Microbiology 151 (2005), 475-489; DOI 10.1099/mic.0.27590-0
Microbiology 151 (2005), 475-489; DOI 10.1099/mic.0.27590-0
© 2005 Society for General Microbiology
Suppression subtractive hybridization as a basis to assess Mycoplasma agalactiae and Mycoplasma bovis genomic diversity and species-specific sequences
Marc S. Marenda1,
Evelyne Sagné1,
François Poumarat2 and
Christine Citti1
1 UMR INRA-ENVT 1225, Ecole Nationale Vétérinaire de Toulouse, 23 Ch des Capelles, 31076 Toulouse, France
2 AFSSA Lyon, 31 Av Tony Garnier BP 7033, 69342 Lyon, France
Correspondence
Marc S. Marenda
m.marenda{at}envt.fr
The phylogenically related Mycoplasma agalactiae and Mycoplasma bovis species are two ruminant pathogens difficult to differentiate and for which a limited amount of sequence data are available. To assess the degree of genomic diversity existing between and within these mycoplasma species, sets of DNA fragments specific for M. bovis type-strain PG45 or for M. agalactiae type-strain PG2 were isolated by suppression subtractive hybridization and used as probes on a panel of M. agalactiae and M. bovis field isolates. Results indicated that approximately 70 % of the DNA fragments specific to one or the other type strain are represented in all field isolates of the corresponding species. Only one M. bovis isolate, which was first classified as M. agalactiae, reacted with 15 % of the PG2-specific probes, while several M. agalactiae isolates reacted with 15 % of the PG45-specific probes. Sequence analyses indicated that most of the genomic diversity observed within one species is related to ORFs with (i) no homologies to proteins recorded in the databases or (ii) homologies to proteins encoded by restriction modification systems. Reminiscent of gene transfer as a means for genomic diversity, a PG45-specific DNA fragment with significant homologies to a central protein of an integrative conjugative element of Mycoplasma fermentans (ICEF) was found in most M. bovis field isolates and in a few M. agalactiae isolates. Finally, sequences encoding part of DNA polymerase III were found in both sets of M. agalactiae- and M. bovis-specific DNA fragments and were used to design a species-specific PCR assay for the identification and differentiation of M. agalactiae and M. bovis.
Abbreviations: MF, membrane filtration; SSH, suppression subtractive hybridization
The GenBank/EMBL/DDBJ accession numbers for the sequences reported in this paper are: AGA-02 to -90, CL844514 to CL844536; AGA-4 to -89, CL844537 to CL844551; BOV-01 to -93, CL844552 to CL844574; BOV-7 to -94, CL844575 to CL844585.
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Copyright © 2005 Society for General Microbiology.
