Kara L. Nelson

Covariation and photoinactivation of traditional and novel indicator organisms and human viruses at a sewage-impacted marine beach.

Sunlight modulates concentrations of Escherichia coli and enterococci in marine waters. However, the mechanism of photoinactivation is poorly understood. Additionally, little is known about photoinactivation of other fecal indicators and human viruses in recreational waters. We sampled nearshore waters at Avalon Beach, California hourly for 72 h for reactive oxygen species (ROS), traditional indicator bacteria (E. coli and enterococci, and QPCR-based detection of enterococci), F+ (DNA and RNA) and somatic coliphages, the human-specific marker in Bacteroidales (HF marker), human enterovirus, and human adenovirus. E. coli and enterococci (regardless of measurement technique) covaried with each other and the coliphages suggesting similar sources and fates. The occurrence of the HF and enterovirus markers was correlated, but their occurrence was not positively correlated with the other indicators. Lower concentrations or occurrence of all microbes, excluding the HF and enterovirus markers, were observed during sunlit as opposed to dark hours, pointing to the importance of photoinactivation. Empirical-deterministic models for a subset of microbial indicators were created to determine field-relevant sunlight inactivation rates while accounting for time dependent sources and sinks. Photoinactivation rates of enterococci and E. coli, enterococci measured by QPCR, and somatic coliphage were estimated at 7, 6, 3, and 28 d(-1) I(-1), respectively, where I is UVB intensity in W/m(2). Average H(2)O(2) was 183 nM and the maximum singlet oxygen steady state concentration was 6.6 fM. Given the clarity of the water, direct genomic damage of bacteria and coliphage, as well as indirect endogenous damage of bacteria, were likely the most important inactivation mechanisms, but we cannot rule out a contribution by indirect mechanisms involving the H(2)O(2) and singlet oxygen produced exogenously.

Inactivation of Escherichia coli by Nanoparticulate Zerovalent Iron and Ferrous Ion

ABSTRACT The mechanism of Escherichia coli inactivation by nanoparticulate zerovalent iron (nZVI) and Fe(II) was investigated using reactive oxygen species (ROS) quenchers and probes, an oxidative stress assay, and microscopic observations. Disruption of cell membrane integrity and respiratory activity was observed under deaerated conditions [more disruption by nZVI than Fe(II)], and OH or Fe(IV) appears to play a role.

Comparing microbial water quality in an intermittent and continuous piped water supply

Supplying piped water intermittently is a common practice throughout the world that increases the risk of microbial contamination through multiple mechanisms. Converting an intermittent supply to a continuous supply has the potential to improve the quality of water delivered to consumers. To understand the effects of this upgrade on water quality, we tested samples from reservoirs, consumer taps, and drinking water provided by households (e.g. from storage containers) from an intermittent and continuous supply in Hubli-Dharwad, India, over one year. Water samples were tested for total coliform, Escherichia coli, turbidity, free chlorine, and combined chlorine. While water quality was similar at service reservoirs supplying the continuous and intermittent sections of the network, indicator bacteria were detected more frequently and at higher concentrations in samples from taps supplied intermittently compared to those supplied continuously (p < 0.01). Detection of E. coli was rare in continuous supply, with 0.7% of tap samples positive compared to 31.7% of intermittent water supply tap samples positive for E. coli. In samples from both continuously and intermittently supplied taps, higher concentrations of total coliform were measured after rainfall events. While source water quality declined slightly during the rainy season, only tap water from intermittent supply had significantly more indicator bacteria throughout the rainy season compared to the dry season. Drinking water samples provided by households in both continuous and intermittent supplies had higher concentrations of indicator bacteria than samples collected directly from taps. Most households with continuous supply continued to store water for drinking, resulting in re-contamination, which may reduce the benefits to water quality of converting to continuous supply.

Solar water disinfection (SODIS) of Escherichia coli, Enterococcus spp., and MS2 coliphage: Effects of additives and alternative container materials

The use of alternative container materials and added oxidants accelerated the inactivation of MS2 coliphage and Escherichia coli and Enterococcus spp. bacteria during solar water disinfection (SODIS) trials. Specifically, bottles made from polypropylene copolymer (PPCO), a partially UVB-transparent plastic, resulted in three-log inactivation of these organisms in approximately half the time required for disinfection in bottles made from PET, polycarbonate, or Tritan(®), which absorb most UVB light. Furthermore, the addition of 125 mg/L sodium percarbonate in combination with either citric acid or copper plus ascorbate tended to accelerate inactivation by factors of 1.4-19. Finally, it was observed that the inactivation of E. coli and enterococci derived from local wastewater was far slower than the inactivation of laboratory-cultured E. coli and Enterococcus spp., while the inactivation of MS2 was slowest of all. These results highlight the importance of UVB in SODIS under certain conditions, and also the greater sunlight resistance of some viruses and of bacteria of fecal origin, as compared to the laboratory-cultured bacteria commonly used to model their inactivation. Furthermore, this study illustrates promising new avenues for accelerating the inactivation of bacteria and viruses by solar disinfection.

