Psyllid Vector Research Articles

Peptide conjugated morpholinos for management of the huanglongbing pathosystem.

'Candidatus Liberibacter asiaticus' (CLas) is the causal agent of the devastating citrus disease Huanglongbing (HLB) and is transmitted by the insect vector Diaphorina citri (Hemiptera: Liviidae). A potential approach for treating CLas infection is the use of synthetic nucleic acid-like oligomers to silence bacterial gene expression. Peptide conjugated morpholinos (PPMOs) targeting essential genes in CLas and the psyllid vector's endosymbiotic bacteria, Wolbachia (-Diaphorina, wDi), were evaluated using in vitro and in vivo assays. Expression of the wDi gyrA gene was significantly reduced following incubation of wDi cells with PPMOs. In addition, the viability of isolated wDi cells was greatly reduced when treated with PPMOs as compared to untreated cells. Feeding D. citri adults with a complementary PPMO (CLgyrA-14) showed significantly reduced (70% lower) expression of the CLas gyrA gene. CLas relative density was significantly lower in the psyllids fed with CLgyrA-14, when compared to untreated insects. Psyllids that were treated with CLgyrA-14 were less successful in transmitting the pathogen into uninfected plants, compared to untreated insects. The expression of essential genes in the D. citri symbiont, wDi and the HLB pathogen were suppressed in response to PPMO treatments. This study demonstrates the potential of PPMOs as a novel strategy for management of bacterial pathogens of fruit trees, such as HLB. © 2020 Society of Chemical Industry.

Effects of Potato Psyllid Vector Density and Time of Infection on Zebra Chip Disease Development after Harvest and during Storage

“Candidatus Liberibacter solanacearum” (Lso) (=“Candidatus Liberibacter psyllarous”) is an uncultured, phloem-limited bacterium that is associated with zebra chip disease (ZC) in potato (Solanum tuberosum L.) and transmitted by the potato psyllid (Bactericera cockerelli (Šulc)). Vector density and timing of infection have been shown to affect ZC prevalence at harvest; however, little work has been done on disease development during storage. Here we confirm with field-cage trials that ZC prevalence at harvest was greater with increased time between inoculation and vine kill. Moreover, we show that with Pacific Northwest growing conditions, ZC can develop over time during storage. Plants inoculated 2 to 3 weeks before vine kill showed little or no ZC symptoms in tubers at harvest, but higher prevalence of symptoms after 3 months in storage. For plants inoculated at 4 to 5 weeks before vine kill, tubers exhibited notable symptoms at harvest, but still showed evidence of symptom development after storage. Plants inoculated within 1 week before vine kill exhibited little or no risk of ZC in tubers at harvest or after storage. Higher vector density tended to contribute to ZC prevalence, but was far less important than timing of infection. These results underscore the potential danger of underestimating ZC prevalence at harvest for tubers being stored long term, and suggest that plants at risk of Lso infection should be protected from potato psyllids until at least 2 weeks before vine kill.

Multi-scale spatial genetic structure of the vector-borne pathogen \u2018Candidatus Phytoplasma prunorum\u2019 in orchards and in wild habitats

Inferring the dispersal processes of vector-borne plant pathogens is a great challenge because the plausible epidemiological scenarios often involve complex spread patterns at multiple scales. The spatial genetic structure of ‘Candidatus Phytoplasma prunorum’, responsible for European stone fruit yellows disease, was investigated by the application of a combination of statistical approaches to genotype data of the pathogen sampled from cultivated and wild compartments in three French Prunus-growing regions. This work revealed that the prevalence of the different genotypes is highly uneven both between regions and compartments. In addition, we identified a significant clustering of similar genotypes within a radius of 50 km or less, but not between nearby wild and cultivated Prunus. We also provide evidence that infected plants are transferred between production areas, and that both species of the Cacopsylla pruni complex can spread the pathogen. Altogether, this work supports a main epidemiological scenario where ‘Ca. P. prunorum’ is endemic in — and generally acquired from — wild Prunus by its immature psyllid vectors. The latter then migrate to shelter plants that epidemiologically connect sites less than 50 km apart by later providing infectious mature psyllids to their “migration basins”. Such multi-scale studies could be useful for other pathosystems.

