Evaluation of the bacteriological quality of underground well water in Oyi local government area of Anambra State, Nigeria

INTRODUCTION

Water is a natural chemical substance which consists of the elements; hydrogen and oxygen in the ratio of two is to one (2:1). It is indispensable for man’s existence on earth as about two-thirds of the human body consists of water and requires between one to seven (1-7) liters of water per day for its appropriate functioning to avoid dehydration.^1,2 In urban and predominantly rural communities in Nigeria, with over 85% of the population living below an average income, traditional drinking water sources such as open reservoirs, springs and open wells are still being used. Water from such sources seldom complies with World Health Organization (WHO) limits for drinking water.³ The quality of water depends on its physical, chemical, and biological characteristics, which determines its utility for different purposes.⁴ Water quality is affected by fecal matter, domestic and industrial sewage and agricultural and pasture runoff, in addition to a lack of awareness and education among the users.⁵

Water fit for human consumption is referred to as potable or drinking water and should be of safe quality, which entails that it does not present any significant health risk over life time consumption; and the only way to tell whether the water is potable is to have it tested.^6,7 Potable water is one that is free from unsafe bacteria and chemical impurities. It must be clear, bright, colourless, and odourless and should contain no suspended matter. Availability of potable public water supply is of great concern to families and communities especially in developing countries where provision of safe drinking water is not available.⁸ Lack of safe drinking water is a threat to public health and well-being of the people and exposes them to risk of waterborne diseases such as diarrheal and dysentery as well as chemical intoxication.^9,10 Therefore, drinking water contaminated from any source is of primary importance due to the danger and risk of waterborne diseases.¹¹

Well water is one of the budding sources of water for human consumption especially in developing countries. It is regarded as a reliable source of water supply because it is often unpolluted, as a result of restricted movement of pollutants in the soil profile in contrast to shallow and permeable water aquifers.^11,12 However, water from wells that are poorly constructed and maintained remains an important source of infectious diseases caused by pathogenic bacteria, viruses and parasites.

There are three broad types of contaminants present in leachates that can pollute groundwater and subsequently affect public health. These are hazardous chemicals, conventional and non-conventional contaminants.¹³ The leachate from some landfills is highly concentrated that small amount of leachate can pollute large amounts of groundwater rendering it unsuitable for use in domestic activities. Drinking contaminated well water can have serious health effects which include diseases such as hepatitis and dysentery which are caused by contamination from septic tank waste. To maintain good health, however, water should be safe to drink, meet the local standards, and ensure the sustainability of national and internal criteria and guidelines established for water quality standards.¹⁴ Safe location of the borehole or well requires careful consideration of factors such as where the borehole or well is about surface drainage and groundwater flow. Pollution of groundwater stems from different sources and these include insanitary conditions during borehole construction, splashing of runoff into open wells, flooding at borehole site, leachate from buried waste pits or latrine, industrial wastes and sewage into the hole through cracks in the aquifer and annular of the hole.^15,16

The detection of bacterial indicators in drinking water suggests the presence of pathogenic organisms that are sources of waterborne diseases.¹⁷ Indicator microorganisms survive better and longer than pathogens, with uniform and stable properties, and may be easily detected using standard laboratory techniques.¹⁸ The occurrence of waterborne illnesses has both economic and social impacts. Consequently, monitoring the levels of contamination and the prevention of disease outbreaks is important from both economic and public health perspectives. Moreover, the need to assess the microbiological quality of water has become imperative because it has a direct effect on the health of individuals.¹⁹

In response to the public health problems caused by contaminated well waters, this study proposes to investigate the bacteriological qualities of underground well water for domestic uses, in the Oyi local Government Area of Anambra state based on World Health Organization (WHO) guidelines for drinking water standards.