Following the Water: A Controlled Study of Drinking Water Storage in Northern Coastal Ecuador

Background To design the most appropriate interventions to improve water quality and supply, information is needed to assess water contamination in a variety of community settings, including those that rely primarily on unimproved surface sources of drinking water. Objectives We explored the role of initial source water conditions as well as household factors in determining household water quality, and how levels of contamination of drinking water change over time, in a rural setting in northern coastal Ecuador. Methods We sampled source waters concurrently with water collection by household members and followed this water over time, comparing Escherichia coli and enterococci concentrations in water stored in households with water stored under controlled conditions. Results We observed significant natural attenuation of indicator organisms in control containers and significant, although less pronounced, reductions of indicators between the source of drinking water and its point of use through the third day of sampling. These reductions were followed by recontamination in approximately half of the households. Conclusions Water quality improved after water was transferred from the source to household storage containers, but then declined because of recontamination in the home. Our experimental design allowed us to observe these dynamics by controlling for initial source water quality and following changes in water quality over time. These data, because of our controlled experimental design, may explain why recontamination has been reported in the literature as less prominent in areas or households with highly contaminated source waters. Our results also suggest that efforts to improve source water quality and sanitation remain important.

Sunlight inactivation of human viruses and bacteriophages in coastal waters containing natural photosensitizers

Sunlight inactivation of poliovirus type 3 (PV3), adenovirus type 2 (HAdV2), and two bacteriophage (MS2 and PRD1) was investigated in an array of coastal waters to better understand solar inactivation mechanisms and the effect of natural water constituents on observed inactivation rates (k(obs)). Reactor scale inactivation experiments were conducted using a solar simulator, and k(obs) for each virus was measured in a sensitizer-free control and five unfiltered surface water samples collected from different sources. k(obs) values varied between viruses in the same water matrix, and for each virus in different matrices, with PV3 having the fastest and MS2 the slowest k(obs) in all waters. When exposed to full-spectrum sunlight, the presence of photosensitizers increased k(obs) of HAdV2, PRD1 and MS2, but not PV3, which provides evidence that the exogenous sunlight inactivation mechanism, involving damage by exogenously produced reactive intermediates, played a greater role for these viruses. While PV3 inactivation was observed to be dominated by endogenous mechanisms, this may be due to a masking of exogenous k(obs) by significantly faster endogenous k(obs). Results illustrate that differences in water composition can shift absolute and relative inactivation rates of viruses, which has important implications for natural wastewater treatment systems, solar disinfection (SODIS), and the use of indicator organisms for monitoring water quality.

The joint effects of efficacy and compliance: A study of household water treatment effectiveness against childhood diarrhea

The effectiveness of household water treatment (HWT) at reducing diarrheal disease is related to the efficacy of the HWT method at removing pathogens, how people comply with HWT, and the relative contributions of other pathogen exposure routes. We define compliance with HWT as the proportion of drinking water treated by a community. Although many HWT methods are efficacious at removing or inactivating pathogens, their effectiveness within actual communities is decreased by imperfect compliance. However, the quantitative relationship between compliance and effectiveness is poorly understood. To assess the effectiveness of HWT on childhood diarrhea incidence via drinking water for three pathogen types (bacterial, viral, and protozoan), we developed a quantitative microbial risk assessment (QMRA) model. We examined the relationship between log(10) removal values (LRVs) and compliance with HWT for scenarios varying by: baseline incidence of diarrhea; etiologic fraction of diarrhea by pathogen type; pattern of compliance within a community; and size of contamination spikes in source water. Benefits from increasing LRVs strongly depend on compliance. For perfect compliance, diarrheal incidence decreases as LRVs increase. However, if compliance is incomplete, there are diminishing returns from increasing LRVs in most of the scenarios we considered. Higher LRVs are more beneficial if: contamination spikes are large; contamination levels are generally high; or some people comply perfectly. The effectiveness of HWT interventions at the community level may be limited by imperfect compliance, such that the benefits of high LRVs are not realized. Compliance with HWT should be carefully measured during HWT field studies and HWT dissemination programs. Studies of pathogen concentrations in a variety of developing-country source waters are also needed. Guidelines are needed for measuring and promoting compliance with HWT.

