Strategy for Structural Elucidation of Polysaccharides: Elucidation of a Maize Mucilage that Harbors Diazotrophic Bacteria.

The recruitment of a bacterial consortium by the host is a strategy not limited to animals but is also used in plants. A maize aerial root mucilage has been found that harbors nitrogen fixing bacteria that are attracted to the carbohydrate rich environment. This synbiotic relationship is facilitated by a polysaccharide, whose complicated structure has been previously unknown. In this report, we present the characterization of the maize polysaccharide by employing new analytical strategies combining chemical depolymerization, oligosaccharide sequencing, and monosaccharide and glycosidic linkage quantitation. The mucilage contains a single heterogeneous polysaccharide composed of a highly fucosylated and xylosylated galactose backbone with arabinan and mannoglucuronan branches. This unique polysaccharide structure may select for the diazotrophic community by containing monosaccharides and linkages that correspond to the glycosyl hydrolases associated with the microbial community. The elucidation of this complicated structure illustrates the power of the analytical methods, which may serve as a general platform for polysaccharide analysis in the future.

Effect of NaHCO3 treatments on the activity of cell-wall-degrading enzymes produced by Penicillium digitatum during the pathogenesis process on grapefruit.

BACKGROUND: This study was performed to clarify the strategies of Penicillium digitatum during pathogenesis on citrus, assessing, on albedo plugs, the effects of treatment with sodium bicarbonate (NaHCO3 ), at two different pH values (5 and 8.3), on cell-wall-degrading enzyme activity over a period of 72 h. RESULTS: Treatment with NaHCO3 , under alkaline pH, delayed the polygalacturonase activity for 72 h, or 48 h in the case of the pectin lyase, compared with the control or the same treatment at pH 5. In contrast, pectin methyl esterase activity rapidly increased after 24 h, in plugs dipped in the same solution. In this case, the activity remained higher than untreated or pH 5-treated plugs up to 72 h. CONCLUSION: The rapid increase in pectin methyl esterase activity under alkaline conditions is presumably the strategy of the pathogen to lower the pH, soon after the initiation of infection, in order to restore an optimal environment for the subsequent polygalacturonase and pectin lyase action. In fact, at the same time, a low pH delayed the enzymatic activity of polygalacturonase and pectin lyase, the two enzymes that actually cleave the α-1,4-linkages between the galacturonic acid residues. © 2018 Society of Chemical Industry.

Uptake, sequestration and tolerance of cadmium at cellular levels in the hyperaccumulator plant species Sedum alfredii.

Sedum alfredii is one of a few plant species known to hyperaccumulate cadmium (Cd). Uptake, localization, and tolerance of Cd at cellular levels in shoots were compared in hyperaccumulating (HE) and non-hyperaccumulating (NHE) ecotypes of Sedum alfredii. X-ray fluorescence images of Cd in stems and leaves showed only a slight Cd signal restricted within vascular bundles in the NHEs, while enhanced localization of Cd, with significant tissue- and age-dependent variations, was detected in HEs. In contrast to the vascular-enriched Cd in young stems, parenchyma cells in leaf mesophyll, stem pith and cortex tissues served as terminal storage sites for Cd sequestration in HEs. Kinetics of Cd transport into individual leaf protoplasts of the two ecotypes showed little difference in Cd accumulation. However, far more efficient storage of Cd in vacuoles was apparent in HEs. Subsequent analysis of cell viability and hydrogen peroxide levels suggested that HE protoplasts exhibited higher resistance to Cd than those of NHE protoplasts. These results suggest that efficient sequestration into vacuoles, as opposed to rapid transport into parenchyma cells, is a pivotal process in Cd accumulation and homeostasis in shoots of HE S. alfredii. This is in addition to its efficient root-to-shoot translocation of Cd.

Co-culturing Chlorella minutissima with Escherichia coli can increase neutral lipid production and improve biodiesel quality.