METHODS

Study area

The study was conducted using underground wells utilized by households in selected 13 wards in Oyi Local Government Area of Anambra state. Oyi is a local Government area in Anambra state, Nigeria with geographic coordinates Latitude: 6° 13' 60.00" N Longitude: 6° 56' 59.99" E. The towns that make up the local government are Nkwelle-Ezunaka, Awkuzu, Ogbunike, Umunya and Nteje. It has an estimated population of 239,700 as of the 2022 projected census and 139.7 km² of land area.²⁰

The study was limited to households in the Oyi local government area of Enugu State faces a number of problems regarding both its drinking and domestic water quality and availability. Wells belonging to households who mainly use well water as their main source of drinking and household water, within the study area were recruited for the study. The selected 13 wards used for this study are designated A-M. Informed consent was obtained from the heads of the households whose wells were recruited as samples for the study, in the course of carrying out the research and presentation of the final work.

Study procedure and sample collection

Clustre sampling method was used for the quantitative study. Here, the households in each ward were divided into 3 multiple groups (clusters), based on the presence and usage of wells as the main source of drinking and domestic usage water source. Next using systematic random sampling, 3 households were picked from each sub-group. This shows that a two-stage sampling method was used to draw 9 samples randomly from each of the selected 13 wards in the local government area, which gave a total of 117 well samples used in the study. Samples were collected in the morning, with sterile bottles. The bottles were also covered with lids, ensuring no air bubbles once the samples were drawn from the different wells.

The sampling was done between the hours of 7.30 am and 9.30 am. The water samples were aseptically collected with 200ml screw-capped, clean, heat-sterilized amber-coloured plastic bottles, from wells whose openings were covered with lids; which were removed and replaced during and after drawing (fetching) from it. The sample bottles were also rinsed with the sample water before filling them; a stone of suitable size was attached to the sampling bottle using a piece of string. The bottle was opened and lowered into the well and was completely immersed in the water without touching the sides of the well and without hitting the body or disturbing any sediment. The bottles were filled and then removed by rewinding the string. The bottles, once filled were covered with their respective screw caps with no air bubbles. All the sampled bottles were immediately labelled with complete details, transported in a light-proof insulated box containing ice packs to the laboratory and analyzed within 6 hours of collection. Sampling was as used.²¹

Microbiological analysis

Bacteriological analysis of water samples was done according to the method of American public health standard methods, for examination of water and wastewater.^22,23

Presumptive coliform identification

A 50 ml of the test sample was transferred into a 50 ml bottle containing 50 ml double strength MacConkey broth. 5 ml of the test sample was transferred into the 5 tubes containing 5 ml of double-strength MacConkey broth. 1 ml of the test sample was transferred into the 5 tubes containing 1 ml of single-strength MacConkey broth. The whole tubes containing the test preparations and Durharm’s tubes were incubated for 18-24 hours at 37 ⁰c. At the end of the process, the tubes were observed for both acid and gas production. Those with acid and gas production give the presumptive coliform count.

Eijkman’s test for E-coli (differential coliform test)

A pair of test tubes containing brilliant green lactose bile broth (BGLBB) and a tube containing tryptone water were used. These preparations were made in two sets. Each tube in each set was inoculated with a drop from the tube which produced acid and gas, while the other set was inoculated with a drop from a tube which did not produce acid and gas from the presumptive test above. The first group was incubated at 37⁰c, while the second group was incubated at 44⁰c for 48 hours. The tubes to be incubated at 44⁰c must be pre-warmed at 44⁰c by placing them in a water bath before sub-culturing. Coliforms produce gas from lactose only at 37⁰c, while E. coli produce gas from lactose at 37⁰c and 44⁰c and also produce indole from tryptophan at 44⁰c.

Confirmatory coliform test

A loopful of presumptive positive and negative tests was sub-cultured on a MacConkey agar plate and incubated at 37 0c for 24 hours. The significant bacterial isolates obtained were inoculated on nutrient agar slopes for biochemical identification.

Biochemical identification of isolates

Conventional microbiology tests; Gram stain, indole test, catalase test, oxidase test, citrate utilization test, and triple iron sugar test were carried out on the isolates.

Urease test: The slants of urea agar base medium were prepared and inoculated with the isolates obtained during early investigation, kept at 37ºC and then examined after 24 hours. The ability of the organisms to produce urease, which breaks down the urea incorporated in the medium; thus liberating ammonia gas, which then increases the pH of the medium to alkalinity was observed. The change in pH was detected by the change of the colour of the indicator (phenol red) used from yellow to reddish pink.

