TY - JOUR
T1 - Prevention of horizontal transfer of laboratory plasmids to environmental bacteria
T2 - comparison of the effectiveness of a few disinfection approaches to degrade DNA
AU - Loret, Suzanne
AU - Habib, Boutaina
AU - Romain, Pierre
AU - Roba, Agnès
AU - Reboul, Angélina
N1 - Funding Information:
This work was supported by a grant from the continued education in Biosafety programmed developed by the prevention department (SerP) of the University of Namur. Boutaina Habib benefited from a PhD grant of the University Mohammed V (Rabat, Morocco), completed by a UNamur Grant for foreign students. Agnès Roba benefited from a PhD grant from the Belgian National Fund for Scientific Research.
Funding Information:
The authors warmly thank Dr. Karim Zouai (Vésale Hospital, Charleroi, Belgium); Salah Eddine Lali and Barbara Marchetti (Marie Curie Hospital, Charleroi, Belgium); Pierre Bogaert, head of the molecular diagnostic laboratory (CHU UCL, Namur, Belgium); and Te-Din Daniel Huang, head of the clinical laboratory (CHU UCL, Namur, Belgium) for the fruitful scientific discussion on our data. We also thank Professor Bouchra Belkadi (Department of Biology of the University Mohammed V, Rabat, Morocco) for his support for Boutaina Habib's research stay at the University of Namur. We are particularly grateful to Professor Xavier De Bolle for allowing us to carry out this study in his laboratory and to benefit from the necessary equipment.
Publisher Copyright:
© 2023, The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.
PY - 2023/8
Y1 - 2023/8
N2 - The routine work of any molecular biology laboratory includes the daily use of microorganisms, including strains of E. coli, transformed with a variety of plasmids expressing at least one antibiotic resistance gene (ARG). Therefore, to avoid the accidental release of ARGs into environmental water, methods for disinfection of liquid laboratory waste must be effective in destroying nucleic acids. In support of this recommendation, the origin of replication of Enterobacteriaceae plasmids has been detected in strains of non-Enterobacteriaceae bacteria isolated from wastewater from laboratories and research institutes, suggesting that interspecific transfer of laboratory plasmids had occurred. Using quantitative polymerase chain reaction, we determined the decimal reduction value (D value, expressed as concentration of disinfectant or length of physical treatment) of several decontamination methods for their DNA degradation effect on cultures of E. coli Top10 transformed with a kanamycin resistant plasmid (pET28A + or pEGFP-C2). The estimated D values were 0.7 M for sulfuric acid, 6.3% for a commercial P3 disinfectant, 25 min for steam sterilization at 121 °C, and 49 min for disinfection by UVC. A 20-min treatment of bacteria cultures with a final concentration of 1–10% sodium hypochlorite was found to be ineffective in completely destroying a bacteria plasmid gene marker (coding for the pBR322 origin of replication). Residual DNA from NaClO-treated cells was 60%, while it decreased under 10% using the commercial disinfectant P3 diluted at 5%. As the degradation was incomplete in both cases, we recommend avoiding discharge of disinfected liquid waste to wastewater (even after chemical neutralization) without additional plasmid destruction treatment, to prevent horizontal transfer of laboratory ARGs to environmental bacteria.
AB - The routine work of any molecular biology laboratory includes the daily use of microorganisms, including strains of E. coli, transformed with a variety of plasmids expressing at least one antibiotic resistance gene (ARG). Therefore, to avoid the accidental release of ARGs into environmental water, methods for disinfection of liquid laboratory waste must be effective in destroying nucleic acids. In support of this recommendation, the origin of replication of Enterobacteriaceae plasmids has been detected in strains of non-Enterobacteriaceae bacteria isolated from wastewater from laboratories and research institutes, suggesting that interspecific transfer of laboratory plasmids had occurred. Using quantitative polymerase chain reaction, we determined the decimal reduction value (D value, expressed as concentration of disinfectant or length of physical treatment) of several decontamination methods for their DNA degradation effect on cultures of E. coli Top10 transformed with a kanamycin resistant plasmid (pET28A + or pEGFP-C2). The estimated D values were 0.7 M for sulfuric acid, 6.3% for a commercial P3 disinfectant, 25 min for steam sterilization at 121 °C, and 49 min for disinfection by UVC. A 20-min treatment of bacteria cultures with a final concentration of 1–10% sodium hypochlorite was found to be ineffective in completely destroying a bacteria plasmid gene marker (coding for the pBR322 origin of replication). Residual DNA from NaClO-treated cells was 60%, while it decreased under 10% using the commercial disinfectant P3 diluted at 5%. As the degradation was incomplete in both cases, we recommend avoiding discharge of disinfected liquid waste to wastewater (even after chemical neutralization) without additional plasmid destruction treatment, to prevent horizontal transfer of laboratory ARGs to environmental bacteria.
KW - Antibiotic resistance gene
KW - Antimicrobial resistance
KW - Biosafety
KW - Decontamination
KW - Horizontal gene transfer
KW - Plasmid transfer
UR - http://www.scopus.com/inward/record.url?scp=85164796513&partnerID=8YFLogxK
U2 - 10.1007/s11356-023-28733-0
DO - 10.1007/s11356-023-28733-0
M3 - Article
SN - 0944-1344
VL - 30
SP - 89369
EP - 89380
JO - Environmental Science and Pollution Research
JF - Environmental Science and Pollution Research
IS - 38
M1 - 10.1007/s11356-023-28733-0
ER -