Lipid productivity and fatty acid composition are important metrics for the production of high quality biodiesel from algae. Our previous results showed that co-culturing the green alga Chlorella minutissima with Escherichia coli under high-substrate mixotrophic conditions enhanced both culture growth and crude lipid content. To investigate further, we analyzed neutral lipid content and fatty acid content and composition of axenic cultures and co-cultures produced under autotrophic and mixotrophic conditions. We found that co-culturing C. minutissima with E. coli under high substrate conditions (10 g/L) increased neutral lipid content 1.9- to 3.1-fold and fatty acid content 1.5- to 2.6-fold compared to equivalent axenic C. minutissima cultures. These same co-cultures also exhibited a significant fatty acid shift away from trienoic and toward monoenoic fatty acids thereby improving the quality of the synthesized fatty acids for biodiesel production. Further investigation suggested that E. coli facilitates substrate uptake by the algae and that the resulting growth enhancement induces a nitrogen-limited condition. Enhanced carbon uptake coupled with nitrogen limitation is the likely cause of the observed neutral lipid accumulation and fatty acid profile changes.

Vascular occlusions in grapevines with Pierce's disease make disease symptom development worse.

Vascular occlusions are common structural modifications made by many plant species in response to pathogen infection. However, the functional role(s) of occlusions in host plant disease resistance/susceptibility remains controversial. This study focuses on vascular occlusions that form in stem secondary xylem of grapevines (Vitis vinifera) infected with Pierce's disease (PD) and the impact of occlusions on the hosts' water transport and the systemic spread of the causal bacterium Xylella fastidiosa in infected vines. Tyloses are the predominant type of occlusion that forms in grapevine genotypes with differing PD resistances. Tyloses form throughout PD-susceptible grapevines with over 60% of the vessels in transverse sections of all examined internodes becoming fully blocked. By contrast, tylose development was mainly limited to a few internodes close to the point of inoculation in PD-resistant grapevines, impacting only 20% or less of the vessels. The extensive vessel blockage in PD-susceptible grapevines was correlated to a greater than 90% decrease in stem hydraulic conductivity, compared with an approximately 30% reduction in the stems of PD-resistant vines. Despite the systemic spread of X. fastidiosa in PD-susceptible grapevines, the pathogen colonized only 15% or less of the vessels in any internode and occurred in relatively small numbers, amounts much too small to directly block the vessels. Therefore, we concluded that the extensive formation of vascular occlusions in PD-susceptible grapevines does not prevent the pathogen's systemic spread in them, but may significantly suppress the vines' water conduction, contributing to PD symptom development and the vines' eventual death.

Spatial imaging of Zn and other elements in Huanglongbing-affected grapefruit by synchrotron-based micro X-ray fluorescence investigation.

Huanglongbing (HLB) is a highly destructive, fast-spreading disease of citrus, causing substantial economic losses to the citrus industry worldwide. Nutrient levels and their cellular distribution patterns in stems and leaves of grapefruit were analysed after graft-inoculation with lemon scions containing 'Candidatus Liberibacter asiaticus' (Las), the heat-tolerant Asian type of the HLB bacterium. After 12 months, affected plants showed typical HLB symptoms and significantly reduced Zn concentrations in leaves. Micro-XRF imaging of Zn and other nutrients showed that preferential localization of Zn to phloem tissues was observed in the stems and leaves collected from healthy grapefruit plants, but was absent from HLB-affected samples. Quantitative analysis by using standard references revealed that Zn concentration in the phloem of veins in healthy leaves was more than 10 times higher than that in HLB-affected leaves. No significant variation was observed in the distribution patterns of other elements such as Ca in stems and leaves of grapefruit plants with or without graft-inoculation of infected lemon scions. These results suggest that reduced phloem transport of Zn is an important factor contributing to HLB-induced Zn deficiency in grapefruit. Our report provides the first in situ, cellular level visualization of elemental variations within the tissues of HLB-affected citrus.

Microplate assay for quantitation of neutral lipids in extracts from microalgae.

Lipid quantitation is widespread in the algae literature, but popular methods such as gravimetry, gas chromatography and mass spectrometry (GC-MS), and Nile red cell staining suffer drawbacks, including poor quantitation of neutral lipids, expensive equipment, and variable results among algae species, respectively. A high-throughput microplate assay was developed that uses Nile red dye to quantify neutral lipids that have been extracted from algae cells. Because the algal extracts contained pigments that quenched Nile red fluorescence, a mild bleach solution was used to destroy pigments, resulting in a nearly linear response for lipid quantities in the range of 0.75 to 40 µg. Corn oil was used as a standard for quantitation, although other vegetable oils displayed a similar response. The assay was tested on lipids extracted from three species of Chlorella and resulted in close agreement with triacylglycerol (TAG) levels determined by thin layer chromatography. The assay was found to more accurately measure algal lipids conducive to biodiesel production and nutrition applications than the widely used gravimetric assay. Assay response was also consistent among different species, in contrast to Nile red cell staining procedures.