Catalase test: The test detects the ability of the organisms to produce the enzyme catalase, which catalyzes the release of Oxygen (O₂) from hydrogen peroxide giving bubbles from the bacterial suspension. The organisms were picked up aseptically with the help of a sterile wire-loop on a slide and then colonies were diluted with distilled water. Further to that, 2-3 drops of 3% hydrogen peroxide were dropped on diluted colonies containing organisms, immediately bubbling took place and the tests were considered to be positive.

Oxidase test: The purpose of this test was to examine the ability of the bacterium to produce the enzyme oxidase, which indicates the bacterium’s ability to exchange electrons with a dye, tetra methyl-p-phenylenediamin di-hydrochloride leading to the reduction of the dye to a deep blue colouration. About 0.1g of tetra methyl-p-phenylenediamine di-hydrochloride (dye) was added to 10 ml distilled water and dissolved. Ascorbic acid (vitamin C) 0.01g was then added to the strips of this reagent, it can also be made by soaking the strips of filter papers in the reagent. For confirmation of this test, a glass rod streak was used to pick a generous amount of the bacterial culture and place it on a moist portion of the filter paper. A positive reaction indicates an intense blue colouration of the paper within 10 seconds.

Indole test: The test organism was inoculated into a broth that contained tryptophan and incubated at 370C for 48 hours. Then 2ml of the broth suspension was transferred to another test tube under aseptic conditions. About 0.5ml of Kovac’s reagent was added to the broth. The mixture was shaken properly to ensure a thorough mixing and then observed for colour reaction. A positive result was indicated by a pink-coloured ring around the interface between the broth suspension and alcohol reagent which rose to the surface.

Sugar fermentation activities of bacterial species

Two grams of different sugars (glucose lactose) were dissolved in 200 distilled water and bottled in a 20 ml bottle. The medium was then sterilized by an autoclave slightly opened. The contents of the 20ml bottle were added in 180ml of the basal medium, mixed well and dispensed in 3ml volumes in bijoux bottles. The medium was sterilized under 15lb pressure for 15 minutes and then stored at room temperature for up to 5-6 consecutive days. A bottle of each medium was inoculated with a streak of solid culture (by a single colony). The inoculated bottles were incubated aerobically at 37ºC for the required period. The change in colour of the indicator from red to yellow indicates acid production organisms. The changes were noted after every 24 hours, for five to six consecutive days.

Data analysis

Water and microbial data analysis were done using statistical software, a statistical package for social sciences (SPSS version 22) developed by IBM Corporation. Means and Standard deviation in the same column with the same superscript were not significantly different at p>0.05. The means were separated using the least significant difference (LSD).

RESULTS

Microbial quality of underground well water

Table 1 shows that Escherichia coli is the highest occurring microbial isolate in the water sample of the wards studied, and most occurrences were noticed in wards A, B, C, E, F and G. Most probable number per 100ml for Proteus spp. ranges from 122.22 MPN/100ml to 233.33 MPN/100ml.

*mean and standard deviation values for Enterobacter spp., Salmonella spp., Bacillus spp., Escherichia coli, Proteus spp., Staphylococcus spp, Shigella spp, Pseudomonas spp., and Streptococcus spp. ^a = p ≤ 0.05, ^ab = p ≥ 0.05

The facultative bacteria identified include Enterobacter spp., Salmonella spp., Bacillus spp., Escherichia coli, Proteus spp., staphylococcus spp, Shigella spp., Pseudomonas sp., and Streptococcus spp. All the bacteria identified were present in high numbers in all the samples and their values were reported as most probable number (MPN). Highest occurrence is E. coli, with most probable number (MPN) of 363.33 per 100 ml of sample was reported in ward F samples. Also lowest occurrence is Shigella spp, with most probable number (MPN) of 120.00 per 100 ml of sample reported in ward D samples.

