Universidade Federal de Santa Maria

Ci. e Nat., Santa Maria, v. 41, e55, 2019.

DOI: http://dx.doi.org/10.5902/2179460X37700

Received: 11/04/2019 Accepted: 23/10/2019

Section Environment

Physical-chemical and microbiological analysis of water of irrigation and microbiological analysis of lettuce (Lactuca sativa) in the Municipality of Ivoti/RS

Camila Corrêa Bierhas^I

Aline Belem Machado^II

Simone Ulrich Picoli^III

Daniela Montanari Migliavacca Osorio^IV

Daiane Bolzan Berlese^V

^I   Feevale University, RS, Brazil – camilabier@yahoo.com.br

^II  Feevale University, RS, Brazil – linebmachado@hotmail.com

^III Feevale University, RS, Brazil – simonepi@feevale.br

^IV Feevale University, RS, Brazil – danielaosorio@feevale.br

^V  Feevale University, RS, Brazil – daianeb@feevale.br

Abstract

The contamination of vegetables by pathogenic microorganisms is directly related to the water quality used in their irrigation. Lettuce is the main vegetable consumed in Brazil and because it does not undergo any processing before its consumption, it is directly affected by the quality of the water used for irrigation. This study analyzed the physical-chemical and microbiological quality of water used in lettuce irrigation and possible microbiological contamination of lettuce. In relation to microbiological analysis, high values were found for heterotrophic bacteria and total coliforms in weirs and vegetables. For fecal coliforms, in almost all water samples, the value found was above that established by the legislation. In lettuce, this occurred in three of twelve samples. No strong correlation was found between water and lettuce contamination. In relation to the physicochemical parameters analyzed, only the value of the turbidity in one of the weirs was above the threshold established by the legislation.

Keywords: Contamination; Microorganism; Vegetable; Water quality

1 Introduction

The main leafy vegetable cultivated in Brazil is lettuce (Lactuca sativa). Because it is an in natura food, as it is consumed raw, it can transmit several diseases. Its contamination is directly related to the quality of the water used in its irrigation (HOLVOET et al., 2014). Water for food production used by small producers is mostly obtained through self-supply options, such as weirs and artesian wells. However, this water can often be contaminated at the point of origin, in the distribution, and especially in the private weirs.

Resolution n. 357/05 of the National Council for the Environment (CONAMA) (BRASIL, 2005) establishes the classification of fresh, brackish and saline water of the National Territory. Water intended for the irrigation of vegetables and fruits that grow on the ground and are eaten raw without peel removal belongs to Class 1 and high-quality parameters are in place.

Thus, the physical-chemical and microbiological evaluation of water becomes extremely important in these cases, which must follow the parameters defined by CONAMA Resolution nº 357/2005 (BRASIL, 2005). Its constant monitoring is important to avoid any kind of contamination in humans related to the consumption of lettuce. Ordinance N. 2914 of the Ministry of Health, which deals with the quality of water for human consumption and its drinking standard, considers the absence of total and fecal coliforms as a standard for drinking water (BRASIL, 2011).

One of the main forms of contamination of vegetables by pathogenic microorganisms is through contaminated water used in garden irrigation (SLIFKO et al., 2000; ROBERTSON & GJERDE, 2001; SIMÕES et al., 2001). Therefore, the sanitary control of the water used for the irrigation of vegetables is very important since they can serve as a vehicle of contamination of these foods, becoming a public health problem (CARVALHO et al., 2003). This contamination can also occur through the use of organic fertilizers with fecal manure or fecal material of human origin. Other factors that may contribute to this occurrence are rainfall levels during the period of analysis, packing, transportation and handling of these vegetables during collection (SOARES, 2004).

For irrigation water, there is no total coliform counting pattern, although it is considered a risk factor for the consumer of in natura vegetables (PERES JUNIOR et al., 2012). Fecal coliforms, on the other hand, are used as standard for the microbiological quality of this water (CETESB, 2001) and their acceptable limit is 200 coliforms per 100 mL (BRASIL, 2005). Among fecal coliforms, Escherichia coli is widely used as an indicator of hygienic-sanitary food quality. This bacterium can cause gastroenteritis or progress to lethal cases in children, the elderly, pregnant women and immunosuppressed patients in general (SIQUEIRA et al., 2010).

The counting of heterotrophic bacteria in the water is directly related to the microbiological quality of the same. Even though there is no standard for counting these microorganisms in the water used for irrigation, this data is a complementary indicator of its quality, since an increase in this number indicates an increase in the presence of organic matter in the water (APHA, 2005).

