Five-year VIM-producing Pseudomonas aeruginosa outbreak in four Belgian ICUs, an investigation report (2019-2023).

BACKGROUND: Verona integron-encoded metallo-ß-lactamase-producing Pseudomonas aeruginosa (VIM-PA) outbreaks are frequently linked to contaminated sink-drains in the intensive care unit (ICU). This study aims to investigate a VIM-PA outbreak occurring at 4 ICUs in a Belgian university center. METHODS: Between 01/01/2019 and 30/07/2023, data were retrospectively retrieved. Whole-genome sequencing of VIM-PA was carried out for available isolates and the core genome multilocus sequencing typing (cgMLST) was used to confirm clonality. New case incidence was estimated by analyzing the weekly data of at-risk and VIM-PA-colonized patients, fitting a regression model. RESULTS: Fifty-one patients were colonized, among them, 32 (63%) were infected by VIM-PA, which contributed to 7 deaths. The outbreak investigation showed that 19 (47%) of the examined sink-drains grew at least once a VIM-PA. Two major clusters were observed by cgMLST: ST111 (59 clones with 40 clinical isolates), and ST17 (8 clones with 6 clinical isolates). The estimated incidence rate of new cases was significantly higher in one unit. CONCLUSIONS: A 5-year prolonged outbreak at the UZ Brussel ICUs was caused by only 2 VIM-PA clones, both linked to sink-drains, with minimal mutations occurring throughout the years. Statistical modeling found different incidence rates between units. Tailored interventions were hence prioritized.

Water Quality Trade-offs for Risk Management Interventions in a Green Building.

Premise plumbing water quality degradation has led to negative health impacts from pathogen outbreaks (e.g., Legionella pneumophila and non-tuberculous mycobacteria), as well as chronic effects from exposure to heavy metals or disinfection by-products (DBP). Common water quality management interventions include flushing, heat shock (thermal disinfection), supplemental disinfection (shock or super chlorination), and water heater temperature setpoint change. In this study, a Legionella pneumophila- colonized Leadership in Energy and Environmental Design (LEED) certified building was monitored to study health-relevant water quality changes before and after three controlled management interventions: (1) flushing at several points throughout the building; (2) changing the water heater set point; and (3) a combination of interventions (1) and (2) by flushing during a period of elevated water heater set point (incompletely performed due to operational issues). Microbial (culturable L. pneumophila, the L. pneumophila mip gene, and cATP) and physico-chemical (pH, temperature, conductivity, disinfectant residual, disinfection by-products (DBPs; total trihalomethanes, TTHM), and heavy metals) water quality were monitored alongside building occupancy as approximated using Wi-Fi logins. Flushing alone resulted in a significant decrease in cATP and L. pneumophila concentrations (p = 0.018 and 0.019, respectively) and a significant increase in chlorine concentrations (p = 0.002) as well as iron and DBP levels (p = 0.002). Copper concentrations increased during the water heater temperature setpoint increase alone to 140°F during December 2022 (p = 0.01). During the flushing and elevated temperature in parts of the building in February 2023, there was a significant increase in chlorine concentrations (p = 0.002) and iron (p = 0.002) but no significant decrease in L. pneumophila concentrations in the drinking water samples (p = 0.27). This study demonstrated the potential impacts of short term or incompletely implemented interventions which in this case were not sufficient to holistically improve water quality. As implementing interventions is logistically- and time-intensive, more effective and holistic approaches are needed for informing preventative and corrective actions that are beneficial for multiple water quality and sustainability goals.

High resolution data visualization and machine learning prediction of free chlorine residual in a green building water system.

People spend most of their time indoors and are exposed to numerous contaminants in the built environment. Water management plans implemented in buildings are designed to manage the risks of preventable diseases caused by drinking water contaminants such as opportunistic pathogens (e.g., Legionella spp.), metals, and disinfection by-products (DBPs). However, specialized training required to implement water management plans and heterogeneity in building characteristics limit their widespread adoption. Implementation of machine learning and artificial intelligence (ML/AI) models in building water settings presents an opportunity for faster, more widespread use of data-driven water quality management approaches. We demonstrate the utility of Random Forest and Long Short-Term Memory (LSTM) ML models for predicting a key public health parameter, free chlorine residual, as a function of data collected from building water quality sensors (ORP, pH, conductivity, and temperature) as well as WiFi signals as a proxy for building occupancy and water usage in a "green" Leadership in Energy and Environmental Design (LEED) commercial and institutional building. The models successfully predicted free chlorine residual declines below 0.2 ppm, a common minimum reference level for public health protection in drinking water distribution systems. The predictions were valid up to 5 min in advance, and in some cases reasonably accurate up to 24 h in advance, presenting opportunities for proactive water quality management as part of a sense-analyze-decide framework. An online data dashboard for visualizing water quality in the building is presented, with the potential to link these approaches for real-time water quality management.