In A, B, and C-wards, all the well water samples had a significant count of microbial content, which does not agree with the value of 0MPN/100ml specified by WHO. In ward A household 5 had the highest count of E. coli at 440 (MPN/100ml) while household 3 had the lowest count at 250 (MPN/100ml); in ward B household 2 had the highest count of Salmonella spp at 340 (MPN/100ml) while household 1 had the lowest count at 200 (MPN/100ml) and also, in ward C household 4 had the highest count of Enterobacter spp. at 270 (MPN/100ml) while household 3 had the lowest count at 170 (MPN/100ml) (Table 2).

In D, E, and F-wards, all the well water samples had a significant count of microbial content, which does not agree with the value of 0MPN/100ml specified by WHO. In ward D households 8 and 9 had the lowest counts of Bacillus. spp at 100 (MPN/100ml) while household 4 had the highest count at 170 (MPN/100ml). In ward E household 2 had the highest count of Streptococcus spp at 280 (MPN/100ml) while households 3 and 7 had the lowest count at 200 (MPN/100ml). Also, in ward F household 5 had the highest count of Proteus spp at 250 (MPN/100ml) while household 4 had the lowest count at 140 (MPN/100ml) (Table 3).

In G, H, and I-wards, all the well water samples had a significant count of microbial content, which does not agree with the value of 0MPN/100ml specified by WHO. In ward G, household 3 had the highest count of Shigella spp at 250 (MPN/100ml) while household 2 had the lowest count at 180 (MPN/100ml). In ward H, household 6 had the highest count of Staphylococcus spp at 300 (MPN/100ml) while households 3 and 8 had the lowest count at 220 (MPN/100ml). And also, in ward I, household 3 had the highest count of Pseudomonas spp at 240 (MPN/100ml) while household 2 had the lowest count at 150 (MPN/100ml) (Table 4).

In G, H, and I-wards, all the well water samples had a significant count of microbial content, which does not agree with the value of 0MPN/100ml specified by WHO. In ward J, household 2 had the highest count of Shigella spp at 230 (MPN/100ml) while households 5 and 6 had the lowest count at 140 (MPN/100ml). In ward k, household 3 had the highest count of Staphylococcus spp at 310 (MPN/100ml) while household 4 had the lowest count at 240 (MPN/100ml). And also, in ward L, household 4 had the highest count of Proteus spp at 160(MPN/100ml) while household 7 had the lowest count at 100(MPN/100ml) (Table 5).

The well water samples examined in M-ward had a significant count of microbial content, which is not within WHO’s acceptable limit of 0MPN/100ml. Household 2 had the highest count of Staphylococcus spp at 280 (MPN/100ml) while households 3 and 6 had the lowest count at 210 (MPN/100ml) (Table 6).

DISCUSSION

Generally, water users in the study area have traditionally relied on hand–dug wells for decades. Local indigenes and dwellers as well, depend mostly on this water source for daily water needs which include cleaning, cooking, bathing, agriculture and occasionally drinking. During the field survey, observations showed that there are many hand dug wells in the study area and almost all were properly caped. Wells with such properly caped conditions are not vulnerable to pollution from pollutants such as insects, small animals, refuse, sediments, and other forms of contaminants that can be prevented from gaining entry into the well water from the surface. Also, the method of abstraction of water from these hand-hand-dugdug wells could introduce pollutants into the wells. Field survey revealed that water for domestic purposes from all sampled hand-dug wells in the area, were drawn using bucket and rope of different types and different sources. These buckets and ropes are most often dirty, and kept in open spaces where contaminants access them; thus, users of these wells are exposed to high levels of contaminants and pollutants which pose threats to their health and livelihoods.