Lettuce is an important microbiological contamination vehicle for its consumers (PERES JUNIOR et al., 2012) because it is an in natura food. This situation can be aggravated by the plant’s own structure, which favors the survival of several pathogenic microorganisms (HOLVOET et al., 2014). Considering that the consumption of water and contaminated food brings severe implications for human health, this study evaluated some physical-chemical and microbiological parameters of the water used in the irrigation of lettuces, as well as possible microbiological contamination of these vegetables.

2 Material and Methods

To analyze the quality of irrigation water of lettuce, physical-chemical parameters (pH, nitrite, nitrate, turbidity, ammonia, and chloride) and microbiological (total and fecal coliforms, heterotrophic bacteria) were used.

One sample was collected per month in each property (weir water) between October 2017 and March 2018. The collection was carried out in two properties that grow vegetables in the rural area of the municipality of Ivoti/RS, in the Picada Feijão region.

In relation to lettuces, two samples were collected per month, in the same period as the water, and microbiological analysis were performed, such as total and fecal coliforms, heterotrophic bacteria and Salmonella sp.

For the collection of water, a sterile bottle with capacity to 1 liter was used. The bottles were identified and transported under refrigerated conditions utilizing an isothermal box containing ice with a temperature of 4 °C, according to APHA (2005). The samples were transported and analyzed at the Analytical Center of Feevale University.

2.1 Physico-chemical analyzes of water

Determination of pH, nitrite, nitrate, turbidity, and chloride through a protocol standardized by the Analytical Center of the Feevale University, according to the “Standard Methods for the Examination of Water and Wastewater” (APHA, 2012).

2.2 Microbiological analysis of water

2.2.1 Determination of the Most Probable Number (MPN) of Total and Thermotolerant Coliforms

For the analysis of total and fecal coliforms, the multiple tube technique was used to determine MPN, according to the “Standard Methods for the Examination of Water and Wastewater” (APHA, 2005).

2.2.2 Presumptive test

In order to verify gas production by fermentation of lactose, three series with 3 tubes each containing 10 mL of lauryl sulfate tryptose broth (LST)/lactose broth (LB) were used with Duran tube inverted inside. Coliforms were considered suggestive of those that presented turbidity and gas production after the incubation period.

2.2.3 Confirmation test for total coliforms

Based on the positive tubes of the presumptive test, the confirmation test for fecal coliforms was performed. Those that presented turbidity and gas production were considered positive. According to the number of positive tubes and using the Hoskins table (APHA, 2005) for three tube series, the MPN of total coliforms per deciliter (dL) of the sample was determined.

2.2.4 Confirmation test for coliforms at 45 °C or fecal coliforms

Based on the positive tubes of the presumptive test, the confirmation test was performed for coliforms at 45 °C or fecal coliforms. According to the number of positive tubes and using the Hoskins table for three tube series, the MPN of fecal coliforms per deciliter (dL) of the sample was determined.

2.2.5 Quantification of mesophilic microorganisms by the Pour Plate method

One mL of the pure water sample was transferred to sterile Petri dishes and 15 - 20 mL of plate count agar (PCA), was melted, cooled and transferred to the same Petri dishes. In addition, 1 mL of the water sample was fractionated in three 0.3 mL aliquots and one 0.1 mL in sterile Petri dishes, adding molten and cooled PCA. All plates were incubated at 35 °C for 48 hours. Afterward, they were evaluated to determine the colony count, which showed the result of mesophilic microorganisms expressed in colony forming units per mL of the sample (SILVA et al., 2017).

2.3 Microbiological analysis of lettuce

For the collection of lettuce, the roots were removed with a disinfected knife at the field, the lettuce were conditioned in sterile bags, identified and transported under refrigerated conditions utilizing isothermal box containing ice with a temperature of 4 °C. The lettuce samples were pooled and processed within 24 h. In the laboratory, external leaves were carefully removed to avoid cross-contamination with soil residues from the field (RODRÍGUEZ et al., 2006).

In the study of total and fecal coliforms, the multiple tube technique was used to determine MPN, according to the “Compendium of Methods for the Microbiological Examination of Foods” (APHA, 2015). A total of 25 + 0.2 g of the sample was weighted, 225 mL of 0.1 % peptone saline was added and homogenized, reaching the dilution 10 - 1, which was the initial dilution for the tests. Subsequently, dilutions 10 - 2 and 10 - 3 were prepared by the serial dilution technique.

2.3.1 Determination of the Most Probable Number (MPN) of Total and Thermotolerant Coliforms

The determination of the MPN of total and fecal coliforms of the lettuce samples was carried out by the same methods used in the analysis of the water.

