such as Brazil and South‐East Asia. Within the Northern Hemisphere, the Iberian peninsula and California are HLB suitable for up to 7 months of the year and are free of HLB currently.Policy implications. We create a thermal niche map which indicates the places at greatest risk of establishment should a crop disease or pest enter these regions. This indicates where surveillance should be focused to prevent establishment. Our mechanistic method can be used to predict new areas for Huanglongbing transmission under different climate change scenarios and is easily adapted to other vector‐borne diseases and crop pests. [ABSTRACT FROM AUTHOR]

BACKGROUND 'Candidatus Liberibacter asiaticus' is transmitted by Diaphorina citri, an insect with a wide range of hosts in Rutaceae. Species related to Citrus occur in Brazilian forests where they may serve as hosts for psyllids and infested citrus orchards. RESULTS: The suitability of plants as hosts of D. citri was classified into four groups. Group I (high suitability): Citrus × aurantium 'Valencia', 'Citrus limonia', Murraya paniculata (syn. Murraya exotica L.) (Aurantioideae: Aurantieae) and Bergera koenigii (Aurantioideae: Clauseneae). Group II (intermediate to low suitability): Citrus (Poncirus) trifoliata 'Pomeroy', Citrus wintersii, Swinglea glutinosa (Aurantieae) and Clausena lansium (Clauseneae). Group III (not suitable): Aegle marmelos, Atalantia buxifolia, Citrus ('Microcitrus') sp. (Aurantieae) and Helietta apiculata (Amyridoideae). Group IV (non‐hosts): Glycosmis pentaphylla (Clauseneae), Balfourodendron riedelianum, Casimiroa edulis, Esenbeckia febrifuga, Esenbeckia leiocarpa, Metrodorea stipularis, Zanthoxylum rhoifolium (Amyridoideae) and Dictyoloma vandellianum (Cneoroideae). Insects survived longer on newly differentiated leaves compared with fully expanded soft leaves. Psyllids either did not develop or did not survive for long on most Group IV species, all of which, with the exception of G. pentaphylla, occur naturally in Brazilian forests. CONCLUSION: Citrus relatives occurring in forests near citrus orchards are not suitable hosts of D. citri and, therefore, do not contribute to huanglongbing spread. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

'Candidatus Liberibacter asiaticus' (CLas) is associated with the most destructive disease of citrus, huanglongbing (HLB). The most widely used methods for detection of CLas are PCR‐based and require purification of DNA from plant samples. Elution of DNA from tissue prints made on nitrocellulose membranes followed by qPCR (TPE‐qPCR) was compared to DNA extraction of plant tissue followed by PCR (X‐PCR) by testing the same tissue samples. The former estimated a higher CLas population in tissue prints than the latter (t‐test; P = 0.009). All extracts prepared for TPE‐qPCR throughout the experiment were also tested by conventional PCR and 80.8% were identified as positive. A similar set of stem and petiole tissue samples was tested by TPE‐qPCR and immunoassay. Although the detection rate by TPE‐qPCR was higher than by immunoassay, about 6% of tissue prints were positive by immunoassay but not by TPE‐qPCR. Thus, a higher detection rate would be achieved by combining TPE‐qPCR with immunoassay. Significant differences were observed in the performance of nitrocellulose membranes from different manufacturers in these assays. Immunotissue prints showed that the spatial distribution pattern of CLas infection varied widely from one sample to another, but the patterns were highly correlated among serial sections from the same sample, suggesting that CLas preferentially colonizes adjacent phloem cells in a vertical rather than horizontal direction. [ABSTRACT FROM AUTHOR]

