Premise plumbing, the pipes and fixtures at the building level, present a unique challenge for maintaining drinking water quality. Of particular concern are opportunistic pathogens, including Legionella pneumophila which can regrow in premise plumbing and cause disease in immunocompromised populations. The goal of this work was to explore engineering methods for control of L. pneumophila and total regrowth. The first line of study involved a series of experiments with simulated glass water heaters (SGWHs) to investigate interactions between specific water chemistry factors and L. pneumophila regrowth, and the second used laboratory grade purified water to investigate the limits of a nutrient control approach for biological stability. Several water chemistry factors including assimilable organic carbon (AOC) content, granular activated carbon (GAC) biofiltration, plumbing materials, copper concentrations and temperature were investigated using SGWHs. AOC is the carbon available for bacteria growth in drinking water. Results indicated that AOC reduction may be a promising method for controlling L. pneumophila and total bacteria regrowth, but there may be a point at which AOC reduction is no longer effective. Prior GAC biofiltration removed organic carbon and was effective in controlling total bacterial regrowth in SGWHs, but actually encouraged L. pneumophila regrowth. A wide variety of materials typically encountered in premise plumbing was investigated and only had limited effect on proliferation of L. pneumophila and total bacteria. The effects were dynamic, even with long-term studies. Copper pipes held promise for control of L. pneumophila, as did copper concentration across a range of pHs. Aqueous copper concentration released from pipes was dependent on temperature, however, and thus this control method may not be applicable in all hot water lines. The peak temperatures for L. pneumophila proliferation fell between 41 and 45 °C, temperatures which could be encountered in a hot water distribution system when the water heater is set to 48 °C, as is often recommended with scalding and energy concerns. A constant temperature of 53 °C seemed to provide control of L. pneumophila, but recolonization is possible even at these high temperatures. Work with laboratory grade water indicated that extreme control of nutrients was not enough to completely control regrowth in premise plumbing. With stagnation in the cleanest conditions, a 2-log increase of a diverse group of bacteria was observed within 10 days. As drinking water can never achieve such nutrient removal, this study presents the limits of nutrient removal as a strategy for regrowth control. This work explored both the potential and the limitations of several mechanisms for controlling regrowth in premise plumbing. Understanding how these water chemistry factors affect L. pneumophila and total bacterial regrowth is critical to identifying the most effective engineering controls. / Master of Science

Nitrification is increasingly of concern in US potable water systems, due to changes from chlorine to chloramine as a secondary disinfectant in order to comply with new regulations for disinfectant by-products. The ammonia that is released from the chloramine decay supports nitrification. A comprehensive literature review systematically examined the complex inter-relationships between nitrification, materials corrosion and metals release. That analysis suggested that nitrification could accelerate decay of chloramine, enhance corrosion of water distribution system materials, and increase leaching of lead and copper to potable water under at least some circumstances. Moreover, that certain plumbing materials would inhibit nitrification, but that in other situations the plumbing materials would enhance nitrification. Experiments verified that nitrification could affect the relative efficacy of chlorine versus chloramine in controlling heterotrophic bacteria in premise plumbing. Without nitrification, chloramine was always more persistent and effective than chlorine in controlling biofilms. But with nitrification and in pipe materials that are relatively non-reactive with chlorine, chloramine was much less persistent and less effective than chlorine. In materials that are reactive with chlorine such as iron pipes, the relative efficacy of chloramine versus chlorine depends on the relative rate of corrosion and rate of nitrification. High rates of corrosion and low rates of nitrification favor the use of chloramine versus free chlorine in controlling bacteria. Plumbing materials had profound impacts on the incidence of nitrification in homes. Effects were due to toxicity (i.e., release of Cu⁺²), recycling of nitrate back to ammonia substrate by reaction (zero-valent iron, lead or zinc materials), or release of nutrients that are essential to nitrification by leaching from concrete or other materials. As a general rule it was determined that concrete and iron materials encouraged growth of nitrifiers in certain oligotrophic waters, materials such as lead, PVC/plastic pipe, glass and surfaces of other materials were readily colonized by nitrifiers, and materials such as copper and brass were very toxic and relatively resistant to nitrifier colonization. Dependent on circumstance, nitrification had no effect, increased or decreased aspects of materials corrosion. Nitrification markedly increased lead contamination of low alkalinity potable water by reducing the pH. In some cases nitrification dramatically decreased leaching of zinc to potable water from galvanized iron, because of lowered dissolved oxygen and reduced pH. Nitrification did not affect copper solubility in low alkalinity water, but is expected to increase copper solubility in higher alkalinity waters. Finally, nitrification in homes plumbed with PVC or plastics can drop the pH and increase leaching of lead from downstream brass materials in faucets. This can explain why some modern homes plumbed with PVC can have more lead in water when compared to homes plumbed with copper pipe. Phosphate had profound impacts on the incidence of nitrification and resulting effects on water quality. While phosphate levels below about 5 ppb could strongly inhibit nitrification due to a nutrient limitation, nitrifiers can obtain sufficient phosphate from plastic, concrete, copper and iron pipe materials to meet nutritional needs. High levels of phosphate inhibitor can reduce the concentration of Cu⁺² ions and make nitrification more likely, but phosphate can also sometimes lower the corrosion rate and increase the stability of disinfectant and its efficacy in controlling nitrifiers. Phosphate plays a key role in determining where, when and if problems with nitrification will occur in a given water distribution system. This work provides some new fundamental and practical insights to nitrification issues through a comprehensive literature review, lab experiments, solubility modeling and field studies. The results and practical tools developed can be used by utilities and consumers to predict nitrification events and resulting water quality problems, and to make rational decisions about practices such as inhibitor dosing, plumbing material selection and use of whole house filters. / Ph. D.

