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A Large-Scale Study of Microbial and Physico-Chemical Quality of Selected Groundwatersand Surface Waters in the North West Province, South Africa
Expanded Title:EXECUTIVE SUMMARY BACKGROUND AND RATIONALE Water from the North West Province (NWP) catchment areas supports prosperous gold and platinum mining, manufacturing industries, agricultural sector as well as a growing urban and rural population. This land-locked province thus contributes generously to the Gross Domestic Product of South Africa. However, water allocation for the province has almost reached the quota available, based on surface water estimates. Furthermore, there are reports that the source water within the catchment may be exposed to pollution from various sources but particularly from economic activities. These reports have demonstrated that several surface waters and groundwaters are faecally contaminated and some with opportunistic pathogenic bacteria. Faecal pollution of surface waters has been blamed on service delivery problems in several peri-urban areas and small towns. There are also reports of poorly treated and in some cases untreated effluent being discharged into rivers and other receiving waters. This practice has health, economic as well as environmental implications. The growth in the agricultural and mining sectors has resulted in increased water requirements as well as pollution. These developments have also played a role in human population distribution dynamics. Such economic activities were responsible for the development of major growth points in the province. Mining activities are mainly centred in the eastern part of the province and the agricultural development across the province. Mining operations were (in some cases still are) responsible for decant of heavy metal-containing water of low pH (acid mine drainage: AMD) into surface water. This has devastating ecological and economics effects. The real impacts of the AMD on groundwater in the North West Province are undetermined. Water pollution from agricultural runoff from livestock farming, including poultry farming and feedlots as well as several other practices in the province is also undetermined. Rainfall in the eastern and central parts of the province is higher than the western part. The latter borders on and is part of the Kalahari Desert. Further social and economic developments as well as climate change will impact on the anticipated water availability, requirements and quality of water. Baseline data in these categories will be important for long-term planning. Water requirements are national priorities and processes such as National Water Resource Strategy are in place. However, detailed large water quality studies have not been conducted. The present study was an effort to address this gap. It extended the WRC-funded work done by Bezuidenhout (2011). AIM 1. To determine the water quality of surface and groundwater in the North West Province from chemico-physical and microbiological perspectives. 2. To determine the profile of selected gastroenteritis-associated pathogens in the contaminated water sources. 3. To investigate the usefulness of molecular fingerprinting data in microbial water quality determination. 4. To determine the cytotoxic effects of associated with consumption of untreated water as well as such effects exerted by some of the indicator mycological and bacterial species isolated from the water sources. 5. To investigate potential risk of consuming such water without any prior treatment. 6. To provide an overview of previous large-scale water quality studies conducted nationally and internationally. 7. To generate a report on whether human health risks could be associated with current trends of water provision in the North West Province and if necessary suggest improvement strategies. METHODOLOGY Water samples were collected at specific time periods from selected groundwater and surface water sources in the North West Province using standard procedures. An effort was made to include dry winter as well as wet summer sampling periods for each of the sites. The groundwater sampling sites were grouped into sampling areas so groundwater locations in this report refer to an area, not specific boreholes. Standard methodologies were used to collect and analyse the water samples. Global positioning data (GPS waypoints) were recorded and used to generate the sampling maps. Certain physical and chemical (temperature, pH, electrical conductivity) parameters were measured on site using multi-probe equipment. Chemical parameters such as chemical oxygen demand, nitrates, phosphates etc. were measured in the laboratory using Hach kits and equipment. The levels of faecal indicators and other microbial organisms were determined using standard or previously-published methods. Standard biochemical and DNA sequencing methods were used for identification of microbes. The presence of E. coli and pathogenic E. coli were determined in water using direct DNA isolation and PCR. Faecal sterol levels were determined by the Szűcs method which involves liquid-liquid