Transmission Of Plant Pathogens Research Articles

Modeling host–microbiome interactions to improve mechanistic understanding of aphid vectored plant pathogens

Insect transmission of plant pathogens involves multi-layered interactions between vectors, viruses, host plants and environmental factors. Adding to the complexity of vector–virus relationships are diverse microbial communities, which are hypothesized to influence pathogen transmission. Although vector–virus interaction research has flourished, the role played by microbes in vector competence and disease epidemiology remains unclear in many pathosystems. We therefore aimed to develop a novel ecological modeling approach to identify environmental drivers of complex vector–virus–microbiome interactions, particularly differences in the abundance of microbial symbionts within vector microbiomes and relationships between symbionts and the probability of virus acquisition. Our approach combines established molecular tools for profiling microbial communities with underutilized Bayesian hierarchical modeling and data integration techniques. We used a globally relevant aphid–virus pathosystem to develop custom vector–microbiome models that incorporate environmental covariates (e.g., temperature, landcover) and applied them to individual vector symbionts to identify the extent to which environmental factors drive changes in microbial communities that then influence virus acquisition by the host aphid. Specifically, we focus on the aphid obligate symbiont (Buchnera) and a wide-spread facultative symbiont (Serratia) as proof of concept to develop models for two major vector species that include a single environmental covariate (i.e., temperature). Overall, we demonstrate how community-level modeling and microbiome profiling can identify candidate microbes and environmental variables associated with vector competence. Our modeling framework can accommodate a range of microbial symbionts with different abundances, overcome spatial misalignment of data streams, and is robust to varying levels of disease incidence. Results show Buchnera relative abundance is strongly negatively associated with virus acquisition by the vector S.avenae, but not R. padi. Although Serratia was not associated with R. padi vector competence, relative abundance was influenced by differences in spring temperatures. This work lays the foundation for developing a broader modeling framework for predicting disease dynamics in agroecosystems and deploying microbiome-targeted pest management tactics.

Disruption of transmission of plant pathogens in the insect order Hemiptera using recent advances in RNA interference biotechnology.

The review discusses current RNA interference (RNAi) biotechnological innovations for crop protection. Special attention is given to the management of insect pests in the order Hemiptera. This insect order has the most members of insects which transmit pathogens on economically important crops. It first briefly summarizes the characteristics of the insects in this order and the type of transmission mechanisms for viral and bacterial plant pathogens. RNAi products developed for other insects are also analyzed. Emphasis was made on the need for innovative management approaches to offset the threat of resistance by both the insect vector to insecticides and the pathogens to microbicides. Subsequently, the RNAi technology is described, which is particularly an ingenious method currently utilized in itself or in combination with other modern biotechnological innovations for managing important vector insects that could provide an additional powerful tool for use in integrated pest control programs. The requirements and recent advances for performing RNAi assays are detailed and an overview is given on how to produce cheaper double-stranded RNA as the main component of RNAi-based biopesticide. Examples of agricultural companies that use RNAi biotechnology in their product development were also discussed.

Bacterial Endosymbionts Identified From Leafhopper (Hemiptera: Cicadellidae) Vectors of Phytoplasmas.