BACKGROUND: The African citrus triozid (ACT) Trioza erytreae Del Guercio (Hemiptera: Triozidae) is one of the most devastating pests of citrus with a well‐known role as a vector of the phloem‐limited bacteria (Candidatus Liberibacter africanus) associated with huanglongbing (citrus greening disease), currently considered the world's most serious disease of citrus. Although the pest can successfully develop and reproduce on non‐citrus host plants, there is no documented information on the geometric morphometry of ACT. We determine the effect of host plants on ACT morphometry under controlled laboratory conditions using traditional and geometric analysis. RESULTS: ACT reared on C. limon and Citroncirus spp. was significantly larger than when reared on the other host plant species. ACT reared on C. anisata and C. tangelo was consistently smaller than that reared on M. koenigii and C. sinensis. Based on warped outline drawings, ACT reared on Citroncirus spp. and Murraya koenigii had narrower wings than when reared on C. anisata, C. limon, and C. sinensis with slightly broader wing patterns. CONCLUSION: This study clearly demonstrates that host plant species affect morphometric variation in ACT, which might have a direct impact on fitness parameters of the pest as well as its potential for dispersion. Wing shape and size appear to be useful in separating populations of ACT into different groups. © 2018 Society of Chemical Industry There is lack of information on Trioza erytreae Del Guercio morphometric variation on host species. The implications of these variations on their potential for dispersion and management strategies is discussed. [ABSTRACT FROM AUTHOR]

BACKGROUND: The Asian citrus psyllid, Diaphorina citri Kuwayama, is the primary vector of the phloem‐inhabiting bacterium Candidatus Liberibacter asiaticus putatively responsible for citrus greening (huanglongbing), a devastating citrus disease. Infestations of D. citri frequently develop on Citrus and other genera within the Rutaceae subfamily Aurantioideae including Murraya and Bergera. The genotype Poncirus trifoliata is also a member of the Aurantioideae and readily hybridizes with Citrus spp., but colonization by D. citri is reduced on this genotype. RESULTS: Working with young potted seedlings grown in a greenhouse, we found that the development of D. citri immatures on four P. trifoliata cultivars, especially 'Kryder 55‐5', was slower compared with the development of immatures on the susceptible Citrus macrophylla. In choice assays, adult psyllids exhibited antixenotic behavior towards accessions of P. trifoliata and laid fewer eggs on this genotype compared with C. macrophylla. CONCLUSIONS: Based on reduced oviposition and delays in development, P. trifoliata exhibits a combination of antixenosis and antibiosis host‐plant resistance to D. citri. A companion plant assay showed that the presence of C. macrophylla stimulated higher oviposition rates on P. trifoliata, but nymph development remained retarded on P. trifoliata. Here, we show that the antixenosis associated with trifoliate accessions can be overcome to some extent by the presence of a preferred susceptible host plant; but in combination with antibiosis P. trifoliata remains an inferior host plant. © 2018 Society of Chemical Industry Asian citrus psyllids are the primary vectors of the bacterium responsible for citrus greening disease in Florida and worldwide. This paper discusses the effects of different types of host‐plant resistance exhibited by Poncirus trifoliata (trifoliate citrus) on oviposition and nymphal development of citrus psyllids. [ABSTRACT FROM AUTHOR]

Citrus huanglongbing (HLB) is a severe disease caused by ‘Candidatus Liberibacter asiaticus’ and vectored by Diaphorina citri. In commercial orchards, where the disease is controlled by removal of trees with symptoms and by frequent vector control, epidemics of HLB are mostly driven by primary infections from inoculum sources external to the orchard. It has been previously shown that the density of D. citri populations is usually higher around the edges than the inner sections of HLB‐affected citrus orchards. Consequently, diseased trees are also concentrated on the edges of orchards. However, there is a lack of quantitative data on HLB gradient dynamics over long periods of time. The objective of this study was to characterize temporal HLB progress and spatiotemporal gradient dynamics over six years in a large citrus farm where the disease was managed according to standard recommendations. Disease incidence and annual disease progress rates were higher at the citrus blocks closer to the edge than those located around inner sections of the farm. A decreasing logistic function provided a good description of the spatiotemporal dynamics of HLB gradients. A fixed slope of disease gradients was estimated over time and diseased trees were observed up to 1500 m from the orchard border. These results confirm that primary infection is the main factor for the progress of HLB epidemics in an orchard under the recommended three‐pronged system management. Therefore, HLB management should be intensified at the blocks located on the edges of the orchards and extended to external sources of inoculum. [ABSTRACT FROM AUTHOR]

