USING REAL-TIME POLYMERASE CHAIN REACTION FOR DETECTION OF LEGIONELLA IN OBJECTS OF THE ENVIRONMENT

Aim. To assess efficacy of using the method of quantitative detection of Legionella in objects of the environment by real-time polymerase chain reaction (RT-PCR). Materials and methods. For the development of the assay, genus-specific primers from gene coding 16S rRNA as well as species-specific primers for detection of Legionella pneumophila on the basis of mip gene sequence. For quantitative detection of L . pneumophila calibration samples of pGEM plasmid containing fragment of the mip gene in known concentration were used. Samples of water and biofilms obtained from cooling stacks of production plants, systems of autonomic water supply, humidification blocks of centralized systems of air conditioning were studied. Results. Correlation of results obtained with RT-PCR and bacteriologic methods was shown during monitoring of potentially dangerous water objects as well as during epidemic outbreak of Legionella infection. Importance of samples preparation stage, during which considerable losses of DNA and inhibition of reaction could occur, is underlined. Disinfection measures on the studied objects significantly influenced on the results of the RT-PCR and can lead to false positive results. Conclusion. Obtained results confirm usefulness of testing of potentially dangerous water objects on the presence of Legionella based on the preliminary screening with RT-PCR for the 24 hours followed by bacteriologic testing of samples for 8 — 12 days.

Legionella, RT-PCR, indication of causative agent, water objects