Impact Of Host Plant Xylem Fluid On Xylella Fastidiosa Multiplication ...

More documents

Recommendations

Info

7061 N I 7172 N I 8946 N I Reverse transcription PCR 0D 1D 1W 2W 3W 4W 6W 8W 10W RealTime PCR Ct-N: 26.3 Ct-I: 22.5 Ct-N: NA Ct-I: 24.2 Ct-N: 28.7 Ct-I: 26.3 Fold induction week 10 1:14 infinite 1:5.2 9353 N I Xf16S N I Actin N I Ct-N: 34.6 Ct-I: 26.7 Ct-N: 36.5 Ct-I: 16.8 Ct-N: 26.2 Ct-I: 25.4 1:239 infinite 1:1.7 Cycle Figure 1. Monitoring of PD-induced genes using conventional reverse transcriptase-PCR and Real Time PCR. Leaf tissue was sampled from growth chamber-grown plants at nine time points (0, 1d, 1w, 2w, 3w, 4w, 6w, 8w, 10w: d-day, w-week) after inoculation. Xylella up-regulated genes identified from in silico analysis are 7061, 7172, 8946, and 9353. Actin serves as a constitutively expressed control. Xf16S = Xylella fastidiosa 16S gene. N; Non-inoculated, I; Inoculated with X. fastidiosa. - 297 -
CONTROL OF PIERCE’S DISEASE THROUGH DEGRADATION OF XANTHAN GUM Project Leader: Donald A. Cooksey Dept. of Plant Pathology University of California Riverside, CA 92521 Researchers: Rosina Bianco Dept. of Plant Pathology University of California Riverside, CA 92521 Cooperator: Neal L. Schiller Division of Biomedical Sciences University of California Riverside, CA 92521 Seung-Don Lee Dept. of Plant Pathology University of California Riverside, CA 92521 Korsi Dumenyo Dept. of Plant Pathology University of California Riverside, CA 92521 Reporting Period: The results reported here are from work conducted between October 2003 and October 2004. ABSTRACT Acinetobacter johnsonii GX123, a Xylella gum-degrading endophyte was co-inoculated with Xylella fastidiosa strain Texas in oleander plants to determine its efficacy as a biocontrol agent in preliminary experiments. Symptoms appeared in both plants inoculated with X. fastidiosa alone and plants co-inoculated with the endophyte. However, symptoms were more severe and appeared earlier in plants inoculated with X. fastidiosa than in those co-inoculated with the endophyte. A. johnsonii GX123 seems to be a promising candidate to control X. fastidiosa. Experiments using a sequential strategy of inoculating the Xylella gum-degrader endophyte prior to X. fastidiosa are ongoing and its effects on symptom expression are still under investigation. INTRODUCTION Pierce’s disease (PD) of grapevine and other leaf scorch diseases caused by Xylella fastidiosa (Xf) are associated with aggregation of bacteria in xylem vessels, formation of a gummy matrix, and subsequent blockage of water uptake. In the closely-related pathogen, Xanthomonas campestris (Xc), xanthan gum is known to be an important virulence factor (Katzen et al, 1998), probably contributing to bacterial adhesion, aggregation, and plugging of xylem. The published genome sequence of Xf (Simpson et al, 2000; Bhattacharyya et al, 2002; Van Sluys et al, 2003) revealed that this pathogen also has genes for producing an exopolysaccharide with a very similar structure to that of xanthan gum. In PD, this Xylella gum is likely to contribute to plugging of the grapevine xylem (Keen et al, 2000) and possibly to the aggregation of the bacterium in the mouthparts of the glassy-winged sharpshooter. Because of its importance as an industrial thickener and emulsifier, xanthan gum synthesis and degradation have been extensively studied (Becker et al, 1998). Bacteria that produce xanthandegrading enzymes have been isolated from soils using enrichment techniques with xanthan gum as the sole carbon source (Sutherland 1987; Ruijssenaars et al, 2000). The purpose of this project is to identify bacteria that produce xanthan-degrading enzymes to target this specific virulence factor of Xf. This approach has the potential to significantly reduce the damage caused by PD in grapes and potentially in other hosts of Xf such as almond and oleander. If the gum is important in the aggregation of the pathogen in the insect vector, then our approach may also reduce the efficiency of transmission of PD. Our first approach will be to develop endophytic bacteria that produce these enzymes in the xylem of grapevines, but another approach is to engineer grape plants to produce these enzymes. Through the cloning and characterization of genes encoding xanthanases and xanthan