Use of suppressive subtractive hybridization to identify Flavobacterium columnare DNA sequences not shared with Flavobacterium johnsoniae.
OBJECTIVE
To identify specific sequences in the fish pathogen Flavobacterium columnare not shared by Flavobacterium johnsoniae.


RESULTS
Suppressive subtractive hybridization (SSH) was used to selectively amplify and clone F. columnare-specific sequences. A highly virulent strain of F. columnare was used as tester and the type strain of F. johnsoniae was used as driver. After library construction, 192 clones were selected and sequenced. From those, 110 clones contained unique F. columnare-specific sequences that were verified using dot blot hybridization. Sequence sizes ranged from 55 to 872 bp with 45,363 bp sequenced in total.


CONCLUSIONS
Specific F. columnare sequences representing all but one (motility related) functional categories were annotated. Several putative virulence factors were identified in F. columnare such as a collagenase, a chondroitinase, proteases, as well as drug resistance and iron transport-related genes.


CONCLUSIONS
Suppressive subtractive hybridization is a cost-effective method for identifying genetic differences between Flavobacterium spp. The number of sequences available from F. columnare has been doubled.
Olivares-Fuster
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O
Department of Fisheries and Allied Aquacultures, Auburn University, Auburn, AL 36849, USA.
Arias
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CR
D000818:Animals; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics*; D005393:Fish Diseases / Q000382:microbiology; D045826:Flavobacteriaceae Infections / Q000382:microbiology / Q000662:veterinary; D005417:Flavobacterium / Q000145:classification* / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D007059:Ictaluridae / Q000382:microbiology; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization / Q000379:methods*; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D004269:DNA, Bacterial