UV reflective properties of magnesium oxide increase attraction and probing behavior of Asian citrus psyllids (Hemiptera: Liviidae)

Asian citrus psyllid (Diaphorina citri) vectors the bacterium Candidatus Liberibacter asiaticus, the causal pathogen of citrus greening disease that is devastating citrus industries worldwide. Suppressing psyllid populations is crucial to prevent disease spread. An attract-and-kill trap based on psyllid behavior would fill a niche for monitoring and control. To optimize visual attraction of psyllids, the ultraviolet (UV) reflective properties of magnesium oxide (MgO) and/or barium sulfate (BaSO4) were assessed for potential application to a trap surface. Under low UV, high UV and natural sunlight conditions, the reflectance, attraction, and probing behaviors of psyllids were evaluated on surfaces containing magnesium oxide or barium sulfate. Magnesium oxide added to yellow sticky traps enhanced visual response of D. citri. Probing assays demonstrated that magnesium oxide alone or as a mixture with a phagostimulant blend, increased the UV reflectance of substrates, as well as, attraction and probing by psyllids. Results demonstrated that psyllids respond to both short (UV) and long (yellow) wavelengths during orientation, and that these inert compounds can increase UV reflectance and improve attractiveness of an attract-and-kill device.

In vitro selection to increase Huanglongbing tolerance of citrusderived from in vitro breeding

Until now, Huanglongbing-HLB disease caused by a simple bacterium (Ca. Liberibacter spp) and distributed by Asian citrus psyllid vector (Diaphorina citri) and clonal propagation (grafting), was a major challenge in the national citrus production even almost all citrus nation production center in Asia, Africa also at the USA. This disease attack at phloem vessels by callouses forming that covering plasmodesmata and could be crop failure. There have been no citrus varieties reported resistant to HLB attack. Plant ploidy increase derived from in vitro breeding through somatic hybridization and endosperm culture expected to increase the plant’s defense system. The aim of this study was to evaluate the tolerance of polyploidy plant to HLB attack. Tolerance of in vitro shoot of citrus regenerated from protoplast fusion (Citrus nobilis + C. unshiu) and culture of tangerine (C. nobilis) endosperm were screened by HLB pathogen suspense as selecting an agent and local lime as a negative control (HLB susceptible citrus). In vitro selection result that in vitro shoots derived from somatic hybridization shows resistance increase to HLB and characterize by recoverability after were selected.

The Enemy is Outside: Releasing the Parasitoid Tamarixia radiata (Hymenoptera: Eulophidae) in External Sources of HLB Inocula to Control the Asian Citrus Psyllid Diaphorina citri (Hemiptera: Liviidae).

Huanglongbing (HLB), the most destructive citrus disease worldwide, was first recorded in Brazil in 2004, and since then, more than 50 million trees identified with this disease have been eliminated. The disease is managed mainly by controlling the psyllid vector Diaphorina citri Kuwayama, 1908 (Hemiptera: Liviidae). Although the presence of the insect in commercial citrus groves is low, HLB infection rates increase in areas bordering the groves. The disease is transmitted by psyllids from host citrus plants in areas outside the managed groves, such as abandoned or organic groves and residential trees, and from orange jasmine plants in urban settings. In order to provide information to support HLB control, this study evaluated the biotic and abiotic variables that affect the dynamics of D. citri populations after releases of the parasitoid wasp Tamarixia radiata (Waterston, 1922) (Hymenoptera: Eulophidae) in external sources of HLB inocula. The study was divided into two parts. After releasing the parasitoids in non-commercial areas, we determined the following: (a) the variables that significantly affected the number of nymphs collected on shoots in the same non-commercial area; (b) the variables that significantly affected the number of adult psyllids collected in a neighboring commercial citrus area. Our results indicated that the number of nymphs in external areas was affected only by the host plant and rainfall. However, periodic parasitoid releases significantly reduced the number of adult psyllids collected in the commercial area. The results indicate that the release of parasitoids in external sources of inocula has the potential to maximize actions for D. citri control, contributing to the reduction of psyllid populations in commercial areas. Consequently, this strategy may help to manage the disease infection without an increase in insecticide use.