2.3.2 Quantification of mesophilic microorganisms by the Pour Plate method

Based on the dilutions 10 - 1 to 10 - 3, 1 mL was transferred to sterile Petri dishes and 15 – 20 mL plate count agar (PCA) was melted, cooled and transferred to the same Petri dishes. Movements in “8” were performed to homogenize the plates and after solidification, they were incubated at 35 °C for 48 hours. After incubation, plaques were counted and the number of colonies found, multiplied by the dilution factor, resulted in the number of mesophilic microorganisms per gram of lettuce (SILVA et al., 2017).

2.3.3 Analysis of Salmonella sp.

The method used for the analysis of Salmonella sp. in lettuce was ISO 6579 for presence/absence of Salmonella sp. in food. A total of 25 g of each sample was added to 225 mL of buffered peptone water. This mixture was homogenized and incubated at 37 °C for 18 hours. After, 0.1 mL was transferred to 10 mL of Rappaport-Vassilidis Soya Broth (RVS) and 1 mL to 10 mL of Muller Kauffmann Novobiocin Tetrathionate Broth (MKTTn). After, the RVS Broth was incubated at 41.5 °C for 24 hours and the MKTTn Broth incubated at 37 °C for the same period. In sequence, the differential plating of each culture was performed, which was incubated again. After this period, the development of typical Salmonella sp. colonies in these differential plating media was observed. When necessary, biochemical and serological confirmation tests were performed (SILVA et al., 2017).

2.4 Statistical Analysis

For the analysis of the data, descriptive statistics were used initially, followed by non-parametric tests due to the non-normality of the data (KOLMOGOROV-SMIRNOV test). For the comparison of the data between the two properties the MANN-WHITNEY test was used, where it was considered as a significant difference when p < 0.05. For the analysis of the weir and lettuce data of the same property the Spearman correlation test was used, considering as significant results those with p < 0.05. Data were processed in the Statistical Package for Social Sciences (SPSS) software, version 24.0.

2.5 Characteristics of properties

2.5.1 Property 1

The weir of property 1 where water samples were collected, receives only rainwater, not suffering from the influence of any other water source. Most of the lettuce produced on this property is sold through a cooperative to some schools in the region. Its weekly production is around 30 dozen per week and the fertilizer used is organic (chicken manure).

2.5.2 Property 2

The weir of property 2 is influenced by the water of a local stream. Its weekly lettuce production is around 500 dozen, which are sold to a large supermarket chain. The fertilizer used is organic (turkey and cattle manure).

3 Results

Table 1 presents the values of the physico-chemical parameters and microbiological data found in the water of the weirs of properties 1 and 2. The physico-chemical parameters are in accordance with the values established by the legislation. Only the value of the turbidity in the weir of property 2 was above the value established by CONAMA nº 375/05 (BRASIL, 2005). In relation to the microbiological analyzes, high values were found for heterotrophic bacteria and total coliforms, both in the water of the weirs and in the vegetables. For fecal coliforms, in eleven of the twelve samples collected, the value found was above the threshold established by CONAMA n° 375/05 (BRASIL, 2005), which is 200 MPN.

Table 1 - Physical-chemical and microbiological data of the water of properties 1 and 2 and parameters of the Resolution nº357/05 of CONAMA
Collection	Weir 1						Weir 2						CONAMA 357/05
	1	2	3	4	5	6	1	2	3	4	5	6
Physical-chemical
Chlorides (mgL^-1)	112.459	120.426	135.213	130.144	122.687	97.362	28.697	38.907	39.913	23.165	54.594	42.134	250
Nitrate (mgL^-1N in NO3)	0.1922	0.1045	0.1007	0.0759	0.0458	0.4601	0.24	0.4896	0.2524	0.142	0.4376	0.1997	10
Nitrite (mgL^-1N in NO2)	n.d	n.d	n.d	0.0401	<LQ	0.096	n.d	0.1953	n.d	0.0379	0.0390	n.d	10
pH	7.24	7.38	7.27	7.3	7.31	7.04	7.25	7.21	7.27	7.04	7.17	6.82	6.0 a 9.0
Turbidity (NTU)	3.3	16.9	14.7	3.8	15.8	12.3	17.1	16.7	13.1	21.2	10.1	74.3	up to 40
Microbiological
Total heterotrophic bacteria (UFC/dL)	>6.0 x 10^5	2.68 x 10^5	2.90 x 10^5	2.80 x 10^5	1,09 x 10^5	>6.0 x 10^5	>6.0 x 10^5	1.07 x 10^3	3.6 x 10^5	5.45 x 10^5	1.75 x 10^5	>6.0 x 10^5
Total coliforms (MPN/dL	>1.100	1.1	>1.100	>1.100	460	>1.100	290	>1.100	>1.100	1.1	>1.100	>1.100
Fecal coliforms (MPN)	>1.100	1.1	290	>1.100	460	>1.100	15	>1.100	1,1	1.1	1.1	>1.100	200
Resolution nº 357/05 of CONAMA (BRASIL, 2005) for Class I water
* <LQ below the limit of quantification. **n.d. not determined by the method.

