Hazard assessment and categorization of microbiological risk in a water treatment and distribution system located in a municipality in the interior of Minas Gerais, Brazil

Consumption of non-potable water is a relevant public health problem due to the possibility of transporting numerous chemical and microbiological contaminants. In 2005, the Brazilian Ministry of Health created the National Program of Surveillance in Environmental Health related to the Quality of Water for Human Consumption (VIGIAGUA), with the primary purpose of managing risks related to water supply to human populations in Brazilian territory. However, VIGIAGUA does not have a methodology capable of characterizing or managing risks. The objective of this research is to create a working model to transform raw data into conceptual data related to low-, mediumand high-risk levels. The data used in the application of the model were obtained through the analyses of water-quality surveillance conducted by the Regional Health Management of Itabira, a municipality with less than 10 thousand inhabitants, whose history of water contamination is alarming. Twelve samples/month were collected between May 2017 and April 2018. The results of the treated water analyses were classified according to the presence of hazards and categorized into risk levels. The results showed 83.3% contamination by total coliforms at the treatment plant and 91.6% in the distribution system. The Escherichia coli contamination was 16.7% in the treatment plant samples and 45.2% in the distribution system. The system was categorized as "High Risk". The analysis of untreated water samples was carried out for the purpose of knowing the contamination pattern of the raw water of the region, finding 100% contamination by total coliforms and 97.2% contamination by E. coli.


INTRODUCTION
Water is an essential resource for human life and its availability on land is increasingly scarce. Of the water resources available on the planet, only 3% are fresh water, and of these, only 0.01% can be used for human consumption, after adequate treatment. This little availability of water is constantly threatened by human action, mainly through runoff resulting from agricultural and industrial activities, which can lead to contamination of groundwater by heavy metals. In addition to chemical contamination, the microbiological quality of the water must be suitable for human consumption .
Global potable water guidelines recommend that faecal indicator bacteria, preferably Escherichia coli, should not be detectable in any 100 mL sample aliquot. However, worldwide water quality reports describe faecal contamination in water sources for human consumption, especially in low-income countries, where supply systems are not sufficient to contain the viability of microorganisms in water, favoring the occurrence of diarrheal diseases (Bain et al., 2014).
In the Brazilian scenario, the rate of diarrhea among children under five years has declined in the last decades, from 1,346,506 records in 1998 to 511,893 in 2015, attributed to better vaccine coverage and optimization of water resources management (Oliveira et al., 2017).
The relevance of efficient water resource management and potable water supply in the global burden of disease was recognized by the World Health Organization in 2013 through goal 7c of the Millennium Development Goals. This goal determined that by 2015 the proportion of the world's population without sustainable access to potable water needed to be reduced to 50.0% (Gnetry-Shields and Bartram 2014).
In Brazil, monitoring of the quality of water distributed to the population is attributed to the Unified Health System (SUS), carried out through the National Program for Monitoring Water Quality for Human Consumption -VIGIAGUA (Brasil, 2005), with the decentralization of actions to the State and Municipal Health Secretariats. In the State of Minas Gerais, the Program is executed by the State Secretariat of Health and Regional Health Units (URS) and in the municipalities, by the Municipal Health Secretariats.
The program aims to carry out risk analysis associated with the consumption of 3 Hazard assessment and categorization of microbiological risk … Rev. Ambient. Água vol. 15 n. 3, e2450 -Taubaté 2020 contaminated water, by monitoring the basic parameters related to the presence of total coliforms/E. coli, turbidity index and residual chlorine. However, it does not have methodology to characterize the risks in the work routine of health surveillance. For the reasons presented and with the perspective of contributing to overcome the challenges of VIGIAGUA and improve the Program, this paper proposes a hazard assessment and categorization of microbiological risks associated with a water supply system, located in a mining municipality with less than 10.000 inhabitants, assigned to the Regional Health Management of Itabira (an agency linked to the State Health Department of Minas Gerais, Brazil), using the model suggested by Carmo et al. (2008).