Immuno-Ultrastructural Localization and Putative Multiplication Sites of Huanglongbing Bacterium in Asian Citrus Psyllid Diaphorina citri.

Huanglongbing, the most destructive citrus disease worldwide, is caused by the bacterium ‘Candidatus Liberibacter asiaticus’ (CLas) and is vectored by the Asian citrus psyllid (ACP). Very little is known about the form and distribution of CLas in infected psyllids, especially at the ultrastructural level. Here, we examined these aspects by transmission electron microscopy, combined with immunogold labeling. In CLas-exposed ACP adults, the CLas bacterial cells were found to be pleomorphic taking tubular, spherical, or flask-shaped forms, some of which seemed to divide further. Small or large aggregates of CLas were found in vacuolated cytoplasmic pockets of most ACP organs and tissues examined, including the midgut, filter chamber, hindgut, Malpighian tubules, and secretory cells of the salivary glands, in addition to fat tissues, epidermis, muscle, hemocytes, neural tissues, bacteriome, and walls of the female spermatheca and oviduct. Large aggregates of CLas were found outside the midgut within the filter chamber and between the sublayers of the basal lamina of the hindgut and Malpighian tubules. Novel intracytoplasmic structures that we hypothesized as ‘putative CLas multiplication sites’ were found in the cells of the midgut, salivary glands, and other tissues in CLas-exposed ACP. These structures, characterized by containing a granular matrix and closely packed bacterial cells, were unbound by membranes and were frequently associated with rough endoplasmic reticulum. Our results point to the close association between CLas and its psyllid vector, and provide support for a circulative-propagative mode of transmission.

Latency and incubation of ‘Candidatus Liberibacter asiaticus’ in citrus after vector inoculation

Huanglongbing (HLB) is a bacterial disease of citrus, transmitted by the Asian citrus psyllid Diaphorina citri. In Brazil, the disease is managed by insecticide spray targeting vector control and removal of trees when they first show HLB symptoms that aims to reduce ‘Candidatus Liberibacter asiaticus’ (CLas) inoculum within the orchard. Albeit symptoms of HLB in field trees appear only months or years after the initial infection, the shortest incubation period (time between pathogen infection and symptom expression) reported for CLas in citrus was about 4 months after inoculation in studies in which this pathogen was graft-inoculated. In this work, an incubation period of 80 days (2.5 months) was recorded for citrus seedlings and 5 months for citrus nursery plants after inoculation with the psyllid vector itself. Psyllids were capable of acquire CLas from the infected seedlings and nursery plants 2.5 months after inoculation. These results show that the latency of CLas in planta is shorter than the incubation. Assessment of the presence of liberibacter in psyllids could be used as a proxy for the presence of HLB bacterium in presymptomatic stage of the plants.

Mycorrhization Mitigates Disease Caused by "Candidatus Liberibacter solanacearum" in Tomato.

Disease caused by the bacterial pathogen “Candidatus Liberibacter solanacearum” (Lso) represents a serious threat to solanaceous crop production. Insecticide applications to control the psyllid vector, Bactericera cockerelli Šulc (Hemiptera: Triozidae) has led to the emergence of resistance in psyllids populations. Efforts to select natural resistant cultivars have been marginally successful and have been complicated by the presence of distinct Lso haplotypes (LsoA, LsoB) differing in symptoms severity on potato and tomato. A potentially promising management tool is to boost host resistance to the pathogen and/or the insect vector by promoting mycorrhization. Here we tested the hypothesis that mycorrhizal fungi can mitigate the effect of Lso infection on tomato plants. The presence of mycorrhizal fungi substantially delayed and reduced the incidence of Lso-induced symptoms on tomato as compared to non-mycorrhized plants. However, PCR with specific Lso primers revealed that mycorrhization did not prevent Lso transmission or translocation to newly formed leaves. Mycorrhization significantly reduced oviposition by psyllids harboring LsoA and survival of nymphs from these eggs. However, mycorrhization had no effect on oviposition by psyllids harboring LsoB or the survival of nymphs from parents harboring LsoB. These findings indicate the use of mycorrhizal fungi is a promising strategy for the mitigation of disease caused by both LsoA and LsoB and warrants additional field testing.

