|Matson, E - IOWA STATE UNIVERSITY|
|Thompson, M - IOWA STATE UNIVERSITY|
Submitted to: American Society for Microbiology
Publication Type: Abstract Only
Publication Acceptance Date: February 22, 2002
Publication Date: May 20, 2002
Citation: Matson, E.G., Humphrey, S.B., Thompson, M.G., Stanton, T.B. 2002. Identification of bacteriophage vsh-1 genes and analysis of their induction of brachyspira hyodysenteriae [abstract]. American Society for Microbiology. p. 179. Technical Abstract: VSH-1 is a mitomycin C-inducible, generalized transducing bacteriophage of the pathogenic spirochete Brachyspira hyodysenteriae (B. hyo). VSH-1 is non-lytic and is incapable of self-replication. VSH-1 virion proteins were purified and sequenced. Based on N-terminal amino acid sequences, PCR primers and oligonucleotide probes were designed to detect, clone, and sequence the VSH-1 genes. RT-PCR was used to analyze VSH-1 transcripts in mitomycin C-treated cultures. Genes encoding VSH-1 virion-associated proteins and sharing the same transcriptional orientation comprised a contiguous14.2 kb region. Three ORFs that included one with 52% amino acid identity to a Treponema pallidum hypothetical glutamate/aspartate transport protein (TpE/D) were located upstream and in the opposite direction of transcription. Five ORFs, located downstream, included a tandem pair with 34-39% amino acid identity to putative methyl accepting chemotaxis proteins (MCPs). RT-PCR demonstrated induced transcription of svp38 (VSH-1 head protein), svp101 (VSH-1 tail protein) and MCP genes 1.5 to 2.5 hours after B. hyo cultures were treated with mitomycin C. In contrast, the B. hyo flagellar gene (flaA1) was constitutively transcribed in non-induced cultures but mRNA levels diminished over time in mitomycin C induced cultures. No increase in transcription was detected for the TpE/D. These methods will be applied to identify and sequence genes involved in regulating VSH-1, defining the organization of VSH-1 transcripts, and screening for alternative inducers of VSH-1.