Quantifying planktonic and particle-associated Legionella in building plumbing and drinking water distribution systems across the U.S.

Professor and J. Neils Thompson Centennial Teaching Fellow in Civil Engineering

University of Texas at Austin

Legionella has been the most frequently reported cause of waterborne disease outbreaks in the U.S. over the last decade. While Legionella in building plumbing has been studied extensively, occurrence data for Legionella spp. in U.S. drinking water distribution systems (DWDSs) are limited. Free-living amoebae (FLA) present in DWDSs have the ability to phagocytose Legionella pneumophila (Lp) without digesting them and to protect them from disinfectants. Furthermore, Lp that have undergone “protozoan priming” (i.e., living intracellular to FLA) exhibit augmented pathogenesis as compared to Lp that are extracellular to FLA. The goal of this work was to quantify planktonic and particle-associated Legionella spp. and Lp from first-draw and flushed-water samples from drinking water taps in 19 cities across the U.S. A total of 114 samples from cold water taps were collected in Summer (August-September 2022), Fall (October-November 2022) and Winter (January 2023). The cities were selected based on geographic diversity, source water type, and secondary disinfectant. The samples were filtered through a 3-μm polycarbonate filter to capture particle-associated bacteria, which could include Legionella intracellular to FLA. The filtrate was filtered through a 0.4-μm polycarbonate filter to capture planktonic bacteria. After DNA extraction, the samples were analyzed for Legionella spp. and Lp via digital polymerase chain reaction; additionally, Legionella-positive DNA samples were sent for Legionella-specific 16S rRNA gene sequencing.

Flushing decreased Legionella spp. concentrations at some, but not all, locations. Source water appears to have a significant effect on Legionella concentration, with higher Legionella spp. concentrations in surface water systems than in groundwater systems (p = 0.020). Most sampled sites had substantially greater concentrations of planktonic than particle-associated Legionella spp. While Legionella spp. were detected in the drinking water of all 19 cities, Lp detection was geographically and temporally sporadic. Besides Lp, the sequencing results demonstrated the occurrence of several other clinically-relevant Legionella spp. in the sampled drinking waters. This work is being continued on a broader scale, where the DWDSs of 50 utilities are being sampled for a variety of opportunistic pathogens, including Lp. The goal is to identify the most important water quality and DWDS factors that control the prevalence of opportunistic pathogens, such that mitigation strategies can be designed to reduce the risk to the consumer.

Mary Jo Kirisits is a professor and the J. Neils Thompson Centennial Teaching Fellow in Civil Engineering in the Environmental and Water Resources Engineering program at The University of Texas (UT) at Austin. She completed her BS degree in Civil Engineering at the State University of New York at Buffalo and her MS and PhD degrees in Environmental Engineering at the University of Illinois at Urbana-Champaign. After concluding a postdoctoral appointment at Northwestern University in the Department of Civil and Environmental Engineering, she joined the faculty at UT. Her research interests include drinking water biofiltration, opportunistic human pathogens, SARS-CoV-2 in wastewater, and the impact of nanomaterials on antimicrobial resistance in bacteria. She currently is leading a multi-institution project funded by the U.S. EPA on opportunistic pathogens and disinfection by-products in drinking water distribution systems.