Suppression-subtractive hybridization as a strategy to identify taxon-specific sequences within the Mycoplasma mycoides Cluster: design and validation of an M. capricolum subsp. capricolum-specific PCR assay.

Laure Maigre; Christine Citti; Marc Marenda; François Poumarat; Florence Tardy

Suppression-subtractive hybridization as a strategy to identify taxon-specific sequences within the Mycoplasma mycoides Cluster: design and validation of an M. capricolum subsp. capricolum-specific PCR assay.

Laure Maigre ^{1, *} Christine Citti ² Marc Marenda ³ François Poumarat ¹ Florence Tardy ¹

* Corresponding author

1 Laboratoire d'études et de recherches en pathologie bovine et hygiène des viandes

2 ENVT - Ecole Nationale Vétérinaire de Toulouse

3 Faculty of Veterinary science

Abstract : The phylogenetically related Mycoplasma capricolum subsp. capricolum and M. mycoides subsp. mycoides biotype Large Colony are two small-ruminant pathogens involved in contagious agalactia. Their respective contributions to clinical outbreaks are not well documented, because they are difficult to differentiate with the current diagnostic techniques. In order to identify DNA sequences specific to one taxon or the other, a suppression-subtractive hybridization approach was developed. DNA fragments resulting from the reciprocal subtraction of the type strains were used as probes on a panel of M. capricolum subsp. capricolum and M. mycoides subsp. mycoides biotype Large Colony strains to assess their intrataxon specificity. Due to a high intrataxon polymorphism and important cross-reactions between taxa, a single DNA fragment was shown to be specific for M. capricolum subsp. capricolum and to be present in all M. capricolum subsp. capricolum field isolates tested in this study. A PCR assay targeting the corresponding gene (simpA51) was designed that resulted in a 560-bp amplification only in M. capricolum subsp. capricolum and in M. capricolum subsp. capripneumoniae (the etiological agent of contagious caprine pleuropneumonia). simpA51 was further improved to generate a multiplex PCR (multA51) that allows the differentiation of M. capricolum subsp. capripneumoniae from M. capricolum subsp. capricolum. Both the simpA51 and multA51 assays accurately identify M. capricolum subsp. capricolum among other mycoplasmas, including all members of the M. mycoides cluster. simpA51 and multA51 PCRs are proposed as sensitive and robust tools for the specific identification of M. capricolum subsp. capricolum and M. capricolum subsp. capripneumoniae.

keyword : 1MYCOPLASME HAFSSA MYCOPLASME

Document type :

Journal articles

Domain :

Life Sciences [q-bio] / Animal biology / Veterinary medicine and animal Health

Life Sciences [q-bio] / Microbiology and Parasitology / Bacteriology

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https://hal-anses.archives-ouvertes.fr/hal-00377609
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Submitted on : Wednesday, April 22, 2009 - 2:51:41 PM
Last modification on : Friday, June 14, 2019 - 6:21:09 PM

Suppression-subtractive hybridization as a strategy to identify taxon-specific sequences within the Mycoplasma mycoides Cluster: design and validation of an M. capricolum subsp. capricolum-specific PCR assay.

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Suppression-subtractive hybridization as a strategy to identify taxon-specific sequences within the Mycoplasma mycoides Cluster: design and validation of an M. capricolum subsp. capricolum-specific PCR assay.

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