Microbiological Effectiveness of Mineral Pot Filters in Cambodia

Oct 3, 2012 - Mineral pot filters (MPFs) are household water treatment (HWT) devices that are manufactured and distributed by the private sector, with...
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Microbiological Effectiveness of Mineral Pot Filters in Cambodia Joe Brown,*,† Ratana Chai,‡ Alice Wang,§ and Mark D. Sobsey§ †

Department of Disease Control, Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London WC1E 7HT, United Kingdom ‡ WaterSHED-Cambodia, Phnom Penh, Cambodia § Department of Environmental Sciences and Engineering, Gillings School of Global Public Health, University of North Carolina, Chapel Hill, North Carolina, United States ABSTRACT: Mineral pot filters (MPFs) are household water treatment (HWT) devices that are manufactured and distributed by the private sector, with millions of users in Southeast Asia. Their effectiveness in reducing waterborne microbes has not been previously investigated. We purchased three types of MPFs available on the Cambodian market for systematic evaluation of bacteria, virus, and protozoan surrogate microbial reduction in laboratory challenge experiments following WHO recommended performance testing protocols. Results over the total 1500 L testing period per filter indicate that the devices tested were highly effective in reducing Esherichia coli (99.99%+), moderately effective in reducing bacteriophage MS2 (99%+), and somewhat effective against Bacillus atrophaeus, a spore-forming bacterium we used as a surrogate for protozoa (88%+). Treatment mechanisms for all filters included porous ceramic and activated carbon filtration. Our results suggest that these commercially available filters may be at least as effective against waterborne pathogens as other, locally available treatment options such as ceramic pot filters or boiling. More research is needed on the role these devices may play as interim solutions to the problem of unsafe drinking water in Cambodia and globally.

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INTRODUCTION Lack of safe drinking water is a major cause of infectious disease in low and middle income countries. A recent study has estimated that 1.2 billion people, 28% of the world’s population, lack access to microbiologically or chemically safe drinking water.1 Inadequate access to safe water contributes to the massive global burden of disease and death resulting from diarrheal and other gastrointestinal illnesses, borne primarily by children in lower-income countries.2 Persistent diarrheal disease can lead to malnutrition and micronutrient deficiencies, cognitive effects, and increased risk of death.3−7 Consistent domestic-level supplies of safe drinking water are needed to deliver the health, economic, and other gains that are associated with access to water.1 Household water treatment and safe storage (HWT) has been proposed as an intermediate strategy for delivering some of the health benefits provided by safe water in places where safe, domestic piped water access is not available. Although a variety of HWT technologies have been developed, tested in laboratory and field studies, and proposed for scale-up, not all are supported by evidence demonstrating high performance in reducing microbial and chemical contaminants, health impact, or adherence (correct, consistent, and sustained use).8,9 The pace of development of new technologies and approaches in HWT is rapid, and critical unknowns about current options limit our understanding of how best to interpret and make use of HWT as a strategy for safe water provision.8,9 The Mineral pot filter (MPF) is common in Cambodia and across East and Southeast Asia. In a market survey of HWT options in Vietnam in 2009, researchers from PATH, an © 2012 American Chemical Society

international nonprofit organization, found one or more MPF devices in 80% of the 440 retail outlets surveyed and 32% of households, suggesting that market penetration of MPFs greatly exceeded other available options with an estimated user population of 7,230,000. A similar PATH survey in Cambodia in 2009 found MPFs in every province and retailer-reported high demand for MPFs in urban and peri-urban locations, with 7% market penetration for an estimated 200,000 users (B. McLaughlin and P. Lennon, PATH, personal communication).10 Several manufacturers in China, Vietnam, and other countries produce the devices, which typically retail from US $15 to US$30 in Cambodia in 2012. The products are marketed with a wide range of claims including water sterilization, arsenic removal, pesticide reduction, prevention or treatment of cancer, improvements in users’ libido, or even supernatural benefits of use.10 Systematic performance testing data on commercially available MPF devices are not publicly available. Anecdotal evidence has suggested they may vary widely in effectiveness and in overall production quality, however (M. Sampson, RDIC, personal communication). Despite the existing unknowns about treatment effectiveness, false marketing claims, and lack of standards or regulatory oversight to protect consumers in these countries, these filters are the only commercial HWT device that can be said to have reached scale in Southeast Asia.10 Received: Revised: Accepted: Published: 12055

