Detection methods for Legionella pneumophila in diverse environmental conditions: A comparative study of FISH, seminested PCR and conventional culture

Abstract

Aims: Legionella bacteria cause Legionnaires' disease and Pontiac fever. It is commonly found in natural water resources and manmade water systems. Environmental conditions such as nutrient deficiency, temperature, pH, disinfectant and the presence of other bacteria can cause Legionella bacteria to pass into the viable but not-culturable (VBNC) phase. This study was aimed to determine appropriate methods to detect Legionella pneumophila bacteria living in water systems with wide temperature and pH ranges threatening human health. Methodology and results: In this study, water samples containing L. pneumophila at a concentration of 10 8 cell/L were exposed to different temperatures (5 °C, 50 °C, 55 °C and 60 °C) and pH (2.2, 5.8, 7.0 and 8.2) values. Conventional culture, FISH and seminested PCR methods were used to detect L. pneumophila . A comparison was made between the methods used in the study to determine the most appropriate method for detecting L. pneumophila bacteria. The results showed that the highest detection rates of L. pneumophila were at 5 °C for 24 h (100%) and at pH 2.2 for 0 th min (100%) by using FISH method. All the samples could be determined by the seminested PCR method. The results of our study showed that the highest detection rates of L. pneumophila were at 5 °C for 24 h (100%) and at pH 2.2 for 0 min (100%) by FISH method. All of the samples could be determined by the seminested PCR method. It was determined that the detection rate was the lowest in the FISH method at 3 min at 60 °C and the highest was 24 h at 5 °C. The lowest detection rate was also observed by using FISH method in the samples exposed to 60 °C for 3 min. Results show that the FISH and seminested PCR methods are the most suitable for detecting L. pneumophila bacteria from water systems exposed to different environmental conditions. Conclusion, significance and impact of study: Different methods (conventional culture, FISH, seminested PCR) used to detect L. pneumophila bacteria were compared in this study. It was concluded that Legionella bacteria passed into the VBNC phase, and compared to molecular methods, the conventional culture method provides a low detection rate of these bacteria. Research findings suggest that it is insufficient to use the conventional culture method alone for the detection of Legionella bacteria from man-made water systems or human samples. This study is important as it is decisive for the determination of the most appropriate method for detecting the human pathogen L. pneumophila bacteria from water samples and the choice for a fast and effective method for the elimination of the infectious agent.