Prevention of horizontal transfer of laboratory plasmids to environmental bacteria: how to select disinfection methods that are also effective in destroying DNA

Abstract

Routine work of any molecular biology laboratory includes the daily use of microorganisms, including non-pathogenic E coli strains, transformed with a variety of plasmids expressing at least one antibiotic resistance gene (ARG). To check the efficacy of neutralization methods, bacteria isolated from wastewater from laboratories and research institutes were identified using the 16S ribosomal RNA sequencing and they were tested for the presence of an origin of replication and several ARGs frequently found in laboratory plasmids. Surprisingly, the origin of replication of Enterobacteriaceae was detected in non-Enterobacteriaceae bacteria strains suggesting that an interspecies transfer of laboratory plasmids had occurred. Using quantitative Polymerase Chain Reaction (qPCR), we determined the Decimal reduction value (D-value, expressed as concentration of disinfectant or length of physical treatment) of several neutralization methods for their DNA-bd effect: i.e. 0.7M for Sulfuric, 3.2% for a commercial disinfectant P3, 25 minutes for steam sterilization at 121°C and 49 minutes for UVC. A 20-minute treatment of laboratory liquid waste (LLW) with 1–10% sodium chlorine is ineffective in destroying completely plasmid gene markers intreated cells. Therefore, to prevent the horizontal transfer of laboratory ARGs to environmental bacteria, bleach-treated liquid waste should not be released in sewage without additional plasmid destruction treatment.