Risk Factors Contributing to Microbiological Contamination of Boreholes and Hand Dug Wells Water in the Vina Division, Adamawa, Cameroon

This study examined the sanitary conditions of public boreholes and hand dug wells water in relation to pathogenic bacteria isolated in water samples. To assess the suitability of drinking water, sanitary inspection of surround-ings of the sources was conducted, membrane filtration technic was used to trap the microorganisms in the water sample and other standard microbiological technics were applied to check the heterotrophic plate counts, total coliforms, fecal coliforms and some pathogenic bacteria in the samples. It was observed that there were no delineated protection zones around many boreholes (61.19%) and in almost all hand dug wells; thus, hand dug wells were pruned to more contamination than boreholes. Heterotrophic plate counts of boreholes were significantly different between (p < 0.0001) zones in dry and rainy seasons. Total coliforms per 100 ml varied significantly (p < 0.001) between the two seasons both in boreholes and hand dug wells. Fecal coliforms were significantly absent in some boreholes, but significantly present in all hand dug wells with the presence of E. coli 157H in some. Risk assessment wells, poor depth and lack of internal lining, uncapped wells, presence of trees near the wells, animal and birds feces, were significant and associated to contamination with Salmonela spp., B. cepaceae, S. aureus E. coli 1 (p < 0.05). Generally water collection points are marked with very poor sanitary conditions and this situation may not change unless the contaminating risk factors are fixed and sanitary measures are taken.


Introduction
There has been encouraging progress with access to safe water and sanitation in both rural and urban areas since the United Nation Water Decade of the 1980s. However, more than 1 billion people around the world still lack access to safe water supply and more than 2.4 billion are without adequate sanitation. In Africa, over one third of the population still remains without access to safe water and sanitation and many of these can only be served by groundwater [1]. The factors leading to contamination are often not well understood, but are frequently ascribed to pollution by on-site sanitation facilities, such as presence pit latrines, as these represent an obvious source of fecal contamination. This is known to be a major cause of disease outbreaks and impact on health [2] [3]. It is further indicated that 1.1 billion people drink water that is at least of "moderate" risk (>10 cfu/100ml Escherichia coli). It is estimated that, by 2030, the African continent may attain 54% of urban proportion [4]. This phenomenal growth has been qualified as sudden and wild to express the uncontrolled nature of urban growth and the implications it may have on the well-being of city-dwellers. Sub-Saharan Africa is ranked among the world's regions which are mostly at a disadvantage. In Cameroon, contamination of improved water sources is usually a result of poor operation, maintenance leading to frequent failures and lack of environmental education [5]. Consequently, these call concerns about the safety of improved water sources used in communities especially in rural areas where Municipalities are failing to maintain good water quality performance. Diarrhoeal diseases are often caused by contaminated water, poor sanitation, and poor hygiene. In Cameroon, diarrhoeal diseases are the most prevalent waterborne diseases among children aged less than five years. In Yaoundé, for example, the prevalence of diarrhoea is increasing as shown in studies conducted in the city among children aged less than five years, the prevalence increased from 10.8% in 1998 to 13.1% in 2004 [6]. In Cameroon and Vina Division in particular, there is lack of monitoring and evaluation strategies of drinking water quality. Hence, poor on-site sanitation and other pollutants are common sources of microbial contamination of groundwater and chemical contaminants. Thus, continuous use of untreated groundwater in the area would pose a public health threat to the  [7]. In the Vina Division, sanitation coverage is low; therefore, it is important to identify the sources of groundwater contamination and to assess their individual and combined impact on ground water quality so that interventions to improve water quality are effective.
For these reasons, we assessed the potential sources of contamination of public selected boreholes and hand-dug wells, the suitability of water for drinking from these groundwater sources by analyzing the fecal indicator bacteria and impact on public health.

Study Site
The study was carried in the Vina Division located in the Adamawa Region be-

Study Design
A cross sectional survey of water points was carried out during the months of December and April in the dry season and in the months of July and October during the rainy season. Sample points were selected based on the fact that an equivalent water point is the hydraulic unit designed to supply a population of about 300 people with drinking water per day (http://www.iwm-network.org/).
Equally distance between the nearest water collection points were taken into consideration. As such 21 public boreholes out of the 364 registered boreholes fulfilled this criterion. Also, 15 hand-dug wells were identified to fulfill this criteria based on a snowball sampling strategy of public wells as most of them were not registered by the Cameroon Ministry of water and energy. The geographic position system of each sampling point was also taken using Garmin etrex 10 GPS.