Legionella pneumophila (Lp) and nontuberculous mycobacteria (NTM) are opportunistic pathogens that can be transmitted via drinking water, when tiny droplets containing the bacteria are aerosolized and inhaled during activities such as showering. The resulting respiratory illnesses, Legionnaires' Disease and NTM lung disease, are among the leading sources of drinking water associated disease in the United States and other parts of the world. Lp and NTM are both difficult to control, because they establish as part of natural biofilms that form within the interiors of pipes and fixtures that deliver drinking water to the point of use. These pathogens are especially problematic within premise (i.e., building) plumbing, where intermittent use throughout the day leads to long periods of stagnation, increased water age, warmer temperatures, and depleted disinfectant residuals that exacerbate bacterial growth. The recent advent of high throughput DNA sequencing has led to the discovery that drinking water microbiomes are diverse, complex, and largely comprised of non-pathogenic microbes. This has further led researchers to hypothesize that the microbial ecology of this diverse microbiome could be harnessed as a natural means to control Lp and NTM, i.e., a "probiotic" approach, but such an approach has not yet been demonstrated. The objective of this study was to assess this hypothesis by utilizing biologically active granular activated carbon (GAC) filters, which are already a widely used drinking water treatment both at the municipal and household scale, as a means to naturally shape the microbial ecology of downstream premise plumbing and inhibit Lp and NTM proliferation. GAC has an extremely high surface area that aids removal of organic carbon via adsorption but also provides an ideal habitat for establishment of biofilms, which removes organic carbon from the water via biodegradation. Convectively-mixed pipe reactors (CMPRs) were used for replicable simulation of premise plumbing distal taps. The CMPRs consisted of four-foot-long closed polyvinyl chloride (PVC) pipe segments with the sealed bottom portion resting in a ~48 °C water bath and with the top portion plugged and exposed to the cooler, ambient atmosphere (25 °C in this study), inducing convective mixing and resulting in an internal water temperature of 37 °C. PVC was chosen because it is common in premise plumbing and generally leaches the least organic carbon among the different types of plastic pipe. Four different influent water conditions were implemented in the experimental design: 1) Untreated, dechlorinated municipal tap water with high organic carbon and low biomass; 2) GAC-treated tap water with low organic carbon and elevated, viable biomass; 3) GAC-treated + 0.22-m pore size membrane-filtered tap water to remove both nutrients and biomass; 4) GAC-treated tap water pasteurized at 70 °C with low nutrients and elevated, killed biomass. The 0.22-m pore size membrane filter simulated the use of a building scale particle filter, while pasteurization simulated water passing through a hot water heater at an elevated temperature recommended for pathogen thermal disinfection. To understand the influence of these experimental conditions on older pipes containing mature biofilms versus new pipes that leach more organics and are being freshly colonized, a set of older pipes colonized with mature ~4-year-old biofilms were compared to newly purchased pipes. Each set of pipes was tested in triplicate for the four different experimental conditions with the full volume replaced three times a week for eight months, simulating infrequently used taps containing warm, continuously mixing water thought to create conditions at a very high risk for opportunistic pathogen growth. In the aged CMPR bulk water effluents, droplet-digital-polymerase-chain-reaction measurements showed a one-log reduction of Lp and NTM when receiving GAC-treated or GAC-treated + particle-filtered influent water versus receiving dechlorinated municipal tap water or GAC-treated + pasteurized water. These findings suggest that decreased biodegradable dissolved organic carbon achieved by GAC filtration acted to suppress Lp and NTM growth, while the additional step of biomass removal by particle filtration provided a more modest benefit. In the CMPRs consisting of new pipes, concentrations of Lp and NTMs in the effluent bulk water were similar among the experimental conditions, except that the CMPRs receiving the GAC-treated + particle-filtered influent water experienced a two-log reduction in NTMs. These results demonstrate that the colonization and proliferation of NTM within premise plumbing can be significantly controlled by limiting nutrients and biomass in the influent water. This work demonstrates the potential of harnessing GAC-treatment as a means to Control Lp and NTM in premise plumbing via nutrient removal. In scenarios where chemical disinfectants have been depleted, off-the-shelf GAC-treatment used as point-of-entry treatment to large buildings with recirculating plumbing could provide benefits that have previously been unrecognized. Alternatively, pasteurization in very hot water heaters could provide a short-term disinfection benefit, but eventually the nutrients embodied in the dead biomass undermine the positive influence of the nutrient removal provided by the GAC-treatment. Improved