extraction and resolution by Gas Chromatography-Mass Spectrometry. Levels of bacteriophages were determined by standard methods and enterovirus by qPCR based methodology. A sequential exploratory mixed-methods design was used for the social water management study. A mixed-methods approach includes the use of both qualitative and quantitative research methods to collect, analyse and interpret the research data. Interviews, focus group discussions and observations (n = 25 participants) were used as the main data gathering methods during the qualitative phase. This was followed by the development of a research questionnaire and a community survey (n = 1 000 participants) during the quantitative phase of the study. RESULTS AND DISCUSSION Microbiological and physico-chemical quality of surface water sources Selected river systems covering the upper middle and lower Vaal river management areas were targeted. Average physico-chemical and microbial levels measured at some sites during the 2010 and 2011 seasons were elevated. Some exceeded the Target Water Quality Range (TWQR) for full and intermediate recreational contact, livestock watering and irrigation. Elevated electrical conductivity when compared to TWQR for drinking water was not at levels likely to cause health impacts. However, this elevated EC could be indicative of increasing salinisation occurring. This may have long-term effects on agriculture activities but also on drinking water production. Electrical conductivity should be flagged as a parameter that is critical to monitor. Enterococcus spp. as well as Escherichia coli were isolated from various surface water sources. Isolation of enterococci such as E. faecium and E. faecalis from surface waters illustrates that any full contact activity in the water is risky and could result in infectious disease. Bacteriophage data supported the general findings of the bacterial analysis, namely that many of the water sources in the North West Province have some form of faecal pollution. The following yeast genera were also isolated and identified in surface waters: Candida, Clavispora, Cryptococcus, Cystofilobasidium Hanseniaspora, Meyerozyma, Pichia, Rhodotorula, Saccharomyces, Sporidiobolis, and Wickerhamomyces. Amongst these are several opportunistic pathogenic species that could cause invasive infections when individuals come into contact with contaminated water. Baseline culture-independent molecular profiling (Polymerase Chain Reaction Denaturing Gradient Gel Electrophoresis (PCR-DGGE) and Next Generation Sequencing (NGS)) data of the dominant bacterial communities in the Vaal River are also provided. Although DGGE profiling has some limitations it allowed for the identification of the bacterial community composition and dynamics in the planktonic component in this river system. Next generation sequencing or high through-put sequencing (HTS) analysis presented a better resolution of the bacterial diversity and dynamics in the Vaal River. This technology was not suitable for detecting faecal indicator bacteria in surface water samples that contained known high levels of E. coli and enterococci. Results presented here demonstrated that some of the wastewater treatment plants (WWTPs) were decanting huge concentrations of enterovirus particles into surface water sources. This was probably due to the operational challenges at the plants. Some other plants in the province were operating in such a manner that up to 99.99% of enteroviruses were removed. Impacts of municipal WWTPs as well as other forms of faecal pollution on virus diversity and dynamics in water sources should be further investigated. Such a study should focus on the percentage of viable viruses that are reaching the water bodies and the epidemiology thereof. In such a virology study the presence of other relevant viruses should also be targeted. Furthermore, detection of bacteriophages in the surface waters that are associated with faecal pollution supports the finding that some of the targeted surface water sources in the North West Province are faecally polluted. Microbiological and physico-chemical quality of groundwater In the North West Province more than 80% of the rural community depend on groundwater for all their water needs. The approach was to mainly target boreholes that provided water for domestic use. This included urban, peri-urban and rural areas. A total of 114 boreholes were sampled. Physico-chemical analyses demonstrated that the pH and EC levels were at acceptable ranges for domestic use. However, it was found that only 28% of the boreholes tested complied with the South African TWQR for nitrate (<6 mg/ℓ), and 43% of the boreholes had nitrate levels greater than 20 mg/ℓ. This study also demonstrated that groundwater from the North West Province is vulnerable to nitrate contamination. In 2009 49% of the 76 boreholes tested were positive for faecal coliforms and 67% for faecal streptococci. Forty seven percent of these boreholes were also positive for presumptive P. aeruginosa and 7% for S. aureus. 