The impact of elevated CO2 concentration on the quality of algal starch as a potential biofuel feedstock.

Cultured microalgae are viewed as important producers of lipids and polysaccharides, both of which are precursor molecules for the production of biofuels. This study addressed the impact of elevated carbon dioxide (CO2) on Chlorella sorokiniana production of starch and on several properties of the starch produced. The production of C. sorokiniana biomass, lipid and starch were enhanced when cultures were supplied with 2% CO2. Starch granules from algae grown in ambient air and 2% CO2 were analyzed by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The granules from algae grown in 2% CO2 were disk-shaped and contained mainly stromal starch; granules from cultures grown in ambient air were cup-shaped with primarily pyrenoid starch. The granules from cells grown in 2% CO2 had a higher proportion of the accumulated starch as the highly branched, amylopectin glucan than did granules from cells grown in air. The rate of hydrolysis of starch from 2% CO2-grown cells was 1.25 times greater than that from air-grown cells and 2-11 times higher than the rates of hydrolysis of starches from cereal grains. These data indicate that culturing C. sorokiniana in elevated CO2 not only increases biomass yield but also improves the structure and composition of starch granules for use in biofuel generation. These modifications in culture conditions increase the hydrolysis efficiency of the starch hydrolysis, thus providing potentially important gains for biofuel production.

Efficient xylem transport and phloem remobilization of Zn in the hyperaccumulator plant species Sedum alfredii.

Sedum alfredii is one of a few species known to hyperaccumulate zinc (Zn) and cadmium (Cd). Xylem transport and phloem remobilization of Zn in hyperaccumulating (HP) and nonhyperaccumulating (NHP) populations of S. alfredii were compared. Micro-X-ray fluorescence (µ-XRF) images of Zn in the roots of the two S. alfredii populations suggested an efficient xylem loading of Zn in HP S. alfredii, confirmed by the seven-fold higher Zn concentrations detected in the xylem sap collected from HP, when compared with NHP, populations. Zn was predominantly transported as aqueous Zn (> 55.9%), with the remaining proportion (36.7-42.3%) associated with the predominant organic acid, citric acid, in the xylem sap of HP S. alfredii. The stable isotope (68)Zn was used to trace Zn remobilization from mature leaves to new growing leaves for both populations. Remobilization of (68)Zn was seven-fold higher in HP than in NHP S. alfredii. Subsequent analysis by µ-XRF, combined with LA-ICPMS (laser ablation-inductively coupled plasma mass spectrometry), confirmed the enhanced ability of HP S. alfredii to remobilize Zn and to preferentially distribute the metal to mesophyll cells surrounding phloem in the new leaves. The results suggest that Zn hyperaccumulation by HP S. alfredii is largely associated with enhanced xylem transport and phloem remobilization of the metal. To our knowledge, this report is the first to reveal enhanced remobilization of metal by phloem transport in hyperaccumulators.

Texture, composition and anatomy of spinach leaves in relation to nitrogen fertilization.

BACKGROUND: The postharvest quality and shelf life of spinach are greatly influenced by cultural practices. Reduced spinach shelf life is a common quandary in the Salinas Valley, California, where current agronomic practices depend on high nitrogen (N) rates. This study aimed to describe the postharvest fracture properties of spinach leaves in relation to N fertilization, leaf age and spinach cultivar. RESULTS: Force-displacement curves, generated by a puncture test, showed a negative correlation between N fertilization and the toughness, stiffness and strength of spinach leaves (P > 0.05). Younger leaves (leaves 12 and 16) from all N treatments were tougher than older leaves (leaves 6 and 8) (P > 0.05). Leaves from the 50 and 75 ppm total N treatments irrespective of spinach cultivar had higher fracture properties and nutritional quality than leaves from other N treatments (P > 0.05). Total alcohol-insoluble residues (AIR) and pectins were present at higher concentrations in low-N grown plants. These plants also had smaller cells and intercellular spaces than high-N grown leaves (P > 0.05). CONCLUSION: Observed changes in physicochemical and mechanical properties of spinach leaves due to excess nitrogen fertilization were significantly associated with greater postharvest leaf fragility and lower nutritional quality.

