Soil chemical properties, leaf mineral status and crop production in a lemon tree orchard irrigated with two types of wastewater

Agricultural Water Management 109 (2012) 54–60

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Soil chemical properties, leaf mineral status and crop production in a lemon tree orchard irrigated with two types of wastewater Francisco Pedrero a,∗ , Ana Allende b , María I. Gil b , Juan J. Alarcón a a b

Irrigation Department, CEBAS-CSIC, P.O. Box 164, Espinardo, Murcia 30100, Spain Research Group on Quality, Safety and Bioactivity, CEBAS-CSIC, P.O. Box 164, Espinardo, Murcia 30100, Spain

a r t i c l e

i n f o

Article history: Received 22 June 2011 Accepted 13 February 2012 Available online 10 March 2012 Keywords: Reclaimed water Phytotoxicity Plant production Fruit quality

a b s t r a c t The effects of applying different types of treated wastewater on citrus trees were studied in Murcia, in the south-east of Spain. Two treatments with wastewater effluents of different quality were applied for three consecutive years. In the first case, the wastewater received a secondary treatment (conventional activated sludge). In the second case, the irrigation water was a mix of well water and wastewater from a tertiary treatment plant (conventional activated sludge with ultraviolet tertiary treatment). The characteristics of the tertiary treated wastewater make it better for irrigation than the secondary treated wastewater. It was considered that high salinity, Cl and B concentration could be the main restrictions associated with treated wastewater irrigation in both cases, although leaf toxicity levels were not observed. The soil nitrate concentration increased over the experimental time period in both water irrigation treatments. The production was affected by the wastewater quality and the total crop yield was lower in the plots irrigated with secondary treated wastewater. However, in these plots, the fruit-quality indexes such as external colour, weight, peel thickness, firmness, soluble solids, pH, total acidity and maturity index were significantly better than those observed in the plots irrigated with tertiary treatment. The soil microbiological analysis revealed an absence of faecal coliforms, Escherichia coli and helminth eggs in the experimental plots irrigated with tertiary treated wastewater, but with secondary treated wastewater the soil accumulation of faecal coliforms exceeded health standards. In both cases, there was an absence of microbiological contamination on fruits. © 2012 Elsevier B.V. All rights reserved.

1. Introduction Wastewater reuse in agriculture is an important management strategy in areas with limited freshwater resources. Such a strategy is important because of the potential economic and environmental benefits. It is therefore necessary to initiate and support wastewater reuse projects all over the world, particularly since the population and demand for food is growing steadily and the fresh water resource will not increase. Several studies have shown the advantages and disadvantages of using wastewater for citrus crops irrigation (Zekri and Koo, 1993, 1994; Aucejo et al., 1997; Morgan et al., 2008; Reboll et al., 2000; Pedrero and Alarcón, 2009; Pedrero et al., 2010; Maurer et al., 1995; Meli et al., 2002; Pereira et al., 2011). Reuse of treated wastewater is a good management option for increasing water supplies

∗ Corresponding author at: Centro de Edafología y Biología Aplicada del Segura (CSIC), P.O. Box 164, E-30100 Espinardo (Murcia), Spain. Tel.: +34 968 396200; fax: +34 968 396213. E-mail addresses: [email protected], [email protected] (F. Pedrero). 0378-3774/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.agwat.2012.02.006

to agriculture. One of its benefits is the plant’s use of both water and nutrients thereby reducing the pollution load that wastewater contributes to the surface water supply (Zekri and Koo, 1994). However, depending upon its sources and degree of treatment, wastewater may contain high concentrations of salts, heavy metals, trace elements, viruses and bacteria. Irrigation with poor quality wastewater may create undesirable effects on soils and plants or pose a potential health threat to the consumer. Improperly treated wastewater can contain food-borne pathogens such as pathogenic bacteria, viruses, protozoa and nematodes (Steele and Odumeru, 2004; Steele et al., 2005). This situation is particularly relevant in some developing countries, where poorly treated wastewater is used for crop irrigation (Rattan et al., 2005). Asano et al. (2007) define reclaimed water as the municipal wastewater that has gone through various treatment processes to meet specific water quality criteria with the intent of being used in a beneficial manner (e.g., irrigation). The term recycled water is used synonymously with reclaimed water. The current water quality criteria for agricultural reuse have been established by USEPA (2004), based mainly on total dissolved solids (TDS), salinity and sodicity (Ayers and Westcot, 1989), and the need for periodic microbiological analysis of the irrigation-water supplies,