Abstract: Huanglongbing (HLB), the most destructive citrus disease worldwide, is associated with the bacteria transmitted by the psyllid Diaphorina citri Kuwayama (Hemiptera: Liviidae). One of the main tactics used to manage this disease is the chemical control of D. citri. In this study, the efficacy of four insecticides commonly used by the citrus growers against the D. citri adults was assessed in two vegetative growth stages of flush: young shoots (~ 10 cm in length) and mature leaves. The bioassays were carried out under the greenhouse and field conditions. The insecticides bifenthrin, dimethoate, imidacloprid and thiamethoxam were tested at the recommended rates of 15, 450, 40 and 25 mg/L of water, respectively. The insecticides were sprayed onto the leaves of each stage until runoff. Psyllid mortality was assessed at different periods after insecticide application (0, 1 and 2 weeks). In both greenhouse and field trees, the period of control was shorter in the young shoots than in the mature leaves. In addition, the efficacy of insecticides rapidly decreased in the young shoots over time, mainly for imidacloprid and bifenthrin (less than 1 week). Thus, these results suggest that the number of spray applications to control D. citri should be more frequent during the flushing period. [ABSTRACT FROM AUTHOR]

Abstract: BACKGROUND: Alternative methods that have the potential to reduce the entry of Diaphorina citri Kuwayama (Hemiptera: Liviidae), the major citrus pest worldwide, into commercial groves could be a viable approach for huanglongbing management. Kaolin is an aluminum silicate that when sprayed on plants forms a white particle film that interferes with host recognition by the insects. Diaphorina citri orients towards the host plants by visual and olfactory cues. The purpose of the study was to determine the effect of processed kaolin on D. citri settling (no‐choice) and probing behavior [electrical penetration graph (EPG) technique] under laboratory conditions, and to study its host plant finding ability and dispersal under field conditions in the absence and presence of young shoots. RESULTS: Under laboratory conditions, kaolin caused an overall reduction of 40% in the number of psyllids settled on treated seedlings; furthermore, the proportion of individuals that were able to reach the phloem was 50% lower on kaolin‐treated plants than on untreated plants. In the field, the plant finding ability of D. citri was disrupted on kaolin‐treated trees (overall reduction of 96%), regardless of the vegetative condition, and psyllid dispersal was slower in kaolin‐treated plots than in the untreated control. CONCLUSION: This study clearly demonstrates that processed kaolin interferes negatively with different aspects of the host plant finding ability of D. citri. These findings suggest that processed kaolin has a high potential to reduce huanglongbing primary infection. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Abstract: In managing plant diseases, there is often tension between a regulator seeking to destroy infected plants to prevent further infection on a national scale and growers seeking to retain infected plants to continue obtaining yield. A high‐profile example is Huanglongbing (HLB) or citrus greening, a bacterial disease that threatens Brazil's citrus industry. To prevent the spread of HLB, especially from abandoned infected groves, the government regulated that if 28% of a plantation unit (grove) is found to be symptomatic, then the whole plantation unit must be destroyed. This decision used the best evidence available in 2008, which suggested that a 28% detectable prevalence corresponded with 100% actual prevalence, the disparity being due to asymptomatic infected trees and imperfect detection methods. Using a mathematical model with parameters estimated from field data, we evaluate the assumptions underlying the 28% threshold. We investigate the effects of spraying insecticide and removing diseased plants on the infectious pressure and potential loss of yield from an infected grove. We find that the relationship between detectable and actual prevalence is much wider than allowed for in the regulations. There is a high probability that groves with detectable levels of symptomatic plants substantially below 28% have a >90% prevalence of infected plants. Paradoxically, in a well‐managed orchard, the threshold of 28% may not be reached at 100% prevalence. Infectious pressure from an infected grove is substantially reduced when growers control disease. Individual growers failing to manage disease, therefore, threaten the wider grower community. Control is likely to increase yield and prolong grove productivity, but in some groves may reduce yield. Policy implications. Current disease thresholds aimed at restricting the spread of the citrus disease, Huanglongbing (HLB), in Brazil allow heavily infected groves to remain in the landscape, but lower thresholds would disadvantage growers who are already controlling disease. There is probably no threshold that is optimal for individual growers and regulators but roguing and spraying is beneficial to both parties. Regulations should focus less on prevalence thresholds, and instead encourage early detection and co‐ordinated spraying among growers to control Huanglongbing on a regional level. [ABSTRACT FROM AUTHOR]