mechanistic understanding of probiotic strategies to opportunistic pathogen control would be needed to overcome inherent limitations to the approaches examined herein, if more effective control is desired in the absence of thermal or chemical disinfection. / Master of Science / Legionella pneumophila (Lp) and nontuberculous mycobacteria (NTM) are bacterial pathogens that are the leading source of drinking water-associated disease in the US. Unfortunately, they are not effectively controlled by protections put in place by the US Safe Drinking Water Act (SDWA). Firstly, they cause respiratory infections, which are spread when tiny droplets of water are inhaled during activities such as showering, whereas the SDWA is specifically designed to protect against ingested pathogens. Secondly, unlike fecal-derived organisms (e.g. E. coli) that are the focus of the SDWA, Lp and NTM grow naturally in drinking water distribution systems, especially in premise (i.e., building) plumbing, where water is warmer and more stagnant. Therefore, even if water leaving the treatment plant is devoid of Lp or NTM, this does not guarantee that the consumer's tap water will be Lp- or NTM-free. Also, even though chlorine or other chemical disinfectant is required by the SDWA to be added to the water leaving the treatment plant to control downstream microbial growth, the disinfectant can be depleted or absent within the premise plumbing itself. Additionally, both Lp and NTM tend to more naturally resist chemical disinfectants than fecal-derived organisms. This research is aimed at overcoming these challenges, opening the door to new approaches to controlling Lp and NTM in premise plumbing. Historically, any microbial growth occurring in drinking water has been viewed as problematic, as it usually indicates the chemical disinfectant is inadequately protecting consumers. However, this work explores whether having an abundant community of beneficial bacteria could improve microbial water quality by competing against pathogens for limited space for attachment and nutrients. Such an approach would be analogous to the use of probiotics in humans, to establish a beneficial gut flora that is less susceptible to pathogen invasion. Granular activated carbon (GAC) filters are often used at drinking water treatment plants and by consumers as a point-of-use (e.g., installed on the kitchen tap or in a refrigerator) or whole-house treatment to remove any contaminants of concern and improve the taste and odor of tap water. The granules within GAC filters have a high surface area that helps remove contaminants, but also provides an environment where microbes can live and thrive. As water enters the filter, beneficial microbes can break down any remaining nutrients in the water (e.g., organic carbon and nitrogen). Additionally, the water leaving the filter carries high levels of microbes that grow on the GAC filter that are shed as water passes through. The resulting water with reduced nutrients and higher concentrations of potentially beneficial microbes could create a competitive environment that alters growth of harmful bacteria, like Lp and NTM, in downstream portions of plumbing. The incoming cold water is also warmed by the building envelope, which increases bacterial growth rates. Thus, the underlying hypothesis of this research is that GAC treatment could provide a combination of reduced nutrients and competitive microbes as water enters downstream premise plumbing and reduce the growth of Lp and NTM. However, GAC-treated water within a building can be further altered by other treatments, like a very hot water heater, which would heat and kill the microbes flowing through it, or a particle filter, which could remove the microbes in the water. This work also seeks to understand how these additional treatments might improve or interfere the nutrient reduction and addition of competitive microbes provided by GAC treatment. This research explores how all these different scenarios affect the growth of Lp and NTM using a lab-scale simulated premise plumbing system constructed out of polyvinyl chloride (PVC) pipe that is a common plumbing material used in homes. Water that was added to the pipes was prepared in four different ways to test the probiotic control hypothesis across distinct experimental conditions that replicate the different influent water scenarios. The four conditions were implemented over the course of eight months with regular chemical and biological analyses conducted to understand the effects of the different influent waters on Lp and NTM. It was discovered that premise plumbing with mature biofilms receiving GAC-treated water or GAC-treated + particle-filtered water contained ~90% less Lp and NTM than premise plumbing receiving non-filtered municipal tap water. However, if the GAC-treated water passes through a water heater, the capacity to limit Lp or NTM growth was lost. While GAC filters are currently thought of as an instantaneous treatment that removes contaminants from water, this work demonstrates how GAC treatment might provide prolonged benefits to water, after it has passed through the filter on its journey to a shower head or faucet. Increased understanding of the exact mechanisms of limited pathogen growth gained by this research can lead to new and effective approaches to protect people from contracting diseases caused by Lp and NTM.