Faecal coliform bacteria are a collection of relatively harmless microorganisms that live in large numbers in the intestines of warm and cold-blooded animals. They aid in the digestion of food. A specific subgroup of this collection is the faecal coliform bacteria, the most common member being Escherichia coli. These organisms may be repeated from the total coliform group by their ability to grow at elevated temperatures and are associated only with the fecal material of warm-blooded animals. The presence of fecal coliform bacteria in aquatic environments indicate that the water has been contaminated with the fecal material of man or other animals. At the time this occurred, the source water may have been contaminated by pathogens or disease-producing bacteria or viruses which can also exist in fecal material. Some water-borne pathogenic diseases include typhoid fever, viral, and bacterial gastroenteritis and hepatitis A. The presence of faecal contamination is an indicator that a potential health risk exists for individuals exposed to this water source. Quite several bacteria species were isolated from well water samples located within the study area. The presence of a group of bacteria known as coliforms in water samples serves as an indicator of pollution.²⁴ Chief among them is Escherichia coli, which was isolated from the samples used in this study and whose presence indicates the possible presence of other intestinal pathogens. Streptococcus spp, Staphylococcus aureus, Shigella spp, Enterococcus spp, Salmonella spp, Pseudomonas spp, Proteus spp and Bacillus spp which were also isolated and identified from the samples are other pathogens of importance that have been linked to gastrointestinal disorders, according to Nwidu et al study.²⁵

One of the occurring microorganism in the samples was Bacillus spp. These are gram-positive aerobic or facultative anaerobes and catalase positive microorganisms.²⁶ They are heat-resistant spore-forming microorganisms that are most often found in soil. Due to their ability to form heat-resistant spores, they can survive and compete with other organisms while secreting metabolites that are antagonistic to other microorganisms in the form of antibodies.²⁷ Although they have been useful in the production of antibodies, their presence in drinking water and even food pose critical health risks that may lead to diseases such as anthrax. Staphylococcus aureus is another gram-positive microorganism that is often associated with nosocomial infections. It is a facultative anaerobe, coagulase and catalase-positive microorganism. Although they are commensals in the mucosa of mammals, reptiles and birds, they can also be opportunistic pathogens. One risk of infection by S. aureus is the issue of its resistance to beta-lactamase antibiotics, including penicillin and methicillin a derivative of penicillin. The presence of these species in water, thus calls for public health concern. Streptococcus spp is a gram-positive catalase-positive microorganism and has been associated with illnesses such as pneumonia and usually upper respiratory tract infections.

Escherichia coli, Proteus spp, Pseudomonas spp, Salmonella spp, Enterobacter spp. and Shigella spp are all gram-negative microorganisms and belong to the group referred to as Enterobacteriaceae. It is worthy of note that Escherichia coli is the highest-occurring microorganism among the wards studied. The gram-negative organisms listed above can cause illnesses such as watery and bloody diarrhoea, dysentery, and urinary tract infection and when introduced into the bloodstream, they can lead to bacteremia. In addition, some strains of Escherichia coli can produce enterotoxins in the small intestine which can also cause diarrhoea if not managed in good time.

The most probable number (MPN) per 100 ml obtained for the well water samples exceeded the standard limit set by WHO. This suggests that the well water samples have been contaminated by potentially dangerous microorganisms and are therefore not fit for drinking purposes. This was confirmed by the characterization of the isolates from the well water samples from the locations under study, which were highly contaminated with more than one bacteria species including Salmonella typhi, Escherichia coli, Enterobacter species and Proteus mirabilis are pathogenic organisms mainly of fecal origin. Any water source used for drinking or cleaning purposes should not contain any organism of faecal origin.²⁸ Presence of enteric coliforms especially Escherichia coli makes the water samples unsuitable for human consumption according to the guidelines set by WHO for the evaluation of the bacteriological quality of drinking water.²⁹ Apart from environmental hygiene and population density, the presence of Salmonella species among the samples in wells in the study area may also be attributed to drainage and flooding from contaminated surface water, which seeps into underground water sources. Findings from this study highlight the non-conformity of well water samples studied with the WHO standard recommendation for safe potable water.³⁰ A situation where enteric pathogens are grossly isolated from sources of water consumed by humans or other animals, is a serious problem which calls for vigilance on the part of the authorities; as it signals possible future outbreaks of waterborne diseases.³¹ The reason for the gross contamination of well water in the study area could be due to poor sanitary conditions around the areas where such wells are located, or drawing water from the wells with contaminated containers, a practice that has been common among the users since individuals bring along their water containers (fetchers) in some cases.