Abstract: The incidence of a bacterium “Candidatus Liberibacter asiaticus” was assessed in a Florida population of Asian citrus psyllid, Diaphorina citri. The bacterium is the presumed causal agent of Asiatic huanglongbing, a serious citrus disease. Adult D. citri were periodically collected between May 2010 and September 2012 in a block of diseased trees located in east‐central Florida. The psyllids were individually subjected to molecular analyses (quantitative polymerase chain reaction assays using HLBaspr primers) to determine whether the bacterium was present and, if so, the population level of the pathogen based on qPCR cycle threshold (CT) values. Significantly greater percentages of females tested positive for the pathogen than males, but there were no significant differences between females and males with respect to population levels of the pathogen within the psyllids. No significant differences were found among the three D. citri colour morphs with respect to percentages of adults testing positive for the pathogen. Among 47 sample dates, a mean of 17.5% of adults per sample date tested positive (CT < 36) for the pathogen with a mean CT value of 31.1. The incidence of the pathogen was generally higher during late fall or early winter and often lower during mid‐ to late summer. There was a significant negative correlation between percentages of D. citri testing positive and air temperature. Increases in the incidence of the pathogen may not necessarily correspond to increases in transmission, as a number of factors both internal and external to D. citri can influence transmission. Transmission rates may be highest during periods when D. citri infestation levels are large, a high percentage of adults carry a high population of the pathogen in their salivary glands, and citrus flush is abundant. [ABSTRACT FROM AUTHOR]

Abstract: Environmental factors that influence flight activity of Diaphorina citri Kuwayama (Hemiptera: Liviidae) may have implications for Huanglongbing spread and management. In this work, four studies were conducted to evaluate the effect of environmental conditions on D. citri take‐off. In the first, insects were transferred to sweet orange seedlings and confined inside an acrylic cage to verify the take‐off periodicity and the effect of environmental factors on this process. In the second, take‐off temperature threshold was estimated by recording the number of insects that initiated flight from a platform when subjected to gradual temperature increases from 15 to 39°C. In the other studies, we evaluated the effect of different photoperiods and temperature regimes (third study) and of constant temperatures (fourth study) on the propensity for D. citri flight. Insects were confined in clear plastic bottle cages with tubes of 50 ml placed on the cab, to collect emerged adults that initiated flights. Results showed that a small portion of the tested population (maximum 10%) tends to take off from plants and this behaviour is more prevalent in the afternoon (14:00–16:00 h), coinciding with daytimes of lower humidity and higher thermal amplitude. Adults that were submitted to lower temperatures (18°C) and short light periods (10 h) showed less propensity to flight. In contrast, at constant 27°C, the insects were more prone to flight, and this result was confirmed when individuals were submitted to increases in temperature, indicating that 27.14°C is the take‐off temperature threshold of D. citri. Results show that temperature plays an important role in the flight activity of D. citri and suggest that control measures of the insect may be more effective in the morning and in temperatures below 27°C, when the probability to take off from a host is lower. [ABSTRACT FROM AUTHOR]

Abstract: The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), is a major citrus pest that transmits the bacteria Candidatus Liberibacter asiaticus and Ca. L. americanus associated with huanglongbing (HLB) disease. Diaphorina citri population densities can affect the effectiveness of its monitoring and sampling methods. Thus, we compared different methods for adult D. citri monitoring in groves with and without insecticide application programmes. Four short‐term experiments were carried out, each one lasting four consecutive weeks. In these experiments, sticky cards with different colours (yellow, light green, green and dark green), sweep net, two suction device models, visual inspection and stem tap sampling were assessed. Two long‐term experiments were conducted for 4.5 and 5 years, in which only yellow sticky card and visual inspection for D. citri monitoring were assessed. For the short‐term experiments, psyllids were detected by all monitoring methods during all sampling periods in areas without chemical control. However, in areas with psyllid control via fortnightly and monthly applications of insecticides, only sticky cards, regardless of their colour, were able to detect the presence of D. citri. Similarly, for the long‐term experiments, yellow sticky cards were more effective than visual inspection for detecting and quantifying D. citri in all areas with or without insecticide application. Therefore, in areas where HLB is present and chemical control of psyllid is required, sticky cards are the most reliable option for monitoring D. citri. [ABSTRACT FROM AUTHOR]