Sanitary Risk Inspection
A sanitary risk survey involving the identification of potential microbial contamination sources was performed for 21 public boreholes and 15 hand-dug wells in the area to assess their risk level to microbial contamination. The sanitary inspection method was adapted from the British Geological Survey as described by Howard et al. (2003) [10], based on a checklist of 12 factors for boreholes and 10 factors for hand dug wells as indicated in Table 1 and Table 2 respectively. The procedure involved physical inspection of water sources followed by inspection of the surrounding environment and scoring of presumed risk factors (Yes = presence or No = absence) and a few questions asked to people using the water point. A final risk score was obtained by summing the score for each water source.

Sample Collection and Microbiological Analysis
Using the membrane filtration technique, each sample was passed through the membrane using a filter funnel and vacuum system. Organisms in samples were then concentrated on the surface of the membrane. Heterotrophic plate count was carried out using a serial dilution and pour plate techniques. M-Endo Les Agar (P.code 101973451, 85766-500G-F) was used for the enumeration of total coliforms, while fecal coliforms were enumerated on m-FC Agar (P.code 101869634, 96961-500G-f). All culture media were prepared following the manufacturer's instructions. E. coli and E. coli 0157.H7 were isolated using Mackonkey sorbitol and confirmed serologically using microgen E. coli 0157 (ref.M44) as rapid confirmatory test for E. coli 0157.

Statistical Analysis
Water collection points were grouped into zones. The risk of contamination score was calculated as the percentage of total positive answers to the questions posed on the checklist. Average concentrations were calculated using Microsoft excels 2018 and Heterotrophic plate counts expressed as logarithmic transformations (log 10 ). Using SPSS, odds ratios were calculated to evaluate relationships between water quality and potential source of contaminations.

Sanitary Inspection Exercise
Results of the assessment of groundwater sources in the area are summarized for key potential contamination risk factors for 21 boreholes and 15 hand dug wells  Table 1 and Table 2 respectively. Seven and eight important risk factors were respectively identified for boreholes and hand dug wells. Results indicated that the quality of water in hand dug wells in the area was more likely to be affected by on-site sanitation facilities than those of the boreholes. The general observation was that there were no delineated protection zones around many boreholes Twelve of the hand dug wells examined (86.6%) did not have proper top-covering protection, in addition to protection like apron walls. Results also showed that in 60% of the wells, latrines and septic tank soak ways were sited on the upstream side of the wells and within distances of between 10 to 30 m. All (100%) of the hand dug wells had no protective fence to block animals and birds from defecating close to the system.
Other important risk factors to contamination observed within the vicinity and in the hand dug wells were complete absence of concrete walls in most of the hand dug wells, or if present were old, with many broken, sticks, old shoes, ropes tied to buckets used for drawing water were normally left at points around the wells that were likely to be source of fecal contamination. Figure 2 illustrates risk contamination of hand dug well and borehole water collection point.
The evaluation of drinking water for presence of bacteria was carried out in both the urban and rural areas of the Vina Division during the dry and rainy seasons. Table 3 and Table 4 indicate respectively the Total Heterotrophic   Bacteria (THB), Total coliform counts and fecal coliform counts from boreholes water and hand dug wells in the dry and rainy seasons. A very significant (p < 0.0001) difference between zones in heterotrophic plate count of bacteria load for boreholes water was observed during the dry season. These values ranged from log 10 42.27 CFU/ml (zone 1) to log 10 46.51 CFU/ml (zone 7), and from log 10 45.14 ± 1.8 CFU/ml (zone 1) to log 10 51.18 ± 0.12 ml (zone 7) during the rainy season. As for hand-dug wells, THB differed very significantly (p < 0.00001) between zones, varying from log 10 47/CFU/ml (zone 1) to log 10 59.72 CFU/(zone 2) in the dry season, while in the rainy season, the highest counts recorded was 61.62 CFU/ml (zone 1), while the lowest was 59.16 CFU/ml (Zone 3). In boreholes water, there was a significant (p < 0.01) difference between the readings of the HTB in the dry season and those of the rainy season. In the hand dug wells, no significance (p = 0.07) difference was noticed in the values obtained between the readings of rainy and dry season.
According to the analysis, the THB was extremely high in zone 3 (46.65

Fecal Coliform Correlation of Hand Dug Wells and Boreholes Water
In the dry season, boreholes in zones 1, 2, 4 and 5 had fecal coliform concentration per 100 ml of more than 100 CFU/100ml. while in the rainy season only zone one had fecal coliform counts out of the range as indicated Table 5. In the hand dug wells both in the dry and rainy season, fecal coliform counts varied very significantly (p < 0.001; p < 0.01), hence all the hand dug wells water were classified at high risk or very high risk to public health (Table 6). Risk classification levels were done based guideline values for bacteriological water quality water control as seen in Table 7.
From this result we can conclude that there is systematic pollution of groundwater in the study area. However, precise recognition of Enteropathogenic E. coli (EPEC), classically associated with outbreaks of infant diarrhea [11] remains  problematic. In this study E. coli (EPEC) was detected in zone one in a borehole water both in the dry and rainy seasons and borehole water in zone 5 were most contaminated with fecal coliforms in the dry season.     Table 8 and Table 9 illustrate Multivariate analysis using odds ratio to assess the association between potential sources of contamination indicated in Table 1 and Table 2

