7.1 Introduction
7.2 Health significance of human viruses in drinking-water
7.3 Occurrence of human viruses in source waters
7.4 Risk management
7.4.1 International approaches
7.4.2 Virus removal by current water treatment processes
7.5 Sampling, testing and data interpretation
7.6 C.t values
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REFERENCES
Figures and Tables
Table 7.1: UV dose requirements for virus inactivation credit
Table 7.2: C.t values for inactivation of viruses by free chlorine, pH 6 – 9
Table 7.3: C.t values for inactivation of viruses by chloramine
Table 7.4: C.t values for inactivation of viruses by chlorine dioxide, pH 6 – 9
Table 7.5: C.t values for inactivation of viruses by ozone
7.1 INTRODUCTION
No maximum acceptable values (MAVs) have been set for human viruses in the Drinking-water Standards for New Zealand (2005). It is likely that a MAV or MAVs will be established in a future edition. This chapter foreshadows such developments.
In the absence of any MAVs for viruses in the current DWSNZ it should be understood that if they are specifically sought, they should not be detected. If detected, advice should be sought from the relevant health authorities.
There are more than 140 different types of human enteric viruses that may contaminate potable source waters. These include several important groups: Hepatitis A virus, Hepatitis E virus, norovirus, enterovirus and adenovirus, that have been associated with waterborne illness and are capable of causing severe, and in some cases fatal, infections.
Human viruses are obligate intracellular parasites, which means they cannot grow or multiply outside their host. Viruses consist of a nucleic acid genome surrounded by a protein capsid and, in some cases, a lipoprotein envelope. These viruses are very small, ranging from 20 – 70 nm in diameter.
Human enteric viruses are shed in the gut, respiratory tract and occasionally urine of an infected person, and are discharged with body wastes into wastewater. Infected people do not always show signs of illness (asymptomatic) but they will still produce virus in their wastes. Specific viruses or strains of viruses are not always present in a community at any one time, but representatives of the large groups (e.g. adenovirus or enterovirus) are generally present on most occasions.
Enteric viruses may be found in high numbers in domestic wastewater. Recent New Zealand studies have shown adenovirus and enteroviruses to be present in concentrations greater 10,000 infectious virus units per litre of wastewater (Watercare Services Surrogate Study 2002). The numbers of viruses in wastewater varies with the level of virus infection in the community but, in general, human viruses will always be present in wastewater, even of small communities, (average human virus load 100 – 1000 infectious viruses/L) and will occasionally reach very high levels (>10,000 infectious virus/L) (Lewis et al 1986). Wastewater treatment processes that do not include a disinfection step are often inefficient in removing viruses (<90% removal) and some viruses may reach potable water supplies.
Human enteric viruses cannot multiply in the environment once outside the host. The viruses are characterised by the ability to survive for days, weeks or more, in the environment depending on the type of water, season and other factors (Hunter 1997).
A large proportion of the human viruses present in source drinking-waters will normally be removed or inactivated by well-operated standard drinking-water treatment processes.
Routine monitoring for viruses in treated water and source water is currently impractical in most situations in New Zealand because of the high cost of sampling and analysis, and problems of detection of a full range of the viruses occurring.
7.2 Health significance of human viruses in drinking-water
Hepatitis A virus, Hepatitis E virus, norovirus, enterovirus and adenovirus may occur in drinking-water where they are present in the source water and when water treatment does not remove them completely. Very few human enteric viruses (1 – 50 virus particles depending on type) are required to produce an infection in a susceptible person (Hunter 1997). The symptoms generally attributed to enteric viruses are gastroenteritis and diarrhoea but they can also cause respiratory, central nervous system, liver, muscular and heart infections. Some waterborne viruses have also been associated with some forms of diabetes, chronic fatigue syndrome and dementia (Nwachcuku and Gerba 2004). The major groups of viruses contaminating water are discussed below but may not represent all the viruses likely to be transmitted by water. It is reasonable to expect that further important groups of waterborne viruses will be detected in the future and that these will most likely cause atypical waterborne disease (Nwachcuku and Gerba 2004).