Table 2 shows the data of the microbiological analysis performed on the lettuces of the two properties. In vegetables, a value above that established by Brazilian Health Regulatory Agency (ANVISA) (2001), which would be up to 100 MPN, was found in only three samples.

Table 2 - Microbiological data of lettuces of properties 1 and 2
Collection	Total heterotrophic bacteria (UFC/g)	Total coliforms (MPN/g)	Fecal coliforms (MPN/g)	Salmonella sp.
Lettuce 1
Collection 1	3.38 x 10^4	<3	<3	absent
Collection 2	1.09 x 10^6	7.3	3.6	absent
Collection 3	1.78 x 10^6	3.6	3.6	absent
Collection 4	3.6 x 10^6	240	150	absent
Collection 5	3.33 x 10^6	43	23	absent
Collection 6	9.5 x 10^6	>1.100	>1.100	absent
Lettuce 2
Collection 1	1.1 x 10^6	43	43	absent
Collection 2	2.72 x 10^6	<100	<100	absent
Collection	1.08 x 10^6	23	23	absent
Collection 4	5.2x 10^6	>1.100	290	absent
Collection 5	6.3 x 10^5	93	93	absent
Collection 6	9.6 x 10^6	93	93	absent

Source: The authors (2018)

* Values in italics are above that recommended by ANVISA (2001) (100 MPN/g)

We observed that, in three lettuce samples, fecal coliform values were higher than allowed by the legislation. Heterotrophic bacteria and total coliforms were also present in all samples. Salmonella sp. was not found in any of the samples.

Table 3 presents the results of the water from the weirs of properties 1 and 2, referring to the six collections performed regarding the median, mean and standard deviation of the physico-chemical analyzes; microbiological analyzes; and the microbiological analyzes of lettuces of properties 1 and 2.

Table 3 - Physical-chemical and microbiological analysis of the water of the weirs of properties 1 and 2, and microbiological analysis of lettuces of properties 1 and 2
Weir	Median	Mean	Standard Deviation
Weir 1
Chloride (mg L-1)	*1.21 x 10	1.19 x 10	0.13 x 10
Nitrate (mg L^-1N in NO3)	0.01 x 10	0.01 x 10	0.01 x 10
Nitrite (mg L^-1N in NO2)	0.0000	0.002 x 10	0.003 x 10
pH	0.72 x 10	0.72 x 10	0.01 x 10
Turbidity (NTU)	1.35 x 10	1.11 x 10	0.60 x 10
Heterotrophic bacteria (UFC/dL)	2.85 x 10^5	3.33 x 10^9	5.16 x 10^9
Total coliforms (MPN/dL)	1.20 x 10^3	1.06 x 10^3	2.96 x 10^2
Fecal coliforms (MPN)	1.15 x 10^3	9.08 x 10^2	4.18 x 10^2
Weir 2
Chloride (mg L-1)	*0.39 x 10	0.37 x 10	0.10 x 10
Nitrate (mg L^-1N in NO3)	0.02 x 10	0.02 x 10	0.01 x 10
Nitrite (mg L^-1N in NO2)	0.009 x 10	1.28 x 10^2	1.98 x 10^2
pH	0.71 x 10	0.71 x 10	0.01 x 10
Turbidity (NTU)	1.69 x 10	2.54 x 10	2.42 x 10
Heterotrophic bacteria (UFC/dL)	4.52 x 10^5	3.33 x 10^9	5.16 x 10^9
Total coliforms (MPN/dL)	1.20 x 10^3	1.03 x 10^3	3.65 x 10^2
Fecal coliforms (MPN)	1.10 x 10^3	9.52 x 10^2	4.61 x 10^2
Leaf Vegetables
Lettuce 1
Heterotrophic bacteria (UFC/dL)	2.21 x 10^6	2.92 x 10^6	3.58 x 10^6
Total coliforms (MPN/dL)	2.51 x 10	2.49 x 10^2	4.74 x 10^2
Fecal coliforms (MPN)	1.33 x 10	2.30 x 10^2	4.78 x 10^2
Lettuce 2
Heterotrophic bacteria (UFC/dL)	1.91 x 10^6	3.38 x 10^6	3.47 x 10^6
Total coliforms (MPN/dL)	7.15 x 10	2.50 x 10^2	4.66 x 10^2
Fecal coliforms (MPN)	7.15 x 10	9.86 x 10	9.78 x 10
* Significant difference (p<0.05), Mann-Whitney

The Spearman test was used to analyze the correlation of the microbiological parameters between irrigation water and lettuce from properties 1 and 2, when the values found were not significant. However, the correlation between weir 2 and lettuce 2 was moderate (R = 0.464), possibly indicating a microbiological contamination of lettuce by irrigation water.

