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

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hosted egg masses but not the other life stages. The suitability of the host plants for these GWSS life stages may be linked to the chemical composition of the xylem fluids (Andersen et al. 1989, 1992, Brodbeck et al. 1990, 1993, 1995, 1996, 1999), data for which will be provided by the xylem analyses. Sowthistle, common groundsel, lambsquarter, cheese weed, primrose and beavertail were not hosting large GWSS numbers, if any, and may be discarded or replaced for next season. This season, the location seems to influence the size of GWSS egg masses (larger egg masses in the south), survival (lower in the south) and parasitism (lower in the south). The underlying factors may be related to temperature and humidity which have been recorded but have not been correlated to the findings yet. The major difference between the coastal and inland locations at similar latitude is the number of second generation adults, and all life stages from the second generation are responsible for most of the location differences. Aside from the egg masses, there are no obvious differences in the other life stages recorded in the coastal and southern location. Further conclusions cannot be drawn without the data that is still being taken in the fecundity and feeding studies and the chemical xylem composition of the host plants. For full understanding of the climatic influences behind these observations, multiple year data are needed and need to be analyzed for temporal and spatial differences, for which two additional years of funding will hopefully be forthcoming from the CDFA. REFERENCES Andersen, P.C., B.V. Brodbeck & R..F. Mizell, III. 1989. Metabolism of amino acids, organic acids and sugars extracted from the xylem fluid of four host plants by adult Homalodisca coagulata. Entomol. Exp. & App. 50: 149-59. Andersen, P. C., B. Brodbeck and R. F. Mizell, III. 1992. Feeding by the leafhopper, Homalodisca coagulata. in relation to xylem fluid chemistry and tension. J. Insect Physiol. 38: 611-622. Brodbeck, B., R. F. Mizell, III & P.C. Andersen. 1990. Amino acids as determinants of host preference for the xylem-feeding leafhopper, Homalodisca coagulata. Oecologia 83:338-345. Brodbeck, B. V., P. C. Andersen & R. F. Mizell, III. 1993. Physiological and behavioral adaptations of three species of leafhoppers in response to the dilute nutrient content of xylem fluid. J. Ins. Physiol. 39:73-81. Brodbeck, B.V., P.C. Andersen & R.F. Mizell, III. 1995. Differential utilization of nutrients during development by the xylophagous leafhopper, Homalodisca coagulata. Ent. Exp. & Appl. 75:279-289. Brodbeck, B., P. C. Andersen & R. F. Mizell, III. 1996. Utilization of primary nutrients by the polyphagous xylophage, Homalodisca coagulata, reared on single host species. Arch. Biochem. and Physiol. 32:65-83. Brodbeck, B.V., P. C. Andersen & R. F. Mizell, III. 1999. The effects of total dietary nitrogen and amino acid balance on the development of xylophagous leafhoppers. Arch. of Ins. Biochem. & Physiol. 42:37-50. FUNDING AGENCIES Funding for this project was provided by the CDFA Pierce’s Disease and Glassy-winged Sharpshooter Board. - 141 -
EXPLORATION FOR FACULTATIVE ENDOSYMBIONTS OF SHARPSHOOTERS Principle Investigator: Alexander H. Purcell Division of Insect Biology University of California Berkeley, CA 94720-3112 purcell@nature.berkeley.edu Cooperators: Eoin Brody ESPM University of California Berkeley, CA 94720-3112 Researcher: Clytia Montllor Curley Division of Insect Biology University of California Berkeley, CA 94720-3112 Chris Carlton Dept. of Entomology Louisiana State University Baton Rouge, LA 70803 Russell Mizell University of Florida NFREC Monticello, FL 32351-5677 Reporting Period: The results reported here are from work conducted from July 1, 2002 to June 30, 2004. ABSTRACT Glassy-winged sharpshooters (GWSS) were collected in California and several states in the southeastern United States in 2002 and 2003 to search for pathogenic or beneficial endosymbiotic bacteria of these insects. Various tissues were examined for the presence of bacteria by PCR: hemolymph, eggs, and bacteriomes. A subset of hemolymph and egg samples were cloned and sequenced based on unique digest patterns of their extracted 16s rDNA, or analyzed by restriction digest patterns of sample compared to known bacterial DNA. Most cloned sequences were identified as Baumannia (one of the primary symbionts of GWSS), and Wolbachia (a common secondary symbiont in a majority of insect taxa investigated). In addition, we isolated bacteria that were most closely related (by 16S rDNA sequence) to the following genera: Acinetobacter, Stenotrophomonas, Pseudomonas, Burkholderia. All are common bacteria that are found in soil, water, or plant surfaces, and also in insect guts or surfaces. INTRODUCTION We have surveyed populations of