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and “mineral stone” cartridges may contain activated carbon, zeolites, or other unspecified media. We tested the three most common MPF brands on the domestic market in Cambodia in 2010, from an informal survey of retail outlets in Phnom Penh undertaken by WaterSHEDAsia: Nova, Korea King, and Seoul. We purchased four of each device and assembled units according to manufacturer instructions. All filters employed ceramic candle filtration followed by filtration through an apparent mixture of unspecified granular media in a packed bed. Product water was stored with a “mineral stone” cartridge in receiving containers where treated water could be dispensed via a tap. Overview of Testing Procedure. We used current WHO (2011) performance testing recommendations in developing laboratory methods.9 We dosed the devices daily with approximately 10 L of test water. We monitored performance over 1500 L of total throughput per test filter. Filters were cleaned weekly and as needed, according to manufacturer instructions on proper use and maintenance. Challenge waters were spiked with microbes daily in the morning, allowed to filter for approximately 6 h, and samples for analysis were taken from the post-treatment storage container as a composite of that day’s filtrate. Filters were challenged with all three microbes daily, but untreated and treated waters were assayed once per week per microbe. A comparison of concentrations in pre- and post-treatment water was used to determine the log10 microbial reductions, the principal outcome measures from testing, as:

Our objective in undertaking this research was to evaluate the performance of MPFs for their ability to remove microbes from water over long-term daily use, under realistic use conditions, and in accordance with recently published guidance and recommendations for microbial performance testing by the World Health Organization.9 The microbiological performance data that we have produced is a necessary first step in a broader assessment of the potential current and future role these previously uncharacterized devices may play in providing safer drinking water in Southeast Asia.

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METHODS MPF Devices. Household-scale MPFs operate by gravity filtration: untreated water in a top container flows through one or more filter units, with treated water dispensed via a tap in the storage container (Figure 1). Treatment elements are a ceramic candle or other microporous, solid filter element followed by granular media filtration. A “mineral stone” cartridge containing additional mixed granular media in contact with product water imparts a distinctive mineral taste to the water. A cloth or fiber prefilter may be used before the ceramic filter. Granular media

log10 reduction value (LRV) = log10(pre‐treatment microbial concentration) − log10(post‐treatment microbial concentration)

Challenge Waters. We used test waters of two types for each filter included in testing: (1) dechlorinated Phnom Penh municipal tap water to model high quality sources with low dissolved organic and particulate matter and (2) dechlorinated tap water supplemented with 1% by volume of sterilized untreated wastewater, of higher organic and particulate content. Characteristics of test waters are presented in Table 1 and are consistent with WHO guidance on appropriate challenge waters for the testing of filtration technologies for household water treatment.9 Test waters were seeded daily over 150 days of testing with known concentrations of test viruses, bacteria, and surrogates for protozoan parasites. The test microbes were (1) Escherichia coli as the model bacterium, a surrogate for waterborne bacterial pathogens such as Salmonella spp., Shigella, spp., Vibrio cholerae, and Campylobacter spp.; (2) the coliphage MS2 as the model virus whose size, shape, and other properties are similar to many human enteric viruses such as noroviruses, infectious hepatitis viruses, and enteroviruses; and (3) spores of Bacillus atrophaeus as a surrogate for Cryptosporidium oocysts and other encysted protozoans. These test microbes were added to test water at concentrations sufficient to determine up to 5 log10 reductions (99.999%). Microbiological Methods. E. coli Testing. E. coli strain CN13 was inoculated in tryptic soy broth (TSB) medium (Difco) and incubated overnight (16 h) at 35 °C. The TSB medium was 3 g of tryptic soy broth per 100 mL of reagent water, sterilized, and allowed to cool to 30 °C or lower before use. Because E. coli CN13 is resistant to the antibiotic nalidixic acid, TSB for growing stocks was supplemented with 1%