Insects often harbor bacterial endosymbionts that provide them with nutritional benefit or with protection against natural enemies, plant defenses, insecticides, and abiotic stresses. Certain endosymbionts may also alter acquisition and transmission of plant pathogens by insect vectors. We identified bacterial endosymbionts from four leafhopper vectors (Hemiptera: Cicadellidae) of 'Candidatus Phytoplasma' species by direct sequencing 16S rDNA and confirmed endosymbiont presence and identity by species-specific conventional PCR. We examined three vectors of Ca. Phytoplasma pruni, causal agent of cherry X-disease [Colladonus geminatus (Van Duzee), Colladonus montanus reductus (Van Duzee), Euscelidius variegatus (Kirschbaum)] - and a vector of Ca. Phytoplasma trifolii, the causal agent of potato purple top disease [Circulifer tenellus (Baker)]. Direct sequencing of 16S identified the two obligate endosymbionts of leafhoppers, 'Ca. Sulcia' and 'Ca. Nasuia', which are known to produce essential amino acids lacking in the leafhoppers' phloem sap diet. About 57% of C. geminatus also harbored endosymbiotic Rickettsia. We identified 'Ca. Yamatotoia cicadellidicola' in Euscelidius variegatus, providing just the second host record for this endosymbiont. Circulifer tenellus harbored the facultative endosymbiont Wolbachia, although the average infection rate was only 13% and all males were Wolbachia-uninfected. A significantly greater percentage of Wolbachia-infected Ci. tenellus adults than uninfected adults carried Ca. P. trifolii, suggesting that Wolbachia may increase this insect's ability to tolerate or acquire this pathogen. Results of our study provide a foundation for continued work on interactions between leafhoppers, bacterial endosymbionts, and phytoplasma.

Special issue: Relationship between insect vectors and plant pathogens

Special issue: Relationship between insect vectors and plant pathogens

Predicting the effects of climate change on the cross-scale epidemiological dynamics of a fungal plant pathogen

The potential for climate change to exacerbate the burden of human infectious diseases is increasingly recognized, but its effects on infectious diseases of plants have received less attention. Understanding the impacts of climate on the epidemiological dynamics of plant pathogens is imperative, as these organisms play central roles in natural ecosystems and also pose a serious threat to agricultural production and food security. We use the fungal ‘flax rust’ pathogen (Melampsora lini) and its subalpine wildflower host Lewis flax (Linum lewisii) to investigate how climate change might affect the dynamics of fungal plant pathogen epidemics using a combination of empirical and modeling approaches. Our results suggest that climate change will initially slow transmission at both the within- and between-host scales. However, moderate resurgences in disease spread are predicted as warming progresses, especially if the rate of greenhouse gas emissions continues to increase at its current pace. These findings represent an important step towards building a holistic understanding of climate effects on plant infectious disease that encompasses demographic, epidemiological, and evolutionary processes. A core result is that neglecting processes at any one scale of plant pathogen transmission may bias projections of climate effects, as climate drivers have variable and cascading impacts on processes underlying transmission that occur at different scales.

Probing behavior of Neophilaenus campestris on various plant species

AbstractThe recent history of Xylella fastidiosa Wells et al. introduction in Europe illustrates how the lack of knowledge about the bacterium‐vector‐host interactions hinders the application of effective containment strategies, with the bacterial spread that currently appears relentless. Vector behavior is a key component in plant pathogen transmission; therefore, detailed knowledge of vector probing behavior on various host plants would furnish useful data for containing the bacterium. Aiming at this goal, we carried out electrical penetration graph (EPG)‐assisted probing behavior observations of the spittlebug, Neophilaenus campestris (Fallen) (Hemiptera: Aphrophoridae), a candidate vector widespread in all the areas where X. fastidiosa is currently established. Spittlebug probing was first characterized on one of its preferred host plants, Bromus madritensis L. (Poaceae), over short (6 h) and long (16 h) time spans. Thereafter, we performed comparative observations of the spittlebug probing on B. madritensis, olive, and grapevine. Overall, the probing behavior of N. campestris, that is, the main characteristic waveforms, did not differ substantially from that of Philaenus spumarius (L.) (Hemiptera: Aphrophoridae); however, here we report and describe a new kind of xylem activity interruption, provisionally termed N2, not previously observed in P. spumarius. Considering the time spent in xylem ingestion as a percentage of the total probing time (host‐suitability indicator used for spittlebugs), grapevine and B. madritensis are the preferred hosts for N. campestris, with olive as the least suitable among the plants tested. Successful probes, that is, probes during which xylem ingestion occurred, on grapevine were 2.5× greater than olive. Therefore, our data on N. campestris probing behavior suggest that the spittlebug role in X. fastidiosa epidemiology in European vineyards should be given more attention. In the present manuscript, we also discuss the current difficulties in interpreting and analyzing the outcomes of EPG studies on spittlebugs.