Nutrient Status and Root Density of Huanglongbing-Affected Trees: Consequences of Irrigation Water Bicarbonate and Soil pH Mitigation with Acidification

The Candidatus Liberibacter asiaticus bacterium, associated with Huanglongbing (HLB) disease of citrus trees, moves downward in the phloem and infects the roots soon after transmission by the Asian Citrus Psyllid (Diaphorina citri) vector into shoots. Before canopy symptoms appear, 30–50% of the roots are damaged. Without aggressive management to reduce abiotic and biotic stress, root loss increases to 70–80%. An extensive survey of HLB-affected groves in central and south-central Florida indicated that a greater decline in fibrous root health as well as a greater expression of HLB symptoms is observed where irrigation water is high in bicarbonates (>100 mg L−1) and soil pH is >6.5. Over three seasons of survey, acidification of irrigation water in the central and south-central citrus growing regions of Florida reduced the decline in root density associated with HLB. Irrigation water treatment with sulfuric acid and soil amendment with elemental sulfur for 36 months to establish a soil pH range from 4.0 to 7.0 increased root growth, soil nutrient availability, and the uptake of Ca, Mg, Mn, and Zn in response to a gradual reduction in soil pH in young and mature Valencia orange groves on Swingle citrumelo rootstock. The reduction in soil pH increased yield and soluble solids in fruit and so would improve citrus production.

A Transcriptomics Approach Reveals Putative Interaction of Candidatus Liberibacter Solanacearum with the Endoplasmic Reticulum of Its Psyllid Vector.

Candidatus Liberibacter solanacerum (CLso), transmitted by Bactericera trigonica in a persistent and propagative mode causes carrot yellows disease, inflicting hefty economic losses. Understanding the process of transmission of CLso by psyllids is fundamental to devise sustainable management strategies. Persistent transmission involves critical steps of adhesion, cell invasion, and replication before passage through the midgut barrier. This study uses a transcriptomic approach for the identification of differentially expressed genes with CLso infection in the midguts, adults, and nymphs of B. trigonica and their putative involvement in CLso transmission. Several genes related to focal adhesion and cellular invasion were upregulated after CLso infection. Interestingly, genes involved with proper functionality of the endoplasmic reticulum (ER) were upregulated in CLso infected samples. Notably, genes from the endoplasmic reticulum associated degradation (ERAD) and the unfolded protein response (UPR) pathway were overexpressed after CLso infection. Marker genes of the ERAD and UPR pathways were also upregulated in Diaphorina citri when infected with Candidatus Liberibacter asiaticus (CLas). Upregulation of the ERAD and UPR pathways indicate induction of ER stress by CLso/CLas in their psyllid vector. The role of ER in bacteria–host interactions is well-documented; however, the ER role following pathogenesis of CLso/CLas is unknown and requires further functional validation.