Norovirus: this group of caliciviruses includes the Norwalk and Norwalk-like viruses. Members of this group are strongly associated with waterborne outbreaks in many parts of the world. Symptoms of infection are mild and self-limiting and include vomiting, diarrhoea and nausea over 24 – 48 hours. Norovirus is quite prevalent in New Zealand and is responsible for a large proportion of viral gastroenteritis reported to health authorities (ESR 2004). This virus is one of the easiest to link to a common source outbreak as the symptoms occur rapidly after contact with the virus (approximately 24 hours).
Hepatitis A and E: Hepatitis A and E have a relatively low occurrence in New Zealand (ESR 2004) but induce quite significant symptoms including fever, malaise anorexia and jaundice. The disease is generally self-limiting but has a 2 percent mortality rate. The infectious doses for these viruses are relatively low (10 – 100 viruses) and symptoms do not occur until 10 – 50 days after infection. Internationally Hepatitis A and E outbreaks have frequently been associated with water.
Enteroviruses and adenoviruses: these two different groups represent the viruses that are most commonly found in contaminated surface water. These viruses produce a very broad range of symptoms including respiratory, skin and eye, nervous system, liver, heart and muscular involvement. Gastroenteritis with vomiting and diarrhoea is a less common outcome of infection with these viruses and is limited to only a few adenovirus and enterovirus types. Reported waterborne outbreaks of these viruses, other than in swimming pools, are very infrequent. It is not clear whether lack of reporting is because the dominant symptoms produced by these viruses are not those traditionally associated with water or food borne disease, or because such outbreaks are indeed rare (Hunter 1997).
Virus infections resulting from treated water have not been reported in New Zealand (ESR 2004) in recent years although internationally such outbreaks are recognised (Hunter 1997). Human viruses have been reported to occur at very low levels (0.1 – 1/100 L) in conventionally treated drinking-water in many countries (Vivier et al 2004) including New Zealand (Kim 2005).
Estimations of viral disease risk using standard risk assessment techniques with a high infectivity virus predict the surprisingly high annual risk of infection of between 1:3 and 1:25 from conventionally treated drinking-water contaminated by viruses at low levels (~1 virus per 100 litres) (Gerba and Rose 1992).
7.3 Occurrence of Human viruses in Source waters
The New Zealand freshwater microbiology study (McBride et al 2002) is the most significant study of human viruses occurrence in surface water in New Zealand to date. This study carried out in collaboration between the Ministries for the Environment, Agriculture and Forestry, and Health tested recreational water locations on 25 rivers and lakes every 2 weeks for 15 months.
Human adenovirus and/or enterovirus were detected, by qualitative molecular methods, in more than 50% of the 275 samples collected. This data suggests that human virus occurs quite frequently in surface waters and in a wide range of source water locations and types.
Subsequent culture based studies of virus occurrence in the Waikato River show that adenovirus and enterovirus levels are generally low, less than 5 per 100 L, but on some occasions may be as high as 10 per 100 L (Watercare Waikato River Monitoring studies 2003 - 2004).
Studies using sensitive qualitative molecular-based virus detection methods suggest that adenovirus occurrence may be 10 times higher than this level on some occasions in the Waikato River (Kim et al 2005) although it is not clear whether all of these viruses are able to produce infections.
International data collated by WHO suggest that typical surface source waters may contain 0 - 10 viruses per litre (WHO 2004).
7.4 Risk management
Potential for disease outbreaks associated with human virus contamination of source waters, and the potential for carry over to treated drinking-water is recognised throughout the developed world. Approaches to controlling the risks are largely through protection of source water quality by control of human activity in reservoir catchments, and through adequate treatment and disinfection of drinking-water. It is now well accepted that bacterial indicators such as E. coli are not adequate surrogates of viral occurrence. Human viruses tend to be more resistant to environmental stresses and water treatment mechanisms than are bacterial indicators, so the absence of the indicator may not equate with absence of the virus contaminant.