F. Pedrero et al. / Agricultural Water Management 109 (2012) 54–60

independently of the water source considered, to minimize negative public health impacts (WHO, 2006). More specific water quality parameters for the use of reclaimed water have been presented by Levine and Asano (2004) and there is considerable interest in the long-term effects of reclaimed water on crops intended for human consumption. In many areas of the world water reuse is viewed increasingly as a means to augment existing water resources against the prospect of continued droughts and water supply shortages. Reclaimed water use projects have been developed in countries facing water shortages (Angelakis et al., 1999). In the United States, for example, California reused 670 hm3 /year in 2003 (330 hm3 /year in 1987) while Florida reused 915 hm3 /year in 2006 (capacity 1900 hm3 /year) (Asano et al., 2007). The use of reclaimed water for irrigation has been progressively adopted by virtually all Mediterranean countries (Lazarova, 2000). Israel was pioneer in this field, soon followed by Tunisia, Cyprus, and Jordan. More recently, European Mediterranean countries started considering water reuse for irrigation (Marecos Do Monte et al., 1996). In Spain, the intensive agriculture is concentrated in the south east, where fresh water is very scarce (Intrigliolo et al., 2011). The present work was conducted in Segura Basin (Murcia), a semiarid Region of Spain, where drought and water deficit are the main factor limiting agricultural production. In this Region, some investigations on precise irrigation scheduling procedures using different plant measurements (García-Orellana et al., 2007) and shading with aluminised-plastic nets (Alarcón et al., 2006), were aimed at improving water use efficiency of lemon trees. The water deficit in the Segura Basin, together with the everincreasing demand due to the continued urban growth in the coastal zone and the major demand from intensive agricultural activity, has made it necessary to use treated wastewater for irrigation. According to the Murcia Regional Ministry of Water and Agriculture, the annual volume necessary to cover the regional agricultural water needs exceeds 880,000 Ml (CARM, 2007). The current volume of treated wastewater in Murcia is 101,800 Ml/year (ESAMUR, 2009), which represents a sixth of renewable resources of the Segura River Basin, and, besides serving other purposes, it supplies 12.8% of the water used for irrigation. In Murcia, the 54.8% of the wastewater treatment plants receive a secondary treatment, and the 42.2% receive a tertiary treatment (ESAMUR, 2009). The major problem associated to reclaimed water use in Murcia is salinity. In this region, 93% of the treated wastewater has an electrical conductivity (EC) higher than 2 dS m−1 and 37% has EC values higher than 3 dS m−1 (ESAMUR, 2005), and it is known that water with EC ≥ 3 dS m−1 requires very intensive management to control adverse salinity effects. The effects of treated wastewater on environmental pollution, plant growth or crop production are rarely studied in field conditions and thus, these types of experimental approaches are scarce (Pedrero et al., 2010). The aim of this work was to study the effects of different reclaimed irrigation waters on citrus tree performance. In particular, the objective of this research was to compare the effects of two types of wastewater, the one obtained through a secondary treatment and the other from a tertiary treatment, and to study their effects on soil chemical properties, leaf mineral status, crop production, fruit quality of citrus and microbiological safety.

