Abstract
BackgroundLegionellosis is an uncommon form of pneumonia. After a clinical encounter, the necessary antibiotic treatment is available if the diagnosis is made early in the illness. Before the clinical encounter, early detection of the main pathogen involved, Legionella pneumophila, in hazardous environments is important in preventing infectious levels of this bacterium. In this study a qualitative test based on combined magnetic immunocapture and enzyme-immunoassay for the fast detection of Legionella pneumophila in water samples was compared with the standard method, in both comparative and collaborative trials. The test was based on the use of anti-Legionella pneumophila antibodies immobilized on magnetic microspheres. The final protocol included concentration by filtration, resuspension and immunomagnetic capture. The whole assay took less than 1 hour to complete.ResultsA comparative trial was performed against the standard culture method (ISO 11731) on both artificially and naturally contaminated water samples, for two matrices: chlorinated tap water and cooling tower water. Performance characteristics of the test used as screening with culture confirmation resulted in sensitivity, specificity, false positive, false negative, and efficiency of 96.6%, 100%, 0%, 3.4%, and 97.8%, respectively. The detection limit at the level under which the false negative rate increases to 50% (LOD50) was 93 colony forming units (CFU) in the volume examined for both tested matrices. The collaborative trial included twelve laboratories. Water samples spiked with certified reference materials were tested. In this study the coincidence level between the two methods was 95.8%.ConclusionResults demonstrate the applicability of this immunosensing technique to the rapid, simple, and efficient detection of Legionella pneumophila in water samples. This test is not based on microbial growth, so it could be used as a rapid screening technique for the detection of L. pneumophila in waters, maintaining the performance of conventional culture for isolation of the pathogen and related studies.
Highlights
Legionellosis is an uncommon form of pneumonia
Comparative trial with natural samples The immunomagnetic method (IMM) test was applicable to detection of L. pneumophila in water samples
A total of 459 water samples, comprising both naturally contaminated and artificially contaminated samples were examined for the presence of L. pneumophila using the reference culture method (ISO 11731-Part 1) and the IMM test in parallel
Summary
Legionellosis is an uncommon form of pneumonia. After a clinical encounter, the necessary antibiotic treatment is available if the diagnosis is made early in the illness. Early detection of the main pathogen involved, Legionella pneumophila, in hazardous environments is important in preventing infectious levels of this bacterium. Patients with immuno-compromised status are susceptible to this atypical pneumonia [7] This pathogen is present in both natural [6] and man-made [7] water environments like cooling towers, evaporative condensers, humidifiers, potable water systems, decorative fountains and wastewater systems (risk facilities). Rapid fluctuations of the concentration of L. pneumophila at risk facilities have been reported [11], as well as persistence of L. pneumophila in drinking water biofilms mostly in a viable but non-culturable state (VBNC) [12], which has been confirmed even after treatments with chlorine used to disinfect cooling towers [13,14]. The effectiveness of treatments on Legionella pneumophila (chlorine, heat, ozone, UV, monochloramine) has been mainly evaluated based on cultivability and that could not be a real indicative of the absence of intact viable cells [16,17,18]
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