Legionella is an opportunistic waterborne pathogen that is difficult to eradicate in colonized drinking water pipes. Legionella control is further challenged by aging water infrastructure and lack of evidence-based guidance for building treatment. This study assessed multiple premise water remediation approaches designed to reduce Legionella pneumophila within a residential building located in an aging, urban drinking water system over a two-year period. Samples (n = 745) were collected from hot and cold-water lines and quantified via most probable number culture. Building-level treatment approaches included three single heat shocks, three single chemical shocks, and continuous low-level chemical disinfection in the potable water system. The building was highly colonized with L. pneumophila with 71 % L. pneumophila positivity. Single heat shocks had a statistically significant L. pneumophila reduction one day post treatment but no significant L. pneumophila reduction at one week, two weeks, and four weeks post treatment. The first two chemical shocks resulted in statistically significant L. pneumophila reduction at two days and four weeks post treatment, but there was a significant L. pneumophila increase at four weeks following the third chemical shock. Continuous low-level chemical disinfection resulted in statistically significant L. pneumophila reduction at ten weeks post treatment implementation. This demonstrates that in a building highly colonized with L. pneumophila, sustained remediation is best achieved using continuous low-level chemical treatment.

Water age in drinking water systems is often used as a proxy for water quality but is rarely used as a direct input in assessing microbial risk. This study directly linked water ages in a premise plumbing system to concentrations of Legionella pneumophila via a growth model. In turn, the L. pneumophila concentrations were used for a quantitative microbial risk assessment to calculate the associated probabilities of infection (Pinf) and clinically severe illness (Pcsi) due to showering. Risk reductions achieved by purging devices, which reduce water age, were also quantified. The median annual Pinf exceeded the commonly used 1 in 10,000 (10-4) risk benchmark in all scenarios, but the median annual Pcsi was always 1-3 orders of magnitude below 10-4. The median annual Pcsi was lower in homes with two occupants (4.7 × 10-7) than with one occupant (7.5 × 10-7) due to more frequent use of water fixtures, which reduced water ages. The median annual Pcsi for homes with one occupant was reduced by 39-43% with scheduled purging 1-2 times per day. Smart purging devices, which purge only after a certain period of nonuse, maintained these lower annual Pcsi values while reducing additional water consumption by 45-62%.

In-refrigerator water dispensing systems are ubiquitous in residential homes with tap water as the inflow. Passage through these systems resulted in significant microbial growth in the water, with the abundance of potential opportunistic pathogens Mycobacterium and Pseudomonas increasing by 8,053- and 221-fold, respectively. Elevated exposure to microbial contaminants linked to in-refrigerator water dispensing systems may represent a significant public health concern.

Although simulated studies have provided valuable knowledge regarding the communities of planktonic bacteria and biofilms, the lack of systematic field studies have hampered the understanding of microbiology in real-world service lines and premise plumbing. In this study, the bacterial communities of water and biofilm were explored, with a special focus on the lifetime development of biofilm communities and their key influencing factors. The 16S rRNA gene sequencing results showed that both the planktonic bacteria and biofilm were dominated by Proteobacteria. Among the 15,084 observed amplicon sequence variants (ASVs), the 33 core ASVs covered 72.8 %, while the 12 shared core ASVs accounted for 62.2 % of the total sequences. Remarkably, it was found that the species richness and diversity of biofilm communities correlated with pipe age. The relative abundance of ASV2 (f_Sphingomonadaceae) was lower for pipe ages 40-50 years (7.9 %) than for pipe ages 10-20 years (59.3 %), while the relative abundance of ASV10 (f_Hyphomonadaceae) was higher for pipe ages 40-50 years (19.5 %) than its presence at pipe ages 20-30 years (1.9 %). The community of the premise plumbing biofilm had significantly higher species richness and diversity than that of the service line, while the steel-plastics composite pipe interior lined with polyethylene (S-PE) harbored significantly more diverse biofilm than the galvanized steel pipes (S-Zn). Interestingly, S-PE was enriched with ASV27 (g_Mycobacterium), while S-Zn pipes were enriched with ASV13 (g_Pseudomonas). Moreover, the network analysis showed that five rare ASVs, not core ASVs, were keystone members in biofilm communities, indicating the importance of rare members in the function and stability of biofilm communities. This manuscript provides novel insights into real-world service lines and premise plumbing microbiology, regarding lifetime dynamics (pipe age 10-50 years), and the influences of pipe types (premise plumbing vs. service line) and pipe materials (S-Zn vs. S-PE).

