Whole-cell repetitive element sequence-based polymerase chain reaction allows rapid assessment of clonal relationships of bacterial isolates

Repetitive element sequence-based polymerase chain reaction (rep-PCR) enables the generation of DNA fingerprints which discriminate bacterial species and strains. We describe the application ofwhole-cell methods which allow specimens from broth cultures or colonies from agar plates to be utilized directly in the PCR mixture. The rep-PCR-generated DNA fingerprints obtained with whole-cell samples match results obtained with genomic DNA templates. Examples with different gram-negative bacteria (e.g., Citrobacter diversus, Escherichia coli, and Pseudomonas aeruginosa) and gram-positive bacteria (e.g., Staphylococcus aureus and Streptococcus pneumoniae) are demonstrated. Rapid specimen preparation methods enable rep-PCR-based fingerprinting to be completed in several hours and, therefore, allows the timely analysis of epidemiological relationships. Discriminatory molecular methods such as plasmid profil-ing, restriction endonuclease digestion of plasmid or chro-mosomal DNA, ribotyping, and multilocus enzyme electro-phoresis (MEE) have been applied retrospectively to study relationships among isolates from apparent clinical out