Polysaccharide compositions of intervessel pit membranes contribute to Pierce's disease resistance of grapevines.

Symptom development of Pierce's disease (PD) in grapevine (Vitis vinifera) depends largely on the ability of the bacterium Xylella fastidiosa to use cell wall-degrading enzymes (CWDEs) to break up intervessel pit membranes (PMs) and spread through the vessel system. In this study, an immunohistochemical technique was developed to analyze pectic and hemicellulosic polysaccharides of intervessel PMs. Our results indicate that PMs of grapevine genotypes with different PD resistance differed in the composition and structure of homogalacturonans (HGs) and xyloglucans (XyGs), the potential targets of the pathogen's CWDEs. The PMs of PD-resistant grapevine genotypes lacked fucosylated XyGs and weakly methyl-esterified HGs (ME-HGs), and contained a small amount of heavily ME-HGs. In contrast, PMs of PD-susceptible genotypes all had substantial amounts of fucosylated XyGs and weakly ME-HGs, but lacked heavily ME-HGs. The intervessel PM integrity and the pathogen's distribution in Xylella-infected grapevines also showed differences among the genotypes. In pathogen-inoculated, PD-resistant genotypes PM integrity was well maintained and Xylella cells were only found close to the inoculation site. However, in inoculated PD-susceptible genotypes, PMs in the vessels associated with bacteria lost their integrity and the systemic presence of the X. fastidiosa pathogen was confirmed. Our analysis also provided a relatively clear understanding of the process by which intervessel PMs are degraded. All of these observations support the conclusion that weakly ME-HGs and fucosylated XyGs are substrates of the pathogen's CWDEs and their presence in or absence from PMs may contribute to grapevine's PD susceptibility.

Cell wall-degrading enzymes enlarge the pore size of intervessel pit membranes in healthy and Xylella fastidiosa-infected grapevines.

The pit membrane (PM) is a primary cell wall barrier that separates adjacent xylem water conduits, limiting the spread of xylem-localized pathogens and air embolisms from one conduit to the next. This paper provides a characterization of the size of the pores in the PMs of grapevine (Vitis vinifera). The PM porosity (PMP) of stems infected with the bacterium Xylella fastidiosa was compared with the PMP of healthy stems. Stems were infused with pressurized water and flow rates were determined; gold particles of known size were introduced with the water to assist in determining the size of PM pores. The effect of introducing trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA), oligogalacturonides, and polygalacturonic acid into stems on water flux via the xylem was also measured. The possibility that cell wall-degrading enzymes could alter the pore sizes, thus facilitating the ability of X. fastidiosa to cross the PMs, was tested. Two cell wall-degrading enzymes likely to be produced by X. fastidiosa (polygalactuoronase and endo-1,4- beta -glucanase) were infused into stems, and particle passage tests were performed to check for changes in PMP. Scanning electron microscopy of control and enzyme-infused stem segments revealed that the combination of enzymes opened holes in PMs, probably explaining enzyme impacts on PMP and how a small X. fastidiosa population, introduced into grapevines by insect vectors, can multiply and spread throughout the vine and cause Pierce's disease.

Xyn10A, a thermostable endoxylanase from Acidothermus cellulolyticus 11B.

We cloned and purified the major family 10 xylanase (Xyn10A) from Acidothermus cellulolyticus 11B. Xyn10A was active on oat spelt and birchwood xylans between 60°C and 100°C and between pH 4 and pH 8. The optimal activity was at 90°C and pH 6; specific activity and K(m) for oat spelt xylan were 350 µmol xylose produced min⁻¹ mg of protein⁻¹ and 0.53 mg ml⁻¹, respectively. Based on xylan cleavage patterns, Xyn10A is an endoxylanase, and its half-life at 90°C was approximately 1.5 h in the presence of xylan.

Strangers in the matrix: plant cell walls and pathogen susceptibility.

Early in infection, pathogens encounter the outer wall of plant cells. Because pathogen hydrolases targeting the plant cell wall are well-known components of virulence, it has been assumed that wall disassembly by the plant itself also contributes to susceptibility, and now this has been established experimentally. Understanding how plant morphological and developmental remodeling and pathogen cell wall targeted virulence influence infections provides new perspectives about plant-pathogen interactions. The plant cell wall can be an effective physical barrier to pathogens, but also it is a matrix where many proteins involved in pathogen perception are delivered. By breaching the wall, a pathogen potentially reveals itself to the plant and activates responses, setting off events that might halt or limit its advance.