2. Materials and methods 2.1. Experimental conditions The experiment was conducted during 2005–2007 in one experimental site planted to lemon trees in the Region of Murcia. The

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experimental orchard size was 12 ha of ‘Fino’ lemon grafted on ‘Macrophyla’ rootstock. The trees were 7 years old and were spaced 7 × 5 m. The water was supplied to trees by drip irrigation with eight compensated pressure emitters per tree, each with a flow rate of 4 L h−1 . The soil was classified as a clay loam soil (32% clay, 32% loam and 36% sand). Two different irrigation wastewater qualities were applied, secondary treated wastewater (STW) was used in one case, while tertiary treated wastewater (TTW) was used in the other case. In this trial, STW was based on the process of Biological Purification of Activated Sludge, and TTW was a mix of well quality groundwater and wastewater from a tertiary treatment plant (conventional activated sludge with ultraviolet tertiary treatment). The experimental design of each treatment was 4 standard experimental plots distributed randomly in blocks. The standard plot was made up of 12 trees, organized in 3 adjacent rows. The two central trees of the middle row were used for measurements and the other 10 trees were guard trees. The irrigation doses were scheduled from January 2005 until December 2007 on the basis of weekly crop evapotranspiration (ETc) estimated from reference evapotranspiration (ETo), calculated with the FAO56 Penman–Monteith equation (Allen et al., 1998), and a monthly crop factor (Castel et al., 1987). Meteorological data at the experimental sites were also used to calculate the water application. The data were collected from a weather station located 2 km from the experimental plots. The total irrigation amounts were measured with inline water flowmeters. The average amount of water applied was 601 mm/year. The average annual precipitation was 318 mm. During the season of the experiment the average annual ETc was 903 mm (Fig. 1). The fertilizers rates of N–P2 O5 –K2 O applied through the drip irrigation system were 240–90–100 (kg ha−1 ). The statistical analysis was performed by weighted analysis of variance (ANOVA) using linear model for SPSS software (version 17.0, SPSS Inc., Chicago, USA). 2.2. Water analysis Three water samples from each irrigation water source were collected monthly between 2005 and 2007 in order to characterize irrigation water quality (36 samples/year). The samples were collected in glass bottles, transported in an ice chest to the lab and stored at 5 ◦ C. The concentration of macronutrients (Na, K, Ca, Mg), micronutrients (Fe, B, Mn) and heavy metals (Ni, Cd, Cr, Cu, Pb, Zn) were determined by Inductively Coupled Plasma (ICP-ICAP 6500 DUO Thermo, England); anions (chloride, nitrate, phosphate and sulphate) were analysed by ion chromatography with a Chromatograph Metrohm (Switzerland); pH was measured with a pH-meter Cryson-507 (Crisom Instruments S.A., Barcelona, Spain); EC and total dissolved solids (TDS) were determined using the multi-range equipment Cryson-HI8734 (Crisom Instruments, S.A., Barcelona, Spain) and turbidity was measured with a turbidity-meter DinkoD-110 (Dinko Instruments S.A., Barcelona, Spain). The microbiological quality of irrigation water was assessed through the detection of total coliforms, faecal coliforms and E. coli by membrane filtration procedure (APHA, 1985). Samples were filtered using a vacuum system through a sterile 0.45-␮m-poresize membrane filters (Millipore, Billerica, USA). Colony formation was obtained after incubation on top of Chromocult agar plates (Merck, Darmstadt, Germany) for 24 h. Incubation temperatures were 37 ◦ C for total coliforms and E. coli, and 44.5 ◦ C for faecal coliforms. Microbial counts were expressed as log cfu ml−1 . The helmint eggs were measured following the Bailenger’s method (Bailenger, 1979). For E. coli O157:H7 detection, enrichments were prepared pouring 25 ml of water samples into sterile stomacher bags and adding 225 ml of mTSB + Novobiocin (Oxoid, Basingstoke,

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160 140

Irrigation Rainfall ETc

mm month-1

120 100 80 60 40 20 0 Jan

Feb

Mar

Apr

May

Jun

Jul

Aug

Sep

Oct

Nov

Dec

Fig. 1. The crop evapotranspiration (ETc), rainfall and irrigation water applied (mm month−1 ). The values are the monthly average from data collected during 2005, 2006 and 2007.

Hampshire, UK). Once homogenized, enrichments were incubated at 37 ◦ C for 24 h. After incubation, enrichments were spread-plated on Sorbitol MacConkey Agar (Scharlau chemie, Barcelona, Spain) containing CT-supplement (Merck KGaA, Darmstadt, Germany), and incubated further for 24 h at 37 ◦ C. Presumptive E. coli O157:H7 colonies (colourless) were selected and stored in eppendorf tubes containing TSB + 10% glycerol at −20 ◦ C, before performing PCR analysis (Manafi and Kneifel, 1989).