Manual flushing of building plumbing is commonly used to address water quality issues that arise from water stagnation. Autonomous flushing informed by sensors has the potential to aid in the management of building plumbing, but a number of knowledge gaps hinder its application. This study evaluates autonomous flushing of building plumbing with online sensor and actuator nodes deployed under kitchen sinks in five residential houses. Online oxidation-reduction potential (ORP) and temperature data were collected for nine weeks during the winter and summer in houses with both free chlorine and chloramine. ORP levels in houses with free chlorine residuals decreased after overnight stagnation. The overnight decrease in ORP was not observed when tap water was automatically flushed for five minutes at 6:00 h every morning. ORP levels in houses with chloramine residuals did not decrease consistently after overnight stagnation, and daily automated flushes did not have an observable effect on the ORP signals. Additional laboratory experiments were carried out to evaluate ORP signals during chlorine decay and after incremental changes in chlorine, as would be expected in building plumbing conditions. Results from the lab and field deployments suggest on-line ORP sensors may be used to detect free chlorine decay due to stagnating water, but are not as effective in detecting chloramine decay. However, field results also suggest ORP may not respond as expected on a timely manner after free chlorine or chloramine have been restored, hindering their applicability in developing control algorithms. In this paper we tested twice-daily five-minute automatic flushing and found that it counteracts water quality degradation associated with overnight stagnation in free chlorine systems. An automatic sensor-based flushing is proposed using online temperature sensor data to determine when flushing has reached water from the main. The results suggest that flushing informed by temperature sensors can reduce the flushing time by 46 % compared to the preset five-minute static flush.

The regrowth and subsequent exposure of opportunistic pathogens (OPs) whilst reopening buildings that have been locked down due to the stay-at-home restrictions to limit the spread of COVID-19, is a public health concern. To better understand such microbiological risks due to lowered occupancy and water demand in buildings, first and post-flush water samples (n = 48) were sampled from 24 drinking water outlets from eight university buildings in two campuses (urban and rural), with various end-user occupancies. Both campuses were served with chlorinated water originating from a single drinking water distribution system in South-East Queensland, situated 14 km apart, where the rural campus had lower chlorine residuals. Culture-dependent and culture-independent methods (such as flow cytometry, qPCR and 16S rRNA gene amplicon sequencing) were used concurrently to comprehensively characterise the OPs of interest (Legionella spp., Pseudomonas aeruginosa, and nontuberculous mycobacteria (NTM)) and the premise plumbing microbiome. Results showed that buildings with extended levels of stagnation had higher and diverse levels of microbial growth, as observed in taxonomic structure and composition of the microbial communities. NTM were ubiquitous in all the outlets sampled, regardless of campus or end-user occupancy of the buildings. qPCR and culture demonstrated prevalent and higher concentrations of NTM in buildings (averaging 3.25 log10[estimated genomic copies/mL]) with extended stagnation in the urban campus. Furthermore, flushing the outlets for 30 minutes restored residual and total chlorine, and subsequently decreased the levels of Legionella by a reduction of 1 log. However, this approach was insufficient to restore total and residual chlorine levels for the outlets in the rural campus, where both Legionella and NTM levels detected by qPCR remained unchanged, regardless of building occupancy. Our findings highlight that regular monitoring of operational parameters such as residual chlorine levels, and the implementation of water risk management plans are important for non-healthcare public buildings, as the levels of OPs in these environments are typically not assessed.

A framework is needed to account for interactive effects of plumbing materials and disinfectants on opportunistic pathogens (OPs) in building water systems. Here we evaluated free chlorine, monochloramine, chlorine dioxide, and copper-silver ionization (CSI) for controlling Pseudomonas aeruginosa and Acinetobacter baumannii as two representative OPs that colonize hot water plumbing, in tests using polyvinylchloride (PVC), copper-PVC, and iron-PVC convectively-mixed pipe reactors (CMPRs). Pipe materials vulnerable to corrosion (i.e., iron and copper) altered the pH, dissolved oxygen, and disinfectant levels in a manner that influenced growth trends of the two OPs and total bacteria. P. aeruginosa grew well in PVC CMPRs, poorly in iron-PVC CMPRs, and was best controlled by CSI disinfection, whereas A. baumannii showed the opposite trend for pipe material and was better controlled by chlorine and chlorine dioxide. Various scenarios were identified in which pipe material and disinfectant can interact to either hinder or accelerate growth of OPs, illustrating the difficulties of controlling OPs in portions of plumbing systems experiencing warm, stagnant water.