MATERIALS AND METHODS
The methodologies used in this work are described in the American Water Works Association (AWWA) and Water Environment Federation (WEF), American Public Health Association (APHA), the American Water Works Association (AWWA) and the Standard Methods for the Examination of Water and Wastewater, as determined in Article 22 of Ordinance/MS 2.914/2011 (Brasil, 2011).
The Brazilian municipalities must comply with a minimum sampling plan for the evaluation of parameters of turbidity, free residual chlorine, total coliforms and E. coli, considered basic indicators of the microbiological quality of water for human consumption. The municipality object of this study has a population between 5,001 and 10,000 inhabitants, so according to VIGIAGUA guidelines, nine monthly samples were analyzed. Priority areas for sample collection were selected according to the points that historically presented the highest frequency of contamination and in accordance with the guidelines of the Brazilian Ministry of Health, prioritizing public areas with large circulation of people (health units, schools, hospitals, etc.) (Brasil, 2016). Two fixed points were defined at the exit of treatment for each of the two existing water treatment systems. Another 14 variable points were sampled alternately each month. Therefore, in the urban area, 9 samples/month were collected, 2 fixed points (treatment plants) and 7 at variable points, alternating between even and odd months. In addition, three additional collections were carried out in a locality considered to be an isolated urban area, whose water supply does not receive any type of treatment, and is only collected, stored and distributed. For these samples, the parameters of turbidity, total coliforms and E. coli were evaluated. The collections were carried out between the months of May/2017 and April/2018.
The technique of collecting and transporting the samples followed the "Item 5.4.4" of the Ezequiel Dias Foundation's Collection Manual (FUNED), under the register "NUMBER: DIOM-DIVISASGA-MQ 0001. Samples for analysis of total coliforms and E. coli were collected in sterile plastic bags containing sodium thiosulfate tablets for neutralization of chlorine. After collection, the pockets were packed in thermal boxes cooled with recyclable ice, kept at a temperature between 2°C to 8°C and transported to the GRS-Itabira laboratory within 22 hours.
The methodology for the identification of total coliforms and E. coli was based on the chromogenic substrate. The technique uses the hydrolyzable substrates ortho-nitrophenyl-β-Dgalactopyranoside (ONPG) and 4-umbelliferyl-β-D-glucuronide (MUG) for simultaneous detection of total coliforms (β-D-galactosidase) and E. coli (β-glucuronidase), identifying the bacteria through the yellow coloration resulting from ONPG hydrolysis, and fluorescence resulting from the hydrolysis of MUG seen under long wavelength UltraViolet light (365nm), (APHA et al., 2012).
The method used to determine free residual chlorine (CRL) is based on the oxidation of N, N-diethyl-p-phenylenediamine (DPD) by chlorine, resulting in a rosy solution with intensity proportional to the concentration of free chlorine Soares et al. (2016). To measure the concentration of CRL, a Policontrol® digital colorimeter was used, the procedures are described in the manufacturer's manual.
The turbidimetric analyses were performed using the Policontrol® digital Turbidimeter, respecting the procedures described in the manufacturer's manual.
Hazard assessment and categorization of the degree of risk was performed as suggested by Carmo et al. (2008) and described in the conclusion item (Table 1). The collected data were evaluated according to collection indicators (CI), related to the minimum sampling plan for the three basic parameters, bacteriological index (BI) (total coliforms and E. coli), turbidity index (TI) and index of free residual chlorine (IFRC) according to the formulas below: Pearson's chi-square test (pValue 0.05) was used to evaluate statistical significance in the dry and rainy periods in relation to the presence of E. coli and total coliforms in samples collected in the distribution system of the urban area (treated water) and treatment plants.
The Wilcoxon-Mann-Whitney test was applied to evaluate the statistical significance (pValue 0.05) between the values of the median turbidity results in the distribution system of 5 Hazard assessment and categorization of microbiological risk … Rev. Ambient. Água vol. 15 n. 3, e2450 -Taubaté 2020 the urban area (treated water) during the rainy and dry periods. In the treatment plants, the test was applied comparing the whole period of analysis and during the rainy and dry periods.