Can CRISPR gene drive work in pest and beneficial haplodiploid species?

Gene drives based on CRISPR/Cas9 have the potential to reduce the enormous harm inflicted by crop pests and insect vectors of human disease, as well as to bolster valued species. In contrast with extensive empirical and theoretical studies in diploid organisms, little is known about CRISPR gene drive in haplodiploids, despite their immense global impacts as pollinators, pests, natural enemies of pests, and invasive species in native habitats. Here, we analyze mathematical models demonstrating that, in principle, CRISPR homing gene drive can work in haplodiploids, as well as at sex‐linked loci in diploids. However, relative to diploids, conditions favoring the spread of alleles deleterious to haplodiploid pests by CRISPR gene drive are narrower, the spread is slower, and resistance to the drive evolves faster. By contrast, the spread of alleles that impose little fitness cost or boost fitness was not greatly hindered in haplodiploids relative to diploids. Therefore, altering traits to minimize damage caused by harmful haplodiploids, such as interfering with transmission of plant pathogens, may be more likely to succeed than control efforts based on introducing traits that reduce pest fitness. Enhancing fitness of beneficial haplodiploids with CRISPR gene drive is also promising.

Dynamic changes in bacterial communities in the recirculating nutrient solution of cucumber plug seedlings cultivated in an ebb-and-flow subirrigation system.

Ebb-and-flow subirrigation systems are highly efficient, water-saving and environmentally friendly. However, one concern with these recirculating systems is the possible transmission of plant pathogens. Here, through 16S rRNA-targeted Illumina sequencing, the bacterial dynamics in a recirculating nutrient solution were characterized for cucumber plug seedlings cultivated in an ebb-and-flow system in summer and winter. Both the bacterial number and diversity in the nutrient solution increased immediately after the first irrigation cycle; then, these values were gradually stable with recirculating irrigation. In summer and winter, different bacterial compositions and changing patterns were observed. In summer, the predominant genera in the nutrient solution included Comamonas, Pseudomonas, Acinetobacter, Reyranella, Sphingobium, Bradyrhizobium, Sphingomonas, and Acidovorax. Of those genera, during recirculating irrigation, the relative abundance of Bradyrhizobium gradually decreased, whereas those of Pseudomonas, Reyranella, Sphingobium, Sphingomonas, and Acidovorax gradually increased. In winter, the bacterial communities were mainly composed of Nevskia, Bosea, Sphingobium, Acidovorax, Pseudomonas, and Hydrocarboniphaga. Of those genera, the relative abundance of Bosea, Sphingobium, and Acidovorax showed an increasing trend, whereas those of Nevskia and Hydrocarboniphaga decreased overall. Furthermore, in both summer and winter, no plant pathogenic bacteria on cucumber could be detected; however, some potentially beneficial bacteria, including Comamonas testosteroni, Acinetobacter baumannii, Pseudomonas aeruginosa, P. koreensis and Sphingobium yanoikuyae, colonized the nutrient solution and exhibited increased relative abundances during irrigation. The colonization of these bacteria might facilitate the plant growth promotion. Inoculation of the microbes from the effluent nutrient solution also promoted the growth of cucumber seedlings, but did not lead to any disease. The present data elucidate the bacterial dynamics in a cucumber cultivation ebb-and-flow system and provide useful information for biological control during cucumber seedling production.