Opportunistic premise plumbing pathogens (OPPPs) have been detected in buildings\' plumbing systems causing waterborne disease outbreaks in the United States. In this study, we monitored the occurrence of OPPPs along with free-living amoeba (FLA) and investigated the effects of residential activities in a simulated home plumbing system (HPS). Water samples were collected from various locations in the HPS and analyzed for three major OPPPs: Legionella pneumophila, nontuberculous mycobacterial species (e.g., Mycobacterium avium, M. intracellulare, and M. abscessus), and Pseudomonas aeruginosa along with two groups of amoebas (Acanthamoeba and Vermamoeba vermiformis). A metagenomic approach was also used to further characterize the microbial communities. Results show that the microbial community is highly diverse with evidence of spatial and temporal structuring influenced by environmental conditions. L. pneumophila was the most prevalent pathogen (86% of samples), followed by M. intracellulare (66%) and P. aeruginosa (21%). Interestingly, M. avium and M. abscessus were not detected in any samples. The data revealed a relatively low prevalence of Acanthamoeba spp. (4%), while V. vermiformis was widely detected (81%) across all the sampling locations within the HPS. Locations with a high concentration of L. pneumophila and M. intracellulare coincided with the highest detection of V. vermiformis, suggesting the potential growth of both populations within FLA and additional protection in drinking water. After a period of stagnation lasting at least 2-weeks, the concentrations of OPPPs and amoeba immediately increased and then decreased gradually back to the baseline. Furthermore, monitoring the microbial population after drainage of the hot water tank and partial drainage of the entire HPS demonstrated no significant mitigation of the selected OPPPs. This study demonstrates that these organisms can adjust to their environment during such events and may survive in biofilms and/or grow within FLA, protecting them from stressors in the supplied water.

As drinking water travels from its source, through various treatment processes, hundreds to thousands of kilometres of distribution network pipes, to the taps in private homes and public buildings, it is exposed to numerous environmental changes, as well as other microbes living in both water and on surfaces. This review aims to identify the key locations and factors that are associated with changes in the drinking water microbiome throughout conventional urban drinking water systems from the source to the tap water. Over the past 15 years, improvements in cultivation-independent methods have enabled studies that allow us to answer such questions. As a result, we are beginning to move towards predicting the impacts of disturbances and interventions resulting ultimately in management of drinking water systems and microbial communities rather than mere observation. Many challenges still exist to achieve effective management, particularly within the premise plumbing environment, which exhibits diverse and inconsistent conditions that may lead to alterations in the microbiota, potentially presenting public health risks. Finally, we recommend the establishment of global collaborative projects on the drinking water microbiome that will enhance our current knowledge and lead to tools for operators and researchers alike to improve global access to high-quality drinking water.

Legionella is an opportunistic waterborne pathogen that is difficult to eradicate in colonized drinking water pipes. Legionella control is further challenged by aging water infrastructure and lack of evidence-based guidance for building treatment. This study assessed multiple premise water remediation approaches designed to reduce Legionella pneumophila (Lp) within a residential building located in an aging, urban drinking water system over a two-year period. Samples (n=745) were collected from hot and cold-water lines and quantified via most probable number culture. Building-level treatment approaches included three single heat shocks (HS), three single chemical shocks (CS), and continuous low-level chemical disinfection (CCD) in the potable water system. The building was highly colonized with Lp with 71% Lp positivity. Single HS had a statistically significant Lp reduction one day post treatment but no significant Lp reduction one, two, and four weeks post treatment. The first two CS resulted in statistically significant Lp reduction at two days and four weeks post treatment, but there was a significant Lp increase at four weeks following the third CS. CCD resulted in statistically significant Lp reduction ten weeks post treatment implementation. This demonstrates that in a building highly colonized with Lp, sustained remediation is best achieved using CCD.