Stem and leaf sequestration of zinc at the cellular level in the hyperaccumulator Sedum alfredii.

* Sedum alfredii is a fast-growing, high-biomass zinc (Zn) hyperaccumulator native to China. Here, the characteristics of in vivo Zn distribution in stems and leaves of the hyperaccumulating (HE) and nonhyperaccumulating ecotypes (NHE) of S. alfredii were investigated by synchrotron radiation X-ray fluorescence (SRXRF) analysis, together with a Zn probe. * Preferential Zn accumulation in leaf and stem epidermis was observed in both ecotypes, but to a much greater extent for HE. Epidermal Zn increased largely in leaves and stems of HE as exposure time was prolonged, while Zn saturation occurred relatively early in HE leaf mesophyll cells and stem vascular bundles. A second peak of Zn enrichment in stem and leaf vascular systems was shown in both ecotypes. However, the proportion of Zn accumulated in stem vascular bundles relative to other tissues was much greater for HE than for NHE. * Leaf and stem distribution patterns of phosphorus (P) and sulphur (S) in the HE were very like that for Zn, while the calcium (Ca) distribution pattern was the reverse of that for Zn. No such relationship was observed in NHE. * Our study mainly suggested that epidermal layers serve as important storage sites for accumulated Zn in the S. alfredii HE.

Identification of endo- and exo-polygalacturonase activity in Lygus hesperus (Knight) salivary glands.

Polygalacturonase (PG) activity found in the salivary gland apparatus of the western tarnished plant bug (WTPB, Lygus hesperus Knight) has been thought to be the main chemical cause of the damage inflicted by this mirid when feeding on its plant hosts. Early viscosity and thermal stability studies of the PG activity in L. hesperus protein extracts were difficult to interpret. Thus, it has been suggested that one or more PG protein(s) with different hydrolytic modes of action are produced by this mirid. In order to understand the quantitative complexity of the WTPB salivary PG activity, PG purification from a protein extract from salivary glands excised from L. hesperus insects was performed using affinity and ion exchange chromatography. To elucidate the qualitative complexity of the purified PGs, the digestion products generated by the PGs were separated using high performance anion exchange chromatography with pulsed amperometric detection. At least five PG proteins were detected; these differing in terms of their glycosylation, mass-to-charge ratios, and/or molecular mass. The characterization of the products generated by these PGs showed that endo- and exo-acting PGs are produced by WTPB. Although none of the PGs was purified to homogeneity, the present work provides biochemical evidence of a multiplicity of PGs that degrade the pectin component of the plant tissue in different fashions. The implications of these findings affect the understanding of WTPB feeding damage and, potentially, help identify ways to control this important crop pest. Arch. Insect Biochem. Physiol. 2008. (c) 2008 Wiley-Liss, Inc.

Characterization of the cell wall of the ubiquitous plant pathogen Botrytis cinerea.

The ascomycete Botrytis cinerea is a destructive and ubiquitous plant pathogen and represents a model organism for the study of necrotrophic fungal pathogens. Higher fungi possess a complex and dynamic multilayer cell wall involved in crucial aspects of fungal development, growth and pathogenicity. Plant resistance to microbial pathogens is determined often by the capacity of the plant to recognize molecular patterns associated with the surface of an interacting microbe. Here we report the chemical characterization of cell walls from B. cinerea during axenic growth. Neutral sugars and proteins constituted most of the mass of the B. cinerea cell walls, although chitin and uronic acids were detected. Glucose was the most abundant neutral sugar, but arabinose, galactose, xylose and mannose also were present. Changes in cell wall composition during culture were observed. As the culture developed, protein levels declined, while chitin and neutral sugars increased. Growth of B. cinerea was associated with a remarkable decline in the fraction of its cell wall material that was soluble in hot alkali. These results suggest that the cell wall of B. cinerea undergoes significant modifications during growth, possibly becoming more extensively covalently cross-linked, as a result of aging of mycelia or in response to decreasing nutrient supply or as a consequence of increasing culture density.