7.4.1 International approaches
The paucity of knowledge on the specific occurrence of human viruses in source waters, and the problems of virus detection and regular monitoring, mean that most guideline documents include only the qualitative requirement that, if tested for, human viruses should not be detected in treated drinking-water.
Where virus guidelines or standard requirements are in place these are stated either in terms of virus occurrence, or as water treatment plant virus removal efficiency. Such values are either derived from acceptable levels of health risk or, pragmatically, reflect virus detection capability.
Recent standard and guideline recommendations have moved towards risk-based evaluation of water treatment requirements. The USEPA Surface Water Treatment Rule includes a virus treatment requirement and requires that treatment of both filtered and unfiltered water sources is sufficient to remove or inactivate 99.99% (4 log) of viruses (USEPA 1994). This requirement is principally based on the acceptable (USEPA 1994) level of waterborne illness in a community (1 case per 10,000 consumers) and the likely level of viruses in surface water. Recent US proposals for surface water disinfection (USEPA 2003a) use the adenovirus group as the target virus.
The WHO Guidelines recognise that water treatment requirements will differ for different communities, and propose a risk-based approach for setting performance targets for surface water treatment plants (WHO 2004).
The risk-based approach takes into account a broad range of factors including virus occurrence and infectivity, water type, community health status and treatment characteristics. Such an approach requires a detailed knowledge of the water supply, water treatment performance and community activities and health status.
Approaches to managing viruses in treated water also recognise that the greatest health risk to a community occurs when water treatment conditions are atypical such as when source water condition is unusual, very high levels of virus occur, or through poor performance, or even failure, within the water treatment process.
7.4.2 Virus removal by current Water treatment processes
Reduction of virus numbers in water as a result of treatment can occur through either virus removal or virus inactivation. Each virus type may react somewhat differently to particular water treatment methods, but the bulk of research to-date suggests that some broad generalisations can be made.
Virus removal can occur by physical association of a virus with other particles. Particle flocs containing viruses are then removed by settlement or filtration. Virus association with particles and flocs may be enhanced by addition of coagulants and certain salts. The extremely small size of viruses means that they are unlikely to be removed if they are not associated with other particles. Water treatment processes such as flocculation, sand filtration, microfiltration and ultrafiltration, and prolonged standing in reservoirs, will result in physical removal of particle-associated viruses. Only reverse osmosis and dialysis membranes have pore sizes small enough to trap virus particles that are not associated with larger particles or flocs.
The effectiveness of virus removal is affected by those factors that act against particle association or floc formation including water condition and pH (LeChevelier and Au 2004).
Virus inactivation occurs through disruption of the external protein coat (capsid), modification of specific surface sites needed for infection (host receptor recognition sites) or major change to the nucleic acid (RNA or DNA). Disinfectants such as chlorine, chlorine dioxide, and ozone will cause disruption of the virus coat and of the exposed nucleic acids (Shin and Sobsey 2003, Tree et al 2003). Ultraviolet light in the range of 200 – 310 nm (antimicrobial range) will disrupt the nucleic acids by causing cross-linking that leaves them unable to replicate.
Viruses can also be inactivated by prolonged holding in reservoirs exposed to sunlight, elevated temperature and extremes of pH (e.g. lime treatment) (Sobsey 1989). Different virus types and strains will show different levels of resistance to chemical or physical inactivation. Adenoviruses are considered to be the most resistant virus group to many disinfection treatments, because of its structure and nucleic acid makeup, and have been used by the USEPA as a model virus for designing UV criteria for surface water treatment (USEPA 2003a).
The potential for virus inactivation by disinfectants is reduced by the presence of other particles or organic matter that will consume disinfectants or of light adsorbing or blocking materials that reduce UV penetration (LeChevelier and Au 2004).
Repair of disinfection damage is unlikely to occur in viruses as they do not have repair mechanisms as such. It has been suggested that some viruses (e.g. adenovirus) may be able to repair their DNA if there is no damage to the virus coat and they are able to infect a human cell (Nwachcuku and Gerba 2004).
