title: Intragenomic heterogeneity in the 16S rRNA genes of Flavobacterium columnare and standard protocol for genomovar assignment.
abstract: Genetic variability in 16S rRNA gene sequences has been demonstrated among isolates of Flavobacterium columnare, and a restriction fragment length polymorphism (RFLP) assay is available for genetic typing of this important fish pathogen. Interpretation of restriction patterns can be difficult due to the lack of a formal description of the expected number and sizes of DNA fragments generated for each of the described genomovars. In this study, partial 16S rRNA gene sequences (ca. 1250-bp fragment) from isolates representing each described genomovar and isolates generating unique restriction patterns were cloned and sequenced. The results demonstrated that some isolates contained up to three different 16S rRNA genes whose sequences generate different RFLP patterns due to intragenomic heterogeneity within HaeIII restriction sites. The occurrence of HaeIII restriction sites within the portion of the 16S rRNA gene used for typing the F. columnare isolates and intragenomic heterogeneity within these sites explained the restriction patterns observed following RFLP analyses. This research provides a standard protocol for typing isolates of F. columnare by RFLP and a formal description of the expected restriction patterns for the previously described genomovars I, II, II-B and III. Additionally, we describe a new genomovar, I/II.
authors: United States Department of Agriculture-Agricultural Research Service (USDA-ARS), Aquatic Animal Health Research Unit, Auburn, AL, USA.
authors: B R
authors: BR
authors: LaFrentz
authors: G C
authors: GC
authors: Waldbieser
authors: T J
authors: TJ
authors: Welch
authors: C A
authors: CA
authors: Shoemaker
mesh_terms: D000818:Animals; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D005393:Fish Diseases / Q000382:microbiology*; D045826:Flavobacteriaceae Infections / Q000382:microbiology / Q000662:veterinary*; D005417:Flavobacterium / Q000235:genetics* / Q000378:metabolism; D018740:Genetic Heterogeneity; D016680:Genome, Bacterial; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012150:Polymorphism, Restriction Fragment Length; D012336:RNA, Ribosomal, 16S / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA / Q000662:veterinary
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: 16S rRNA; Flavobacterium columnare; columnaris disease; genomovar; intragenomic heterogeneity; restriction fragment length polymorphism