Disassembly of the fruit cell wall by the ripening-associated polygalacturonase and expansin influences tomato cracking.

Fruit cracking is an important problem in horticultural crop production. Polygalacturonase (SlPG) and expansin (SlEXP1) proteins cooperatively disassemble the polysaccharide network of tomato fruit cell walls during ripening and thereby, enable softening. A Golden 2-like (GLK2) transcription factor, SlGLK2 regulates unripe fruit chloroplast development and results in elevated soluble solids and carotenoids in ripe fruit. To determine whether SlPG, SlEXP1, or SlGLK2 influence the rate of tomato fruit cracking, the incidence of fruit epidermal cracking was compared between wild-type, Ailsa Craig (WT) and fruit with suppressed SlPG and SlEXP1 expression (pg/exp) or expressing a truncated nonfunctional Slglk2 (glk2). Treating plants with exogenous ABA increases xylemic flow into fruit. Our results showed that ABA treatment of tomato plants greatly increased cracking of fruit from WT and glk2 mutant, but not from pg/exp genotypes. The pg/exp fruit were firmer, had higher total soluble solids, denser cell walls and thicker cuticles than fruit of the other genotypes. Fruit from the ABA treated pg/exp fruit had cell walls with less water-soluble and more ionically and covalently-bound pectins than fruit from the other lines, demonstrating that ripening-related disassembly of the fruit cell wall, but not elimination of SlGLK2, influences cracking. Cracking incidence was significantly correlated with cell wall and wax thickness, and the content of cell wall protopectin and cellulose, but not with Ca2+ content.

Enzyme-Catalyzed Production of Potato Galactan-Oligosaccharides and Its Optimization by Response Surface Methodology.

This work shows an optimized enzymatic hydrolysis of high molecular weight potato galactan yielding pectic galactan-oligosaccharides (PGOs), where endo-ß-1,4-galactanase (galactanase) from Cellvibrio japonicus and Clostridium thermocellum was used. For this, response surface methodology (RSM) by central composite design (CCD) was applied. The parameters varied were temperature (°C), pH, incubation time (min), and enzyme/substrate ratio (U/mg). The optimized conditions for the production of low degree of polymerization (DP) PGOs were obtained for each enzyme by spectrophotometric assay and confirmed by chromatography. The optimal conditions predicted for the use of C. japonicus galactanase to obtain PGOs of DP = 2 were T = 51.8 °C, pH 5, E/S = 0.508 U/mg, and t = 77.5 min. For DP = 3, they were T = 21 °C, pH 9, E/S = 0.484 U/mg, and t = 12.5 min; and for DP = 4, they were T = 21 °C, pH 5, E/S = 0.462 U/mg, and t = 12.5 min. The efficiency results were 51.3% for substrate hydrolysis. C. thermocellum galactanase had a lower yield (35.7%) and optimized conditions predicted for PGOs of DP = 2 were T = 60 °C, pH 5, E/S = 0.525 U/mg, and time = 148 min; DP = 3 were T = 59.7 °C, pH 5, E/S = 0.506 U/mg, and time = 12.5 min; and DP = 4, were T = 34.5 °C, pH 11, E/S = 0.525 U/mg, and time = 222.5 min. Fourier transformed infrared (FT-IR) and nuclear magnetic resonance (NMR) characterizations of PGOs are presented.

Xylella fastidiosa requires polygalacturonase for colonization and pathogenicity in Vitis vinifera grapevines.

Xylella fastidiosa is the causal agent of Pierce's disease of grape, an economically significant disease for the grape industry. X. fastidiosa systemically colonizes the xylem elements of grapevines and is able to breach the pit pore membranes separating xylem vessels by unknown mechanisms. We hypothesized that X. fastidiosa utilizes cell wall degrading enzymes to break down pit membranes, based on the presence of genes involved in plant cell wall degradation in the X. fastidiosa genome. These genes include several beta-1,4 endoglucanases, several xylanases, several xylosidases, and one polygalacturonase (PG). In this study, we demonstrated that the pglA gene encodes a functional PG. A mutant in pglA lost pathogenicity and was compromised in its ability to systemically colonize Vitis vinifera grapevines. The results indicate that PG is required for X. fastidiosa to successfully infect grapevines and is a critical virulence factor for X. fastidiosa pathogenesis in grapevine.