Dirty water has a profound effect on the lives of the global majority. Waterborne disease takes lives directly (1), and further weakens or kills those affected by HIV/AIDS, malaria, and malnutrition (2, 3). Viral, bacterial, prion, fungal, and protozoan waterborne diseases can be eliminated simultaneously with the provision of clean drinking water (3). In regions where reliable sanitation systems do not yet exist, local environmental conditions may dictate which pathogens will be more abundant. This article will focus on the cholera pathogen Vibrio cholerae, a bacterium that causes extreme physical pain with high mortality, primarily in Southeast Asia, Africa, and Latin America, although it occurs throughout the world (4, 5, 6).
V. cholerae is a gram-negative bacterium responsible for 180,000-500,000 cases of cholera infection annually (7, 8). V. cholerae has at least 155 serogroups, and of these, only two serogroups contain strains that are currently responsible for epidemic and endemic cholera (i.e. O1 and O139) (9). Strains within these two serogroups vary in production of cholera-toxin, colonization factors, surface antigens, and polysaccharides contributing to chlorine-resistance (10). Since 1817, eight pandemics of cholera have taken lives across the world, spanning all continents except Australia and Antarctica (6).
The exchange of V. cholerae between regions can occur by several vectors, including by infected individuals, or contaminated food or water (6, 11). Pathogenic strains of V. cholerae can remain viable in fresh, brackish, or salt water for months before disease outbreaks actually occur, which is typically when water temperatures rise (12). In these aquatic systems, the bacteria can attach to many different organisms including zooplankton (e.g. copepods) (13), phytoplankton (14), or amoebas (15), and benthic organisms such as oysters (11). The bacteria can also form biofilms on these hosts (16, 17). Influxes of rain or nutrients may cause seasonal maxima first of phytoplankton populations and then copepods (6, 11). With a bloom of organisms that serve as V. cholerae attachment sites, and with additional resources provided directly to the bacteria, the bacterial populations thrive (6). Untreated water therefore has a greater probability of containing sufficient toxin-producing cells to initiate an infection, especially for children, the elderly, or those with compromised immune systems (3, 8).
Once an infectious dose of approximately 104-106 cells is ingested (6), the production of cholera toxin acts primarily on the epithelial cells of the intestine (8, 18). After binding to a cellular receptor, the toxin’s most significant effect is inhibition of the guanosine triphosphatase (GTPase) activity of a signaling protein subunit (Gαs) (19, 20). As part of the signaling pathway, when Gαs is bound to GTP (Gαs -GTP), it activates adenylate cyclase which then produces cyclic adenosine 3′,5′-monophosphate (cAMP) (18). The secondary messenger cAMP activates chloride export by the cystic fibrosis transmembrane conductance regulator (CFTR) and inhibits sodium absorption (8). The cells therefore continue to lose salts, which are followed by water through osmosis. Normally, the hydrolysis of GTP on Gαs stops the signaling pathway, however the cholera toxin has inhibited the GTPase activity. The result is the rapid onset of diarrhea, with patients losing 10-30 liters of fluids in the first three days (5). The severe volume depletion and electrolyte imbalance that follow can result in dehydration, shock, and acidosis (8). Acidosis occurs after the loss of water and therefore blood volume, which then impairs normal kidney function (21). The tubule cells decrease in efficiency of sodium reabsorption, lowering blood anionic base concentration. The tubule cells also decrease in the efficiency of ammonium (i.e. net hydrogen) excretion. The result is the overall lowering of blood pH. Each step along the GTPase signaling cascade represents a target for cholera treatment for those with access to healthcare (8). Possibilities include the inhibition of the cellular receptor for the cholera toxin, deactivation of the CFTR protein, or most importantly, oral rehydration solutions. Without proper medical treatment or electrolyte replacement, the mortality rate is approximately 50% (5).
Modern science has the tools to prevent and treat cholera and other waterborne diseases, but for approximately 2.5 billion individuals worldwide, economics prevents those technologies from ever reaching them (1). Even if international aid for water supply and sanitation increases fifty-fold, universal access to clean water will not be achieved until after 2025 (22, 23). Until that day, makeshift technologies will be necessary in reducing waterborne disease (3). Simple boiling is often effective, but fuel is too expensive for many households. With enough finances, an intermediate water treatment system may address waterborne disease at the scale of local communities, e.g. sedimentation tanks followed by slow-sand filtration (24); however when community-level systems are inaccessible, cost-effective technologies addressing specific, local pathogens such as V. cholerae are useful on a household level (7, 25). A review of four general household prevention techniques is presented below.
The Accessible, Cost-effective Prevention of Cholera
When fuel for boiling drinking water is too expensive, a useful cholera prevention technique is filtration through a household fabric, specifically the Bangladeshi sari cloth (7). When the cloth is folded so that water is filtered through four layers, anything over 20 μm is removed, including particles and plankton to which the cholera pathogen may be attached. In the study by Colwell et al. (2003), the household use of filtration through sari cloth reduced clinical cholera cases by 48% in a study of approximately 133,000 individuals in Bangladesh (7). Filtration through the cloth is an accessible technology for many communities with endemic cholera, and could be a vital part of an effective cholera-prevention program by removing V. cholerae reservoirs from drinking water. The major drawback is that free-swimming bacteria, including V. cholerae and other pathogenic species, are not removed from the water; however, this could be solved by using simple filtration in combination with one of the techniques discussed below.
