Identification of plasmids by PCR-based replicon typing

• Published in: Journal of microbiological methods Vol. 63; no. 3; pp. 219 - 228
• Main Authors: Carattoli, Alessandra, Bertini, Alessia, Villa, Laura, Falbo, Vincenzo, Hopkins, Katie L., Threlfall, E. John
• Format: Journal Article
• Language: English
• Published: Shannon Elsevier B.V 01.12.2005; Elsevier Science
• Subjects: Bacterial Typing Techniques - methods; Bacteriological methods and techniques used in bacteriology; Bacteriology; Base Sequence; Biological and medical sciences; Biotechnology; DNA, Bacterial - genetics; Drug Resistance, Multiple, Bacterial - genetics; Enterobacteriaceae; Enterobacteriaceae - drug effects; Enterobacteriaceae - genetics; Escherichia coli - classification; Escherichia coli - drug effects; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Genetic engineering; Genetic technics; Humans; In vitro gene amplification. Pcr technique; Methods. Procedures. Technologies; Microbiology; Molecular Epidemiology; PCR; Plasmids; Plasmids - genetics; Polymerase Chain Reaction - methods; Replicon; Replicon - genetics; Salmonella enterica; Salmonella enterica - classification; Salmonella enterica - drug effects; Salmonella enterica - genetics; Salmonella enterica - isolation & purification; Serotyping; Plasmids; PCR; Replicon; Plasmid; Polymerase chain reaction; Typing; Bacteria; Identification; Oligonucleotide; Enterobacteriaceae
• ISSN: 0167-7012; 1872-8359
• DOI: 10.1016/j.mimet.2005.03.018

The epidemiological importance of tracing plasmids conferring drug resistance prompted us to develop a PCR method based on replicons (inc/rep PCR) of the major plasmid incompatibility groups among Enterobacteriaceae. Eighteen pairs of primers were designed to perform 5 multiplex- and 3 simplex-PCRs, recognizing FIA, FIB, FIC, HI1, HI2, I1-Iγ, L/M, N, P, W, T, A/C, K, B/O, X, Y, F, and FIIA. The specificity of the method was tested on a collection of 61 reference plasmids and on 20 Salmonella enterica strains of different serotypes isolated in Italy. Results indicated that the inc/rep PCR method demonstrates high specificity and sensitivity in detecting replicons on reference plasmids and also revealed the presence of recurrent and common plasmids in epidemiologically unrelated Salmonella isolates of different serotypes. These results suggest that the method is potentially applicable to a large number of strains to trace the diffusion of specific multi-drug resistance plasmids in different environments.