Figure 1. Typical gravity-driven MPF design showing top container of approximately 10 L capacity (A), untreated water (B), porous ceramic candle filter (C), cylindrical mixed media packed bed filtration cartridge (D), product water (E), cylindrical “mineral stone” element containing additional mixed granular media in contact with product water (F), and tap for dispensing drinking water (G). 12056

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Table 1. Challenge Water Characteristics parameter

challenge water 1

challenge water 2

Phnom Penh municipal tap water, dechlorinated with sodium thiosulfate mean pH, n = 211 6.7 (95% CI 6.6−6.7) mean turbidity, 2.4 (95% CI 2.3−2.6) NTU, n = 224

Phnom Penh municipal tap water, dechlorinated with sodium thiosulfate, supplemented with 1% by volume untreated municipal wastewater autoclaved at 121 °C for 20 min 6.6 (95% CI 6.5−6.7) 2.7 (95% CI 2.6−2.9)

Table 2. Summary of Testing Data on MPF Effectiveness against Test Microbes over 1500 Liters log10 reduction over 1500 liters throughput, arithmetic means (95% CI) dechlorinated tap water

dechlorinated tap water with 1% sterilized untreated wastewater

parameter

n

Nova

Korea King

Seoul

Nova

Korea King

Seoul

E. coli MS2 B. atrophaeus turbidity

49 28 52 224

5.6 (5.0−6.1) 3.0 (2.7−3.3) 2.5 (1.9−3.1) 0.74 (0.67−0.80)

4.2 (3.6−4.9) 3.1 (2.9−3.4) 1.3 (1.0−1.6) 0.64 (0.58−0.69)

4.7 (4.1−5.3) 3.0 (2.7−3.2) 1.6 (1.3−2.0) 0.58 (0.49−0.66)

4.2 (3.6−4.9) 2.0 (1.6−2.3) 1.9 (1.4−2.4) 0.67 (0.61−0.74)

4.2 (3.6−4.7) 2.2 (1.9−2.6) 0.93 (0.76−1.1) 0.74 (0.69−0.80)

4.1 (3.5−4.7) 2.0 (1.8−2.3) 1.2 (0.86−1.5) 0.62 (0.57−0.68)

mL. MS2 bacteriophages in pre- and post-treatment water were enumerated by the plaque technique on tryptic soy agars containing appropriate antibiotics (streptomycin/ampicillin) using the double agar layer (DAL) or spot titer agar medium− host lawn plate techniques,15−18 with host E. coli F-amp.19 The two methods were not significantly different in preliminary comparison tests (data not shown), although the spot titer method does not have as low a detection limit as the DAL method due to the small sample volumes assayed.20 Plaques were counted and bacteriophage concentrations are expressed as plaque forming units (pfu) per 100 mL. Microbiological Methods. B. atrophaeus. Bacillus spp. spores have been suggested as experimental surrogates for Cryptosporidium oocysts in treatment process and transport modeling.9,21−24 We used spores of B. atrophaeus as surrogates for protozoa potentially present in untreated drinking water. Bacillus atrophaeus spores were produced on AK Agar #2 (sporulating agar), with methods for stock production, spore harvest, and storage based on published protocols.21,23 In order to assay only spores and not vegetative cells that might also be present in samples, the samples were pretreated by heat exposure at 70 °C for 20 min before culturing at 37 °C for 24 h. We used a membrane filtration-based assay that has been developed for rapid enumeration of Bacillus spp. spores.25 Physical−Chemical Parameters. Turbidity of all water samples was measured in triplicate using a Hach turbidimeter (model 2100P), and the average values were reported as nephelometric turbidity units (NTU). Sample pH was measured in triplicate using a Hach pH meter (model HQ11d). We also collected pre- and post-treatment samples for a future analysis of metals leaching. Statistical Analysis. E. coli, MS2, and B. atrophaeus concentrations in samples were calculated on the basis of a minimum of two dilutions and three replicates per dilution. Log10 reductions were calculated for each test with detectable microbes in matched untreated and treated water samples. Descriptive statistics were used to characterize water quality testing results. Parametric and nonparametric statistical tests were used to compare results when data were normally and non-normally distributed, respectively, as determined by a Shapiro-Wilk normality test.26 All statistics were interpreted using an a priori significance level of α = 0.05. All statistical testing was performed in Stata version 12.1 (StataCorp, College Station, TX, USA).

