DAO 65:209-216 (2005)  -  doi:10.3354/dao065209

Polymorphisms in 16S rRNA genes of Flavobacterium psychrophilum correlate with elastin hydrolysis and tetracycline resistance

Marilyn Soule1, Stacey LaFrentz1, Kenneth Cain2,3, Scott LaPatra4, Douglas R. Call1,3,*

1Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, Washington 99164-7040, USA
2Department of Fish and Wildlife Resources, University of Idaho, Moscow, Idaho 83844-1136, USA
3WSU and UI Center for Reproductive Biology, Pullman, Washington 99164-4231, USA
4Research Division, Clear Springs Foods, PO Box 712, Buhl, Idaho 83316, USA
*Corresponding author. Email:

ABSTRACT: Flavobacterium psychrophilum is the etiological agent of bacterial coldwater disease, which causes significant problems to aquaculture worldwide. A recent study (Soule M, Cain K, LaFrentz S, Call DR [2005] Infect Immun 73:3799–3802) identified two 16S rRNA gene sequence variants (6 base differences) within the variable stem-loop region 3 for F. psychrophilum strains ATCC 49418 and CSF 259-93. That study also hypothesized that F. psychrophilum is composed of at least 2 distinct genetic lineages (I and II) described by a microarray-based comparative genomics study. In the present study, we augmented an existing 16S rDNA microarray to detect both 16S rRNA sequence variants from F. psychrophilum. Subsequent microarray experiments showed that CSF 259-93 hybridized as expected, but ATCC 49418 was positive for both sequence variants. We then developed a PCR-restriction fragment length polymorphism (RFLP) assay (MnlI and MaeIII) to distinguish between the 2 sequences. Gel isolation of PCR-RFLP products, cloning, and sequencing confirmed that ATCC 49418 harbors both 16S rRNA sequences. Microarray experiments showed that 11 of 14 strains from genetic Lineage I harbor both the CSF 259-93 and ATCC 49418 16S rRNA sequence variants, whereas all 15 Lineage II strains were only positive for the CSF 259-93 sequence (p < 0.0001). Elastin hydrolysis and tetracycline resistance were most closely associated with the latter strains (p < 0.0001 and p = 0.024, respectively). These data support the hypothesis that F. psychrophilum is composed of at least 2 distinct genetic lineages that are closely associated with host origin.


KEY WORDS: 16S rRNA · Flavobacterium psychrophilum · Bacterial coldwater disease · Elastin hydrolysis · Microarray hybridization · Tetracycline resistance


Full text in pdf format