Source-to-tap investigation of the occurrence of nontuberculous mycobacteria in a full-scale chloraminated drinking water system.

Nontuberculous mycobacteria (NTM) in drinking water are a significant public health concern. However, an incomplete understanding of the factors that influence the occurrence of NTM in drinking water limits our ability to characterize risk and prevent infection. This study sought to evaluate the influence of season and water treatment, distribution, and stagnation on NTM in drinking water. Samples were collected source-to-tap in a full-scale, chloraminated drinking water system approximately monthly from December 2019 to November 2020. NTM were characterized using culture-dependent (plate culture with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry [MALDI-TOF MS] isolate analysis) and culture-independent methods (quantitative PCR and genome-resolved metagenomics). Sampling locations included source waters, three locations within the treatment plant, and five buildings receiving water from the distribution system. Building plumbing samples consisted of first draw, 5-min flush, and full flush cold-water samples. As the study took place during the COVID-19 pandemic, the influence of reduced water usage in three of the five buildings was also investigated. The highest NTM densities source-to-tap were found in the summer first draw building water samples (107 gene copies/L), which also had the lowest monochloramine concentrations. Flushing was found to be effective for reducing NTM and restoring disinfectant residuals, though flush times necessary to improve water quality varied by building. Clinically relevant NTM species, including Mycobacterium avium, were recovered via plate culture, with increased occurrence observed in buildings with higher water age. Four of five NTM metagenome-assembled genomes were identified to the species level and matched identified isolates.IMPORTANCENTM infections are increasing in prevalence, difficult to treat, and associated with high morbidity and mortality rates. Our lack of understanding of the factors that influence NTM occurrence in drinking water limits our ability to prevent infections, accurately characterize risk, and focus remediation efforts. In this study, we comprehensively evaluated NTM in a full-scale drinking water system, showing that various steps in treatment and distribution influence NTM presence. Stagnant building water contained the highest NTM densities source-to-tap and was associated with low disinfectant residuals. We illustrated the differences in NTM detection and characterization obtained from culture-based and culture-independent methods, highlighting the complementarity between these approaches. We demonstrated that focusing NTM mitigation efforts in building plumbing systems, which have the highest NTM densities source-to-tap, has potential for immediate positive effects. We also identified steps during treatment that increase NTM levels, which provides beneficial information for utilities seeking to reduce NTM in finished water.

Reduction of peritoneal dialysis associated infections using a novel exit-site care practice.

INTRODUCTION: Peritoneal dialysis (PD) related infections are a significant obstacle leading to PD discontinuation. Since catheter related infections (CRI), defined as exit site infections and/or tunnel infection, can progress to peritonitis, vigorous efforts are implemented in CRI prevention. Following an increased CRI rate in our institution, partially related to environmental organisms found in water distribution systems, we hypothesized that exit site care that includes prevention of water exposure-related pathogens may reduce CRI. METHODS: In this prospective single center study, we compared a contemporary cohort consisting of PD patients who implemented the modified exit-site care protocol, mainly including water avoidance during shower with stoma bag usage and local Mupirocin ointment against a historical control group before the protocol implementation. The historical cohort was allowed water exposure and used local gentamicin ointment. The primary outcome was the development of a CRI. Secondary outcomes were PD associated peritonitis and infection related outcomes. RESULTS: There were 55 patients in contemporary cohort and 58 in historical group. The CRI rate was significantly lower in study group (0.11/episodes per patient year [EPP]) compared to control group (0.71 EPP), p < 0.001. A multivariate Cox regression analysis demonstrated a protective effect of being in the contemporary cohort compared to historical group (HR for first CRI = 12.0 95%CI: 4.0-35.7, p < 0.001). Peritonitis rate was significantly lower in contemporary cohort (0.19/EPP) compared to the historical group (0.40/EPP), p = 0.011. Transfer to hemodialysis was significantly lower in contemporary cohort than historical group (7.3% vs 31.0% in contemporary and historical group respectively, HR = 0.2, 95%CI; 0.05-0.6, p = 0.001). CONCLUSION: An exit site care protocol that includes water avoidance and local Mupirocin use reduced substantially both CRI and peritonitis rate in patients treated with PD.

Two-year evaluation of Legionella in an aging residential building: Assessment of multiple potable water remediation approaches.

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.

Increased flushing frequency of a model plumbing system initially promoted the formation of viable but non culturable cells but ultimately reduced the concentration of culturable and total Legionella DNA.

