Title

Detection and Monitoring of Environmental Triclosan Degradation Gene Expression in Situ

Item Type

Presentation

Location

Elaine Langone Center, Forum

Session

Ecology and Water Quality

Start Date

12-11-2016 1:30 PM

End Date

12-11-2016 4:00 PM

Keywords

water quality, plasmid, microbiology, antibiotic

Description

Triclosan (TCS) is a broad-spectrum antimicrobial that blocks the active site of the enoyl-acyl carrier protein reductase enzyme, therefore preventing the formation of fatty acids in bacteria. Due to TCS being a potential environmental and health hazard, it was recently included on a list of antibacterial compounds banned by the FDA. Sphingomonas sp. RD1 is known to degrade TCS utilizing a TCS-inducible oxygenase encoded by the genes tcsAB. In order to detect to use of tcsAB for degradation of TCS in situ, a PCR-produced DNA fragment containing the tcsAB promoter region was inserted upstream of the lux cassette of pUCD615, thus creating a plasmid that will luminesce when activated by TCS presence. The plasmid was transformed into RD1 creating a TCS presence reporter strain for application in wastewater discharge. In addition, environmental presence of this pathway was investigated by two approaches: 1) BLAST searches of publicly available metagenomes of sewage and activated sludge and 2) tcsA PCR on DNA extracted from wastewater treatments in south central Pennsylvania. Although no instances of tcsA were detected in metagenomes, PCR (confirmed by sequencing) indicated the presence of tcsA in one sewage sludge sample. Attempts to isolate the tcsA-containing bacteria from that sludge sample have resulted in one putative tcsA-containing strain.

Language

eng

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Nov 12th, 1:30 PM Nov 12th, 4:00 PM

Detection and Monitoring of Environmental Triclosan Degradation Gene Expression in Situ

Elaine Langone Center, Forum

Triclosan (TCS) is a broad-spectrum antimicrobial that blocks the active site of the enoyl-acyl carrier protein reductase enzyme, therefore preventing the formation of fatty acids in bacteria. Due to TCS being a potential environmental and health hazard, it was recently included on a list of antibacterial compounds banned by the FDA. Sphingomonas sp. RD1 is known to degrade TCS utilizing a TCS-inducible oxygenase encoded by the genes tcsAB. In order to detect to use of tcsAB for degradation of TCS in situ, a PCR-produced DNA fragment containing the tcsAB promoter region was inserted upstream of the lux cassette of pUCD615, thus creating a plasmid that will luminesce when activated by TCS presence. The plasmid was transformed into RD1 creating a TCS presence reporter strain for application in wastewater discharge. In addition, environmental presence of this pathway was investigated by two approaches: 1) BLAST searches of publicly available metagenomes of sewage and activated sludge and 2) tcsA PCR on DNA extracted from wastewater treatments in south central Pennsylvania. Although no instances of tcsA were detected in metagenomes, PCR (confirmed by sequencing) indicated the presence of tcsA in one sewage sludge sample. Attempts to isolate the tcsA-containing bacteria from that sludge sample have resulted in one putative tcsA-containing strain.