Sanitary Inspection
Epidemiologic studies indicate that sanitary surveys have played an important T. B. Viban et al.
role in determining pollution sources in water bodies. It provides the foundation required to design and implement an effective water quality sampling program and provides valuable information to assist in the interpretation of water quality data. In particular, it provides public health authorities with information to aid the selection of sampling locations, times and frequencies, in order to estimate more accurately water quality, and therefore, to allow for sound risk management decisions [12]. As part of the comprehensive and complementary risk-based assessment of drinking water quality, the purpose of the on-site visit is to identify and evaluate all existing and potential sources of microbiological contamination that could affect the safety of water in the area [13]. A higher score represents a greater risk that drinking water is contaminated by fecal pollution from the area immediately surrounding the well. The principal public health concern is the use of vulnerable groundwater aquifers and surface water without purification or disinfection measures for drinking purposes. Adamawa region is known to have 53.3% of domestic water from boreholes and 42.4% of water from hand dug wells with 364 public boreholes distributed in the seven sub-divisions of the Vina Division.
The results of risk assessment of microbial contamination in this study have shown that the locations of boreholes and hand dug wells are not adequate to prevent harvested water from contamination. And there were high counts of fecal coliform in hand dug wells water. Most of the hand dug wells sampled were within distances of <10 m from potential sources of groundwater contamination. There was therefore a possibility of constant inflow of leachate of effluents from the bottom of pit latrines into the nearby wells and some boreholes water Improper sanitary practices close to water collection points resulted in a dense concentration of water sources with continuous inoculation of bacteria and other microbial pathogens into the aquifers of the area. These pathogens can spread within the groundwater system through adventive transport by flowing groundwater. Abstraction of groundwater for various uses will induce forced convection and facilitate inflow of contaminants from nearby contamination sources. A similar study was carried out by Taonameso, (2019) [14] who assessed the potential health risk to rural borehole water communities in South Africa and concluded that there was a need for environmental education.

Bacteria Load in Water Samples
Biochemical contamination in groundwater might be enhanced by anthropogenic activities such as improper disposal of waste materials. In addition, the application of fertilizers, pesticides, and metal-containing paints play a significant contribution in the perspective of groundwater pollution. Also water sources are threatened by contamination due to increased urbanization, poor wastewater drainage and other environmental pollution caused by the lack of proper garbage collection set up. Heterotrophs are broadly defined as microorganisms that require organic carbon for growth. They include bacteria, yeasts and molds [13]. In this study, there was a variation in heterotrophic plate counts analyzed for boreholes water and hand dug wells in both seasons from all sampled locations. The total heterotrophic plate counts for all water samples were generally high exceeding the limit of 1.0 × 10 2 cfu/mL which is the standard for drinking water [15]. In boreholes and hand dug wells the THB counts did not fall within the standards prescribed by WHO. Similar results have been reported in total heterotrophic plate counts of boreholes water in some states of northern Nigeria [16] [17] [18]. The high values obtained could be due to poor environmental conditions and the presence of stagnant water around the boreholes, which provide an excellent breeding ground for bacteria. The presence of heterotrophic bacteria exceeding the WHO limits indicates that water samples were contaminated with bacteria that could make water unsafe for drinking. This finding agrees with similar studies by Okezi, (2019) [19] who evaluated the THB of ground water in northern states of Nigeria. The higher number of heterotrophic bacteria recorded during the rainy season could probably be due to increased surface area of the volume of water and increased carbon intake which exposes the ground water runoffs. During dry season, water contamination by heterotrophic bacteria could be caused by human activities like, washing, dipping of dirty legs or hands and cans inside the wells while fetching water or increased in population since water usually become scarce in the dry season.
Ground water which is believed to be the purest form of water because of the purification properties of the soil was found to be contaminated, due to improper construction, shallowness, animal wastes, proximity to toilet facilities, sewage, refuse dump sites, seepage or discharge from septic tanks, and various human activities [20].