Legionella is the causative agent of Legionnaires' disease, and its prevalence in potable water is a significant public health issue. Water stagnation within buildings increases the risk of Legionella. However, there are limited studies investigating how stagnation arising through intermittent usage affects Legionella proliferation and the studies that are available do not consider viable but non culturable (VBNC) Legionella. This study used a model plumbing system to examine how intermittent water stagnation affects both VBNC and culturable Legionella. The model plumbing system contained a water tank supplying two biofilm reactors. The model was initially left stagnant for ≈5 months (147 days), after which one reactor was flushed daily, and the other weekly. Biofilm coupons, and water samples were collected for analysis at days 0, 14 and 28. These samples were analysed for culturable and VBNC Legionella, free-living amoebae, and heterotrophic bacteria. After 28 days, once-a-day flushing significantly (p < 0.001) reduced the amount of biofilm-associated culturable Legionella (1.5 log10 reduction) compared with weekly flushing. However, higher counts of biofilm-associated VBNC Legionella (1 log10 higher) were recovered from the reactor with once-a-day flushing compared with weekly flushing. Likewise, once-a-day flushing increased the population of biofilm-associated Vermamoeba vermiformis (approximately 3 log10 higher) compared with weekly flushing, which indicated a positive relationship between VBNC Legionella and V. vermiformis. This is the first study to investigate the influence of stagnation on VBNC Legionella under environmental conditions. Overall, this study showed that a reduction in water stagnation decreased culturable Legionella but not VBNC Legionella.

Can free chlorine residuals entering building plumbing systems really be maintained to prevent microbial growth?

Secondary disinfection aims to prevent microbial regrowth during distribution by maintaining disinfectant residuals in water systems. However, multi-factorial interactions contribute to free chlorine decay in distribution systems, and even more so in building plumbing. Assembling 1737 samples from nine large institutional buildings, a meta-analysis was conducted to determine whether building managers can actively rely on incoming free chlorine residuals to prevent in-building microbial amplification. Findings showed that free chlorine concentrations in first draws met the 0.2 mg/L common guide level in respectively 26 %, 6 % and 2 % of cold, tepid and hot water samples, whereas flushing for 2-60 min only significantly increased this ratio in cold water (83 %), without reaching background levels found in service lines. Free chlorine was significantly but weakly (R≤ 0.2) correlated to adenosine triphosphate, heterotrophic plate count and total and intact cell counts, thus evidencing that residuals contributed to decreased culturable and viable biomass. Detection of culturable Legionella pneumophila spanning over a 4-log distribution solely occurred when free chlorine levels were below 0.2 mg/L, but no such trend could be distinguished clearly for culturable Pseudomonas aeruginosa. Water temperatures below 20 °C and >60 °C also completely prevented L. pneumophila detection. Overall, the majority of elevated microbial counts were measured in distal sites and in tepid and hot water, where free chlorine is less likely to be present due to stagnation and increased temperature. Therefore, building managers cannot solely rely on this chemical barrier to mitigate bacterial growth in bulk water.

Quantitative Microbial Risk Assessment Framework Incorporating Water Ages with Legionella pneumophila Growth Rates.

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%.

Linkage of in-refrigerator water dispensing systems to elevated exposure to microbial contamination at the microbiome scale.

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.

Building size and disinfectant type influence the detection and concentration of Mycobacterium spp. in hot water plumbing.

The number of nontuberculous mycobacteria (NTM) lung disease cases is increasing in the United States (US). This respiratory disease is primarily caused by three NTM species: Mycobacterium avium, M. intracellulare, and M. abscessus. Since disease transmission could occur through water aerosolization, this study investigated these three species' occurrence (sporadic and persistent) in hot water samples collected from residences (n = 70) and office buildings (n = 30) across the US. A longitudinal survey design was used. Three quantitative Polymerase Chain Reaction (qPCR) assays were used to measure the mycobacterial species in the water samples. Additionally, the water's disinfectant residual was measured. A structure's age and square footage were evaluated to predict mycobacterial contamination. Also, the seasonal occurrence of each species was assessed by structure type. Residences had a 43 % (30/70), and office buildings had a 77 % (23/30) detection frequency of one or more Mycobacterium spp. in their hot water. The age of the structure influenced M. intracellulare detection frequency but not M. avium and M. abscessus. The structure's square footage affected M. avium and M. intracellulare detection frequency but not M. abscessus. In chlorinated water, M. intracellulare was detected 1.4× more often in office buildings' hot water than in chloraminated water. In chloraminated water, the Mycobacterium spp. were detected 2-2.5× more often in residences, while M. avium and M. abscessus were detected 1.5-2.3× more often in office buildings, compared to chlorinated water. Each Mycobacterium spp. had a different trend associated with the type of structure and disinfectant. Further research is needed to better understand NTM occurrence in the built environment to improve public health.

Bacterial communities of planktonic bacteria and mature biofilm in service lines and premise plumbing of a Megacity: Composition, Diversity, and influencing factors.