nalidixic acid 100× stock concentrate (1 g of nalidixic acid sodium salt dissolved in 100 mL of reagent water, filter sterilized via a 0.22 μm pore size membrane filter assembly, added at 0.1 mL of nalidixic acid to 10 mL of TSB for a final concentration of 100 mg/L).11 After overnight incubation, 1 mL of E. coli culture was transferred aseptically to 30 mL of fresh TSB medium (with nalidixic acid) in a shaker flask and incubated at 35 °C for 3−4 h at 35 °C, until absorbance was measured to be approximately 1.5 units at 520 nm. Once cultures reached the desired growth phase, 20 mL samples were taken and centrifuged at 4800g for 20 min. The supernatant was discarded, and the pellet of E. coli cells was washed 3 times and resuspended in 20 mL of deionized (DI) water. One mL of this cell suspension was added per 10 L of each challenge water. The final concentration of E. coli CN13 was 105−107 cfu/mL in challenge waters. E. coli in pre- and post-filter samples was enumerated by filtering undiluted and diluted samples through 47 mm diameter, 0.45 μm pore size cellulose ester filters in standard, sterile magnetic membrane filter funnels, and membranes were incubated on agar or broth media-soaked absorbent pads. Agar and broth media (Rapid HiColiform media, HiMedia, M1465/M1453) detected total coliform (TC) bacteria and E. coli by cleavage of a chromogenic β-galactoside substrate to detect total coliforms and a fluorogenic βglucuronide substrate to detect E. coli, producing distinctive color TC colonies and blue fluorescing E. coli colonies under long-wave UV light at 366 nm.12−14 Plates were incubated for 20−24 h at 35 °C. These methods conform to EPA Approved Method 1604,11 except HiMedia M1465 and M1453 were substituted for the more costly MI medium used in the EPA method. In preliminary studies in which samples were plated on both media, MI and M1465 or M1453, E. coli detection was comparable (data not shown). E. coli concentrations were expressed as colony forming units (cfu) per unit volume of water. Microbiological Methods. MS2 Testing. High titer stocks of the F+RNA coliphage MS2 were produced through confluent lysis on soft agar medium plates with phages, logphase host (E. coli F-amp), streptomycin, and ampicillin and incubated at 35 °C for 24 h. The lysate−agar mixture was subjected to chloroform extraction. Phage stocks were assayed to determine infectivity titer using standard plaque assay techniques.15,16 Stocks of high titer bacteriophage were spiked into each challenge water to concentrations of 105−108 pfu/ 12057

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RESULTS

Results from challenge testing are summarized in Table 2 and in Figures 2−4. Over the 1500 L cumulative testing period per filter, all filters reduced E. coli by a mean of 99.99% (4.0 log10) or greater, MS2 by a mean of 99% (2.0 log10) or greater, and B.

