Effect of Intracellular Resuscitation of Legionella pneumophila in Acanthamoeba polyphage Cells on the Antimicrobial Properties of Silver and Copper

Abstract

The property of Legionella pneumophila entering into a viable but noncultivable (VBNC) state under drinking water conditions (50 mL, pH 7.0, and 25 degrees C) and the intracellular resuscitation in Acanthamoeba polyphage cells were investigated. Then, the survival profiles of L. pneumophila residing in the planktonic phase and the endosymbiosis phase against antimicrobial silver and copper reagents were differentially compared with the case of Pseudomonas aeruginosa. The number of L. pneumophila in a cultivable state was rapidly reduced to below the detection limit (5.0 log reduction) within 30 days of incubation in synthetic drinking water, while the number of L. pneumophila in a viable state varied in only 0.1 log reduction during the same period, and the levels were sustained constantly for 190 days; in contrast, P. aeruginosa multiplied even in drinking water and continuously maintained its cultivability and viabilityfor 190 days. Distinctively, the numbers of E. coli in both cultivable and viable states were simultaneously diminished as 3.0 log and 1.6 log reduction. The cultivability of L. pneumophila in the VBNC state was recovered and started to multiply after coincubation with A. polyphage in the same environment (initial population of inoculated amoeba was adjusted as 1.0 x 10(5) amoeba/ mL), and P. aeruginosa also multiplied in amoeba cells. Finally, the populations of L. pneumophila in the planktonic phase after 10 days coincubation were detected at 1.7 x 10(7) CFU/mL, and this population was considered to have originated from the release of bacteria residing inside amoeba caused by the destruction of amoeba cells. Bacteria in the planktonic phase that were exposed to silver and copper were completely inactivated (more than 7 log reduction) within 30 min, while bacteria in the endosymbiosis phase showed much higher resistance against the exposure to the same concentrations of silver and copper. L. pneumophila and P. aeruginosa in A. polyphage cells survived to levels of 5.6 x 10(1) and 1.1 x 10(1) CFU/mL at the silver exposure (0.1 mgAg/L) and 7.3 x 10(3) and 6.1 x 10(4) CFU/ mL at the copper exposure (1.0 mgCu/L), respectively, after 7 days.