Detection of Diverse Sequence Types of Legionella pneumophila by Legiolert Enzymatic-Based Assay and the Development of a Long-Term Storage Protocol

Legiolert is a rapid culture-based enzymatic method for the detection and quantification of Legionella pneumophila in potable and nonpotable water samples. We aimed to assess the ability of this assay to detect diverse sequence types and validated a simple method to preserve samples. We used this assay on 253 potable and 165 nonpotable cooling tower water samples from various buildings in Quebec, Canada, and performed sequence-based typing on 96 isolates. Six sequence types were identified, including ST1, ST378, ST1427, ST2859, ST3054, and ST3069. Whole-genome sequencing revealed that ST2859 was a member of the L. pneumophila subspecies fraseri. Additional tests with pure isolates also found that subspecies Pascullei and Raphaeli could be detected via Legiolert. Eight storage methods, including the current recommendation to store Legiolert trays at 4 degrees C, were evaluated for their ability to preserve viable cultures. Of those, storage of Legiolert culture with 10% glycerol at -80 degrees C produced the best results, fully preserving culturable Legionella for at least 12.5 months. We incorporated these findings into a standard procedure for processing Legiolert packets. Overall, Legiolert captures a variety of common and new STs in addition to important L. pneumophila subspecies and can be easily stored, which allows the conservation of a population of isolates for later characterization.

Automated summary

The study assessed the capability of Legiolert to detect diverse sequence types of Legionella pneumophila in water samples and validated a simple preservation method. The results demonstrated that Legiolert can detect multiple sequence types and important L. pneumophila subspecies, and it can be easily stored for further characterization.