RESULTS AND DISCUSSION
Twelve monthly samples were collected between May 2017 and April 2018, totaling 144 samples, divided into 108 treated and 36 untreated. Among the 108 (N) treated water samples, 89.8% (n = 97) were positive for total coliforms and 38.9% (n = 42) for E. coli, (Tables 1 and  2). All samples showed non-compliance for the IFRC parameter (values <0.2 mg/Lrecommended value: 0.2 -2.0 mg/L) and turbidity compliance except School 1, Department Public Administration 1 and (5 turbidimetric units -Tu) as a function of the sample standard deviation ( Table 3). The microbiological standard determined by Administrative Rule no. 2,914/2011 is the absence of total coliforms in 100 mL of sample at the exit of the treatment, and in the distribution system (for systems that supply less than 20,000 inhabitants), tolerance of presence of coliforms in a sample/month. For presence of E. coli there is no tolerance in the exit and system or distribution. In addition to contamination reaching all sampled collection points, the most alarming result is the contamination ratio between the treatment outflows and the distribution system. For total coliforms, there was 83.3% contamination of the samples at the treatment exit, for the two treatment stations (TS-1 and TS-2) and 91.6% in the distribution system (Table 1). For E. coli, there was a 16.7% contamination of the samples at the treatment exit, for the two stations (TS-1 and TS-2), and 45.2% in the distribution system (Table 2). The presence of total coliforms and E. coli in the water supply and distribution system persisted throughout the study months, and from July 2017 total coliform contamination was present in all samples analyzed. For E. coli, the highest frequencies of contamination were observed during the rainy season, between November and March (Santos and Garcia, 2016), especially in November/2017, January and February 2018, where percentage of contamination were 55.5%, 100.0% and 88.9%, respectively. The result is in agreement with the fact that there is a consensus that the worsening tendency of water quality in rainy periods is a consequence of the drag of organic matter and various particles, which protect the microorganisms from contact with chemical disinfectants, which favors its viability (Gleason and Fagliano, 2017). The increase in the presence of particulate matter in the rainy season was also observed in the turbidity results, whose average values, considering all collection points in the urban zone, were lower in the driest months of the year (May to October 2017 and April of 2018 -7 months) when compared to the rainy season months (November 2017 to March 2018 -5 months) (Figure 1).  chlorine concentrations below 1 mg/L favor the formation of biofilms (Liu et al., 2015). A similar result was demonstrated in the study by Yousefi et al. (2018), who identified an increase in the viability of total coliforms due to the decrease in the amount of free residual chlorine, in water for human consumption, in villages in the city of Poldasht, Iran. In addition, the particulate material, transported through the distribution system as a result of non-existent or inadequate filtration processes, can deposit in the bottom of the tubes, and the inlaying of particles and materials with nutritive potential also favor the formation of biofilms, facilitating the viability and dissemination of potentially pathogenic agents (Liu et al., 2013).
Although Administrative Rule no. 2,914/2011 recommends that the pH of the water be maintained in the range of 6.0 to 9.5 in the distribution system, the ideal value for disinfection with chlorinated derivatives is 8.0, because at this value there is an availability of hypochlorous acid around 35%. When the pH assumes values of 8.5; 9.0 and 9.5; hypochlorous acid becomes available in the respective percentages of 12%, 5% and 2%, which is insufficient for the disinfection process (Antonio and Macedo, 2004). During this study, three pH measurements were performed at a temperature of 26.9°C. Two in samples from treatment station (TS-1), one before treatment and another post-treatment, and one sample in the distribution system (School 2), whose values were respectively 10.31; 9.10 and 8.95. These results demonstrate that even with the use of chlorinated disinfectants, if there is no adequate chemical correction of pH in the treatment plants, the microbiological protection will not be effective.
The structure and functioning of the treatment plants have direct relevance in the results found. The lack of complete cycle in the treatment (coagulation, flocculation, sedimentation and filtration), in the case of TS-1, which has only a chlorination process in the Parshall gutter, does not guarantee the final quality of the product. TS-2 has the physical structure to perform the complete treatment cycle, but does not use the chemicals necessary for the process, as well as to promote the dilution of the chlorinated water by TS-1, by opening the floodgates of its filters, transferring water without treatment for TS-1 reservoirs. The situation is aggravated by the meeting in the distribution system of the water coming from the two treatment plants, which makes it difficult to identify the origin and exact water flow.
When comparing the percentage of total coliforms and E. coli of all treated water samples collected in the urban area between the rainy and dry periods, these were significantly higher for the two parameters in the rainy season (Figures 2A and 2B). The same statistically significant difference occurred for the values of the turbidity medians between the two periods of the year, both for the samples collected from the distribution system ( Figure 2C) and those collected at the exit of the treatment plants ( Figure 2D). However, for the annual values of the turbidity medians between TS-1 and TS-2, there is no statistical difference between them ( Figure 2E), being considered an undesirable result due to oscillations in the rainy season. As the structural quality of the distribution system is fundamental for the production of drinking water, knowledge about the ecology of the bioindicator (E. coli) should be considered in the planning of actions to protect the source. This fact is important because it was initially believed that this bacterium inhabited exclusively the lower intestinal tract of warm-blooded animals at a concentration (per gram of feces) between 10 7 -10 9 CFU in humans and 10 4 -10 6 CFU in domestic animals. However, recent studies have shown that E. coli can survive for long periods of time in soil, sand, sediment and water, in tropical, subtropical and temperate climates, which facilitates its dissemination in water supply systems and explains the increased contamination during rainy periods (Jang et al., 2017). In addition, Frick et al. (2018) evaluated the abundance of E. coli in several groups of animals (homeothermic and poikilothermic), in an alluvial backwater in Austria, whose results showed the average concentrations of this bacterium in poikilothermic excreta, close to those found in homeothermic plants, confirming that its occurrence it is not exclusive to warm-blooded animals.
The results show the inability of the municipal supply system to contain the presence of bacteria and other potentially pathogenic agents, such as viruses and protozoa, which constitute a microbiological hazard for the transmission of these microorganisms (Nabeela et al., 2014).  Therefore, the high level of microbiological contamination present in the water of human consumption of this municipality can be directly associated with the absence of conventional treatment, inadequate structural flow of the treatment system, also considering the lack of use of chemical products like flocculants and pH correctors , besides the physical conditions of the pipes in the distribution, which do not have routine maintenance/prevention of corrosion or periodic replacement of old pipes, conditioning the formation of biofilms.