Spatiotemporal dynamics and quantitative analysis of phytoplasmas in insect vectors

Phytoplasmas are transmitted by insect vectors in a persistent propagative manner; however, detailed movements and multiplication patterns of phytoplasmas within vectors remain elusive. In this study, spatiotemporal dynamics of onion yellows (OY) phytoplasma in its vector Macrosteles striifrons were investigated by immunohistochemistry-based 3D imaging, whole-mount fluorescence staining, and real-time quantitative PCR. The results indicated that OY phytoplasmas entered the anterior midgut epithelium by seven days after acquisition start (daas), then moved to visceral muscles surrounding the midgut and to the hemocoel at 14–21 daas; finally, OY phytoplasmas entered into type III cells of salivary glands at 21–28 daas. The anterior midgut of the alimentary canal and type III cells of salivary glands were identified as the major sites of OY phytoplasma infection. Fluorescence staining further revealed that OY phytoplasmas spread along the actin-based muscle fibers of visceral muscles and accumulated on the surfaces of salivary gland cells. This accumulation would be important for phytoplasma invasion into salivary glands, and thus for successful insect transmission. This study demonstrates the spatiotemporal dynamics of phytoplasmas in insect vectors. The findings from this study will aid in understanding of the underlying mechanism of insect-borne plant pathogen transmission.

Economic injury levels for the potato yellow vein disease and its vector, Trialeurodes vaporariorum (Hemiptera: Aleyrodidae), affecting potato crops in the Andes

The Economic Injury Level (EIL) for pest insects is conventionally expressed in terms of the number of insects, because the level of pest injury to plants is usually difficult to quantify. However, when insect injury is mainly caused indirectly by transmission of plant pathogens, the number of insects may not be an ideal measure to calculate the EIL. The greenhouse whitefly (GWF) Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae) is the only known vector of the Potato yellow vein virus (PYVV), the causal agent of potato yellow vein disease (PYVD), but the direct injury caused by insect feeding may not be significant since potato is not a preferred host for the GWF. Here, we quantified the effect of GWF feeding and PYVD incidence on potato yield, as compared to potato market value and control costs to determine the EIL for this system. We found that the direct injury caused by GWF feeding does not affect potato yield. However, an exponential reduction in crop yield was observed with the increase of PYVD incidence. Thus, the criteria to decide upon applying a control measure ranges between 100 and 1200 infected plants/ha depending mainly on potato market price. We suggest that disease incidence should be used to calculate the EIL for PYVD management, instead of focusing on estimations of vector population size. Furthermore, we stress that potato market price is fundamental in defining an appropriate, case specific EIL for decision-making in PYVD management.

Plasticity of mycangia in Xylosandrus ambrosia beetles

Insects that depend on microbial mutualists evolved a variety of organs to transport the microsymbionts while dispersing. The ontogeny and variability of such organs is rarely studied, and the microsymbiont's effects on the animal tissue development remain unknown in most cases. Ambrosia beetles (Coleoptera: Curculionidae: Scolytinae or Platypodinae) and their mutualistic fungi are an ideal system to study the animal-fungus interactions. While the interspecific diversity of their fungus transport organ-mycangia-is well-known, their developmental plasticity has been poorly described. To determine the ontogeny of the mycangium and the influence of the symbiotic fungus on the tissue development, we dissected by hand or scanned with micro-CT the mycangia in various developmental stages in five Xylosandrus ambrosia beetle species that possess a large, mesonotal mycangium: Xylosandrus amputatus, Xylosandrus compactus, Xylosandrus crassiusculus, Xylosandrus discolor, and Xylosandrus germanus. We processed 181 beetle samples from the United States and China. All five species displayed three stages of the mycangium development: (1) young teneral adults had an empty, deflated and cryptic mycangium without fungal mass; (2) in fully mature adults during dispersal, the pro-mesonotal membrane was inflated, and most individuals developed a mycangium mostly filled with the symbiont, though size and symmetry varied; and (3) after successful establishment of their new galleries, most females discharged the bulk of the fungal inoculum and deflated the mycangium. Experimental aposymbiotic individuals demonstrated that the pronotal membrane invaginated independently of the presence of the fungus, but the fungus was required for inflation. Mycangia are more dynamic than previously thought, and their morphological changes correspond to the phases of the symbiosis. Importantly, studies of the fungal symbionts or plant pathogen transmission in ambrosia beetles need to consider which developmental stage to sample. We provide illustrations of the different stages, including microphotography of dissections and micro-CT scans.