Opportunistic premise (i.e., building) plumbing pathogens (OPPPs) and antibiotic resistant bacteria are emerging microbial concerns in drinking water. OPPPs, such as Legionella pneumophila, are the leading cause of drinking water disease in many developed countries. Contributing factors include the relative success in controlling fecal pathogens, the presence of complex building plumbing systems that create habitats for OPPPs, and the relative resistance of OPPPs to disinfectants, and aging populations that are susceptible to infection. Concurrently, drinking water is increasingly being scrutinized as a potential environment that is conducive to horizontal gene transfer of antibiotic resistance genes (ARGs), selection pressure for enhanced survival of resistant bacteria, and a route of transmission of antibiotic resistant pathogens. While maintaining a disinfectant residual is an established approach to controlling OPPPs in premise plumbing, some studies have indicated that co-resistance and cross-resistance to disinfectants can increase the relative abundances of resistant bacteria and ARGs. Thus, there may be trade-offs to controlling both OPPPs and antibiotic resistance in premise plumbing that call for controlled study aimed at optimizing residual disinfection application for this purpose. A critical review of the scientific literature in Chapter 2 revealed that premise plumbing is a biologically and chemically complex environment, in which the choice of pipe material has cascading effects on water chemistry and the corresponding premise plumbing microbiome. This, in turn, has broad implications for the control of OPPPs, which need to be elucidated through controlled experiments in which worst case premise plumbing conditions are held constant (e.g., warm temperature), while other variables are manipulated. Chapter 3 introduces the convectively-mixed pipe reactors (CMPRs) as a novel low-cost, small footprint approach to replicably conduct such experiments. The CMPRs were demonstrated to effectively simulate key chemical and biological phenomena that occur in distal reaches of premise plumbing. In Chapter 4, the CMPRs were leveraged to study the interactive effects of four disinfectants (chlorine, monochloramine, chlorine dioxide, and copper-silver ionization) and three pipe materials (PVC copper, and iron). The CMPRs were inoculated with two antibiotic-resistant OPPPs: Pseudomonas aeruginosa and Acinetobacter baumannii. It was found that pipe-material (PVC or PVC combined with iron or copper) profoundly impacted the water chemistry in a manner that dictated disinfection efficacy. In Chapter 5, we applied shotgun metagenomic shotgun sequencing to evaluate effects of the combination of pipe material and disinfectant type on the wider microbial community, especially their ability to select for or reduce ARGs. In Chapter 6, we used CMPRs and metagenomic sequencing in a study comparing Dutch drinking water practices to our prior testing in an American system. Dutch drinking water is of interest because of lack of historical use of disinfectants was hypothesized to result in a microbial community that is relatively depleted of ARGs or mobile genetic elements, which can enhance spread of ARGs as disinfectants are applied. Generally, it was found that OPPPs required higher doses of disinfectants for inactivation than the general microbial community, sometimes concentrations approaching the regulatory limits in the US (e.g., 4 mg/L of total chlorine). Even successful reductions were modest, typically ~1-log, and failed to eliminate either P. aeruginosa or A. baumannii. Moreover P. aeruginosa, A. baumannii, and non-tuberculous mycobacteria varied substantially in their preference for pipe material and susceptibility to disinfectants. We found that disinfectants tended to increase the relative abundance of OPPPs, ARGs, and mobile genetic elements. Disinfectants were sometimes associated with net increases in levels of these pathogens and genes when applied at low levels (e.g., 0.1 mg/L of monochloramine), which effectively acted to reduce competition from less resistant and non-pathogenic taxa. When a low dose of monochloramine was applied to PVC CMPRs in the US, we estimated from metagenomic sequencing data that this water contained roughly 100,000 cells per milliliter of taxa known to contain pathogenic members. The Dutch drinking water exhibited more diverse microbial communities and lower relative abundances of taxa containing pathogens. ARGs were two times proportionally more abundant in CMPRs operated in the US without disinfectant than in the corresponding CMPRs operated in the Netherlands. The findings of this dissertation can help to optimize the application of in-building disinfectant addition for addressing concerns related both to OPPPs and antibiotic resistance. The studies herein highlight the necessity of developing comprehensive OPPP and antibiotic resistance control strategies that emphasize not just disinfectant dose, but other key control parameters such as contact time, hydraulics, and temperature. The functional