Validation of the Legionella pneumophila SG1 DETECT Kit for Quantification of Legionella pneumophila Serogroup 1 Bacteria in Potable Waters, Process Waters, and Surface Waters: AOAC Performance Tested MethodSM 052002

Abstract

The L.p.SG1 DETECT Kit is a rapid, quantitative method for the detection and enumeration of Legionella pneumophila serogroup 1 (L.p. SG1) bacteria from different water matrixes. The method is based on a combination of immunomagnetic separation (IMS) and flow cytometric (FCM) quantification. To this end, the method employs magnetic particles conjugated to anti-L.p. SG1 antibodies for the IMS of the target bacteria from environmental matrices and fluorescently labelled anti-L.p. SG1 antibodies for subsequent quantification by FCM. The IMS can be performed either manually with a magnetic rack (rqmicro.MIMS) or automated with the rqmicro.STREAM sample preparation instrument. Compared to the reference method ISO 11731:2017, which is based on culturing and enumeration of colony forming units (CFU) on agar plates, and can take up to 10 days until results are available, analysis with the L.p. SG1 DETECT Kit is culture-independent and delivers results within 2 h. This Performance Tested Method validation study demonstrates a robust method with recoveries exceeding 69%, inclusivity of 100%, exclusivity of 97.2% and a shelf life of at least 6 months at 4 °C or 40 days at 25 °C. The Limit of Detection (LoD) was determined at 21 CFU/L and the Limit of Quantitation (LoQ) at 80 CFU/L for potable water using the rqmicro.STREAM. The matrix study across 3 different types of water matrixes (potable, surface and industrial process water), demonstrates superior repeatability and reproducibility, as well as equivalent or even superior detection of L.p. SG1 bacteria compared to the standard ISO 11731 method.

Culture-Independent Quantification of Legionella pneumophila in Evaporative Cooling Systems Using Immunomagnetic Separation Coupled with Flow Cytometry

“…IMS uses the interaction of cell surface antigens and antibodies attached to magnetic beads, and cells can be extracted by placing the bead cell suspension into a magnetic field [24]. The combination of IMS and FCM was used in earlier studies to quantify L. pneumophila Sg 1 in tap water [25,26] and bioaerosols [27]. Based on these results, the analytical system rqmicro.COUNT has been further developed so that IMS and FCM can be implemented automatically and simultaneously in one device (Figure 1) using a specific panel of antibodies against L. pneumophila Sg 1-15 for the selective detection of viable cells.…”

Evaluation of a Most Probable Number Method for Detection and Quantification of Legionella pneumophila

“…At the same time, it also requires expensive professional equipment and trained operators. Other methods have also been reported for the detection of L. pneumophila in water but are less common, including IMS [ 29 , 30 ], electrochemical genosensors [ 31 ] and biosensor-based detection methods [ 19 ].…”