Nonpathogenic bacteria associated with potato stems cross-react withCorynebacterium sepedonicum antisera in immunofluorescence

Crowley, C. F.; Boer, S. H. De

doi:10.1007/bf02854878

Cited by 28 publications

(6 citation statements)

References 6 publications

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“…Inexpensive, less specific polyclonal antibodies are sometimes used to increase the sensitivity of serological methods. However, this method frequently produces false positive results due to cross-reactivity with other bacteria [47] and cannot be used to detect Cms strains without EPS layers [6,48]. …”

Section: Discussionmentioning

confidence: 99%

Development of a Sensitive and Specific Polyclonal Antibody for Serological Detection of Clavibacter michiganensis subsp. sepedonicus

Przewodowski

Przewodowska

2017

PLoS ONE

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The quarantine bacterium Clavibacter michiganensis subsp. sepedonicus (Cms) causes bacterial ring rot (BRR) in potato but is difficult to detect, hampering the diagnosis of this disease. ELISA immunoassays have not been widely used to detect Cms because commercially available anti-Cms antibodies detect mainly EPS-producing bacteria and can fail to detect strains that do not produce EPS. In the current study, we developed a new type of polyclonal antibody that specifically detects Clavibacter michiganensis subsp. sepedonicus bacteria irrespective of their EPS level. We first found that the presence of bacterial EPS precluded quantitative measurement of bacteria by currently available immunoenzymatic methods, but that washing Cms cells with acidic and basic buffers to remove EPS before analysis successfully standardized ELISA results. We used a mix of three strains of Cms with diverse EPS levels to generate antigen for production of antibodies recognizing Cms cells with and without an EPS layer (IgG-EPS and IgG-N-EPS, respectively). The resulting IgG-N-EPS recognized almost all Cms strains tested in this work regardless of their mucoidal level. The availability of this new antibody renders immunological diagnostics of Cms more sensitive and reliable, as our newly developed antibodies can be used in many type of immunoassays. This work represents an important step forward in efforts to diagnose and prevent the spread of BRR, and the methods and solutions developed in this work are covered by six Polish, one European and one US patents.

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Section: Discussionmentioning

confidence: 99%

Development of a Sensitive and Specific Polyclonal Antibody for Serological Detection of Clavibacter michiganensis subsp. sepedonicus

Przewodowski

Przewodowska

2017

PLoS ONE

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“…Diagnostic kits are available for some subspecies, but false positive results and limited sensitivity remain problems for serological detection of Cm (Crowley and De Boer 1982;De Boer and Wieczorek 1984;Mills et al 1997;Pastrik 2000). Identification of Cm has also been based on fatty acid methyl ester analysis (FAME).…”

Section: Introductionmentioning

confidence: 99%

Polymorphism analysis of housekeeping genes for identification and differentiation of Clavibacter michiganensis subspecies

Waleron

Kamasa

et al. 2011

Eur J Plant Pathol

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The utility of polymorphism analysis was determined for differentiation of the following subspecies of the Gram-positive plant pathogenic bacterium, Clavibacter michiganensis: C. m. subsp. michiganensis, C. m. subsp. sepedonicus, C. m. subsp. insidiosus C. m. subsp. nebraskensis, and C. m. subsp. tessellarius. Specific primers designed for amplification of the housekeeping genes recA, rpoB, and rpoD generated 827-, 1037-, and 862-bp DNA fragments, respectively. PCR products obtained from 40 C. michiganensis strains were analysed using RFLP with four restriction endonucleases, and those PCR products with specific RFLP patterns were sequenced. The genotypes discriminated after PCR-RFLP were specific for each subspecies and also allowed for differentiation of C. m. subsp. michiganensis strains. Sequence analysis of the recA, rpoB, and rpoD gene fragments also distinguished C. michiganensis subspecies and was useful for phylogenetic analysis of all subspecies. For rapid, inexpensive, and effective differentiation of the five subspecies in this research, we recommend the amplification of recA and/or rpoD gene fragments and digestion of the PCR products with the restriction endonuclease FnuDII.

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“…Whereas immunofluorescence with polyclonal antisera was too nonspecific for accurate identification of a single bacterial pathogen (De Boer 1982, Crowley & De Boer 1982, the monoclonal antibody was highly specific for C. sepedonicum (DeBoer & Wieczorek 1984). With the setup used in this work, positive fluorescing cells could be adequately discriminated from other fluorescence in the video image.…”

Section: Resultsmentioning

confidence: 99%

An automated microscope system for estimating the population of Corynebacterium sepedonicum cells labelled with monoclonal antibodies in immunofluorescence

Boer

Hall

1988

Canadian Journal of Plant Pathology

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Nonpathogenic bacteria associated with potato stems cross-react withCorynebacterium sepedonicum antisera in immunofluorescence

Cited by 28 publications

References 6 publications

Development of a Sensitive and Specific Polyclonal Antibody for Serological Detection of Clavibacter michiganensis subsp. sepedonicus

Development of a Sensitive and Specific Polyclonal Antibody for Serological Detection of Clavibacter michiganensis subsp. sepedonicus

Polymorphism analysis of housekeeping genes for identification and differentiation of Clavibacter michiganensis subspecies

An automated microscope system for estimating the population of Corynebacterium sepedonicum cells labelled with monoclonal antibodies in immunofluorescence

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