2.3. Soil analysis Twelve soil samples collected at a depth of 0–20 cm (sampled midway between emitters spaced 90 cm apart) were taken every three months at both irrigation treatments between January 2005 and December 2007. The soil was dried at room temperature for 1 week, ground and sieved through a 2 mm nylon mesh before analysis. Correction for dry mass was obtained from a separate portion by drying at 105 ◦ C for 24 h. Organic matter (OM) and total N were analysed using an automatic microanalyser Flash EA 1112 Series (England) and Leco Truspec (Sant Joseph, USA). The macro-elements, microelements and heavy metals were determined by Inductively Coupled Plasma (ICP-ICAP 6500 DUO Thermo, England) after nitric–perchloric acid (2:1) digestion (Thompson, 1982). Replicate samples (0.25 g) were digested by aqua regia acid HCl/HNO3 . Anions were analysed by ion chromatography with a Chromatograph Metrohm (Switzerland) after using a standard soil:distilled water ratio at 1:2.5 (w:w). The pH was determined on saturated soil-paste samples with a pH-meter Cryson-507 (Crisom Instruments S.A., Barcelona, Spain). The electrical conductivity of the saturated paste extract (ECe) was measured with a multi-range Cryson-HI8734 electrical conductivity meter (Crison Instruments, S.A., Barcelona, Spain). Soluble Ca and Mg were measured using the EDTA tritation method and Na was measured using a flame photometer (Richards, 1954). For soil microbiological analysis, twelve soil samples every three months were taken at each irrigation treatment. Each sample was placed in sterile closed containers suitable for isolation of the environment. Samples were transported in an ice chest to the lab and stored at 5 ◦ C before being processed. Soil samples were diluted 1:10 in sterile 0.1% peptone water and homogenized by hand in sterile lab stomacher bags. To determine the microbial quality of soil samples, 25 g of soil were homogenized in a 1:10 dilution of sterile 0.1% buffered peptone water using sterile filter stomacher bags (Seward Limited, London, UK) and a stomacher (IUL Instrument, Barcelona, Spain) for 90 s and plated on chromocult plates to determine the loads of total and thermo-tolerant coliforms and E. coli as previously described. Microbial counts were expressed as log cfu g−1 .

2.4. Leaf analysis Spring flush leaves from non-fruiting branches were sampled every three months during 2005, 2006 and 2007. Twenty leaves were sampled from 12 trees at each irrigation treatment and time. Leaves were washed with a detergent (alconox 0.1%), rinsed in tap water, cleaned with a dilute solution of 0.005% HCl and finally rinsed in distilled water, left to drain on a filter paper and oven dried for at least 2 days at 65 ◦ C. Dried leaves were ground and a nitric-perchloric acid (2:1) digestion (Thompson, 1982) was executed. Replicate samples (0.25 g) were digested by aqua regia acid HCl/HNO3 . The concentration of macro-elements, microelements and heavy metals were determined by Inductively Coupled Plasma (ICP-ICAP 6500 DUO Thermo, England). Later anions were analysed by ion chromatography with a Chromatograph Metrohm (Switzerland) after using a standard leaf:distilled water ratio at 1:2.5 (w:w). Total N and C concentrations were measured using an automatic micro-analyser Flash EA 1112 Series (England) and Leco Truspec (Sant Joseph, USA).