lyases we will facilitate possible efforts to transform grapevines to produce these enzymes. Previously, we used modified xanthan gum that mimics Xylella gum from a Xc mutant as the sole carbon source for enrichment culture from infected grapevines and oleanders. The Xylella gum biosynthetic operon in the Xf genome is different than the one in Xc from which the commercial xanthan gum is obtained. Since it is not feasible to produce Xylella gum for our studies from the slow-growing Xf, we genetically modified a strain of Xc to produce a modified xanthan gum that is predicted to have the same chemical structure as that from Xf. This was accomplished by deleting the gumI gene from the biosynthetic operon. Over 100 bacterial strains were initially recovered from enrichment experiments, and 11 were subsequently confirmed to effectively degrade Xylella gum. These strains were then tested for cellulase activity. Degradation of the cellulosic backbone of the gum polymer would be desirable, but we do not want enzymes that recognize and degrade plant cellulose. One particular strain (GX123) with high gum-degrading activity but no cellulase activity isolated from oleander was identified as Acinetobacter johnsonii (Aj), and characterized in more detail. In vitro, growth and biofilm production by GX123 were enhanced by Xylella gum as a substrate and by cells of Xf added to a minimal medium. The gum was degraded rapidly during log-phase growth of this endophyte, and viscosity was reduced almost to non-detectable levels. GX123 colonized stems and leaves of oleander systemically (10 4 -10 5 cfu/g of plant tissue 20 days after inoculation), and systemic colonization was enhanced by co-inoculation with Xf. The effect of using GX123 as an endophyte to reduce the ability of Xf to produce disease symptoms in oleander was studied. - 298 -
Page 1 and 2:
IMPACT OF HOST PLANT XYLEM FLUID ON
Page 3 and 4:
0.18 600 Optical density 0.16 0.14
Page 5 and 6:
OPTIMIZING MARKER-ASSISTED SELECTIO
Page 7 and 8:
esistant and susceptible genotypes
Page 9 and 10:
MAP BASED IDENTIFICATION AND POSITI
Page 11 and 12:
esistant genotypes are in process a
Page 13 and 14:
BREEDING PIERCE’S DISEASE RESISTA
Page 15 and 16:
Progeny from crosses of field resis
Page 17 and 18:
a = (1=low, 4= high); b = (1=green,
Page 19 and 20:
- 78 -
Page 21 and 22:
- 80 -
Page 23 and 24:
v.8) for differences due to the pla
Page 25 and 26:
SHARPSHOOTER FEEDING BEHAVIOR IN RE
Page 27 and 28:
correlated with all other findings
Page 29 and 30:
EFFECTS OF FEEDING SUBSTRATE ON RET
Page 31 and 32:
REFERENCES Bextine, B. and T. Mille
Page 33 and 34:
will also provide insights into the
Page 35 and 36:
OBJECTIVES 1. Determine glassy-wing
Page 37 and 38:
GWSS per 30 s sweep (seasonal avera
Page 39 and 40:
ELISA for the presence of GWSS egg
Page 41 and 42:
Project Leader: Thomas P. Freeman E
Page 43 and 44:
Freeman, T. P., R. A. Leopold, D. R
Page 45 and 46:
pathogen, when they move into viney
Page 47 and 48:
A NOVEL IMMUNOLOGICAL APPROACH FOR
Page 49 and 50:
1.6 GWSS Adults That Fed on Protein
Page 51 and 52:
Hagler, J.R. & S.E. Naranjo. 2004.
Page 53 and 54:
RESULTS: Oviposition Survey Wild gr
Page 55 and 56:
Host specificity testing: No-choice
Page 57 and 58:
currently employed morphological ch
Page 59 and 60:
increase our present understanding
Page 61 and 62:
BIOLOGY AND MORPHOMETRIC ANALYSIS O
Page 63 and 64:
Table 1. Mean a developmental durat
Page 65 and 66:
EFFECTS OF USING CONSTANT AND CYCLI
Page 67 and 68:
REFERENCES Gautam, R. D. 1986. Effe
Page 69 and 70:
PARASITISM OF THE GLASSY-WINGED SHA
Page 71 and 72:
Table 1. Parasitism by G.ashmeadi o
Page 73 and 74:
Project Leader: Robert F. Luck Dept
Page 75 and 76:
A more interesting analysis using t
Page 77 and 78:
MYCOPATHOGENS AND THEIR EXOTOXINS I
Page 79 and 80:
POPULATION DYNAMICS AND INTERACTION
Page 81 and 82:
No egg masses were recorded on olea
Page 83 and 84:
EXPLORATION FOR FACULTATIVE ENDOSYM
Page 85 and 86:
Although Baumannia and Wolbachia we
Page 87 and 88:
EFFECTS OF SUBLETHAL DOSES OF IMIDA
Page 89 and 90:
Table 2. Mortality and flight perfo
Page 91 and 92:
A NOVEL METHOD TO INDUCE OVIPOSITIO
Page 93 and 94:
REFERENCES Brodbeck, B. V., P. C. A
Page 95 and 96:
Cohorts H. coagulata neonates were
Page 97 and 98:
REFERENCES Blua, M. J. and D. J. W.