33% of the boreholes had heterotrophic plate bacteria exceeding 1000 cfu/ml. In 2010, 38 boreholes were sampled. Of these 55% were positive for faecal coliforms and 63% for faecal streptococci. During this sampling period 55% of the boreholes were positive for P. aeruginosa. Detection of faecal indicators was higher in the warm wet seasons than the cold dry season. This was observed in both the number of positive results as well as levels detected. Members of the Enterobacteriaceae family identified included E. coli and K. pneumonia. Based on MLGA results, 34% of the boreholes sampled in 2010 were positive for E. coli. However, using multiplex PCR on DNA directly isolated from water samples it was found that E. coli was present in 47% of boreholes. This indicates that the membrane filtration approach underestimated the presence of E. coli in borehole water in the NWP. Using FC/FS (faecal coliform/faecal streptococci) ratio determination, it was estimated that at least 14% of the boreholes tested had FC/FS ratio >2, indicating potential human faecal contamination. Next generation sequencing (or high through-put sequencing) results provided data on the microbial diversity in borehole water of the NWP. This method is rapid and provided information about the bacterial community structure in these water samples. It was demonstrated that the winter and summer bacterial compositions were different. In the summer samples there were Gamma and Beta proteobacteria as well as Actinobacteria present. Enterobacter sp., Proteus sp., Stenotrophomonas maltophilia are species that were always detected among the Gamma proteobacteria. Sequences from E. coli as well as Pseudomonas sp. and S. aureus were also detected in the summer samples. These results are consistent with the results from the culture dependent method. Next generation sequencing (or HTS) thus holds promise as a method for monitoring bacterial indicator species in groundwater. This, however, needs to be further tested. The results presented here indicated that microbiological quality of groundwater in the North West Province may be of concern as more than 75% of boreholes were positive for faecal pollution. This indicates that groundwater should be tested and treated before being supplied to communities. It was, however, demonstrated that 23% of the boreholes tested negative for both faecal coliforms and faecal streptococci. This result indicates that there are boreholes where no faecal pollution has occurred. This could be due to protection of the borehole and thus the aquifer. Management practices should be put in place to prevent pollution of aquifers. Faecal sterol determination in water quality applications In this study the Szűcs method, an established method for detecting faecal sterols in water, was used to detect six target sterols (coprostanol, cholesterol, dehydrocholesterol, stigmasterol, β-sitosterol, and stigmastanol) in water samples from the NWP. Wastewater treatment plant influent and effluent samples were collected and analysed for human faecal sterol biomarkers. Environmental water samples were spiked with faeces from cattle, chickens, horses, pigs, and sheep. All the samples were subjected to liquid-liquid extraction, silylation and derivatisation. Derivatised samples were analysed by GC-MS. The method was evaluated for quantitation and differences between the water samples from each species. Standard curve assays were linear up to 160 ng/ℓ and the limit for quantification was 20 ng/ℓ. Coprostanol was the human faecal sterol biomarker, while herbivore profiles were dominated by terrestrial sterol biomarkers (stigmasterol and stigmastanol). Differences in sterol fingerprints and concentrations between various animals and humans provide the opportunity to determine the origin of faecal pollution. Water samples collected from boreholes and rivers were analysed for faecal sterols, physico-chemical properties and bacteriological quality. Coprostanol and cholesterol were detected in one groundwater sample. These results were in some cases supported by bacteriological data and in other cases not. Sterols may become bound to soil particles. It is therefore not expected that sterols will be detected in groundwater. Their detection in groundwater in this study should therefore be further explored. Surface water samples were also analysed for faecal sterols using the Szűcs method. Cholesterol was detected in some samples and was the only faecal sterol detected. This indicates that the faecal pollution at these sites were likely to be from animals, not humans. Faecal sterol analysis is a powerful method and has potential to distinguish between faecal pollution from various animal as well as human sources. Potential human health effects The main human health concern is the high level of nitrates in some of the groundwater sources, some exceeding the 20 mg/ℓ level that could cause methaemaglobinemia. This was not the case with surface water sources in associated areas. Signs of salinisation were observed for some of the surface water samples. However, at the moment this cannot be linked to human health concerns. Faecal indicator and opportunistic bacteria were regularly