A real-time PCR method for quantifying viable ascaris eggs using the first internally transcribed spacer region of ribosomal DNA.

ABSTRACT Worldwide, 1.4 billion people are infected with the intestinal worm Ascaris lumbricoides. As a result, Ascaris eggs are commonly found in wastewater and sludges. The current microscopy method for detecting viable Ascaris eggs is time- and labor-intensive. The goal of this study was to develop a real-time quantitative PCR (qPCR) method to determine the levels of total and viable Ascaris eggs in laboratory solutions using the first internally transcribed spacer (ITS-1) region of ribosomal DNA (rDNA) and rRNA. ITS-1 rDNA levels were proportional to Ascaris egg cell numbers, increasing as eggs developed from single cells to mature larvae and ultimately reaching a constant level per egg. Treatments causing >99% inactivation (high heat, moderate heat, ammonia, and UV) eliminated this increase in ITS-1 rDNA levels and caused decreases that were dependent on the treatment type. By taking advantage of this difference in ITS-1 rDNA level between viable, larvated eggs and inactivated, single-celled eggs, qPCR results were used to develop inactivation profiles for the different treatments. No statistical difference from the standard microscopy method was found in 75% of the samples (12 of 16). ITS-1 rRNA was detected only in samples containing viable eggs, but the levels were more variable than rDNA levels and ITS-1 rRNA could not be used for quantification. The detection limit of the rDNA-based method was approximately one larvated egg or 90 single-celled eggs; the detection limit for the rRNA-based method was several orders of magnitude higher. The rDNA qPCR method is promising for both research and regulatory applications.

Human virus and bacteriophage inactivation in clear water by simulated sunlight compared to bacteriophage inactivation at a southern California beach.

Few quantitative data exist on human virus inactivation by sunlight and the relationship between human and indicator viruses under sunlit conditions. We investigated the effects of sunlight on human viruses (adenovirus type 2, poliovirus type 3) and bacteriophages (MS2, Q-Beta SP, Fi, M13, PRD1, Phi-X174, and coliphages isolated from Avalon Bay, California). Viruses were inoculated into phosphate buffered saline or seawater, exposed to a laboratory solar simulator for ≤12 h, and enumerated by double agar layer or cell culture to derive first-order inactivation rate constants (k(obs), h(-1)). The viruses most resistant to sunlight were adenovirus type 2 (k(obs)= 0.59 ± 0.04 h(-1)) and bacteriophage MS2 (k(obs)= 0.43 ± 0.02 h(-1)), which suggests MS2 may be a conservative indicator for sunlight resistant human viruses in clear water when sunlight inactivation is the main removal mechanism. Reasonable agreement was observed between somatic coliphage inactivation rates measured in the solar simulator (k(mean) = 1.81 h(-1)) and somatic coliphages measured in the surf zone during a field campaign at Avalon Bay during similar sunlight intensity (k = 0.75 h(-1) at log-RMSE minimum; k(range) = 0.54 h(-1) to >1.88 h(-1); Boehm, A. B. et al. Environ. Sci. Technol. 2009, 43, (21), 8046-8052). Hence, measuring sunlight inactivation rates of viruses in the laboratory can be used to estimate inactivation in the environment under similar sunlight and water quality conditions.

Inactivation of MS2 Coliphage by Ferrous Ion and Zero-Valent Iron Nanoparticles

This study demonstrates the inactivation of MS2 coliphage (MS2) by nano particulate zerovalent iron (nZVI) and ferrous ion (Fe[II]) in aqueous solution. For nZVI, the inactivation efficiency of MS2 under air-saturated conditions was greater than that observed under deaerated conditions, indicating that reactions associated with the oxidation of nZVI were mainly responsible for the MS2 inactivation. Under air-saturated conditions, the inactivation efficiency increased with decreasing pH for both nZVI and Fe(II), associated with the pH-dependent stability of Fe(II). Although the Fe(II) released from nZVI appeared to contribute significantly to the virucidal activity of nZVI, several findings suggest that the nZVI surfaces interacted directly with the MS2 phages, leading to their inactivation. First, the addition of 1,10-phenanthroline (a strong Fe(II)-chelating agent) failed to completely block the inactivation of MS2 by nZVI. Second, under deaerated conditions, a linear dose-log inactivation curve was still observed for nZVI. Finally, ELISA analysis indicated that nZVI caused more capsid damage than Fe(II).