First report of Candidatus Phytoplasma prunorum, the European stone fruit yellows phytoplasma on peach trees on the territory of Canton of Geneva, Switzerland

In recent years, ‘Ca. P. prunorum’, the agent of ESFY was reported from several apricot orchards of Canton of Wallis, the main apricot production region in Switzerland (Genini and Ramel, 2004). The psyllid vector Cacopsylla pruni was also found in several locations in the Lake Geneva area (Ackermann et al., 2006). The presence of the disease and of its proven vector C. pruni at the eastern part of the Lake Geneva area, as well as the existing risk of dissemination of ESFY to other stone fruit orchards along the Lake Geneva are the reasons for which ESFY needs to be studied further. ESFY and its possible dissemination through C. pruni on the territory of Canton of Geneva, bordering France, has never been yet studied. In 2016, visual observations were conducted in several stone fruit orchards near Geneva. Plant material was obtained from peach trees, displaying some of the typical symptoms such premature leaf colouration, leaf-roll, tree decline (Sabaté et al., 2015), in the autumn, when the concentration of the phytoplasma in the upper parts of the trees is the highest. Phloem was prepared from branches and was extracted with a CTAB-based adapted protocol (Lefort and Douglas, 1999). PCR amplification of phytoplasma DNA was achieved with the universal primers: fP1/rP7 (Deng and Hiruki, 1991; Schneider et al., 1995). All positive samples were tested with the ESFY-specific non-ribosomal primers ECA1/ECA2 (Jarausch et al., 1998). Typical symptoms of ESFY, like premature leaf colouration, leaf yellowing with reddish edges, leaf-roll, severe chlorosis, die-back of top branches and partial or, complete decline of the trees (Figure 1) were found in a peach orchard (GPS coordinates: 46°15'17.4"N 6°12'40.7"E) located in the area of Collonge-Bellerive, close to Geneva city. Ten trees were sampled in this orchard. The presence of ‘Ca. P. prunorum’ was confirmed in two of them (Figure 2 and Figure 3). At the beginning of spring 2017, the monitoring of the ESFY symptoms in the infested orchard was resumed. Early bud break was found on the two infected trees and the infection was confirmed again by PCR (Figure 4 and Figure 5). The same symptoms were observed in many trees of this orchard, which correlated with the previous autumn observations. Trapping confirmed the presence of the insect vector Cacopsylla pruni (Bodnár et al., 2018) in the infected area. In order to understand more about the origin and the diversity of ESFY phytoplasma in this area, additional plant and insect samples will be analyzed. The correlation between symptoms and infected plants will be studied further. We report here for the first time on the occurrence of ‘Candidatus Phytoplasma prunorum’ (‘Ca. P. prunorum’) the agent of ESFY on the territory of Canton of Geneva.

Prevalence of 'Candidatus Liberibacter solanacearum' Haplotypes in Potato Tubers and Psyllid Vectors in Idaho From 2012 to 2018.

'Candidatus Liberibacter solanacearum' (Lso) is an uncultured, phloem-associated bacterium causing a severe tuber disease in potato called zebra chip (ZC). Seven haplotypes of Lso have been described in different hosts, with haplotypes A and B found associated with infections in potato and tomato. In the field, Lso is transmitted by the potato psyllid (Bactericera cockerelli), and between 2011 and 2015, a significant change in Lso haplotype prevalence was previously reported in Idaho: from exclusively A haplotype found in tested psyllids in 2012 to mainly B haplotype found in collected psyllids in 2015. However, prevalence of Lso haplotypes in Idaho was not analyzed in potato tubers exhibiting symptoms of ZC. To fill in this knowledge gap, prevalence of Lso haplotypes was investigated in potato tubers harvested in southern Idaho between 2012 and 2018, and it was found to change from exclusively A haplotype in the 2012 season to an almost equal A and B haplotype distribution during the 2016 season. During the same period, haplotype distribution of Lso in psyllid vectors collected using yellow sticky traps also changed, but in psyllids, the shift from A haplotype of Lso to B haplotype was complete, with no A haplotype detected in 2016 to 2018. The changes in the haplotype prevalence of the Lso circulating in potato fields in southern Idaho may be, among other factors, responsible for a decrease in the ZC incidence in Idaho potato fields between an outbreak of the disease in 2012 and a very low level of ZC afterward.