Water treatment plants will normally include both virus removal and virus inactivation processes that act as multiple barriers.
Virus removal and inactivation efficiencies for a range of water treatment processes are reviewed in the WHO (2004) Guidelines (chapter 7), and by LeChevallier and Au (2004).
7.5 Sampling, testing and data Interpretation
The determination of virus removal efficiency within a water treatment plant, or occurrence in treated water, is dependant on the ability to reliably detect and enumerate the viruses. Determination of the health risk that viruses pose to the community using the water further depends on the ability to demonstrate or infer that the viruses detected are capable of causing human infection.
Virus detection and enumeration. No single method allows detection of all virus types and strains. Traditionally viruses have been concentrated from water samples using filtration or adsorption based techniques with subsequent detection by culture in a permissive human or primate cell line. Many of the virus concentration techniques were developed using poliovirus or other enterovirus types and it is not clear how effectively these work for other virus types particularly norovirus.
Virus concentration from large volumes of water is laborious and time consuming and adds significantly to the cost of virus analysis. Not all virus types are culturable in cell lines, again norovirus has not been cultured routinely and is not detectable using traditional methods.
Viruses (culturable and non-culturable) can be detected at very low levels using polymerase chain reaction (PCR) based molecular methods that target novel DNA or RNA sequences in the genetic information of the virus. Virus assay using PCR can target individual viruses or groups of viruses and multiple analyses would be required to investigate all the relevant viruses in a particular sample (Greening et al 2002). Recent advances in real-time PCR have made these methods both rapid and quantitative and potentially quite routine. PCR based methods are around 10-fold less sensitive that culture based methods for virus detection (Lewis et al 2000). Quantitative PCR based molecular methods are also significantly less expensive than traditional culture methods. It is unusual for a virus concentration and detection method to consistently recover more than 50 - 60% of the virus present in a sample (Lee and Jeong 2004).
Virus sampling strategies. Relatively few viruses are needed for an infection to occur in a susceptible person so low numbers of viruses must be quantified in relatively large volumes of finished water. For example, if source waters contain 5000 viruses per 100 L it would be necessary to sample and analyse at very least 200 L of finished water to demonstrate a 4 log reduction in viruses. Typically, source water sample volumes should be 10 – 100 L, partially treated waters 50 – 200 L, and finished, disinfected water sample volumes 100 – 200 L.
The current cost of virus analysis may make regular monitoring beyond the means of many groups responsible for drinking-water treatment.
Specific short-term studies of virus occurrence and inactivation/removal within a plant are feasible but should be designed carefully to allow adequate interpretation of the data.
Determination of virus infectivity. Molecular methods for virus detection do not specifically show whether viruses are still infectious. Detection of viruses using a cell-culture based technique shows that the viruses are infectious and pose a risk of illness to water consumers. Infectivity of a virus can however be inferred for certain RNA viruses (norovirus, enteroviruses, Hepatitis A and E) from molecular detection data where the viruses are subjected to chemical disinfection, but not UV disinfection (Greening et al 2002). Virus viability is inferred whenever virus nucleic acid is detected because the nucleic acids (single stranded RNA) are extremely susceptible to degradation in the environmental.
Interpretation of virus detection and occurrence data. Where viruses are detected in finished drinking-water the response to the data should be based, in consultation with relevant health authorities, on a risk evaluation incorporating the type and number of virus detected, the reproducibility of the result, and the health status and vulnerability of the community.
7.6 C.t VALUES
Appendix C of the LT1ESWTR Disinfection Profiling and Benchmarking Technical Guidance Manual (USEPA 2003b) includes C.t tables for disinfection of viruses by various disinfectants. These tables are referenced to AWWA (1991) but in the text of USEPA (1991) it would appear that it was a USEPA publication. Because the 2003 publication still uses the 1991 tables it is assumed that the data the 1991 tables were based on is still the latest information!