CONCLUSION
The results demonstrated the inability of the water treatment system analyzed to neutralize the presence of bacteria and other potentially pathogenic agents, like viruses and protozoa, which constitutes a microbiological hazard for their transmission. Therefore, the development of the methodology presented in this study is necessary to categorize the levels of risk associated with the hazard and to advise on the identification of flaws in the water treatment and distribution systems of this municipality, in order to promote corrective actions in water supply (Chart 1). The methodology used can be applied in the work routine of public health surveillance in other municipalities, as an instrument for monitoring water contamination for human consumption and allowing quick and adequate responses in the planning of governmental actions. Chart 1. Categorization of the risk associated to water consumption in the studied municipality, evaluated according to the presence or absence of hazards.

45.2% Presence
Evaluation of water quality: turbidity 24 samples analyzed. 6 samples above 1Tu = 25% 2 samples above 5Tu Evaluation of water quality: free residual chlorine Does not attend Does not attend Aspects of a general nature -Statistical treatment Statistically significant difference for the presence of total coliforms in the distribution system in treated water area during rainy/dry periods

Unwanted outcome
Statistically significant difference for the presence of E. coli in the distribution system in treated water area in the rainy/dry periods

Unwanted outcome
Statistically significant difference for turbidity in the distribution system in the area with treated water in the rainy/dry periods

Unwanted outcome
Statistically significant difference for turbidity between TS-1 and TS-2 in rainy/dry periods Unwanted outcome Statistically significant difference for turbidity between TS-1 and TS-2 (May/ 17 to Apr/18) Unwanted outcome

High risk
Subtitle:

Low risk
Attendance to all basic parameters of potability in the treatment exit and distribution system

Medium Risk
Attendance to all the basic parameters of potability at the treatment exit, associated to the no attendance of the parameter for total coliforms in the distribution system.

High risk
Non-compliance with the defined parameter for E. coli.