4 Discussion

The physical-chemical parameters analyzed in the water of the two weirs studied in this study were in accordance with the current legislation, with the exception of turbidity, which in the last collection at property 2 was above permitted. This probably happened due to rainfall the day before the collection. A similar result was described in the study by Franco & Hernandez (2009), in which the turbidity values of the stream increased at all collection points during the rainy season. This increase in turbidity is justified by the erosion of the riverbanks that occurs in the rainy season (ANA, 2018).

In relation to the microbiological analysis of water used for irrigation, heterotrophic bacteria were found in all samples, as reported in the studies of Araújo et al. (2015) and Silva et al. (2016). Although there is no standard for counting these microorganisms in irrigation water, their count is directly related to their quality.

Total and fecal coliforms were also found in all water samples, for the former we do not have reference values in the legislation, but for fecal coliforms, there is an established limit (200 MPN) and only in one sample this value was not exceeded. In the study by Scherer et al. (2016), these same microorganisms were also found in all water samples, and the fecal coliforms were above the values established by the CONAMA Resolution n° 357/2005 (BRASIL, 2005) in all of them. The water of the weirs was considered inappropriate for use in the cultivation of vegetables. Similar results were described in the study by Araújo et al. (2015), in which only one sample was in agreement with the legislation. In another study, by Santos et al. (2010), 89% of irrigation water samples were positive for total coliforms and 87% for fecal coliforms. These results demonstrate the importance of controlling the quality of the water used in the irrigation, in order to avoid the contamination of the vegetables irrigated by it and also of the cultivated soil (ARAÚJO et al., 2015).

In the studied lettuce, heterotrophic bacteria and total coliforms were found in all samples, and although there are no parameters for counting these bacteria, these results should be considered a risk factor for the consumer (PERES JUNIOR et al., 2012). According to Scherer and collaborators (2016), total coliforms were found in all the studied vegetable samples, as well as in the study by Arbos and collaborators (2010), who analyzed organic lettuce. Bergamo & Gandra (2016) also found total coliforms in all samples of cultivated lettuce, both in the traditional and in the hydroponic mode.

Fecal coliforms were also found in all lettuce samples, but only three of them were above the value allowed by ANVISA (2001). However, the medians found in both properties were above the reference value. According to a study carried out by Frittoli & Rodrigues (2014), in 35% of the samples of minimally processed vegetables, the value of fecal coliforms was higher than that established by the legislation. Another study, carried out by Santos et al. (2010), found an increased contamination of these bacteria mainly in lettuce and coriander. According to Arbos and collaborators (2010), of the thirteen samples studied, three of them (two of lettuces and one of carrot) presented values for fecal coliforms above that allowed by the legislation. The presence of fecal coliforms in 100 % of the samples of this study can be related to the cross-contamination from the lack of hygiene and sanitary conditions of the manipulators, water, and/or utensils used by the producers (BERGAMO & GANDRA, 2016).

Santana & collaborators (2006) studied lettuces grown in three cultivation systems and found high concentrations of fecal coliforms in all samples, but the highest frequency of contamination was observed in organic cultivation, followed by traditional and hydroponic cultivations. According to Bergamo & Gandra (2016), Escherichia coli (fecal coliform) was found in 22.2% of the traditionally grown lettuce samples, in 11.1% of the hydroponic cultivars and in none of the organic cultivars, however, this difference was not statistically significant. The presence of fecal coliforms in water is related to the presence of fecal contamination, which represents a health risk to humans once lettuce is eaten raw (WHO, 2006).

In the present study, the presence of Salmonella sp. was not detected in any of the collected lettuce samples, as well as in the study by Costa et al. (2012) who analyzed conventional and organic lettuces. Also, the presence of this microorganism was not found in any of the lettuce samples of the 10 self-service restaurants analyzed in the State of Tocantins by Peres Junior et al. (2012). Similarly, Santana et al. (2006), as in the other studies, also did not detect the presence of Salmonella sp. in any of their samples.

