Detection of Xanthomonas campestris pv. pelargonii in geranium and greenhouse nutrient solution by serological and PCR techniques

Springer Science and Business Media LLC - Tập 103 - Trang 555-563 - 1997
S. Chittaranjan1, S.H. De Boer2
1Pacific Agriculture Research Centre, Vancouver, Canada
2Centre for Animal and Plant Health, Charlottetown, Canada

Tóm tắt

Specificity of a new monoclonal antibody, 2H5, to Xanthomonas campestris pv. pelargonii, causal agent of geranium bacterial blight, was determined by enzyme-linked immunosorbent assay (ELISA) and immunofluorescence tests on 14 strains of X. c. pelargonii, 12 strains of other X. campestris pathovars, 3 strains of other Xanthomonas spp., 3 strains of other plant pathogens, and 43 saprophytic bacteria isolated from geranium. X. c. pelargonii was detected in tissue from symptomatic and asymptomatic geraniums sampled from commercial growers, and artificially inoculated plants, by monoclonal antibody-based tests. The intensity of response in ELISA was only moderately correlated (r = 0.56) with symptom severity, while symptom severity was not correlated (r = 0.16) with the number of fluorescing cells in immunofluorescence. The bimodal frequency distribution of ELISA and immunofluorescence results served to validate arbitrarily chosen positive/negative threshold values. Most positive ELISA and immunofluorescence test results were confirmed by the polymerase chain reaction (PCR) using published primers (Manulis et al., 1994. Appl. Environ. Microbiol 60, 4094-4099). In contrast to plant tissue, the bacterium was detected in greenhouse nutrient solution with greater sensitivity by immunofluorescence and PCR than by ELISA. Sensitivity of detection was enhanced 100-fold by concentration of the bacteria by centrifugation.

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Tài liệu tham khảo

Anderson MJ and Nameth ST (1990) Development of a polyclonal antibody-based serodiagnostic assay for the detection of Xanthomonas campestris pv. pelargonii in geranium plants. Phytopathology 80: 357–360

Atmatjidou RF, McMahon R and Hoitink H (1991) Transmission of plant pathogens in ebb and flow. Greenhouse Grower. pp 20–21

Benedict AA, Alvarez AM and Pollard LW (1990) Pathovar-specific antigens of Xanthomonas campestris pv. begoniae and X. campestris pv. pelargonii detected with monoclonal antibodies. Appl Environ Microbiol 56: 572–574

Berthier Y, Verdier V, Guesdon JL, Chevrier D, Denis JB, Decoux G and Lemattre M (1993) Characterization of Xanthomonas campestris pathovars by rRNA gene restriction patterns. Appl Environ Microbiol 59: 851–859

De Boer SH (1990) Immunofluorescence for bacteria. In: Hampton R, Ball E and De Boer S (eds) Serological Methods for Detection and Identification of Viral and Bacterial Plant Pathogens (pp 295–298) APS Press, St. Paul, MN

De Boer SH, Stead DE, Alivizatos AS, Janse JD, Van Vaerenbergh J, De Haan TL and Mawhinney J (1994) Evaluation of serological tests for detection of Clavibacter michiganensis subsp. sepedonicus in composite potato stem and tuber samples. Plant Disease 78: 725–729

De Boer SH, Ward LJ, Li X and Chittaranjan S (1995) Attenuation of PCR inhibition in the presence of plant compounds by addition of BLOTTO. Nuc Acids Res 23: 2567–2568

DeParasis J and Roth DA (1990) Nucleic acid probes for identification of phytobacteria: identification of genus-specific 16s rRNA sequences. Phytopathology 80: 618–621

Digat B (1978) Mise en evidence de la latence epiphylle du Xanthomonas pelargonii (Brown) Starr et Burkholder chez le Pelargonium. Ann Phytopathol 10: 61–66

Digat B (1987) Methodologie de la detection des bacteries pathogenes du pelargonium et organisation generale de la certification sanitaire de cette culture en France. EPPO Bull 17: 281–286

Jordan RL (1990) Strategy and techniques for the production of monoclonal antibodies. In: Hampton R, Ball E and De Boer S (eds) Serological Methods for Detection and Identification of Viral and Bacterial Plant Pathogens (pp 55–76) APS Press, St. Paul, MN

Kado CI and Heskett MG (1970) Selective media for isolation of Agrobacterium, Corynebacterium, Erwinia, Pseudomonas, and Xanthomonas. Phytopathology 60: 969–976

Lazo GR and Gabriel DW (1987) Conservation of plasmid DNA sequences and pathovar identification of strains of Xanthomonas campestris. Phytopathology 77: 448–453

Manulis S, Valinsky L, Lichter A and Gabriel DW (1994) Sensitive and specific detection of Xanthomonas campestris pv. pelargonii with DNA primers and probes identified by random amplified polymorphic DNA analysis. Appl EnvironMicrobiol 60: 4094–4099

McGuire RJ, Jones JB and Sasser M (1986) Tween media for semiselective isolation of Xanthomonas campestris pv. vesicatoria from soil and plant material. Plant Disease 70: 887–891

McLaughlin RJ and Chen TA(1990) ELISA methods for plant pathogenic prokaryotes. In: Hampton R, Ball E and De Boer S (eds) Serological Methods for Detection and Identification of Viral and Bacterial Plant Pathogens (pp 197–204) APS Press, St. Paul, MN

Schaad NW (1988) A laboratory guide for identification of plant pathogenic bacteria (2nd ed.) APS Press, St. Paul, MN. 158 pp

Strider DL (1985) Geranium. In: Strider DL (ed) Diseases of Floral Crops Vol. 2 (pp 142–156) Praeger Publishers, New York

Sutula CL, Gillett, JM, Morrissey SM and Ramsdell DC (1986) Interpreting ELISA data and establishing the positive-negative threshold. Plant Disease 70: 722–726

Tuinier JE and Stephens CT (1989) Use of serology to detect Xanthomonas campestris pv. pelargonii in aqueous extracts of geranium plants. Plant Disease 73: 875–878