detected in surface and groundwater sources of the NWP. It was shown that pathogenic E. coli may also be present among the environmental E. coli population. Various known opportunistic pathogenic enterococci were regularly detected in surface waters. Pseudomonas spp. were also regularly detected in groundwaters. Among the yeasts isolated several opportunistic pathogenic species were also regularly detected. These studies were conducted over two separate sampling periods, about one year apart. The results suggest that these bacteria (and in the case of surface waters also yeast) could be ubiquitous. This is cause for concern and regular monitoring of such sites is proposed. It could indicate that pathogenic microorganisms such as viruses (enterovirus, adenovirus and hepatitis A & B) and bacteria (Vibrio cholerae, Shigella spp. Yersina spp. and Enterocolitica spp.) may also be present in these water sources. This study has also detected bacteriophages associated with faecal pollution. The phages are surrogates for human viruses. What the study has further demonstrated is that entoviruses are being discharged into receiving surface waters. This study did not determine whether the viruses were viable. However, if one considers that faecal bacteria detected in the water sources were all viable there is a chance that the viruses may also be viable. The detection of virus genetic material in a water source is thus a cause for concern. All these findings demonstrate that people in the North West Province that directly use untreated water for household purposes or recreation may be exposed to several pathogenic or opportunistic pathogens. Such exposures imply that the health of these individuals may be compromised. Amongst the enterococci isolates from surface waters, up to 20% displayed β-haemolysis, a character that demonstrates the production of an enzyme that is associated with virulence. Furthermore, dominant multiple antibiotic resistance patterns were observed for faecal streptococci isolates at most sites in both years. Between 40 and 55% of the isolates were resistant to selected β-lactam antibiotics including Penicillin G. Not all the isolates that were resistant to Penicillin G were resistant to Ampicillin or Amoxillin. A large proportion was also resistant to Vancomycin. Up to 68% was resistant to Neomycin and 56% to Ciprofloxacin. Most of the isolates (up to 99%) were susceptible to Streptomycin. All β-haemolysis positive isolates (potential pathogenic) were resistant to 3 and more antibiotics from various antibiotic groups. Similar results were observed for both study periods Several of the faecal coliform isolates from surface water were resistant to multiple antibiotics, especially β-lactam antibiotics. Table 1 is a summary of the percentage of faecal coliforms that were resistant to antibiotics. Table 1: Antibiotic resistance profiles (%) among faecal coliforms isolated in 2009 and 2010. 2009 2010 Amoxyllin 54 21 Ampicillin 41 15 Cephalothin 40 42 Oxytetracyclin 30 22 Trimetroprim 11 8 Fewer isolates in 2010 were resistant to β-lactam antibiotics compared to those from 2009. However, resistance to Cephalothin, a cephem β-lactam, was similar for both sampling periods. These results are of concern, particularly if one considers the potential of the isolates to cause invasive infections in sensitive individuals. In such cases antibiotics may not be effective. The cause of the resistance is unknown. A future investigation into the likely causes may be of value. A colorimetric method which uses duodenum cells was developed to determine cytoxicity of polluted water. This method has the potential to provide direct answers on the cytotoxic potential of untreated water within 24 hours. It makes use of a cytotoxic index that was developed in this study in which the percentage cytotoxicity is determined and then compared to a negative control. In the present preliminary study the percentage cytotoxicity of tap water and water from a protected borehole was very low compared to untreated river and dam water, untreated and treated sewage. This method is still under development. The refined method will be tested in follow-up studies. A social and water management study Five different themes emerged during the qualitative phase of the study of the interactions of communities with water, namely: perceptions about water, with water quality, availability and management as three sub-themes; beliefs or attitudes towards water, with ‘water should be free’ and spiritual and cultural connections as two sub-themes; sources of water, where most participants produced a list of available sources; uses of water, with physical (physiological) needs, everyday household use, recreational purposes and religious/spiritual and cultural purposes as four sub-themes; and water management. The quantified and verified results indicate that there are no meaningful differences between the two districts (Dr Keneth Kaunda and Bophirima) regarding their perceptions about the quality of their water or its availability. The majority of the participants perceive the quality of their water to be