diversity of OPPPs, antibiotic resistant bacteria, and the background premise plumbing microbiome further necessitates broad, holistic programs for monitoring and control. / Doctor of Philosophy / Efforts to provide safe drinking water face two emerging threats: the rise of pathogens that thrive in the plumbing environment that delivers water to the tap and the rise of antibiotic resistance. In the US and many other parts of the world, opportunistic pathogens are the predominant agents responsible for disease spread by tap water. Opportunistic pathogens tend to infect aged or immunocompromised individuals (hence, 'opportunistic') and grow well in in-building plumbing. Globally, antibiotic resistance is on the rise and becoming a fundamental threat to modern medicine. Pathogenic bacteria become resistant to antibiotics used to treat infections when they acquire antibiotic resistance genes (ARGs), which can happen either by mutation or from other resistant bacteria sharing ARGs. Overuse or misuse of antibiotics can impose selection pressure that stimulates horizontal gene transfer and enhance survival of bacteria that are resistant. Prior studies have suggested that under some circumstances, disinfectants used to control pathogens in drinking water can also select for antibiotic resistant bacteria. Thus, the overarching goal of this research was to optimize the type and dose of disinfectant used, depending on building-level factors such as pipe material, for effectively controlling proliferation of both opportunistic pathogens and antibiotic resistance. This dissertation largely focuses on in-building plumbing systems, which are home to potentially tens of thousands of bacterial cells per milliliter of water or per square centimeter of internal pipe surfaces. These bacteria interact not only with each other and other microbes, but also with features of the plumbing environment, such as the water chemistry or the pipe materials. Building plumbing systems are highly intricate ecosystems that can undermine the effectiveness of disinfectants provided by utilities. One major contribution of this research is the development of the convectively-mixed pipe reactors (CMPRs) as a simple and easy-to-use test system that recreates combinations of features of interest encountered in in-building plumbing. We applied the CMPRs to study two common residual disinfectants (chlorine and monochloramine) supplied by water utilities, and two other disinfectants (chlorine dioxide and copper-silver ionization) which are commonly dosed by building operators, especially in hospitals and other buildings housing individuals susceptible to infection. These four disinfectants were applied to CMPRs consisting of PVC, copper, and iron pipe. Chemical, culture, and DNA methods were used to understand how these disinfectants affected the microbes and their ecology. We then took the opportunity to set up CMPRs in the Netherlands, where there has been no historical exposure to chlorine because their water quality regulations emphasize limiting nutrients in the water and elevating the hot water line temperatures as means to control microbial growth. The CMPRs effectively produced worst-case plumbing scenarios, where opportunistic pathogens were especially difficult to control through residual disinfection. Dosed disinfectants tended to be no longer measurable in the water after five hours. The CMPRs also showed that the disinfectant most effective for one pathogen could be the least effective for another. If doses were applied near regulatory limits, the concentrations of pathogens and antibiotic resistance genes decreased. However, opportunistic pathogens tended to survive better than background populations of bacteria. Bacteria carrying ARGs also survived some disinfectant conditions better as well. Thus, if doses were applied at levels that could inactivate some microbes, but not the opportunistic pathogens, pathogen abundances sometimes increased. These results were largely confirmed in the experiment with Dutch drinking water. Here, chlorine appeared to be more problematic than monochloramine in terms of enriching pathogens and antibiotic resistance. We also noted that Dutch waters garnered more diverse microbial communities, with fewer DNA markers for pathogens and antibiotic resistance. In general, this research takes a key step towards optimizing application of residual disinfectants for control of both opportunistic pathogens and antibiotic resistance. Because disinfectants can have negative impacts on drinking water microbial communities when supplied insufficiently, it is important that the other features of in-building plumbing, such as the selection of pipe material or the hydraulics, facilitate disinfectants reaching all portions of plumbing and at the necessary concentrations. It is recommended that the selection process for disinfectant type and dose considers the plumbing materials and other conditions such that disinfection can be aimed towards controlling multiple opportunistic pathogens, which can vary in their susceptibility, and antibiotic resistance.