Figure 3. MS2 challenge test results showing mean log10 reductions over 1500 L.

atrophaeus by 88% (0.93 log10) or greater, in both challenge waters. Moderate variability in effectiveness is apparent between filter units (Table 2), as indicated by 95% confidence intervals and over time (Figures 2−4). Nova was more effective in reducing E. coli in dechlorinated tap water (p = 0.003), and all filters were more effective against MS2 (p < 0.001, p < 0.001, p < 0.001, respectively) using

Figure 2. E. coli challenge test results showing mean log10 reductions over 1500 L. 12058

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0.048) in both dechlorinated tap water and in dechlorinated tap water supplemented with wastewater (p = 0.001 and p = 0.032, respectively). All filters reduced E. coli to a greater extent than MS2 in a comparison of within-filter mean log10 reductions (p < 0.001 in each case; Table 2). Similarly, all filters reduced MS2 to a greater extent than B. atrophaeus (p < 0.001 in each case; Table 2). All filters reduced turbidity significantly from pretreatment levels (Table 2), although mean turbidity did not exceed 5 NTU even in waters supplemented with 1% sterile wastewater. Pretreatment water pH did not change significantly following treatment (data not shown).

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DISCUSSION Results from testing suggest that at least one filter (Nova) could meet WHO recommended performance levels for the “Protective” level9 but not consistently across test waters. The three filters were as effective or more effective than other locally available drinking water treatment options, including ceramic filters,27,28 biosand filters,29,30 and boiling.31 Fluctuation in performance over time, evident in Figures 2−4, may be related to unmeasured changes in water chemistry, variations in pretreatment concentration, or other factors which may also vary under use conditions. As measured by mean performance, however, our results indicate that the MPF devices we tested have potential to deliver microbiologically safer drinking water to users over extended use. The reasons for relatively low reductions of B. atrophaeus spores observed in this study are uncertain. It is possible that because the spores are relatively hardy, they were unaffected by any biological activity or antimicrobial chemical agents (if any) in the filters. Because they are relatively hydrophobic, they may not have been amenable to removal and retention by electrostatic interaction with the filter media. Furthermore, retained spores could have possibly germinated, propagated, and then resporulated in the filter medium resulting in overall low net spore reductions by the filter system and may have contributed to the evident variable reduction of B. atrophaeus over the course of testing (Figure 4). Further studies would be needed to determine if these proposed explanations are valid. In this study, we assessed MPF effectiveness in reducing microbes only and did not evaluate the capacity of the devices for removing chemicals that may be present in untreated water. An unpublished report32 by researchers at Resource Development International−Cambodia and the Royal University of Phnom Penh concluded that MPFs were not effective against arsenic or fluoride, removing a mean 30% of arsenic in laboratory testing. Further chemical testing is warranted in light of the effectiveness claims made by manufacturers in reducing arsenic, pesticides, and other chemicals. A commonly expressed concern about MPFs is that uncharacterized “mineral stone” media may leach unsafe levels of chemical contaminants into product water (personal communication, M. Sampson, A. Shantz, RDIC; B. McLaughlin and P. Lennon, PATH). Our study did not examine this issue, although we did take and preserve samples for metals analyses for further work. Some MPF manufacturers claim that mineral stones contain exotic materials such as germanium which are advertised as having effects as diverse as preventing cancer or increasing sex drive. The uncharacterized and often unlabeled use of minerals of unknown composition or purity as filtration media or in contact with product water is a potential cause for concern. Chemical analyses of treated water for undesirable

Figure 4. B. atrophaeus challenge test results showing mean log10 reductions over 1500 L.

dechlorinated tap water than dechlorinated tap water supplemented with 1% sterilized wastewater. The Korea King filter was more effective in reducing B. atrophaeus in dechlorinated tap water than sewage-supplemented dechlorinated tap as well (p = 0.017), but Nova and Seoul were not (p = 0.23 and p = 0.055, respectively). Nova reduced B. atrophaeus to a greater extent than Korea King (p = 0.001) or Filter 3 (p = 12059