Insect Transmission of Plant Pathogens: a Systems Biology Perspective.

Insect-vectored pathogens pose one of the greatest threats to plant and animal, including human, health on a global scale. Few effective control strategies have been developed to thwart the transmission of any insect-transmitted pathogen. Most have negative impacts on the environment and human health and are unsustainable. Plant pathogen transmission by insect vectors involves a combination of coevolving biological players: plant hosts, insect vectors, plant pathogens, and bacterial endosymbionts harbored by the insect. Our ability to help growers to control vector-borne disease depends on our ability to generate pathogen- and/or disease-resistant crops by traditional or synthetic approaches and to block pathogen transmission by the insect vector. Systems biology studies have led to the reexamination of existing paradigms on how pathogens interact with insect vectors, including the bacterial symbionts, and have identified vector-pathogen interactions at the molecular and cellular levels for the development of novel transmission interdiction strategies.

Diaphorina citri Induces Huanglongbing-Infected Citrus Plant Volatiles to Repel and Reduce the Performance of Propylaea japonica.

Transmission of plant pathogens through insect vectors is a complex biological process involving interactions between the host plants, insects, and pathogens. Simultaneous impact of the insect damage and pathogenic bacteria in infected host plants induce volatiles that modify not only the behavior of its insect vector but also of their natural enemies, such as parasitoid wasps. Therefore, it is essential to understand how insects such as the predator ladybird beetle responds to volatiles emitted from a host plant and how the disease transmission alters the interactions between predators, vector, pathogens, and plants. In this study, we investigated the response of Propylaea japonica to volatiles from citrus plants damaged by Diaphorina citri and Candidatus Liberibacter asiaticus through olfactometer bioassays. Synthetic chemical blends were also used to determine the active compounds in the plant volatile. The results showed that volatiles emitted by healthy plants attracted more P. japonica than other treatments, due to the presence of high quantities of D-limonene and beta-ocimene, and the lack of methyl salicylate. When using synthetic chemicals in the olfactory tests, we found that D-limonene attracted P. japonica while methyl salicylate repelled the predator. However, beta-ocimene attracted the insects at lower concentrations but repelled them at higher concentrations. These results indicate that P. japonica could not efficiently search for its host by using volatile cues emitted from psyllids- and Las bacteria-infected citrus plants.

RNA Interference in Insect Vectors for Plant Viruses.

Insects and other arthropods are the most important vectors of plant pathogens. The majority of plant pathogens are disseminated by arthropod vectors such as aphids, beetles, leafhoppers, planthoppers, thrips and whiteflies. Transmission of plant pathogens and the challenges in managing insect vectors due to insecticide resistance are factors that contribute to major food losses in agriculture. RNA interference (RNAi) was recently suggested as a promising strategy for controlling insect pests, including those that serve as important vectors for plant pathogens. The last decade has witnessed a dramatic increase in the functional analysis of insect genes, especially those whose silencing results in mortality or interference with pathogen transmission. The identification of such candidates poses a major challenge for increasing the role of RNAi in pest control. Another challenge is to understand the RNAi machinery in insect cells and whether components that were identified in other organisms are also present in insect. This review will focus on summarizing success cases in which RNAi was used for silencing genes in insect vector for plant pathogens, and will be particularly helpful for vector biologists.

Editorial

Editorial

Deep Characterization of the Microbiomes of Calophya spp. (Hemiptera: Calophyidae) Gall-Inducing Psyllids Reveals the Absence of Plant Pathogenic Bacteria and Three Dominant Endosymbionts.