average (72%), while 25% thought that their water is good, 1% that their water is very good/ excellent and only 3% that it is of poor quality. In addition, the majority of the participants in each of the two districts perceives their water to be a limited resource (Dr Keneth Kaunda district = 85%; and Bophirima = 85%) but differed in their opinion regarding how their water is managed. In Bophirima, almost half of the participants (47%) said that they think their water is not managed correctly, while the minority of participants in the Dr Kenneth Kaunda district (28%) thought their water is managed incorrectly. Ninety percent of participants felt that they should not have to pay for water. They also felt that water must be used sparsely and be conserved. With regard to their beliefs and attitudes towards water, almost half of the participants said that they have a spiritual connection with water (45%) and that they use water for example to cleanse themselves or others after a funeral ceremony (75%) or to make contact with their ancestors (45%). The study also confirmed that the majority of households make use of municipal water (77%). Sources of water did not differ greatly between the two districts, except in regard to the harvesting of rain water. In the Bophirima district, communities make more use of rain water harvesting (32%), compared to the Dr Kenneth Kaunda district where only 11% harvest water. Other sources of water such as a borehole with a windmill (21%), or a borehole with an electrical pump (10%), seasonal pans (6%), fountains (5%), dams and rivers (4%) and wells (4%) are used to a lesser extent. A number of uses differed statistically and practically between the two districts. In the Bophirima district, traditional uses such as using water to drive out evil spirits (cleansing themselves or members of their family or house) differed from the Dr Keneth Kaunda district (Table 2). It demonstrates that more individuals in the Bophirima district uses their water for the purposes listed compared to the Dr Kenneth Kaunda district participants. Table 2: Recreational and traditional uses of water (%). Bophirima Dr Kenneth Kaunda Drive out spirits 75 50 Make traditional medicine 81 57 Recreational purposes 75 55 Harvest food 43 20 Livestock watering 70 28 Other popular uses of water that scored above 88% include the use of water for house building or other physical structures (for example in combination with soil) (94.2%), to cook their food (99%), to drink (98%), and to flush their toilet (96%). More than 90% indicated that water helps them when they are fasting (92%). Ninety seven percent indicated that water is used during religious ceremonies (e.g. washing their own or other’s feet before church) Almost all the participants indicated that they use water to wash their goods (cleaning of physical objects other than themselves) (99.3%). More than 19% indicated that they use water for gardening (domestic plants) (93.2). Eighty eight percent indicated that they use water to make traditional beer (umqhombothi). The use of water for self-cleansing (enema) seemed less popular (68%). Seventy six percent indicated that they use water to initiate the traditional healer/s in their community. More than 60% indicated that they use water to wash themselves or others after a funeral service. Amongst all the participants 28% were farmers using water on a large scale to produce crops. The majority of households still appear to make use of a tap in their yard. Fewer had access to piped water in their houses. A section of the communities was still using a communal tap for which they have to travel less than 50 meters or to a lesser extent more than 50 meters. Finally, most households appear to store water inside their homes (65%) where it is cooled down in most cases (83%). However, communities in the Bophirima district tend to store water more often in containers outside their house (56%) compared to those participants from the Dr Keneth Kaunda district (34%). CONCLUSIONS The purpose of the research was to conduct a large-scale study of microbial and physico-chemical quality of selected surface waters and groundwaters in the North West Province, South Africa. The sampling period was from 2009 to 2011. Results have shown various trends. • Nitrates in groundwater and salts in surface water are the main physico-chemical hazards. • A number of the groundwater and surface water sources in the North West Province are polluted with faecal matter. The faecal pollution was demonstrated by various methods including standard culture methods, direct DNA isolation and PCR, faecal sterols, bacteriophages and in the case of groundwater, also next generation sequencing. • Bacteriophage and enterovirus data indicated that the sources may also contain viruses that could be human pathogens. • Large numbers of faecal coliforms and enterococci isolated in this study were resistant to several antibiotic groups. This is cause for concern as it may eventually have human and animal health as well as plant pathology implications. • A cytoxicity test to determine the impacts of microorganisms on