Consumer exposure to opportunistic pathogens in potable water systems poses a significant challenge to public health as manifested by numerous cases of pneumonia, non-tuberculosis lung disease, and keratitis eye infections. Water utilities have extensive understanding in control of heterotrophic and coliform bacteria re-growth in water distribution systems via disinfection, control of assimilable organic carbon (AOC), and biologically degradable organic carbon (BDOC). However, much little is known about the effect of AOC on the proliferation of heterotrophic bacteria and pathogens within premise plumbing. This thesis is the first systematic examination of opportunistic pathogen persistence and amplification in simulated glass water heaters (SGWH) as a function of influent organic matter concentration. The role of plumbing conditions that may internally generate AOC is critically examined as part of this evaluation. Strong correlations were often observed between influent organic matter and heterotrophic bacteria in effluent of SGWH as indicated by 16S rRNA gene abundance (average R2 value of 0.889 and 0.971 for heterotrophic organisms and 16S rRNA respectively). The correlation was strongest if water turnover was more frequent (every 48-72 hours) and decreased markedly when water changes were less frequent (stagnation up to 7 days). No simple correlations were identified between the concentration of pathogenic bacteria (L. pneumophila, M. avium, A. polyphaga, and H. vermiformis) and AOC, although correlations were observed between M. avium and TOC over a limited range (and only for a subset of experiments). Indeed, there was little evidence that Legionella and Acanthamoeba proliferated under any of the conditions tested in this work. Parallel experiments were conducted to examine the extent to which factors present in premise plumbing (e.g. sacrificial magnesium anode rods, cross-linked polyethylene, nitrifying bacteria, and iron) could influence water chemistry and influence growth of bacteria or specified pathogens. Although these factors could strongly influence pH, dissolved oxygen concentrations, and levels of organic matter (e.g. iron, magnesium, nitrifying), there was no major impact on effluent concentrations of either heterotrophic bacteria or premise plumbing pathogens under the conditions investigated. While additional research is needed to confirm these findings, at present, there is no evidence of correlations between organic matter and pathogen concentrations from SGWH under conditions tested. Substantial effort was also invested in attempting to identify SGWH and oligotrophic nutrient conditions that would consistently support L. pneumophila and A. polyphaga amplification. A review of the literature indicates no prior examples of large scale amplification of these microorganisms at nutrient levels commonly found in synthesized potable water. It is likely that a complex combination of abiotic and biotic factors (i.e. micronutrients, necrotrophic growth, ambient water temperature, disinfectant type and dose, plumbing materials, water usage patterns), which are not yet fully understood, control the amplification and viability of these pathogenic organisms in premise plumbing systems. / Master of Science

No description available.

No description available.

Residential water heating is linked to the primary source of waterborne disease outbreaks in the United States, and accounts for greater energy demand than the combined water/wastewater utility sector. To date, there has been little research that can guide decision-making with regards to water heater selection and operation to minimize energy costs and the likelihood of waterborne disease. We have outlined three types of systems that currently dominate the marketplace: 1) a standard hot water tank with no hot water recirculation (STAND), 2) a hot water tank with hot water recirculation (RECIRC), and 3) an on-demand tankless hot water system with no hot water recirculation (DEMAND). Not only did the standard system outperform the hot water recirculation system with respect to temperature profile during flushing, but STAND also operated with 32 – 36% more energy efficiency. Although RECIRC did in fact save some water at the tap, when factoring in the energy efficiency reductions and associated water demand, RECIRC actually consumed up to 7 gpd more and cost consumers more money. DEMAND operated with virtually 100% energy efficiency, but cannot be used in many circumstances dependent on scaling and incoming water temperature, and may require expensive upgrades to home electrical systems. RECIRC had greater volumes at risk for pathogen growth when set at the lower end of accepted temperature ranges, and lower volumes at risk when set at the higher end when compared to STAND. RECIRC also tended to have much lower levels of disinfectant residual (40 -850%), 4-6 times as much hydrogen, and 3-20 times more sediment compared to standard tanks without recirculation. DEMAND had very small volumes of water at risk and relatively high levels of disinfection. A comparison study of optimized RECIRC conditions was compared to the baseline modes of operation. Optimization increased energy efficiency 5.5 – 60%, could save consumers 5 – 140% and increased the disinfectant residual up to 560% higher disinfectant residual as compared to the baseline RECIRC system. STAND systems were still between 3 – 55% more energy efficient and could save consumers between $19 - $158 annual on water and electrical costs. Thus, in the context of “green” design, RECIRC systems provide a convenience to consumers in the form of nearly instant hot water, at a cost of higher capital, operating and overall energy costs. / Ph. D.