Bacteria associated with sap-feeding insect herbivores include not only symbionts that may increase their hosts’ fitness but also harmful plant pathogens. Calophya spp. gall-inducing psyllids (Hemiptera: Calophyidae) are being investigated for their potential as biological control agents of the noxious weed, Brazilian peppertree (Schinus terebinthifolia), in Florida. Although there are no examples of plant pathogen transmission by members of the family Calophyidae, several insects in the superfamily Psylloidea are known to transmit pathogenic bacteria in the genera Candidatus Liberibacter and Candidatus Phytoplasma. To determine whether Calophya spp. harbor potentially harmful plant pathogenic bacteria, we sequenced small subunit (SSU) ribosomal RNA (rRNA) gene amplicons generated from individuals from four Calophya spp. populations. All microbial SSU gene sequences fell into the bacterial domain, with 98-99% belonging to the Proteobacteria. The Calophya microbiomes contained a relatively simple community, with 49-79 operational taxonomic units (OTUs; 97%) detected, and only 5-8 OTUs with greater than 1% abundance. Candidatus Carsonella showed the highest relative abundance, with OTUs from this candidate genus representing between 51 – 65% of all recovered sequences. The next most abundant clade observed was an unclassified Enterobacteriacae group closely related to bacteria from the genera Buchnera and Blochmannia that ranged from 20-31% in relative abundance. Wolbachia populations were the third most abundant group and represented 7-27% of the diversity in microbial OTUs. No SSU rRNA gene sequences from putative pathogenic bacteria from the genera Ca. Liberibacter or Ca. Phytoplasma were detected in the microbiomes of the four Calophya populations. The probability that infected psyllids were present in our colonies, but were not sampled, was extremley low (1.39 x 10-10). As far as we are aware, our study is the first to characterize the microbiome of a candidate biological control agent, and coupled with previous work demonstrating a high degree of host specificity and absence of plant viruses, suggests that releasing Calophya spp. in United States poses minimal risk to non-target plants.

Transcriptome analysis of the salivary glands of Nephotettix cincticeps (Uhler)

The green rice leafhopper (GRH), Nephotettix cincticeps, is one of the most important pests of rice in temperate Asian countries. GRH, a vascular feeder, secretes watery and gelling saliva in the process of feeding on phloem and xylem sap. It is known that GRH saliva contains several bioactive proteins, including enzymes such as laccase and beta-glucosidase. In this study, we performed transcriptome analysis of salivary glands of GRH using Illumina paired-end sequencing. Of 51,788 assembled contigs, 16,017 (30.9%) showed significant similarity to known proteins in the NCBI nr database, while 34,978 (67.5%) could not be annotated by similarity search, Pfam, or gene ontology (GO). Contigs (905) with predicted signal peptides and no putative transmembrane domains are suggested to represent secreted protein coding genes. Among the 76 most highly expressed putative secretory protein contigs, 68 transcripts were found to be salivary gland-specific or at least -dominant, but not expressed in stomach or Malpighian tubules. However, 45 of the 68 transcripts were unknown proteins. These findings suggest that most of the GRH transcripts encoding secreted proteins expressed in salivary glands are species and/or tissue specific. Our results provide a fundamental list of genes involved in GRH–Poaceae host plant interactions including successful feeding and plant pathogen transmission.

Eriophyoidea (Acari: Trombidiformes; Prostigmata) fauna of Shiraz County, Iran

1 Depertment of Plant Protection, Faculty of Agriculture, Shiraz University, Shiraz, Iran; E-mails: n.doryanizadeh@gmail.com, akrami@shirazu.ac.ir 2. Agricultural and Natural Resources Research Center of Khorasan Razavi, Mashhad, Iran; E-mail: hashemkamali@gmail.com * Corresponding author Mites belong to Eriophyoidea are important due to their feeding damage, their role in the transmission of plant pathogens and as potantial biological control agents against weeds (Monfreda et al. 2010). It is estimated that world fauna of eriophyoid mites could be increased to 35000 to 50000 species. Thus, discovery of many new species is not surprising (Amrine et al. 2003). During 2010–11, 16 species of Eriophyoidea belonging to two families and eight genera were collected from 16 plant species from Shiraz (Fars Province). One species is a new record to mite fauna of Iran (marked by *) and three are unknown species (marked by **). The species are listed in Table 1. The mites were separated by an insect pin, cleared in a droplet of glacial lactic acid, and then mounted on a microscopic slide using Hoyer’s medium (de Lillo & Skoracka 2010). Table 1. List of Eriopyoidea species of Shiraz county.