New Ca. Liberibacter psyllaurous haplotype resurrected from a 49-year-old specimen of Solanum umbelliferum: a native host of the psyllid vector

Over the last century, repeated emergence events within the Candidatus Liberibacter taxon have produced pathogens with devastating effects. Presently, our knowledge of Ca. Liberibacter diversity, host associations, and interactions with vectors is limited due to a focus on studying this taxon within crops. But to understand traits associated with pathogen emergence it is essential to study pathogen diversity in wild vegetation as well. Here, we explore historical native host plant associations and diversity of the cosmopolitan species, Ca. L. psyllaurous, also known as Ca. L. solanacearum, which is associated with psyllid yellows disease and zebra chip disease, especially in potato. We screened tissue from herbarium samples of three native solanaceous plants collected near potato-growing regions throughout Southern California over the last century. This screening revealed a new haplotype of Ca. L. psyllaurous (G), which, based on our sampling, has been present in the U.S. since at least 1970. Phylogenetic analysis of this new haplotype suggests that it may be closely related to a newly emerged North American haplotype (F) associated with zebra chip disease in potatoes. Our results demonstrate the value of herbarium sampling for discovering novel Ca. Liberibacter haplotypes not previously associated with disease in crops.

African Citrus Greening Disease in East Africa: Incidence, Severity, and Distribution Patterns.

African citrus greening disease (ACGD) is considered as one of the major diseases of citrus threatening citrus production in East Africa. Our study aimed for the first time to assess the incidence, severity, and distribution patterns of ACGD in Kenya and Tanzania. In total, 105 citrus orchards were assessed in 13 regions representing low, mid, and high altitude areas. In each backyard and orchard, trees were randomly selected and rated for visual ACGD symptoms; then leaves and insect samples collected for analysis of 'Candidatus Liberibacter africanus' (CLaf), the presumptive causal agent of ACGD. Endpoint PCR, sequencing, and molecular phylogenetic tools were employed to confirm the identity of potential circulating pathogens. Incidence and severity of ACGD varied significantly among the different regions. Both Trioza erytreae (Del Guerico) (Hemiptera: Triozidae) and the invasive Asian citrus psyllid vector Diaphorina citri (Kuwayama) (Hemiptera: Liviidae) were found to co-occur in upper and lower midland regions. Molecular characterization identified 'Candidatus Liberibacter africanus spp. Clausenae' (CLafCl) as the main causal agent of ACGD in most of the citrus plants and insect samples. No instances of Candidatus Liberibacter asiaticus infection were found. These findings provide valuable insights into understanding and management of ACGD by employing stringent and early disease detection tools to curb the spread of the disease.

The infection of its insect vector by bacterial plant pathogen "Candidatus Liberibacter solanacearum" is associated with altered vector physiology

Many bacterial and viral plant pathogens are transmitted by insect vectors, and pathogen-mediated alterations of plant physiology often influence insect vector behavior and fitness. It remains largely unknown for most plant pathogens whether, and how, they might directly alter the physiology of their insect vectors in ways that promote pathogen transmission. Here we examined whether the presence of “Candidatus Liberibacter solanacearum” (“Ca. L. solanacearum”), an obligate bacterial pathogen of plants and of its psyllid vector alters the physiochemical environment within its insect vector, the potato psyllid (Bactericera cockerelli). Microelectrodes were used to measure the local pH and oxygen tension within the abdomen of “Ca. L. solanacearum”-free psyllids and those infected with “Ca. L. solanacearum”. The hemolymph of infected psyllids had higher pH at 9.09 ± 0.12, compared to “Ca. L. solanacearum”-free psyllids (8.32 ± 0.11) and a lower oxygen tension of 33.99% vs. 67.83%, respectively. The physicochemical conditions inside “Ca. L. solanacearum”-free and –infected psyllids body differed significantly with the infected psyllids having a higher hemolymph pH and lower oxygen tension than “Ca. L. solanacearum”-free psyllids. Notably, the bacterial titer increased under conditions of higher pH and lower oxygen tension values. This suggests that the vector’s physiology is altered by the presence of the pathogen, potentially, resulting in a more conducive environment for “Ca. L. solanacearum” survival and subsequent transmission.