However, as a limitation, this study did not investigate potential sources of contamination such as the sanitary features of the well samples in relation to well water quality in the study area.

CONCLUSION

The bacteriological properties of all the samples from the 13 wards of the study area indicate a significant count of microorganisms, especially E. coli. This indicates recent faecal contamination in well water in the study area which signifies the presence of pathogenic microorganisms in the well water. It is therefore evident that there is a high incidence of contamination of well waters by pathogenic organisms in most well waters from the Study area. Underground well water sources to be utilized for domestic purposes such as cooking should therefore be treated or disinfected before use, either by boiling and filtration or by chemical sterilization or a combination of both. In addition, responsible bodies should pay attention to the scarcity of potable drinking water in most Local Government Areas and should work towards the provision of alternative safe water sources.

ACKNOWLEDGEMENTS

We would like to thank the technologists at the Institute of Public Health, University of Nigeria, Enugu Campus, Enugu, Nigeria.

Funding: No funding sources

Conflict of interest: None declared

Ethical approval: The study was approved by the Institutional Ethics Committee

REFERENCES

1. 1. Okonkwo TJN, Okorie O, Okonkwo CJO. Public health risk status of the water supply frame work at Kwame Nkrumah (Postgraduate) Hall, University of Nigeria, Nsukka and Environ. Afr J Environ Sci Technol. 2011;5(7):522-9.
2. 2. Mbaenyi-Nwaoha IE, Egbuche NI. Microbiological evaluation of sachet water and street vendored yoghurt and “Zobo” drinks sold in Nsukka Metropolis. Int J Biol Chem Sci. 2012;6(4):1703-17.
3. 3. Peter-Ikechukwu A, Omeire GC, Okafor DC, Eluchie C, Odimegwu NE, Nze SN, et al. Assessment of the quality of borehole water sample in federal housing estate and sites and services areas of Owerri, Imo State, Nigeria. Food Sci Qual Manag. 2015;42(1):5-12.
4. 4. Adhikary PP, Chadrasekharan H, Chadkraborty D, Kanibe K. Assessment of groundwater pollution in west Delhi, India using geostatistical approach. Environmental Monitoring Assessment. 2010;167(1-4):599-613.
5. 5. Nogueira G, Nakamura CV, Tognim MCB, Abreu Filho BA, Dias Filho BP. Microbiological quality of drinking water of urban and rural communities, Brazil. Revista de Saude Publica. 2003;37(2):232-6.
6. 6. World Health Organization. Guidelines for Drinking-Water Quality. 1st Addendum to 3rd Edition Vol 1 Recommendations, 2006. Available at: https://www.who.int/publications-detail-redirect/9241546743. Accessed on 26 November 2023.
7. 7. Ngwa NR, Chrysanthus N. Bacteriological analysis of well water sources in the Bambui student residential area. J Water Res Protect. 2013;5(11):1013-7.
8. 8. Okonko IO, Ogunjobi AA, Adejoye AD, Ogunnusi TA, Olasogba MC. Comparative studies and microbial risk assessment of different water samples used for processing frozen seafood in Ijoraolopa, Lagos State Nigeria. Afr J Biotech. 2008;7(16):2902-7.
9. 9. Hughes JM, Koplan JP. Saving lives through global safe water. J Emerg Infect Dis. 2005;11(10):1636-7.
10. 10. Agbabiaka TO, Sule IO. Bacteriological assessment of selected borehole water samples in ilorin metropolis. Int J Appl Biol Res. 2011;2(2):31-7.
11. 11. Gambo JB, James Y, Yakubu MB. Physicochemical and bacteriological analysis of well water at crescent road Poly Quarters, Kaduna. Int J Eng Sci. 2015;4(11):11-7.
12. 12. Adejuwon JO, Mbuk CJ. Quality of underground water. J Geol Min Res. 2011;3(5):161-8.