Foods of animal origin are the main transmitters of this microorganism, but other studies have already detected its presence in products of plant origin, as in the study of Scherer and collaborators (2016), who found the presence of Salmonella sp. in five of the nine samples of lettuce studied. According to a study carried out by Takayanagui et al. (2000), this bacterium was found in 3.1% of the vegetables of 129 horticultural gardens surveyed. Rodrigues (2007) analyzed common, organic and hydroponic lettuce in the capital city Brasília and found that of the thirty samples studied, Salmonella was found in three common lettuces, two hydroponic lettuces, and two organic lettuces, indicating the lack of care in the process.

Despite the unsatisfactory microbiological quality of irrigation water analyzed in the present study, no strong correlation was found between it and the lettuce contamination index, only a moderate but not significant correlation (probably due to sample size) indicating a possible contamination of the lettuce by water. In contrast, Takayanagui et al. (2000) verified the contamination of lettuce, chicory, and watercress by fecal coliforms from irrigation water in vegetable gardens in the interior of the State of São Paulo. Another study carried out by Arbos et al. (2010) reports that water contaminated with human fecal material and the use of organic fertilizer without adequate composting time are the main forms of contamination of vegetable gardens.

According to Guimarães et al. (2003), some studies in Brazil have demonstrated the contamination of vegetables by fecal coliforms that may have been transmitted by irrigation water. This is demonstrated by Silva and collaborators (2016), who present, in their study, a strong correlation between the index of contamination of water and vegetables. In another study by Abreu et al. (2010) no signs of contamination were found in the soil or organic fertilizers used, and there was a strong indication that lettuce contamination was caused by irrigation water. Also, the fact that rural properties often have animal husbandry, and due to the producer’s own dwelling that can lead to some type of contamination (CHAVES et al., 2010; ARAÚJO et al., 2015; VOLKWEIS et al., 2015), can contribute to the contamination of the irrigation water and the lettuce.

According to Jensen et al. (2015), the use of contaminated water, together with inadequate handling and transport, is responsible for the entry of E. coli into the food chain. In another study, Rodrigues (2007), also states that the contamination of vegetables can occur from planting to commercialization and consumption. According to Mogharbel (2005), due to the potential risk of diseases conveyed by vegetables that are consumed raw, the correct orientation of the producers is very important and manipulators of these foods, in relation to personal hygiene practices and also on the importance of an adequate washing and disinfection of lettuce leaves before consumption. Nascimento (2002) reports that because the hygiene process is the only treatment received by lettuce between the cultivation and the consumption if it is not carried out properly the transmission of several diseases may occur.

Therefore, it is essential to analyze and control the quality of water used in the irrigation of vegetables that are consumed raw as lettuce, since there are still many possibilities for contamination until they reach the final consumer. The use of contaminated water together with inadequate transport and handling favors the contamination of the people who eat them and can cause serious health problems, especially in children, the elderly, pregnant women and immunosuppressed persons.

5 Conclusions

According to the results obtained in this study, the water used for irrigation of lettuce did not have an adequate microbiological quality. However, the physical-chemical parameters analyzed were in accordance with the legislation in almost all samples.

In relation to the studied lettuce samples, heterotrophic bacteria and total coliforms were found in all samples, but the fecal coliforms were found only in some of them. However, Salmonella sp. was not detected in any sample.

Although a moderate correlation has been found between contamination of irrigation water and lettuce, many other studies demonstrate a strong correlation between them.

Thus, it is essential to monitor the quality of irrigation water used in vegetables that will be consumed in natura. The correct handling and sanitization of these foods to avoid potential contamination of their consumers are also fundamental.

References

ABREU IMO, JUNQUEIRA AMR, PEIXOTO JR, OLIVEIRA SA. Qualidade microbiológica e produtividade de alface sob adubação química e orgânica. Ciência e Tecnologia de Alimentos. 2010;30(1):108 – 118.

AGÊNCIA NACIONAL DE ÁGUAS (ANA). Indicadores de qualidade: Índice de qualidade das Águas (IQA) [Internet]. Brasília: 2018 [cited 10 march 2019]. Available from: http://pnqa.ana.gov.br/indicadores-indice-aguas.aspx.

AGÊNCIA NACIONAL DE VIGILÂNCIA SANITÁRIA (ANVISA). RDC n. 12, de 02 de janeiro de 2001. Regulamento técnico sobre padrões microbiológicos em alimentos. Diário Oficial [da] República Federativa do Brasil, Poder Executivo, Brasília: ANVISA, 2001.

