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.

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.

Assimilable organic carbon (AOC) is a suspected contributor to growth of microbes, including pathogens, in plumbing systems. Two phases of research were completed to improve knowledge of AOC and other forms of organic carbon in premise plumbing. In the first phase, the AOC Standard Method 9217B was compared to a new luminescence-based AOC in terms of time, cost, convenience, and sources of error. The luminescence method was generally more accurate, as it better captured the peak growth of the test organisms. It was also less expensive and less time-consuming. A few approaches to improving the accuracy of the method and detect possible errors were also presented. In the second phase of research, the possibility of AOC generation in premise plumbing was reviewed and then tested in experiments. It has been hypothesized that removal of AOC entering distribution systems might be a viable control strategy for opportunistic premise plumbing pathogens (OPPPs), but if AOC was generated in premise plumbing systems this approach would be undermined. Possible sources of AOC creation in premise plumbing, which is herein termed "distribution system derived biodegradable organic carbon (DSD-BDOC)," include: leaching of organic matter from cross linked polyethylene (PEX) pipes, autotrophic oxidation of H2 generated from metal corrosion (e.g. sacrificial magnesium anode rods and iron pipes), rendering of humic substances more biodegradable by sorption to oxides such as Fe(OH)3, and accumulation of AOC on filters and sediments. The potential for various plumbing and pipe materials to generate AOC was compared in controlled simulated water heater experiments. Under the worst-case condition, generation up to 645 µg C/L was observed. IT was not possible to directly confirm the biodegradability of the generated organic carbon, and there were generally no correlations between suspected generation of organic carbon and either heterotrophic plate counts (HPC) or of bacterial 16S rRNA genes. DSD-BDOC was also explored in a simulated distribution system with two disinfectant types (chlorine and chloramine) and three pipe materials (PVC, cement, and iron). TOC increased with water age, probably due to leaching of organics from PVC and possibly the aforementioned DSD-BDOC due to autotrophic reactions of nitrifiers and iron-related bacteria. As before, relationships between the higher levels of organic carbon and either HPC or 16S were not observed. / Master of Science

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.

<p>Significant seasonal changes in chemical and microbiological water quality can occur in buildings at different fixture locations due to temperature and time dependent reaction rates. Here, a series of calibrated plumbing hydraulic-water quality models were developed for the extensively monitored Retrofitted Net-zero Energy, Water & Waste (ReNEWW) house in West Lafayette, Indiana USA. Knowledge gaps that inhibited higher resolution water quality modeling were also identified. The eight new models predict the absolute level of free chlorine, total trihalomethanes (TTHM), Cu (Copper), Fe (Iron), Pb (lead), NO₃^–(nitrate-nitrogen), heterotrophic plate count (HPC), and <i>Legionella spp.</i> concentration at each fixture for plumbing use, operational characteristics, and design layouts of the plumbing system. Model development revealed that the carrying capacity to describe Legionella spp. growth (and other organisms) under water usage and plumbing design conditions is lacking in the literature. This information needed for higher resolution modeling. Reducing building water use by 25% prompted increased absolute concentrations of HPC and Legionella, each increasing by a factor of about 10⁵. When the service line length was increased, Legionella spp. concentrations increased by up to 10⁶gene copies /L in the Summer season. The proposed modeling framework can be used to support better planning, design, analysis, and operational decision-making.</p>