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).

A methodology to simulate peak water demand for water supply systems using semi-direct methods: A case study for a residential building in Mexico.

Historically, approaches for determining peak water demand in buildings have been based on probabilistic methods. Extensive research has shown that these methods lack accuracy because of the human factor in the probability of use. Inaccuracy in the calculation of peak water demand is the main cause of oversized water supply systems in buildings. This has led to unfavorable effects such as: 1) increasing the building carbon footprint due to the use of more construction materials, and 2) engendering health hazards due to the stagnation of water causing microbial growing. This paper presents a step-by-step methodology that serves to calculate the peak water demand by simulating the use of plumbing fixtures based on data obtained from standardized flowrate. With the implementation of the methodology, the peak water demand estimated was 2.6 times lower in comparison to traditional methods. The main conclusion drawn from the research is the potential of the methodology to easily simulated peak water demand in residential buildings in the short term. Thus, it reveals a hotspot for peak water demand calculation and can serve as routes for future research.

Multidrug-Resistant Bacteria Contaminating Plumbing Components and Sanitary Installations of Hospital Restrooms.

Antimicrobial resistance (AMR) poses several issues concerning the management of hospital-acquired infections, leading to increasing morbidity and mortality rates and higher costs of care. Multidrug-resistant (MDR) bacteria can spread in the healthcare setting by different ways. The most important are direct contact transmission occurring when an individual comes into physical contact with an infected or colonized patient (which can involve healthcare workers, patients, or visitors) and indirect contact transmission occurring when a person touches contaminated objects or surfaces in the hospital environment. Furthermore, in recent years, toilets in hospital settings have been increasingly recognised as a hidden source of MDR bacteria. Different sites in restrooms, from toilets and hoppers to drains and siphons, can become contaminated with MDR bacteria that can persist there for long time periods. Therefore, shared toilets may play an important role in the transmission of nosocomial infections since they could represent a reservoir for MDR bacteria. Such pathogens can be further disseminated by bioaerosol and/or droplets potentially produced during toilet use or flushing and be transmitted by inhalation and contact with contaminated fomites. In this review, we summarize available evidence regarding the molecular features of MDR bacteria contaminating toilets of healthcare environments, with a particular focus on plumbing components and sanitary installation. The presence of bacteria with specific molecular traits in different toilet sites should be considered when adopting effective managing and containing interventions against nosocomial infections potentially due to environmental contamination. Finally, here we provide an overview of traditional and new approaches to reduce the spreading of such infections.

Toward the autonomous flushing of building plumbing: Characterizing oxidation-reduction potential and temperature sensor dynamics.

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.

Inactivation efficacy and mechanisms of wavelength-specific UV sources for various strains of Legionella pneumophila serogroup 1.

Infections of Legionnaires' disease in the United States caused by Legionella have increased ninefold between the years 2000-2018. Legionella harbored in biofilms or inside amoeba within premise plumbing can be more resistant to disinfectants, thus causing treatment challenges. Ultraviolet-light emitting diodes (UV-LEDs) are an emerging water disinfection technology with several advantages over conventional UV lamps. In this study, we evaluated the effects of UV-LEDs (255, 265, and 285 nm), a low-pressure (LP) mercury UV lamp (254 nm), and a bandpass filtered medium-pressure (MP) mercury UV lamp (220 nm) on properties and inactivation of three strains of L. pneumophila serogroup 1. The UV-LEDs emitting at 255 and 265 nm showed greater inactivation performance against all the strains compared to the UV-LED at 285 nm and the LP UV lamp at 254 nm. Our results showed that strains of the same serogroup exhibited different UV sensitivities. Analyses of DNA and protein damage revealed that UV exposure using 254, 255, and 265 nm predominantly causes DNA damage, while protein damage is predominant at 220 nm. Both DNA and protein damage were observed at 285 nm, but the extent of DNA damage was relatively less significant compared to the other wavelengths. Electric energy consumption analysis showed that water treatment using UV-LEDs is currently unsatisfactory compared to conventional LP UV lamps due to the mediocre wall plug efficiency (WPE) of UV-LEDs. However, recent studies indicate that the WPE of UV-LEDs is continuously improving. Overall, our study highlights that UV-LEDs are a promising technology for inactivating waterborne pathogens and have the potential to replace existing UV mercury lamps for water disinfection applications.

Influence of pipe materials on in-building disinfection of P. aeruginosa and A. baumannii in simulated hot water plumbing.

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.

Extended water stagnation in buildings during the COVID-19 pandemic increases the risks posed by opportunistic pathogens.

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.

Assessing residential activity in a home plumbing system simulator: monitoring the occurrence and relationship of major opportunistic pathogens and phagocytic amoebas.

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.