BACKGROUND Control of the Asian citrus psyllid Diaphorina citri Kuwayama, the most important pest of citrus worldwide, is based on the use of insecticides, though unsatisfactory results have recently been reported. In this study, insecticide resistance of D. citri to three insecticides (bifenthrin, malathion, and chlorpyrifos) was examined. RESULTS Three populations (designated Dci-CParácuaro, Dci-El Junco, and Dci-Antúnez) of both adults and fourth-instar D. citri individuals were collected in 2014 at two different times and on one occasion, respectively, from three locations (Crucero de Parácuaro, El Junco, and Antúnez). These locations represent the major commercial Mexican lemon production areas in the Apatzingán Valley in the state of Michoacán, Mexico. The three populations of D. citri adults and fourth-instar nymphs at the different collection times showed low levels of resistance (≤7-fold) to bifenthrin, but were very resistant to malathion (≤345- and ≤432-fold for adults and fourth instars, respectively) and chlorpyrifos (≤2435- and ≤1424-fold for adults and fourth instars, respectively). CONCLUSION Resistance levels to the tested insecticides were highly variable but homogeneous among seasons and localities. Resistance management programmes that include crop sanitation, use of biological and cultural control practices, and rotation of insecticide classes should be established, particularly in areas where D. citri has developed resistance to malathion and chlorpyrifos. © 2017 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

The use of proper management strategies for citrus huanglongbing ( HLB), caused by ' Candidatus Liberibacter asiaticus' (Las) and transmitted by Asian citrus psyllid ( ACP) ( Diaphorina citri), is a priority issue. HLB control is based on healthy seedlings, tolerant rootstock cultivars and reduction of ACP populations. Here, dynamic populations of Las in different citrus hosts and each instar of ACP were studied, together with the seasonal growth and distribution of Las in different tissues, using conventional and TaqMan real-time PCR. Different levels of susceptibility/tolerance to HLB were seen, resulting in different degrees of symptom severity and growth effects on hosts or rootstocks. Troyer citrange, Swingle citrumelo and wood apple were highly tolerant among 11 rootstock cultivars. Regarding distribution and seasonal analysis of Las, mature and old leaves contained high concentrations in cool temperatures in autumn and spring. Las was detected earlier through psyllid transmission than through graft inoculation, and the amounts of Las ( AOL) varied in different hosts. Thus, different AOL (104-107 copy numbers μL−1) and Las-carrying percentages ( LCP; 40-53.3%) were observed in each citrus cultivar and on psyllids, respectively. Furthermore, both AOL and LCP were lower in nymphs than in adult psyllids, whereas the LCP of psyllids were not affected by increasing the acquisition-access time. The present study has significant implications for disease ecology. The combination of early detection, use of suitable rootstocks and constraint of psyllid populations could achieve better management of HLB disease. [ABSTRACT FROM AUTHOR]

Asian citrus psyllid Diaphorina citri is the vector of the citrus Huanglongbing ( HLB) associated bacterial agent ' Candidatus Liberibacter asiaticus' ( C Las). The molecular interactions between C Las and D. citri remain unclear. In the present study, protein profiles of mitochondrial, microsomal and cytosolic fractions from uninfected and CLas-infected adult D. citri are investigated using two-dimensional gel electrophoresis. The comparative analysis reveals a total of 18, 24 and 20 protein spots that are unique or differentially expressed in mitochondrial, microsomal and cytosolic proteins fractions respectively. These proteins are successfully identified by mass spectrometry. Among the 62 identified proteins, 30 are up-regulated, whereas 32 are down-regulated. These proteins include important components in energy metabolism such as ATP synthase, ATPase, ATP/ ADP carrier protein, etc.; host stress responses such as heat shock proteins; host detoxification processes (i.e., cytochrome P450 and glutathione S-transferase); and the cytoskeleton (such as actin, tubulin, myosin and tropomyosin). These data suggest that, after C Las infection, several proteins of D. citri, especially energy metabolism and protein biosynthesis, are altered, and extensive host defence responses are induced. In conclusion, the present study reports proteomic information that is helpful in understanding the vector-pathogen relationship between C Las and D. citri, and could be used to identify potential targets for limiting the spread of C Las, as well as to provide new insights into HLB management. [ABSTRACT FROM AUTHOR]