the glassy-winged sharpshooter (GWSS), Homalodisca coagulata, for bacterial symbionts that might be exploited to manipulate the biology of this insect vector of <strong>Xylella</strong> fastidiosa (Xf) (Purcell and Feil 2001). Pathogens or other microbial associates of GWSS have not been employed to date as biological control agents or contributors to the control of these pests largely because none are known, although some efforts to discover viruses of GWSS have been made. Although endosymbiotic bacterial associates of leafhoppers are little-understood and unexploited to date, their potential importance is well worth exploring. The first step has been to look for and identify any naturally occurring bacteria in GWSS populations from a wide geographical range. <strong>Of</strong> particular interest to us in this study were bacterial associates that are facultative (also referred to as “secondary”), i.e., that occur in some individuals or populations but are not required by their hosts; and that could be introduced into, or augmented in pest populations. We use the term symbiont here in the biological sense of “living together” and do not imply mutual benefit (Douglas 1994). Facultative bacterial associates have been described in a variety of homopterans including leafhoppers (Swezy and Severin 1930, Schwemmler 1974, McCoy et al. 1978, Purcell et al. 1986). The only leafhopper facultative symbiont studied in some depth is BEV, a bacterium that occurs in Euscelidius variegatus in France, but apparently not in California (Purcell et al. 1986). Uninfected females of E. variegatus inoculated with cultures of BEV transmitted the bacteria transovarially (“vertically”) to their offspring, with resulting deleterious effects (Purcell et al. 1986, Purcell and Suslow 1987). This bacterium could also be transmitted horizontally between leafhoppers feeding on the same plant; hence it could persist in the population in spite of its negative fitness effects. It is clear from our studies of facultative bacteria in aphids (Chen et al. 2000, Montllor et al. 2002) as well as from the study of BEV, that endosymbiotic associations are complex and have critically important effects, both positive and negative, on the physiology, population biology and vector potential of their hosts. Some of the most extensive studies on the effects of facultative symbionts on insect hosts involve Wolbachia, a transovarially transmitted bacterium that occurs in 20-76% of investigated insect species (Weeks et al. 2002) with a range of interesting effects (e.g.,Werren 1994, Stouthammer et al.1999). Wolbachia has recently been described from GWSS (Moran et al. 2003), though its effects remain unknown. Although Wolbachia has “helped raise the awareness of the potential contribution of endosymbionts...it is important not to discard other alternatives” (Weeks et al. 2002). Our approach was to investigate whether other alternatives existed for GWSS. - 142 -
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IMPACT OF HOST PLANT XYLEM FLUID ON
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0.18 600 Optical density 0.16 0.14
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OPTIMIZING MARKER-ASSISTED SELECTIO
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esistant and susceptible genotypes
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MAP BASED IDENTIFICATION AND POSITI
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esistant genotypes are in process a
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BREEDING PIERCE’S DISEASE RESISTA
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Progeny from crosses of field resis
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a = (1=low, 4= high); b = (1=green,
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v.8) for differences due to the pla
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SHARPSHOOTER FEEDING BEHAVIOR IN RE
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correlated with all other findings
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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: No egg masses were recorded on olea
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
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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
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RESULTS Construction of cDNA Librar
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CONCLUSIONS Genetic resistance and
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Mutagenesis of Xylella The EZ::TN T
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ISOLATION OF BACTERIOPHAGES SPECIFI
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Project Leader: Michele M. Igo Sect
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outer membrane fractions using two-
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1995; Purcell and Saunders, 1999).