Opportunistic pathogens (OPs) can naturally colonize premise (i.e., building) plumbing and are the leading cause of disease associated with potable water in the U.S. and many other countries. While secondary disinfectant is added by utilities prior to water distribution through pipes, the residual in water at the property line is sometimes insufficient to suppress OP growth. Conditions encountered in premise plumbing can further diminish disinfectant in water after it crosses the property line. This dissertation examines how multiple factors at play in drinking water distribution systems and premise plumbing influence OP growth in order to inform development of rational guidance to reduce incidence of waterborne illness. Operating an at-scale cross-linked polyethylene (PEX) plumbing system with one water flush per day, influent chloramine always decayed within four hours in stagnant pipes containing mature biofilms, which is 2-3 orders of magnitude faster than in the same water not contacting pipes. Chloramine often followed second order decay kinetics, though decay rate coefficients were highly variable with some taps eventually transitioning from second to first order decay over time or with increasing influent chloramine concentration. The rate of chloramine decay was unexpectedly reduced in the water heater tank compared to room temperature pipes, possibly due to lower surface-area-to-volume ratio and higher temperature within the tank. A complementary glass jar experiment confirmed that, contrary to expectations, chloramine could decay slower at the higher temperature of 37-39°C maintained in the water heater, compared to the cooler 19-30°C typical of the pipes. These findings demonstrate the need for disinfectant decay models specific to conditions encountered in premise plumbing. Nitrification, a key microbial process that can catalyze chloramine decay, was typically complete within 24 hours after water entered the stagnant pipes. Counterintuitively, the water heater had a relatively lower rate of nitrification along with some detectable denitrification. This work also showed that oxygen, essential for aerobic microbial growth, can permeate through walls of PEX pipe and enter into the water from the atmosphere of the building. Considering the unique array of conditions that were found to influence the persistence of disinfectants in premise plumbing, a new approach was proposed for managing OP risk, referred to herein as the "CT framework." CT was defined as the integral of the chlorine concentration (C) at a point in the premise plumbing versus water retention time (T). Legionella pneumophila was not detectable in pipes with a CT > 78 mg*min/L over a 24 hour period, which is comparable to reported CT thresholds for 3-log inactivation of biofilm-associated L. pneumophila in batch experiments. There was a tradeoff between control of L. pneumophila and Mycobacterium avium in the water heater, as M. avium increased by >1 log as influent chloramine and CT increased, while L. pneumophila decreased by >1.5 logs. Further research is needed to elucidate the influence of factors such as water storage tank hydrodynamics and sediment on the persistence of different OPs. Building water retention time was also found to be an overarching variable that governs microbial growth in some circumstances in premise plumbing. Total cell counts and L. pneumophila occurrence mirrored expected trends based on the classic microbial growth curve with phases of lag, exponential growth, stationary growth, and decay. The location in the plumbing system where each phase dominated depended on water retention time, disinfectant level, and temperature. The microbial growth curve considerations add an additional dimension to the CT framework for predicting L. pneumophila growth potential in premise plumbing. Specifically, elevated heat or chloramine, was able to temporarily suppress or even eliminate growth, but the phases of classic microbial growth could be restarted once disinfectant or very high temperatures were absent. Total cell counts and L. pneumophila typically peaked at a building water retention time of 7 days, demonstrating that once a week flushing guidance to protect public health may not be advantageous in all situations. Collectively, this work offers fundamental and practical insights into factors driving disinfectant decay and microbial proliferation in premise plumbing, offering a modified CT and microbial growth concept framework to help guide the management of OPs in premise plumbing. / Doctor of Philosophy / Access to safe drinking water is fundamental to human health and wellbeing and is considered to be a human right by some agencies. Opportunistic pathogens (OPs) can grow in some drinking water systems and cause deadly diseases, such as Legionnaires' Disease. Legionnaires' Disease and illnesses caused by other OPs are now the leading cause of drinking water-associated disease in the U.S. and many other countries. Chlorine or chloramine are disinfectants required to be present in treated drinking water in the U.S. before it is piped through the distribution systems to consumers. This helps to limit growth of OPs and other microbes in the distribution systems. However, the concentration of disinfectant that remains in water as it crosses the property line is sometimes inadequate to suppress OP growth. Even if the amount of disinfectant entering a building is boosted, there are some plumbing materials and circumstances that can quickly reduce the disinfectant. These challenges are sometimes worsened by water and energy conservation efforts, which extend the time water spends in a building and presents tradeoffs with preventing OP growth. This dissertation examines how multiple factors at play in drinking water distribution systems and building plumbing individually and collectively influence OP growth, with a goal of developing rational guidance to reduce incidence of waterborne illness. Experiments were conducted using a large at-scale building plumbing system. These experiments revealed new insights into the relationship among factors such as how long the water stagnates in pipes, water temperature, the disinfectant concentration at each tap, and the level of specific OPs of concern. Chloramine was gone within four hours of stagnation in plastic cross-linked polyethylene (PEX) pipes containing a mature biofilm, which is 100-1000× faster than observed in the same