Use of low-frequency-cleavage restriction endonucleases for DNA analysis in epidemiological investigations of nosocomial bacterial infections

Abstract

Epidemiological investigations of bacterial infections are generally based on multiple phenotypic markers that are often difficult to verify. A more general and reliable method is genomic DNA analysis by restriction endonucleases. However, the commonly used endonucleases produce too many fragments for correct separation by agarose electrophoresis. In contrast, simple electrophoretic patterns are obtained after genomic DNA digestion by low-frequency-cleavage restriction endonucleases and pulsed-field gel electrophoresis, making it easier to compare numerous strains from the same species. This technique was used to investigate an Acinetobacter calcoaceticus outbreak in a urologic department and bronchial colonization of artificially ventilated patients by Pseudomonas aeruginosa in an intensive care unit. The method allowed a clear distinction between epidemic and self-contaminating strains in these different epidemiological situations.

Comparison of four different methods for epidemiologic typing of Acinetobacter baumannii

“…3, lanes 2 to 4 and 5 to 7). Among isolates representing PFGE type h, two variants were detected that differed in one band (data not 1 Table 1; 1 and 2, biotype 9 isolates from hospital C (plasmid type V); 3 and 4, biotype 9 isolates from hospital C (plasmid type Va); 5 and 6, biotype 9 isolates from hospital C (plasmid type VI); 7 and 8, biotype 9 isolates from hospital C (plasmid type VIa); 9 and 10, biotype 6 isolates from hospital A (plasmid type I); 11 Table 1; 2 to 5, biotype 1 isolates from hospital A (PFGE type a); 6 to 9, biotype 2 isolates from hospital C (PFGE type e); 10 to 12, biotype 9 isolates from hospital C (PFGE type g); 13 to 16, biotype 9 isolates from hospital C (PFGE type f); 17 to 19, biotype 9 isolates from hospital C (PFGE type h).…”

“…None of these techniques is capable of typing all strains. More recently, genotypic methods such as analysis of plasmid profiles (13,17), ribotyping (7,9), analysis of chromosomal DNA by pulsed-field gel electrophoresis (PFGE) (1,11), and fingerprinting by arbitrarily primed PCR (12) have been introduced and have improved epidemiological typing of A.…”

Is biofilm formation related to the hypermutator phenotype in clinicalEnterobacteriaceaeisolates?

“…Patients with polymicrobial bacteraemia were excluded. Potential clonality of the isolates was studied using pulse-field gel electrophoresis (PFGE) (Allardet-Servent et al, 1989). PFGE patterns were compared by calculating the Dice correlation coefficients using GELCOMPAR II software (Applied Maths, St-Martens-Latem, Belgium) and were clustered into a dendrogram using the unweighted pair group method (tolerance, 2.0%).…”

Subtracted restriction fingerprinting – a new typing technique using magnetic capture of tagged restriction fragments

“…Macrorestriction analysis describes the digestion of whole cell DNA with a rare‐cutting endonuclease and subsequent analysis of the relatively small number of large fragments by pulsed‐field gel electrophoresis (PFGE) [39]. Macrorestriction analysis is currently the method of choice for typing Salmonella [9–11,18,20] and other pathogens in many laboratories.…”