human cell cultures was also developed as part of this study. It may be useful in future studies where water quality and suitability for human consumption is determined. The adaptations proposed in the study make it more rapid and precise but also, at the moment, more costly to conduct. • Baseline data on social aspects of source and drinking water management was also provided for 6 communities from 2 of the 4 districts of the North West Province. The study presented some data on how members from these communities interact with and manage water. It also demonstrated that certain perceptions, beliefs and behaviours are associated with these interactions. The results demonstrated that water has important social and cultural meaning and importantly, that future education programmes could build on knowledge existing with these communities. RECOMMENDATIONS FOR FUTURE RESEARCH • Sources of high EC values should be investigated as this could have serious implications on soil quality in the province and by implication also food security. It may eventually also impact on drinking water purification processes. Long-term studies are thus necessary to generate data that would be useful for predictive modelling studies. Such data would also be useful to provide advice to agricultural and water purification authorities. • Boreholes should be identified that may be vulnerable to certain types of contamination. Data from the present study will be useful in identifying areas for further investigation. Once identified appropriate interventions or treatment options could then be put in place before such water is supplied as safe for human or animal consumption. • Sources of faecal pollution in surface water should also be identified and programmes be put in place to ensure that this type of pollution is prevented. Data from the present study could be used to identify specific areas in the selected rivers that should be further investigated for faecal pollution sources. • Results from this study have demonstrated that a full health risk assessment should be conducted on some of the water sources in the province. This is needed particularly within the context of the large section of the community of the NWP, particularly in rural and peri-urban centres, that are immuno-compromised. At present health effects of consuming the faecally-contaminated water could be under-reported and a detailed study is thus necessary • The cause of multiple antibiotic resistance and the mechanisms involved should be further investigated. This was not established in the present study but should be determined. Broader understanding of antibiotic resistance may provide means to predict the spread of the resistance and in the process provide tools to curb the spread. As more pathogens become resistant to available drugs the search for newer more effective drugs may drain resources. This could result in a vicious circle in the treatment of human, animal and plant infectious diseases. • Future studies to determine the presence of E. coli and other indicators in the water should be conducted also using PCR (qPCR) methods instead of only culture based methods. Quantitative PCR (qPCR) is more accurate and rapid compared to plating methods. Analysis times will be decreased and larger numbers of samples could be analysed. • In-depth HTS (and culture-based) analysis of the microbial diversity and water quality in the surface and groundwater is recommended with the focal point on the identification of specific species within the major key groups. This should be conducted for viruses, bacteria, yeasts, fungi and other eukaryotes that may have health implications. Culture dependent methods are time consuming and a limited number of samples are normally analysed. These methods also require specific media types for specific microorganisms and many important microorganisms may not be detected in samples. Methods based on analysis of genetic material (HTS, qPCR) are more efficient than culture dependent methods. • More knowledge about the social context and management of water at a community level in the North West Province should also be obtained. In the present study data were collected from only 6 communities in 2 of the 4 districts of the province. These communities represent only a small section of the NWP and should thus be extended to include more communities but also communities from the other two districts. Data contained in these reports must also be responsibly translated into information that the affected communities could relate to and use.
Date Published:03/04/2013
Document Type:Research Report
Document Subjects:Water Resource Management/IWRM - Catchment Management
Document Keywords:Hydrology, Water Quality
Document Format:Report
Document File Type:pdf
Research Report Type:Standard
WRC Report No:1966/1/13
ISBN No:978-1-4312-0400-7
Authors:Bezuidenhout CC
Project Leader:Bezuidenhout CC
Organizations:North West University
Document Size:2 915 KB
Attachments:Ground water DATA_CCB.xls
Social Study NWP_water_18Apr_2013_CCB.xls
SW_bac_Yeast DATA_CCB2.xls
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