13. 13. Adeyemi O, Oloyede OB, Oladiji, AT. Physico-Chemical and Microbial Characteristics of Leachate Contaminated Groundwater. A J Biochem. 2007;2(5):343-8.
14. 14. Ajagu KC, Ajiwe VIE. Physicochemical assessment of water from selected wells in enugu south local government area of Enugu State, Nigeria. Int J Pure Appl Biosci. 2017;5(1):1-10.
15. 15. Rajesh B, Rattan L. Essential of Medical Microbiology: Microbiology and Immunology. 4^th ed. India: Brothers Medical Publishers; 2004:516.
16. 16. Onwughara IN, Nnorom IC, Kanno OC. Issues of roadside disposal habit of municipal solid waste, environmental impacts and implementation of sound management practices in developing Country, Nigeria. Int J Environm Sci Develop. 2010;1(5):409-17.
17. 17. Yassin MM, Abu Amr SS, Al-Najar HM. Assessment of microbiological water quality and its relation to human health in Gaza Governorate, Gaza Strip. Public Health. 2006;120(12):1177-87.
18. 18. Chatterjee SN, Das D, Roy M, Banerjee S, Dey P, Bhattacharya T. Bacteriological examination of drinking water in Burdwan India Concerning Coliforms. Afr J Biotechnol. 2007;6(22):2601-2.
19. 19. Anyanwu CU, Okoli EN. Evaluation of the bacteriological and physicochemical quality of water supplies in Nsukka, Southeast, Nigeria. Afr J Biotechnol. 2012;11(48):10868-73.
20. 20. John AO, Anarah SE, Sani M. GIS-based assessment of smallholder farmers’ perception of climate change impacts and their adaptation strategies for maize production in Anambra State, Nigeria. In the book: Corn-Production and Human Health in Changing Climate; 2018:25.
21. 21. Chukwuma, OU, Uchenna, OM, Ogonna, US, Chinedu, OS. The effects of effluents’ discharge from some paint industries on soil’s physicochemical properties and bioattenuation of polluted soil. Ind Domest Waste Manag. 2022;2(2):46-60.
22. 22. APHA (American Public Health Association). Standard Methods for the Examination of Water and Wastewater. 18th Edition, American Public Health Association (APHA), American Water Works Association (AWWA) and Water Pollution Control Federation (WPCF), Washington DC, 1992. Available at: https://www.scirp.org/(S(351jmbntvnsjt1aadkposzje))/reference/ReferencesPapers.aspx?ReferenceID=1818549. Accessed on 26 November 2023.
23. 23. Agu KC, Orji MU, Onuorah SC, Egurefa SO, Anaukwu CG, Okafor UC, et al. Influence of solid waste dumps leachate on bacteriological and heavy metals contamination of groundwater in Awka. Am J Life Sci Res. 2014;2(4):450-7.
24. 24. Lukubye B, Andama M. Physico-chemical quality of selected drinking water sources in Mbarara municipality, Uganda. J Water Res Protect. 2017;9:707-22.
25. 25. Nwidu LL, Obeh B, Okoriye T, Vaikosen NA. Assessment of water quality and prevalence of water-borne diseases in Amassoma, Niger Delta, Nigeria. Afr J Biotechnol. 2008;7(17):2993-7.
26. 26. Waites MJ, Morgan NL, Rockey JS, Higton G. Industrial microbiology: an introduction. London: Blackwell Publisher, School of Applied Science, South Bank University, London, UK; 2008:302.
27. 27. Kuta FA. Antifungal effects of calotropis procera stem bank extract against Trichoplyton gypseun and Epiderinoplyton Flocosum. Afr J Biotechnol. 2008;7(13):2116-8.
28. 28. World Health Organization . Guidelines for drinking - water quality [ electronic resource ] : incorporating 1 st and 2 nd addenda , Vol.1, recommendations, 3rd ed., 2008. Available at: https://iris.who.int/handle/10665/204411. Accessed June 4, 2019.
29. 29. Lechevalier MW, Cawthon CD, and Lee RG. Factors promoting survival of bacteria in chlorinated water supplies. Appl Environm Microbiol. 1987;88:649-554.
30. 30. Okafor UC, Ezeanochie PE, Obubu M. Microbial assessment of some selected fish ponds in Awka, Anambra State: comparative study and modelling. Agri Biolog Sci J. 2020;6(2):91-9.