Attraction and oviposition responses of the fungus gnat Bradysia impatiens to microbes and microbe‐inoculated seedlings in laboratory bioassays

AbstractLaboratory tests were conducted to examine preferences of Bradysia impatiens Johannsen (Diptera: Sciaridae) larvae and adults for various microbes associated with greenhouse crops. Fungus gnat larvae and adults exhibited a preference for cultures of Pythium spp. over the medium used to grow the pathogens. Larvae also exhibited a preference for geranium seedlings infected with pathogenic Pythium spp. [P. aphanidermatum (Edson) Fitz., P. ultimum Trow, and P. irregulare Buis. (Oomycota: Peronosporales)] over non‐inoculated plants. Adult fungus gnats exhibited a strong ovipositional preference for the aforementioned Pythium spp. as well as a variety of other microorganisms, including the pathogenic fungus Thielaviopsis basicola (Berk. & Br.) (Ascomycota: Microascales), the geranium‐infecting bacterium Xanthomonas campestris pv. pelargonii (Brown) Dye (Proteobacteria: Xanthomonadales), the non‐pathogenic species Pythium torulosum Coker & P. Patt. and Pythium graminicola Subramaniam, the pathogen‐suppressive fungus Trichoderma harzianum Rifai (Ascomycota: Hypocreales), and the insect pathogenic fungus Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreales). Our study is the first to demonstrate that fungus gnats are attracted to and/or stimulated to oviposit by a wide array of living microorganisms both in pure culture and in association with plant seedlings. These findings have important implications with respect to the potential role of fungus gnats in plant pathogen transmission.

Salivary enzymes are injected into xylem by the glassy-winged sharpshooter, a vector of Xylella fastidiosa

A few phytophagous hemipteran species such as the glassy-winged sharpshooter, Homalodisca vitripennis, (Germar), subsist entirely on xylem fluid. Although poorly understood, aspects of the insect’s salivary physiology may facilitate both xylem-feeding and transmission of plant pathogens. Xylella fastidiosa is a xylem-limited bacterium that causes Pierce’s disease of grape and other scorch diseases in many important crops. X. fastidiosa colonizes the anterior foregut (precibarium and cibarium) of H. vitripennis and other xylem-feeding vectors. Bacteria form a dense biofilm anchored in part by an exopolysaccharide (EPS) matrix that is reported to have a β-1,4-glucan backbone. Recently published evidence supports the following, salivation-egestion hypothesis for the inoculation of X. fastidiosa during vector feeding. The insect secretes saliva into the plant and then rapidly takes up a mixture of saliva and plant constituents. During turbulent fluid movements in the precibarium, the bacteria may become mechanically and enzymatically dislodged; the mixture is then egested back out through the stylets into plant cells, possibly including xylem vessels. The present study found that proteins extracted from dissected H. vitripennis salivary glands contain several enzyme activities capable of hydrolyzing glycosidic linkages in polysaccharides such as those found in EPS and plant cell walls, based on current information about the structures of those polysaccharides. One of these enzymes, a β-1,4-endoglucanase (EGase) was enriched in the salivary gland protein extract by subjecting the extract to a few, simple purification steps. The EGase-enriched extract was then used to generate a polyclonal antiserum that was used for immunohistochemical imaging of enzymes in sharpshooter salivary sheaths in grape. Results showed that enzyme-containing gelling saliva is injected into xylem vessels during sharpshooter feeding, in one case being carried by the transpiration stream away from the injection site. Thus, the present study provides support for the salivation-egestion hypothesis.