Plants can defend themselves against herbivores through activation of defensive pathways and attraction of third-trophic-level predators and parasites. Trophic cascades that mediate interactions in the phytobiome are part of a larger dynamic including the pathogens of the plant itself, which are known to greatly influence plant defenses. As such, we investigated the impact of a phloem-limited bacterial pathogen, Candidatus Liberibacter asiaticus ( CLas), in cultivated citrus rootstock on a well-studied belowground tritrophic interaction involving the attraction of an entomopathogenic nematode ( EPN), Steinernema diaprepesi, to their root-feeding insect hosts, Diaprepes abbreviatus larvae. Using belowground olfactometers, we show how CLas infection interferes with this belowground interaction by similarly inducing the release of a C12 terpene, pregeijerene, and disconnecting the association of the terpene with insect presence. D. abbreviatus larvae that were not feeding but in the presence of a CLas-infected plant were more likely to be infected by EPN than those near uninfected plants. Furthermore, nonfeeding larvae associated with CLas-infected plants were just as likely to be infected by EPN as those near noninfected plants with D. abbreviatus larval damage. Larvae of two weevil species, D. abbreviatus and Pachnaeus litus, were also more attracted to plants with infection than to uninfected plants. D. abbreviatus larvae were most active when exposed to pregeijerene at a concentration of 0.1 μg/μl. We attribute this attraction to CLas-infected plants to the same signal previously thought to be a herbivore-induced plant volatile specifically induced by root-feeding insects, pregeijerene, by assessing volatiles collected from the roots of infected plants and uninfected plants with and without feeding D. abbreviatus. Synthesis. Phytopathogens can influence the structuring of soil communities extending to the third trophic level. Field populations of EPN may be less effective at host-finding using pregeijerene as a cue in citrus grove agroecosystems with high presence of CLas infection. [ABSTRACT FROM AUTHOR]

Diaphorina citri, known as the Asian citrus psyllid ( ACP), is the insect vector of a devastating citrus disease (huanglongbing; HLB), which has caused billions of dollars in damage in Florida since its detection in 2005. Data from long-term monitoring programs in two Florida citrus groves were used to assess ACP demography and population ecology, which is needed to implement more effective management strategies for HLB. Seasonal patterns and correlations between ACP density estimates and a suite of environmental and community indicators, previously shown to influence ACP demography, were described and interpreted. Data were evaluated for evidence for spatial clustering and density-dependent recruitment of ACP in a major outbreak event using Taylor power law analysis and by fitting a stochastic Beverton-Holt recruitment model using state-space approach. Strong evidence for density-dependent recruitment and spatial clustering was found and should be considered for future modelling work as it can greatly influence ACP populations by affecting their growth rates, dispersal behaviour and morphology, as well as HLB transmission. Observations of ACP density and dispersion in space and time, along with the estimated parameters from the Beverton-Holt model, suggested that intraspecific competition for resources may initiate both local, then widespread dispersal process, thus affecting grove-wide and area-wide HLB transmission. When these results were synthesised with those of parallel studies, the removal of several of these regulating factors in a single year could lead to widespread disease of the entire crop in a grove, and likely surrounding groves as well. We provide the first field evidence of the consequences of back-to-back ACP colonisation on the rapid spread of the HLB in a grove (a mechanism previously demonstrated in laboratory settings). We stress the importance of proper integrated pest management and area-wide management to prevent such outbreaks. [ABSTRACT FROM AUTHOR]

The Asian citrus psyllid Diaphorina citri Kuwayama (Hemiptera: Liviidae) is an economically important pest of citrus throughout Asia and the Americas because it transmits Candidatus Liberibacter asiaticus ( CLas), which is the presumed causal agent of citrus greening disease., We investigated whether biotic and abiotic characteristics can be used to predict Diaphorina citri population abundance and assessed whether agricultural intensity explained the distribution of D. citri populations during winter dormant periods (December to March)., Over two consecutive winters, we examined the abundance of D. citri in groves throughout Florida in response to four different management regimes, defined as: conventional, intermittent, unmanaged and organic., During both years, the winter abundance of D. citri in groves with intermittent management was greater than in groves with other management regimes. Latitude and row orientation both had a significant effect on psyllid density during winter. Diaphorina citri abundance was higher when more than 20% of the surrounding landscape was urbanized., These findings suggest that only conventional management of groves reduced D. citri populations during winter periods. By contrast, intermittent management was associated with higher D. citri populations. These results might be of some concern in light of the economic and environmental costs of repeated insecticide applications and the dramatic decline of citrus production in the U.S.A. [ABSTRACT FROM AUTHOR]