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DEVELOPMENT OF SSR MARKERS FOR GENO
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Strain Name Host of Origin County o
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ROLE OF ATTACHMENT OF XYLELLA FASTI
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wild-type cells was not conclusive
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DETERMINATION OF GENES CONFERRING H
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S1 S2 S3 S4 Nb123456789bb123456789b
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MULTILOCUS SEQUENCE TYPING TO IDENT
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GENOME-WIDE IDENTIFICATION OF RAPID
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Americas and since most of the plan
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EFFECTS OF CHEMICAL MILIEU ON ATTAC
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CONCLUSIONS Our overall objective i
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RESULTS Objective 1. We conducted t
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Redak, R. A., Purcell, A. H., Lopes
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In parallel, an “activating trans
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PATTERNS OF XYLELLA FASTIDIOSA INFE
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% of vessels 100 80 60 40 20 Mugwor
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DOCUMENTATION AND CHARACTERIZATION
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Magnolia002 showed more identity (9
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PLASMID ADDICTION AS A NOVEL APPROA
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GENETIC VARIABILITY OF XYLELLA FAST
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probing activities). In preliminary
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the slow release valve was opened a
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12. Hopkins DL (1977) Diseases caus
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The almost complete absence of info
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QUANTIFYING LANDSCAPE-SCALE MOVEMEN
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Table 1. The mean (±SD) ELISA read
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EPIDEMIOLOGICAL ASSESSMENTS OF PIER
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multiply to relatively high (easily
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SPATIAL DATABASE CREATION AND MAINT
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Project Leader: Thomas M. Perring D
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Figure 2. Individual leaves from a
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The state variables, process functi
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DEVELOPMENT OF A FIELD SAMPLING PLA
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Figure 1. Three main dispersion pat
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OBJECTIVES 1. Track the movement of
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REFERENCES 1. Beard, C.B., Cordon-R
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cases, positive phloem samples were
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EXPLOITING XYLELLA FASTIDIOSA PROTE
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Figure 2. Polymer disk accumulation
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CHARACTERIZATION OF NEONICOTINOIDS
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EVALUATION OF RESISTANCE POTENTIAL
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Project Leader: Doug Cook Dept. of
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Based on the in silico analysis, de
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CONTROL OF PIERCE’S DISEASE THROU
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Figure 1. Oleander ‘White’ afte
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RESULTS During the reporting period
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DEVELOPMENT OF AN ARTIFICIAL DIET A
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DESIGN OF CHIMERIC ANTI-MICROBIAL P
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Neutrophil elastase Cecropin B Chim
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and WTXb is very low (0.00-0.40%) a
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100 100 0.056 North America 100 0.0
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GENETIC DIFFERENTIATION AMONG GEOGR
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Table 1. Single-populations descrip
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MOLECULAR DISTINCTION BETWEEN POPUL
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These novel observations strongly s
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SEQUENCE DIVERGENCE IN TWO MITOCHON
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Table 3. Pairwise sequence distance
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DEVELOPMENT OF MOLECULAR DIAGNOSTIC
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A. B. Relative Density of SCAR Band
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THE ALIMENTARY TRACK OF GLASSY-WING
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We have dissected and identified al
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REALIZED LIFETIME PARASITISM AND TH
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REPRODUCTIVE AND DEVELOPMENTAL BIOL
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Mean adult longevity (days) 25 20 1
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the proposed exploratory work; thes
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SEARCHING FOR AND COLLECTING EGG PA
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REFERENCES Hoddle, M. S. and S. V.
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some Gram(+) bacteria, but are inac
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7. Hultmark, D., et al., Insect Imm
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Isolation of Fungal Pathogens Soil
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IDENTIFICATION OF MECHANISMS MEDIAT
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concentrations of 1.5 x 10 7 bacter
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anchor it in the outer membrane (sh
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SYMBIOTIC CONTROL OF PIERCE’S DIS
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MANAGEMENT OF PIERCE’S DISEASE OF
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pfF mutants are taken up by insects
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Simpson, A. J. G., F. C. Reinach, P
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Al-Wahaibi, A.K., Owen, and J. G. M
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patterns differed among the lines,
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Punja, Z.K. 2001. Genetic engineeri
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mind, we conducted an additional st
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REFERENCES Toscano, N., Byrne, F.,
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RESULTS AND CONCLUSIONS The program
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COMPATIBILITY OF INSECTICIDES WITH
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More tests are in progress to addre
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Impact Of Host Plant Xylem Fluid On Xylella Fastidiosa Multiplication ...

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