Validation of Polymerase Chain Reaction Method for the Detection of Legionella pneumophila
Keywords:
Polymerase chain reaction, Legionella pneumophila, Legionaires’ disease, mip gene, Water sourcesAbstract
Legionella pneumophila, the main cause of Legionnaires’ disease, is found in various water sources. The bacterium is spread to humans through the inhalation of contaminated aerosols. Surveillance of pathogens in water sources is therefore essential. The standard method for the detection
of L. pneumophila is culture. This requires an expert to read the results, and it takes a long time. This study aimed to develop and validate a PCR method for rapid detection of L. pneumophila with high specificity but without the requirement for specialists. Many samples were tested using the macrophage infectivity potentiator (mip) gene as the target, and the results were compared with the culture method to measure consistency. The PCR product of the developed PCR method was 265 bp, with a limit of detection equal to 102 CFU/mL, without cross-reactivity with 100 isolates of Legionella and 10 common waterborne bacteria. Comparing the effectiveness with culture using water samples, the results showed a well-correlated agreement (kappa = 0.81) and a negative predictive value of 100%. In addition, when bacterial colonies suspected of being L. pneumophila were used, the comparison results indicated a perfect agreement (kappa = 1.00). However, the PCR method provided the results eight times faster than the culture method. Thus, the developed PCR method is suitable as an alternative method for the surveillance and the control of L. pneumophila contamination in water resources. It can also be used as a method for the confirmation of suspected L. pneumophila bacterial colonies in addition to the existing culture method for faster analysis results.
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