Study of Legionella pneumophila treatment with copper in drinking water by single cell-ICP-MS

Legionella pneumophila is a persistent opportunistic pathogen that poses a significant threat to domestic water systems. Previous studies suggest that copper (Cu) is an effective antimicrobial in water systems. A rapid and sensitive quantification method is desired to optimize the conditions of L. pneumophila treatment by Cu and to better understand the interaction mechanisms between Cu and cells. In this study, we developed a highly sensitive single cell (SC)-ICP-MS method to monitor L. pneumophila cell concentration and track their uptake of Cu. The SC-ICP-MS method showed excellent sensitivity (with a cell concentration detection limit of 1000 cells/mL), accuracy (good agreement with conventional hemocytometry method), and precision (relative standard deviation < 5%) in drinking water matrix. The cupric ions (Cu²⁺) treatment results indicated that the total L. pneumophila cell concentration, Cu mass per cell, colony-forming unit counting, and Cu concentration in supernatant all exhibited a dose-dependent trend, with 800-1200 µg/L reaching high disinfection rates in drinking water. The investigation of percentages of viable and culturable, viable but nonculturable (VBNC), and lysed cells suggested there always were VBNC present at any Cu concentration. Experimental results of different Cu²⁺ treatment times further suggested that L. pneumophila cells developed an antimicrobial resistant mechanism with the prolonged Cu exposure. This is the first quantification study on the interactions of Cu and L. pneumophila in drinking water using SC-ICP-MS.