title: Genetic characterization of plasmid-associated triphenylmethane reductase in Listeria monocytogenes.
abstract: The enzyme triphenylmethane reductase (TMR) reduces toxic triphenylmethane dyes into colorless, nontoxic derivatives, and TMR-producing microorganisms have been proposed as bioremediation tools. Analysis of the genome of Listeria monocytogenes H7858 (1998-1999 hot dog outbreak) revealed that the plasmid (pLM80) of this strain harboring a gene cassette (bcrABC) conferring resistance to benzalkonium chloride (BC) and other quaternary ammonium disinfectants also harbored a gene (tmr) highly homologous to TMR-encoding genes from diverse Gram-negative bacteria. The pLM80-associated tmr was located two genes downstream of bcrABC as part of a putative IS1216 composite transposon. To confirm the role of tmr in triphenylmethane dye detoxification, we introduced various tmr-harboring fragments of pLM80 in a pLM80-cured derivative of strain H7550, from the same outbreak as H7858, and assessed the resistance of the constructs to the triphenylmethane dyes crystal violet (CV) and malachite green. Transcriptional and subcloning data suggest that the regulation of TMR is complex. Constructs harboring fragments spanning bcrABC and tmr were CV resistant, and in such constructs tmr transcription was induced by sublethal levels of either BC or CV. However, constructs harboring only tmr and its upstream intergenic region could also confer resistance to CV, albeit at lower levels. Screening a panel of BC-resistant L. monocytogenes strains revealed that all those harboring bcrABC and adjacent pLM80 sequences, including tmr, were resistant to CV and decolorized this dye. The findings suggest a potential role of TMR as a previously unknown adaptive attribute for environmental persistence of L. monocytogenes.
authors.affiliation: Departments of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA vikrant.dutta@gmail.com.
authors.forename: Vikrant
authors.initials: V
authors.lastname: Dutta
authors.affiliation: Biomanufacturing Training and Education Center, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Driss
authors.initials: D
authors.lastname: Elhanafi
authors.affiliation: Department of Statistics, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Osborne
authors.affiliation: Departments of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Mira Rakic
authors.initials: MR
authors.lastname: Martinez
authors.affiliation: Departments of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Sophia
authors.initials: S
authors.lastname: Kathariou
mesh_terms: D001548:Benzalkonium Compounds / Q000378:metabolism / Q000633:toxicity; D001711:Biotransformation; D004251:DNA Transposable Elements; D023061:Gene Order; D005840:Gentian Violet / Q000378:metabolism / Q000633:toxicity; D008089:Listeria monocytogenes / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids; D012394:Rosaniline Dyes / Q000378:metabolism / Q000633:toxicity; D014320:Triphenylmethyl Compounds / Q000378:metabolism*
chemical_list: D001548:Benzalkonium Compounds; D004251:DNA Transposable Elements; D012394:Rosaniline Dyes; D014320:Triphenylmethyl Compounds; C005095:malachite green; C046945:triphenylmethane; D010088:Oxidoreductases; D005840:Gentian Violet