Submitted to: Anaerobe
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/19/2007
Publication Date: 4/19/2007
Citation: Matson, E.G., Zuerner, R.L., Stanton, T.B. 2007. Induction and transcription of VSH-1, a prophage-like, gene transfer agent of Brachyspira hyodysenteriae. Anaerobe. 13(2):89-97. Interpretive Summary: Our previous research discovered a novel bacterial virus in the swine pathogenic bacterium known as Brachyspira hyodysenteriae. We named this virus VSH-1. VSH-1 can transfer genes, including antibiotic resistance genes, between B. hyodysenteriae cells. We are identifying the conditions that make VSH-1 grow within B. hyodysenteriae cells so we can understand how antibiotic resistance genes spread and are maintained in the animal intestinal tract. In this study, we investigated the growth of VSH-1 inside B. hyodysenteriae by measuring molecules that VSH-1 uses to makes its proteins. These molecules are known as messenger RNA or mRNA. We found that VSH-1 mRNA is made as a long molecule and identified the start of that mRNA molecule. We found that mRNA made by VSH-1 can be detected two hours after the start of the growth cycle of VSH-1. This knowledge is important because it has enabled us to develop molecular assays to detect chemicals, environmental conditions, and swine cell products that stimulate the growth of VSH-1 and the spread of antibiotic resistance genes. Characterizing novel gene transfer mechanisms like VSH-1 and identifying conditions that stimulate the spread of antibiotic resistance in the intestinal tract are important steps towards reducing antibiotic resistant bacteria. These findings are interesting and useful for representatives of the pharmaceutical industry and research scientists investigating antibiotic resistance.
Technical Abstract: Brachyspira hyodysenteriae is an anaerobic spirochete and the etiologic agent of swine dysentery. B. hyodysenteriae cells harbor VSH-1, a mitomycin C-inducible prophage that packages random 7.5 kb fragments of host chromosomal DNA and mediates generalized transduction of host genes. VSH-1-like entities are widely distributed in Brachyspria species and have likely contributed to shaping the population structure of B. hyodysenteriae. VSH-1 late-genes for viron structures and host-cell escape were recently identified in a 16.3 kb region of the B. hyodysenteriae chromosome, which allowed the current analyses of VSH-1 transcription. RT-PCR and hybridization experiments using RNA collected from B. hyodysenteriae cells at various time points during VSH-1 induction determined that the abundance of VSH-1 mRNA increased dramatically between 2 and 4 h in cells treated with mitomycin C and similarly with H2O2. In contrast, the mRNA level of a B. hyodysenteriae housekeeping gene, flaA1, concurrently decreased when cells were exposed to these inducing agents. Chloramphenicol co-treatment appeared to abrogate the effects of mitomycin C. VSH-1 genes are co-transcribed as a polycistronic operon that initiates near the beginning of the prophage sequence. Information from these transcription analyses will likely be useful for the identifying proteins and genes that regulate VSH-1 production and as well as environmental inducers of this novel gene transfer agent.