AMERICAN PUBLIC HEALTH ASSOCIATION (APHA). Microbiological examination. In: American Public Health Association (APHA). Standard methods for the examination of water and wastewater. 21. ed. Washington: APHA, 2005.

AMERICAN PUBLIC HEALTH ASSOCIATION (APHA). Microbiological examination. In: American Public Health Association (APHA). Standard methods for the examination of water and wastewater. 22. ed. Washington: APHA, 2012

AMERICAN PUBLIC HEALTH ASSOCIATION (APHA). Microbiological examination. In: American Public Health Association (APHA). Compendium of methods for the microbiological examination of foods. Washington: APHA, 2015.

ARAÚJO FV, VIEIRA L, JAYME MMA, NUNES MC, CORTÊS M. Avaliação da qualidade da água utilizada para irrigação na bacia do Córrego Sujo, Teresópolis, RJ.  Cadernos Saúde Coletiva. 2015;23(4):380-385.

ARBOS KA, FREITAS RJS, STERTZ SC, CARVALHO LA. Segurança alimentar de hortaliças orgânicas: aspectos sanitários e nutricionais. Ciência e Tecnologia de Alimentos. 2010;30(1):215-220.

BERGAMO G, GANDRA EA. Avaliação microbiológica de alface cultivada sob as formas tradicional, orgânica e hidropônica. Brazilian Journal of Food Research. 2016;7(3):82-93.

BRASIL. Conselho Nacional do Meio Ambiente (CONAMA). Resolução 357, de 17 de março de 2005. Dispõe sobre a classificação dos corpos de água e diretrizes ambientais para o seu enquadramento, bem como estabelece as condições e padrões de lançamento de efluentes, e dá outras providências. Diário Oficial [da] República Federativa do Brasil, Poder Executivo, Brasília (Brasil): 2005.

BRASIL. Portaria n° 2914 de 12 de dezembro de 2011. Ministério da Saúde, Brasília (Brasil): 2011.

CARVALHO JB, NASCIMENTO ER, NETO JFN, CARVALHO IS, CARVALHO LS, CARVALHO JS. Presença de ovos de helmintos em hortaliças fertilizadas com iodo da lagoa de estabilização. Revista Brasileira de Análises Clínicas. 2003;35(2):101-103.

CHAVES KF, SILVA NBN, VIEIRA TB, MENDES ACG, GRAVINA CS, MARTINS ADO, MARTINS ML. Avaliação microbiológica da água empregada em laticínios da região de Rio Pomba-MG. UNOPAR Científica. Ciências Biológicas e da Saúde. 2010;12(4):5-8.

COMPANHIA AMBIENTAL DO ESTADO DE SÃO PAULO (CETESB). Variáveis de qualidade das águas [Internet]. São Paulo:2001 [cited 08 march 2019]. Available at: http://www.cetesb.sp.gov.br/ Agua/rios/variaveis.asp.

COSTA EA, FIGUEIREDO EAT, CHAVES CS, ALMEIDA PC, VASCONCELOS NM, MAGALHÃES IMC, MORAES AF, PAIXÃO LMN. Avaliação de alfaces e eficiência da sanitização. Revista Alimentos e Nutrição. 2012;23(3):387-392.

FRANCO PRAM, HERNANDEZ FBT. Qualidade da água para irrigação na microbacia do Coqueiro, Estado de São Paulo. Revista Brasileira de Engenharia Agrícola e Ambiental. 2009;13(6):772-780.

FRITTOLI RB, RODRIGUES LH. Analysis of fecal coliform and Salmonella sp. in samples of minimally processed vegetables. Revista Científica da FHO- UNIARARAS. 2014;2(2):14 – 20.

GUIMARÃES AM, ALVES EGL, FIGUEIREDO HCP, COSTA GMD, RODRIGUES LDS. Frequência de enteroparasitas em amostra de alface (Lactuca sativa) comercializada em Lavras, Minas Gerais. Revista da Sociedade Brasileira de Medicina Tropical. 2003;36(5):621-623.

HOLVOET K, SAMPERS I, SEYNNAEVE M, JACXSENS L, UYTTENDAELE M. Agricultural and management practices and bacterial contamination in greenhouse versus open field lettuce production. International Journal of Environmental Research and Public Health. 2014;12(1):32-63.

JENSEN DA, FRIEDRICH LM, HARRIS LJ, DANYLUK MD, SCHAFFNER DW. Cross contamination of Escherichia coli O157:H7 between lettuce and wash water during home-scale washing. Food Microbiology. 2015;46:428-433.

MOGHARBEL ADI, MASSON ML. Perigos associados ao consumo da alface, (Lactuca sativa), in natura. Revista Alimentos e Nutrição. 2005;16(1):83-88.