In areas with high incidence of solar radiation, household batch reactors (i.e. containers constructed from recycled household materials) can utilize the bactericidal effects of both temperature and ultraviolet light (UV) to clean water. Martin-Dominguez et al. (2005) constructed batch reactors with two-liter plastic water bottles over a reflective metal (e.g. aluminum) that deactivated 100% of coliform bacteria after four hours in the sun (25). Bottles painted half-black maintained higher temperatures by 15-25oC and may further deactivate pathogens by pasteurization (25). Mani et al. (2006) showed that conditions of low solar radiation require the use of a fully transparent bottle over a reflective surface (26), which could be particularly important, for example, when using batch reactors during the monsoon season in areas of Southeast Asia with endemic cholera. With sub-optimal solar radiation, the water’s exposure to UV must be optimized because the bactericidal properties of UV are more important than thermal inactivation (26). V. cholerae is vulnerable both to UV and pasteurization (27, 28); however, further research should investigate the exact V. cholerae deactivation capabilities for a given type of batch reactor, before the prototype is presented to a community. Batch reactors are an accessible, cost-effective technology that can be useful in areas without clean drinking water (25), and could be useful in areas with endemic V. cholerae. UV radiation deactivates viruses, bacteria, and protozoa (29, 30); therefore, these batch reactors may be useful in confronting waterborne pathogens simultaneously.
Halogen-releasing agents, specifically those containing chlorine, have long been important in sanitizing water (3), particularly in areas with endemic cholera (31). Chlorine-releasing agents include N-chloro compounds, chlorine dioxide, and sodium hypochlorite, and although they act similarly, these compounds deactivate microbial contaminants by slightly different mechanisms (32). Sodium hypochlorite (NaOCl) is found in household bleach (3-6% NaOCl by mass), and is therefore most useful because bleach is cheap and abundant even in rural communities. In water of pH 4-9, NaOCl ionizes to Na+ and OCl- and is found in equilibrium with hypochlorous acid (HOCl). HOCl is the source of active chlorine. In the presence of a bacterial cell, HOCl will chlorinate nucleotide bases, disrupt oxidative phosphorylation, and prevent growth and proliferation by inhibiting up to 96% of DNA synthesis, and between 10-30% of protein synthesis (32). Some experts have attributed the 1991 cholera epidemic in Peru partially to an anti-chlorination campaign based on NaOCl’s carcinogenic properties (31). Chlorination of drinking water using household bleach is regularly practiced in many rural Peruvian communities, with government signs proclaiming “No hay diarrhea con agua clorada” [there is no diarrhea with chlorinated water], alongside directions to add two drops of bleach per liter of water. Chlorination can produce hazardous disinfection byproducts (DBPs; e.g. trihalomethanes) that are linked to cancers and birth defects (33); however, in many regions, chlorination programs have successfully prevented waterborne diseases and saved lives for many years (3, 32). When used properly (i.e. 2-4 drops of 5.25% NaOCl per liter water), chlorination is more reliable than solar radiation batch reactors because it acts as a residual disinfectant for viruses, bacteria, and protozoa (3). Because V. cholerae can be found in biofilms (17), for which chlorine is only able to deactivate the external layer (16), filtration pre-disinfection is an essential complement to this technique. If drinking water is especially murky, additional chlorine can be safely added until there is a slight chlorine odor, but total chlorine should never exceed 5 mg/liter (34). Chlorination, or another halogen-releasing agent, is vital in the rapid sanitation of water, and is an important element even in advanced water cleaning systems.
Vaccines for cholera have also been used for over a century, but with minimal efficacy in general because the duration of immunogenic response is very short-lived, especially for children under five (8). Further difficulties arise from the diversity of pathogenic strains within the O1 and O139 serogroups (35). Cholera vaccine research currently focuses on the development of a single dose vaccine that could be affordable and rapidly immunogenic for all ages, from infant to adult (35, 36). The most promising candidate is Peru-15, a live oral vaccine of the El Tor Inaba strain in the O1 serogroup. In the 2007 study of Qadri et al. (35), 84% of Bangladeshi children 2-5 years old given the large-dose vaccine developed a vibriocidal antibody, and that immune response occurred within seven days. The vaccine has already been shown to be safe and immunogenic in adults (36), and together, the two findings suggest that the Peru-15 vaccine could be useful in high risk areas before or during a cholera outbreak. Further research will be necessary in determining the duration of this immune-response, and its effectiveness against different strains of V. cholerae (35). Currently, cholera vaccines may not be sufficiently accessible or long-lasting to be primary components of cholera prevention programs. In the future, the advanced understanding of immunological mechanisms may enable the rational synthesis of a universal cholera vaccine that elicits a stronger and longer lasting immune response (37). Vaccinology is a dynamic science that will continue to evolve with the understanding of immunological mechanisms, but as of yet, there are no universal vaccines (i.e. covering ~90% of the target population) for many bacterial pathogens, including V. cholerae.
Conclusion
Every year, V. cholerae infects hundreds of thousands of people with cholera, a rapid and debilitating disease. In areas without formal water sanitation or adequate medical supplies, there are effective, accessible technologies that can prevent the disease. Simple filtration, solar radiation batch reactors, and household bleach can save lives in regions of endemic cholera, while at the same time removing other waterborne pathogens from drinking water. Education programs currently are and will continue to be essential in bringing these technologies to high-risk areas until the development of permanent sanitation infrastructure.
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