Page 99 and 100:
Figure 2 shows the average numbers
Page 101 and 102:
REPRODUCTIVE BIOLOGY AND PHYSIOLOGY
Page 103 and 104:
REFERENCES Blua, M. J., Phillips, P
Page 105 and 106:
- 164 -
Page 107 and 108:
- 166 -
Page 109 and 110:
RESULTS Some plant families had no
Page 111 and 112:
Table 5. Winter, spring and summer
Page 113 and 114:
16S rDNA is by far the most sequenc
Page 115 and 116:
DNA MICROARRAY AND MUTATIONAL ANALY
Page 117 and 118:
Inhibition of biofilm is BSA concen
Page 119 and 120:
CULTURE-INDEPENDENT ANALYSIS OF END
Page 121 and 122:
Borneman, J., M. Chrobak, G. Della
Page 123 and 124:
P (CFU P OBJECTIVE 1. Determine the
Page 125 and 126:
Purcell, AH and SR Saunders. 1999a.
Page 127 and 128:
Figure 1: Southern blot of tolC::ap
Page 129 and 130:
Project Leader: David Gilchrist Dep
Page 131 and 132:
III. Production of transgenic plant
Page 133 and 134:
RESULTS Construction of cDNA Librar
Page 135 and 136:
CONCLUSIONS Genetic resistance and
Page 137 and 138:
Mutagenesis of Xylella The EZ::TN T
Page 139 and 140:
ISOLATION OF BACTERIOPHAGES SPECIFI
Page 141 and 142:
Project Leader: Michele M. Igo Sect
Page 143 and 144:
outer membrane fractions using two-
Page 145 and 146:
1995; Purcell and Saunders, 1999).
Page 147 and 148:
DEVELOPMENT OF SSR MARKERS FOR GENO
Page 149 and 150:
Strain Name Host of Origin County o
Page 151 and 152:
ROLE OF ATTACHMENT OF XYLELLA FASTI
Page 153 and 154:
wild-type cells was not conclusive
Page 155 and 156:
DETERMINATION OF GENES CONFERRING H
Page 157 and 158:
S1 S2 S3 S4 Nb123456789bb123456789b
Page 159 and 160:
MULTILOCUS SEQUENCE TYPING TO IDENT
Page 161 and 162:
GENOME-WIDE IDENTIFICATION OF RAPID
Page 163 and 164:
Americas and since most of the plan
Page 165 and 166:
EFFECTS OF CHEMICAL MILIEU ON ATTAC
Page 167 and 168:
CONCLUSIONS Our overall objective i
Page 169 and 170:
RESULTS Objective 1. We conducted t
Page 171 and 172:
Redak, R. A., Purcell, A. H., Lopes
Page 173 and 174:
In parallel, an “activating trans
Page 175 and 176:
PATTERNS OF XYLELLA FASTIDIOSA INFE
Page 177 and 178:
% of vessels 100 80 60 40 20 Mugwor
Page 179 and 180:
DOCUMENTATION AND CHARACTERIZATION
Page 181 and 182:
Magnolia002 showed more identity (9
Page 183 and 184:
PLASMID ADDICTION AS A NOVEL APPROA
Page 185 and 186:
GENETIC VARIABILITY OF XYLELLA FAST
Page 187 and 188: - 246 -
Page 189 and 190: probing activities). In preliminary
Page 191 and 192: the slow release valve was opened a
Page 193 and 194: 12. Hopkins DL (1977) Diseases caus
Page 195 and 196: The almost complete absence of info
Page 197 and 198: QUANTIFYING LANDSCAPE-SCALE MOVEMEN
Page 199 and 200: Table 1. The mean (±SD) ELISA read
Page 201 and 202: EPIDEMIOLOGICAL ASSESSMENTS OF PIER
Page 203 and 204: multiply to relatively high (easily
Page 205 and 206: SPATIAL DATABASE CREATION AND MAINT
Page 207 and 208: Project Leader: Thomas M. Perring D
Page 209 and 210: Figure 2. Individual leaves from a
Page 211 and 212: The state variables, process functi