NASCIMENTO MS. Avaliação comparativa de tratamentos químicos na sanitização de frutas e verduras. (Dissertação Mestrado). Faculdade de Ciências Farmacêuticas, Universidade Estadual Paulista “Júlio de Mesquita Filho”, Araraquara.  2002.

PERES JUNIOR J, GONTIJO EEL, SILVA MG. Perfil parasitológico e microbiológico de alfaces comercializadas em restaurantes self-service de Gurupi-TO. Revista Científica do ITPAC. 2012;5(1):33-37.

ROBERTSON LJ, GJERDE B. Occurrence of parasites on fruits and vegetables in Norway. Journal of Food Protection. 2001;64:1793-1798.

RODRIGUES CS. Contaminação microbiológica em alface e couve comercializadas no varejo de Brasília-DF. Monografia (monography). Brasilia:Universidade de Brasília/UnB; 2007.

RODRÍGUEZ C, LANG L, WANG A, ALTENDORF K, GARCÍA F, LIPSKI A. Lettuce for human consumption collected in Costa Rica contains complex communities of culturable oxytetracycline- and gentamicin-resistant bacteria. Applied and Environmental Microbiology. 2006;72(9):5870-5876.

SANTANA LRR, CARVALHO RDS, LEITE CC, ALCÂNTARA LM, OLIVEIRA TWSD.  Qualidade física, microbiológica e parasitológica de alfaces (Lactuca sativa) de diferentes sistemas de cultivo. Revista Ciência e Tecnologia de Alimentos. 2006;26(2):264-269.

SANTOS YO, GUIMARÃES AG, ALMEIDA PF. Hygienic sanitary quality of vegetables and evaluation of treatments for the elimination of indigenous E. coli and E. coli O157:H7 from the surface of leaves of lettuce (Lactuca sativa L.). Revista Ciência e Tecnologia de Alimentos. 2010;30(4):1083-1098.

SCHERER K, EICHELBERGER GRANADA C, STÜLP S, SPEROTTO RA. Avaliação bacteriológica e físico-química de águas de irrigação, solo e alface (Lactuca sativa L.) Revista Ambiente & Água. 2016;11(3):665-675.

SILVA N, JUNQUEIRA VCA, SILVEIRA NFA. Manual de métodos de análise microbiológica de alimentos e água. 5. ed. São Paulo: Editora Blucher. 2017

SILVA A, LIMA C, QUEIROZ JJF, JÁCOME P, JÁCOME-JÚNIOR AT. Análise bacteriológica das águas de irrigação de horticulturas. Revista Ambiente & Água. 2016;11(2).

SIMÕES M, PISANI B, MARQUES EGL, PRANDI MAG, MARTINI MH, CHIARINI PFT, ANTUNES JLF, NOGUEIRA AP. Hygienic-sanitary conditions of vegetables and irrigation water from kitchen gardens in the municipality of Campinas, SP. Brazilian Journal of Microbiology. 2001;32(4):331-333.

SIQUEIRA LPD, SHINOHARA NKS, LIMA RMTD, PAIVA JDED, LIMA FILHO JLD, CARVALHO ITD. Avaliação microbiológica da água de consumo empregada em unidades de alimentação. Ciência & Saúde Coletiva. 2010;15:63-66.

SLIFKO TR, SMITH HV, ROSE JB. Emerging parasites zoonoese associated with water and food. International Journal for Parasitology. 2000;30:1379-1393.

SOARES B. Ocorrência de estruturas parasitárias em hortaliças [dissertation]. Florianópolis: Universidade Federal de Santa Catarina; 2004.

TAKAYANAGUI OM, FEBRÔNIO LH, BERGAMINI AM, OKINO MH, SILVA AAMC, SANTIAGO R, CAPUANO DM, OLIVEIRA MA, TAKAYANAGUI AM. Fiscalização de hortas produtoras de verduras do município de Ribeirão Preto, SP. Revista da Sociedade Brasileira de Medicina Tropical. 2000;33(2):169-174.

VOLKWEIS DSH, LAZZARETTI J, BOITA ERF, BENETTI F. Qualidade microbiológica da água utilizada na produção de alimentos por agroindústrias familiares do município de Constantina/RS. Revista Eletrônica em Gestão, Educação e Tecnologia Ambiental. 2015;19:18-26.

WORLD HEALTH ORGANIZATION (WHO). Guidelines for the Safe Use of Wastewater, Excreta and Greywater. Wastewater Use in Agriculture. V. 2, Geneva: WHO, 2006.