Fecal Coliform Correlation of Hand Dug Wells and Boreholes Water
Results from groundwater samples from all wells and some boreholes indicated fecal contamination with indicator organism (fecal coliforms). Fecal contamination from cows and chickens is an issue for many water sources. Many hand dug wells are near houses which are cheaply provided to satisfy the immediate need season. The emergence of communicable disease outbreaks related to water in many rural areas remains a major challenge, even though there is a vast improvement of infrastructure dedicated to accessing safe drinking water to these communities [21].
The presence of a typical Enteropathogenic E. coli (EPEC) water used in the Vina Division is a major concern because of its ability to cause acute infections. Low-economic communities are known to use water of poor quality and this is recognized internationally as an issue of critical concern to public health. The operation and management of these water supplies may be inadequate due to the limited resources and lack of awareness of factors affecting water quality [22].
Based on the WHO, (2004) [23] requirement the presence of E. coli cells in 100 mL of drinking water, is considered as not suitable for human consumption, unless treated. Similar studies were carried out at Dodowa in Ghana and identified high level contamination of boreholes and hand dug wells water with high levels of fecal coliforms [24]. Other studies were carried out by Mushi, (2012) [18] in Tanzania correlated risk factor contamination to E. coli detected in hand dug wells and boreholes water samples. Similar study indicated high level contamination of boreholes and hand dug wells water at Yola Nigeria [25]. To this effect we attributed high level of fecal coliforms counts in the dry season in our area of study to increased anthropogenic activities with relatively lower groundwater tables and conditions that enable accumulation of fecal material in water. It is reported that groundwater resources are at less risk of being contaminated by animal feces when livestock densities are low or when livestock are spread out over a wide area of land [26]. Boreholes and hand dug wells in the Vina-Division were observed to have poor drainage systems and stagnant water accumulated around the water collection points so that, underground contamination through seepage was eminent. The penetration of surface water carrying animal wastes or sewage to groundwater abstraction wells could lead to gastrointestinal illnesses as faecal material could carry various pathogenic microbes. Enterohaemorrhagic E. coli (EPEC) or Shiga toxin-producing E. coli found in humans, cattle and goats has been found in water and causing of bloody diarrhea has previously reported [27]. Each year, (EPEC) reportedly causes more than 200 million cases of diarrhoea and 380,000 deaths monthly in children in developing countries [22].
The results of this study indicate that even though boreholes are viewed as improved drinking water sources and are expected to provide safe drinking water, some of the boreholes are of very poor microbiological quality. As reported by Onda et al. (2012) [28], pathogenic bacteria and other substances from excreta in pit latrines can leach through the sub-surface soils and contaminate groundwater as seen in zone 1, both in the dry and rainy seasons.
Other observations indicated that Environmental Impact Assessment (E.I.A)  [29]. In Uganda where in risk factors contributing to Microbiological Contamination of shallow groundwater did not show any strong association with fecal coliforms. These data emphasized the vulnerability of the shallow and groundwater in the Vina division to contamination and a rapidly emerging public health risk following environmental contamination of ground water. In relation to this, antibiotic resistance profile of bacteria isolated from hand dug wells and boreholes water in Vina division is very high [30] indicating a public health concern. Some of the areas in which we sampled have been known for the endemic prevalence of typhoid fever and sporadic presence of cholera epidemics. Studies have shown that Immunity resulting from repeated exposure from pathogen sources, such as drinking water, has been considered to be an important factor in the control of outbreak of many some diseases, in this area [31]. It is possible that the residents of very poor sanitary and low level of education municipalities are frequently exposed to pathogens causing typhoid fever, increasing the breeding of mosquitoes and the development of other water parasites increasing their immunity A study conducted in Conakry, Guinea, showed that there was widespread contamination of shallow dug wells and suggested that microbiological contamination was associated with poor maintenance of the wells [32].

Conclusion
Field work and laboratory experiments were conducted to identify potential sources and contamination of selected boreholes and hand-dug wells water in the Vina Division. The microbial water quality was tested through heterotrophic wells and boreholes are heavily loaded with fecal matter. This situation will not change unless the contaminating risk factors as we found in Table 1 and Table 2 are fixed and sanitary measures are taken. Hand-dug wells in the area seem at even greater risk to contamination compared to boreholes as indicated by multivariate analysis. Based on the WHO drinking water standards, groundwater from many sampling points was not suitable for drinking and needs to be treated before consumption. So far, a relatively limited data is available in literature for Sub Saharan Africa on potential sources of ground water contamination and sanitation and there is little or no systematic monitoring of key microbiological and chemical drinking-water-quality parameters.

Limitations
This study had difficulties in sampling repetitions, because some boreholes got damaged between sampling periods, as such could not be repeated. To solve this problem, another borehole close to the sampling area and with same geological settings was used as replacement.

Recommendations
This study shows that the microbiological contamination is high in drinking water that comes from groundwater sources in urban and rural areas. Groundwater remains an important source of water for people of this community, but measures need to be taken urgently to avoid consumption of such untreated water. It is important for the local governing body, the local councils, to come out with regulations for setting of wells and boreholes to ensure adequate distances between the sources and potential point sources of pollution and that local management committees involving women are created at each location point. A law is needed that will enforce the provision of protection zones around facilities used in abstracting groundwater for domestic use in the area. On top of that, it is important to do regular repair of broken wells and boreholes. The community needs to be educated both on activities around the drinking water sources that impair the microbial quality of groundwater and on safe handling of water obtained from these sources.