Page 213 and 214: DEVELOPMENT OF A FIELD SAMPLING PLA
Page 215 and 216: Figure 1. Three main dispersion pat
Page 217 and 218: - 276 -
Page 219 and 220: - 278 -
Page 221 and 222: OBJECTIVES 1. Track the movement of
Page 223 and 224: REFERENCES 1. Beard, C.B., Cordon-R
Page 225 and 226: cases, positive phloem samples were
Page 227 and 228: EXPLOITING XYLELLA FASTIDIOSA PROTE
Page 229 and 230: Figure 2. Polymer disk accumulation
Page 231 and 232: CHARACTERIZATION OF NEONICOTINOIDS
Page 233 and 234: EVALUATION OF RESISTANCE POTENTIAL
Page 235 and 236: Project Leader: Doug Cook Dept. of
Page 237: Based on the in silico analysis, de
Page 241 and 242: Figure 1. Oleander ‘White’ afte
Page 243 and 244: RESULTS During the reporting period
Page 245 and 246: DEVELOPMENT OF AN ARTIFICIAL DIET A
Page 247 and 248: DESIGN OF CHIMERIC ANTI-MICROBIAL P
Page 249 and 250: Neutrophil elastase Cecropin B Chim
Page 251 and 252: and WTXb is very low (0.00-0.40%) a
Page 253 and 254: 100 100 0.056 North America 100 0.0
Page 255 and 256: GENETIC DIFFERENTIATION AMONG GEOGR
Page 257 and 258: Table 1. Single-populations descrip
Page 259 and 260: MOLECULAR DISTINCTION BETWEEN POPUL
Page 261 and 262: These novel observations strongly s
Page 263 and 264: SEQUENCE DIVERGENCE IN TWO MITOCHON
Page 265 and 266: Table 3. Pairwise sequence distance
Page 267 and 268: DEVELOPMENT OF MOLECULAR DIAGNOSTIC
Page 269 and 270: A. B. Relative Density of SCAR Band
Page 271 and 272: THE ALIMENTARY TRACK OF GLASSY-WING
Page 273 and 274: We have dissected and identified al
Page 275 and 276: REALIZED LIFETIME PARASITISM AND TH
Page 277 and 278: REPRODUCTIVE AND DEVELOPMENTAL BIOL
Page 279 and 280: Mean adult longevity (days) 25 20 1
Page 281 and 282: the proposed exploratory work; thes
Page 283 and 284: SEARCHING FOR AND COLLECTING EGG PA
Page 285 and 286: REFERENCES Hoddle, M. S. and S. V.
Page 287 and 288: some Gram(+) bacteria, but are inac
Page 289 and 290:
7. Hultmark, D., et al., Insect Imm
Page 291 and 292:
Isolation of Fungal Pathogens Soil
Page 293 and 294:
IDENTIFICATION OF MECHANISMS MEDIAT
Page 295 and 296:
concentrations of 1.5 x 10 7 bacter
Page 297 and 298:
anchor it in the outer membrane (sh
Page 299 and 300:
SYMBIOTIC CONTROL OF PIERCE’S DIS
Page 301 and 302:
MANAGEMENT OF PIERCE’S DISEASE OF
Page 303 and 304:
pfF mutants are taken up by insects
Page 305 and 306:
Simpson, A. J. G., F. C. Reinach, P
Page 307 and 308:
Al-Wahaibi, A.K., Owen, and J. G. M
Page 309 and 310:
patterns differed among the lines,
Page 311 and 312:
Punja, Z.K. 2001. Genetic engineeri
Page 313 and 314:
mind, we conducted an additional st
Page 315 and 316:
REFERENCES Toscano, N., Byrne, F.,
Page 317 and 318:
RESULTS AND CONCLUSIONS The program
Page 319 and 320:
COMPATIBILITY OF INSECTICIDES WITH
Page 321 and 322:
More tests are in progress to addre
show all

Impact Of Host Plant Xylem Fluid On Xylella Fastidiosa Multiplication ...

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?