water that did not contact pipes. The rate at which chloramine disappeared changed with conditions from tap to tap, or with time at a given tap, in ways that were unexpected based on prior assumptions. Further, the hydraulic characteristics and low temperature of the water heater influenced chloramine decay in the tank in a way that increased survival and release of OPs. We found that other microbes residing in pipes, such as nitrifying microbes, can also play a role in decay of disinfectant and their activity also is controlled by the water retention time and temperature in the system. These findings reinforce the need to thoroughly understand how chemical, biological, and hydraulic factors combine to influence OP growth in buildings. To account for the array of factors that contribute to the decay of disinfectant, we introduce premise plumbing "CT" as a new integrative framework to guide management of OPs. We define CT as the integral of the disinfectant concentration (C) at a stagnant point in the building plumbing verses the time (T) water has resided at that point, to characterize the ability of the water to kill or suppress growth of bacteria. If the calculated CT values in the at-scale plumbing system were high enough, Legionella pneumophila, the OP that causes Legionnaires' Disease, was never detected in pipes. However, if CT was too low, L. pneumophila was not controlled. Oddly, M. avium, another problematic OP, exhibited a contradictory trend within the water heater. This indicates that the CT concept may not control M. avium in chloraminated water heaters with complex water flow patterns and sediment. Higher chloramine caused lower L. pneumophila and higher M. avium in the water heater, but this tradeoff did not occur in cold water pipes when the room temperature was below that required for OP growth, indicating that room temperature setpoint could be a significant factor for OP control in buildings. Building water retention time, which is the time that water takes to move through the plumbing before it is consumed from a tap, was identified in this research to be a key driver of microbial growth that can be readily controlled by building managers. Trends of total microbial cell count and L. pneumophila in the premise plumbing system and complementary experiments followed all the phases of growth associated with bacteria in a simple glass jar, including a lag, rise, peak, and then decay of cells. Elevated heat or chloramine was able to temporarily suppress growth or even kill cells, but the phases of growth were again observed once the chemical or thermal disinfectant was removed. In any building, there is likely a frequency of flushing water at a given tap that is "worst case" for bacterial growth. In the absence of disinfectant, bacteria in pipes that are frequently supplied with nutrients through fresh water can be expected to have sustained growth, but if bacteria are starved of nutrients, there is some die off. In our system, total microbial cell counts and L. pneumophila peaked at a water retention time of about one week. Thus, this work suggests that current advice to flush building pipes once a week might sometimes create issues with microbial growth rather than solve them. Collectively, this research advances both fundamental and practical understanding of the factors driving disinfectant decay and microbial proliferation in premise plumbing. The premise plumbing CT and microbial growth concept framework is introduced to help inform better management of building water systems to prevent or remediate the growth of pathogens and reduce risk of human infection.

Opportunistic premise plumbing pathogens (OPPPs) are responsible for many infections linked to drinking water. The annual cost of disease caused by these waterborne pathogens is $850 million. Key characteristics of these opportunistic waterborne pathogens include: disinfectant- resistant, biofilm formation, thermal-tolerance, desiccation-resistant, growth in amoebae and growth in low oxygen conditions. Methylobacterium spp. have been recognized as an emerging OPPP, so the purpose of this study was to investigate these waterborne bacteria in more detail to determine whether they have all characteristics of OPPPs. Seven Methylobacterium spp. strains were studied to measure growth in laboratory broth medium and drinking water, measure hydrophobicity on surfaces found in household plumbing, measure adherence and biofilm formation to surfaces found in household plumbing and measure susceptibility to hot water heater temperatures. Methylobacterium spp. were found to aggregate in lab broth medium and drinking water, hydrophobic on different surfaces in household plumbing, adhere readily and form biofilm on different surfaces and thermal-tolerant to water heater temperatures. These results support and identify Methylobacterium spp. as opportunistic premise plumbing pathogens. / Master of Science / Opportunistic premise plumbing pathogens (OPPPs) are microbial residents of drinking water systems and premise plumbing that cause infection. Premise plumbing includes water pipes in hospitals, houses, apartment buildings or office buildings. OPPPs share a number of characteristics that contribute to their growth and survival in drinking water systems. In this study, <i>Methylobacterium</i> spp., an emerging OPPP, were studied to see if they share all of the characteristics of OPPPs. Seven <i>Methylobacterium</i> spp. strains were studied to measure growth in laboratory broth medium and drinking water, measure hydrophobicity (ability to repel water) on surfaces found in household plumbing, measure adherence to surfaces found in household plumbing and measure susceptibility to high temperatures. <i>Methylobacterium</i> spp. were found to form clusters of cells in lab broth medium and drinking water, hydrophobic on different surfaces in household plumbing, adhere readily on different surfaces and resistant to high temperatures. These results support <i>Methylobacterium</i> spp. are opportunistic premise plumbing pathogens. This is important because there is now a better understanding of how <i>Methylobacterium</i> spp. survive in drinking water systems to prevent its growth and persistence. This study was also able to determine which pipe surfaces support the least amount of <i>Methylobacterium</i> spp. growth to be used be used by plumbers and homeowners to reduce exposure to <i>Methylobacterium</i> spp.