Screening Sticky Cards as a Simple Method for Improving Efficiency of Diaphorina citri (Hemiptera: Liviidae) Monitoring and Reducing Nontarget Organisms.

Management of Diaphorina citri Kuwayama (Hemiptera: Liviidae) populations is one of the major strategies for reducing the spread and incidence of huanglongbing (HLB). HLB is putatively caused by Candidatus Liberibacter spp. (Rhizobiales: Phyllopbacteriaceae) that are transmitted to citrus by psyllid vectors. Diaphorina citri population monitoring is done to detect its presence and inform on management decisions. Various methods are used for detecting and estimating D. citri densities but trapping with yellow or lime-green sticky cards has proven to be the most effective method. These sticky cards rely on the color preference of adult D. citri, but many flying organisms are attracted to the same color spectrum as psyllids. Hence, in field situations, sticky traps are hampered by large numbers of bycatches of nontarget organisms and debris. Here, we described a method using a mesh laid on the surface of traps as a sift to catch mainly psyllids, while reducing bycatches. By filtering D. citri through this mesh, they can be counted more rapidly and accurately. Although mesh-covered traps captured 5-15% less D. citri relative to uncovered ACP traps, both types of traps statistically agreed on D. citri detection and population densities. The effectiveness of mesh-covered traps did not vary with season. In addition, mesh-covered traps eliminated >90% of nontarget organisms and allowed for quicker enumeration of D. citri. We expect this method will become an important component of redesigning integrated pest management programs in citrus groves by reducing unintended impacts of beneficial arthropods during large scale D. citri monitoring.

Monitoring psyllid vectors of apple proliferation in South Tyrol (Italy): a multi-year survey

Monitoring psyllid vectors of apple proliferation in South Tyrol (Italy): a multi-year survey

The chemistry of multitrophic interactions in phytoplasma disease systems and advances in control of psyllid vectors with semiochemicals

Phytoplasmas causing the three most important diseases on fruit crops, Candidatus Phytoplasma mali (apple proliferation), Ca. P. pyri (pear decline), and Ca. P. prunorum (European stone fruit yellows (ESFY)), have been investigated for the chemistry of their interactions in a multitrophic context. The chemically mediated ecological interactions of the phytoplasmas with vector insects (psyllids), their (alternate) host plants, pollinators and insect antagonists (entomopathogenic fungi) were studied. Attractive and repellent compounds were identified and tested in laboratory studies and field surveys. Additionally, a new entomopathogenic fungus was isolated from psyllid host. The identified new chemical compounds, blends and antagonists are used for the development of biotechnical control methods using the complete spectrum of available methods and materials for application. Traps and dispensers, microencapsulated volatiles, and also nanofibers are used for the development of appropriate formulations for field applications of semiochemicals for phytoplasma vector control.

Genome Analysis of Haplotype D of Candidatus Liberibacter Solanacearum.

Candidatus Liberibacter solanacearum (Lso) haplotype D (LsoD) is a suspected bacterial pathogen, spread by the phloem-feeding psyllid Bactericera trigonica Hodkinson and found to infect carrot plants throughout the Mediterranean. Haplotype D is one of six haplotypes of Lso that each have specific and overlapping host preferences, disease symptoms, and psyllid vectors. Genotyping of rRNA genes has allowed for tracking the haplotype diversity of Lso and genome sequencing of several haplotypes has been performed to advance a comprehensive understanding of Lso diseases and of the phylogenetic relationships among the haplotypes. To further pursue that aim we have sequenced the genome of LsoD from its psyllid vector and report here its draft genome. Genome-based single nucleotide polymorphism analysis indicates LsoD is most closely related to the A haplotype. Genomic features and the metabolic potential of LsoD are assessed in relation to Lso haplotypes A, B, and C, as well as the facultative strain Liberibacter crescens. We identify genes unique to haplotype D as well as putative secreted effectors that may play a role in disease characteristics specific to this haplotype of Lso.