The USEPA Surface Water Treatment Rule required (inter alia) that treatment of both filtered and unfiltered sources remove or inactivate 4 log (99.99%) of viruses. This requirement was enacted in 1989. Presumably the 1991 tables were developed to assist water suppliers assess the degree of disinfection of viruses.
USEPA’s LT2ESWTR (2003a) includes a table showing the C.t values for disinfecting viruses using UV light. The proposed UV doses for inactivation of viruses were based on the dose-response of adenovirus because, among viruses that have been studied, it appears to be the most UV resistant and is a widespread waterborne pathogen. Health effects of adenovirus are described in Embrey (1999).
It is doubtful that this same approach was used in developing the 1991 tables; viruses are simply referred to collectively, and viruses are not defined in the 1991 information provided. Some viruses require a much higher C.t than others. Nor is it explained whether the data relate to studies in single virions or cell-associated virions – the latter require a higher C.t.
Table 7.1 shows the UV doses that water suppliers must apply to receive credit for up to 4 log inactivation of viruses. This is Table IV – 21 in USEPA (2003a).
Table 7.1: UV dose requirements for virus inactivation credit
| Log Credit | Virus1 UV Dose (mJ/cm2) |
| 0.5 | 39 |
| 1.0 (90% removal) | 58 |
| 1.5 | 79 |
| 2.0 (99% removal) | 100 |
| 2.5 | 121 |
| 3.0 (99.9% removal) | 143 |
| 3.5 | 163 |
| 4.0 (99.99% removal) | 186 |
1 based on adenovirus studies
Tables 7.2, 7.3, 7.4, 7.5 have been taken from Appendix C of USEPA (2003b) and copied from the 1991 publication, i.e. they refer to undefined viruses.
Based on Table 7.2, a free available chlorine content of 0.20 mg/L after 30 minutes retention time is equivalent to a C.t of 6. This would achieve 4 log inactivations at 10°C. At 5°C the minimum retention time should be 40 minutes, or if that cannot be achieved, the residual free chlorine content should be increased to 0.30 mg/L.
Table 7.2: C.t values for inactivation of viruses by free chlorine, pH 6 – 9
| Log inactivation | 1°C | 5°C | 10°C | 15°C | 20°C | 25°C |
| 2 | 5.8 | 4.0 | 3.0 | 2.0 | 1.0 | 1.0 |
| 3 | 8.7 | 6.0 | 4.0 | 3.0 | 2.0 | 1.0 |
| 4 | 11.6 | 8.0 | 6.0 | 4.0 | 3.0 | 2.0 |
Table 7.3: C.t values for inactivation of viruses by chloramine
| Log inactivation | 1°C | 5°C | 10°C | 15°C | 20°C | 25°C |
| 2 | 1243 | 857 | 643 | 428 | 321 | 214 |
| 3 | 2063 | 1423 | 1067 | 712 | 534 | 356 |
| 4 | 2883 | 1988 | 1491 | 994 | 746 | 497 |
Table 7.4: C.t values for inactivation of viruses by chlorine dioxide, pH 6 – 9
| Log inactivation | 1°C | 5°C | 10°C | 15°C | 20°C | 25°C |
| 2 | 8.4 | 5.6 | 4.2 | 2.8 | 2.1 | 1.4 |
| 3 | 25.6 | 17.1 | 12.8 | 8.6 | 6.4 | 4.3 |
| 4 | 50.1 | 33.4 | 25.1 | 16.7 | 12.5 | 8.4 |
Table 7.5: C.t values for inactivation of viruses by ozone
| Log inactivation | 1°C | 5°C | 10°C | 15°C | 20°C | 25°C |
| 2 | 0.9 | 0.6 | 0.5 | 0.3 | 0.25 | 0.15 |
| 3 | 1.4 | 0.9 | 0.8 | 0.5 | 0.40 | 0.25 |
| 4 | 1.8 | 1.2 | 1.0 | 0.6 | 0.50 | 0.30 |
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