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levels of toxic compounds associated with the filter or cartridge media would be a useful next step in evaluating these technologies. The apparent diversity of mineral stone formulations means that product water should be assayed for a wide range of potentially harmful chemicals under a variety of challenge conditions to ensure that treated water is safe for users. There is a wide variety of MPFs now on the market locally and across the region, so general statements about the effectiveness of these devices across all manufacturers and models are not possible. Source waters that are of higher turbidity or different pH than our laboratory challenge waters are not uncommon in Cambodia, and these differences may affect performance in situ. User behaviors, including unsafe water handling and device maintenance, may also be associated with recontamination of treated water, so performance in the household may differ from the data we report. For these reasons, field-based studies of these filters as they are actually used may be helpful. We stress the need for caution in interpreting and generalizing these results. Nevertheless, these results are encouraging and underscore the need for further characterization of technologies that are being used widely for HWT. Additional studies may support a more complete and comprehensive understanding of the range of effectiveness of these filters and their modes of action for microbial reduction under laboratory and household use conditions, revealing common features that either contribute to or undermine microbial reduction performance. Gaining such insights could lead to recommendations for MPF properties that maximize microbial reductions and minimize possible leaching of undesirable chemicals from the filter media. The significant and expanding market penetration of MPFs in Southeast Asia has demonstrated that appealing, userfocused products for water treatment can reach scale without subsidies, even when the initial investment at the household level is significant. MPFs are designed to appeal to consumers’ aesthetic sensibilities and are supported by savvy marketing, a strategy that has drawn attention from designers and technology developers, donors, and implementers who seek to expand access to HWT through enterprise development and commercial marketing. Since 2010, the Seattle-based international nonprofit organization PATH has been working with manufacturers in China to develop and market HWT devices based on the MPF model, an acknowledgment of the benefits of starting with a technology and design that people will use and demand and working on making it more effective, reliable, and available to the people who could most benefit from it. The reverse model, starting with an effective technology and then working out how to persuade people to purchase and use it, has been and continues to be the operational model followed by many manufacturers and implementers of technologies,31 which may be one factor that has limited the success of efforts to scale up HWT.33,34 Recent evidence from Bangladesh35,36 and elsewhere underscores the difficulty of generating demand for and sustained use of water treatment products, despite the evidence that adherence plays a critical role in achieving health gains expected of HWT.37,38 Further analysis of the performance of the spectrum of MPFs available to consumers and further consideration of how to achieve effective performance with MPFs and other HWT products that consumers desire and are willing to pay for is recommended. The development of effective, practical, and uniform policies, regulations, certification criteria, and perform-

ance evaluation protocols and procedures is also needed at national levels, where uncharacterized water treatment technologies are being sold to the public. Finally, this study represents the first application of the new WHO performance testing recommendations for HWT,9 which have been conceived to be flexible and adaptable to basic microbiological facilities in resource-limited settings and to provide country-specific testing data on locally available technologies. While Cambodia does not yet have a national certification program for HWT, the development of this testing laboratory and the demonstration that systematic and rigorous evaluations of HWT performance are locally possible are two steps toward that important goal.

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AUTHOR INFORMATION

Corresponding Author

*Tel: +44 (0)7923 166 928; e-mail: [email protected]. Author Contributions

The manuscript was written by J.B. with contributions from all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This study was supported by the UNU and GIST Joint Programme (IERC) Gwangju Institute of Science and Technology and WaterSHED-Asia, a Global Development Alliance supported by USAID and awarded to the University of North Carolina−Chapel Hill. We gratefully acknowledge laboratory assistance from Sina Ngov, Mark Elliott, and Doug Wait. Thanks also to Andrew Shantz of Resource Development International−Cambodia and PATH, who provided insightful comments on drafts of this manuscript.

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REFERENCES

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dx.doi.org/10.1021/es3027852 | Environ. Sci. Technol. 2012, 46, 12055−12061