2.5. Fruit analysis For each plot, 25 fruits (100 fruits per treatment) were picked at commercial maturity from each irrigation treatment (STW and TTW) in 2006 and 2007. Fruits were harvested randomly from the outer canopy of selected trees in order to obtain a homogeneous sample. A first statistical analysis was made with no significance differences between plots in each treatment. Quality indexes: Titratable acidity (TA), pH and soluble solid content (SSC) were evaluated on fruit samples collected at irrigation treatment. TA was determined by titration of 10 ml of juice with 0.1 mol l−1 NaOH to pH 8.1. The pH values were measured using a pH meter and SSC with an Atago N1 handheld refractometer (Tokyo, Japan). Microbial analyses: Fruit samples were taken from the peel of the fruit by a cork borer of 7.1 cm2 . At least three peel disks from each fruit were taken and each sample was composed of five fruits. Samples were placed in a 250 ml sterile flask containing 50 ml of sterile 0.1% buffered peptone (BPW, AES Laboratoire, Combourg, France) and vigorously shaken using a IKA-VIBRAX-VXR mixer for 5 min. The peptone wash solution was diluted in sterile 0.1% peptone and plated on appropriate media. Total aerobic mesophilic bacteria were enumerated by using plate count agar (PCA) (Scharlau Chemie S.A., Barcelona, Spain) after incubation for 48 h at 30 ◦ C. Total and thermotolerant coliforms and E. coli were isolated by using Chromocult agar (Merck, Darmstadt, Germany) after incubation for 24 h at 37 ◦ C for total coliforms and E. coli and 44.5 ◦ C for thermotolerant coliforms. Yeast and moulds were enumerated in Rose Bengal agar

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(Scharlau Chemie S.A., Barcelona, Spain) by spread-plating 100 ␮l of the appropriate sample dilution and incubated at 30 ◦ C for 48–72 h. Microbial counts were expressed as log cfu cm−2 . Identification of Escherichia coli O157:H7 by immunochromatographic rapid test: The GLISA Singlepath® E. coli O157 (Merck) was used for rapid test for E. coli O157:H7 in fruit samples. To perform the test, samples of 25 g of lemon peel were homogenized with 225 ml of EHEC (Enterohemorrhagic E. coli) enrichment broth (EEB) [tryptic soy broth (TSB) + bile salts (1–5 g l−1 ) (No. 3) + VCC (vancomycin, cefixime and cefsulodin) − selective supplement] (Oxoid). Five ml of post-enrichment EEB culture were transferred to a polypropylene cap tube and placed in a boiling water bath for 20 min. After cooling to room temperature, three-falling drops (150 ␮l) were placed in the Singlepath test device sample port, and after 15 min, results were read and scored according to the description of the test. Samples were considered positive when red lines appeared on both test and control zones at or prior to 20 min. 3. Results 3.1. Irrigation water quality The physical–chemical and microbiological parameters measured during three years for both sources of irrigation-water are shown in Table 1. The quality of the two sources was significantly different in terms of sodium, chlorine and boron concentrations as well as EC, TDS and pathogen contents (thermotolerant coliforms, E. coli and helmith eggs). The values of these parameters were significantly higher in the irrigation-water coming from STW than that taken from TTW (Table 1). The average chloride concentration registered over the three years was 15.86 and 10.60 meq l−1 in STW and TTW sources respectively. The average level of boron concentration detected in the water of STW was twice (0.96 ppm) the average level measured in TTW (0.50 ppm). The average value of the electrical conductivity over the experimental period was 2.96 in STW and 2.13 dS m−1 in TTW. The microbiological quality of the irrigation water used in STW showed higher levels of thermotolerant coliforms than those used in TTW. E. coli O157:H7 was not detected in any of the irrigation water samples (Table 1). 3.2. Soil chemical composition The results of chemical analysis of ion concentrations in soil samples collected from STW and TTW are shown in Table 2. The concentrations of Na, K, Ca and Mg did not show significant difference between the soils of both irrigation treatments and were almost constant during the experiment. The electrical conductivity of the soil saturated paste extract in STW increased along the experiment, reaching the value of 2.2 dS m−1 in 2007. The concentrations of boron and chloride measured in STW were, respectively, 4 and 1.5 folds higher than those measured in TTW. The soil nitrate concentration showed a slight increase over the experimental period and in both wastewater irrigation treatments (Table 2). It was also observed that the poor microbial quality of the irrigation water used in STW caused an increase in the microbial levels in soil samples collected at this treatment, represented by an increase in the thermotolerant coliforms loads (Table 2). 3.3. Leaf mineral analysis The leaf concentrations of Na, K, Ca, Mg and sulphates did not show significant differences between years and treatments (Table 3). The leaf Cl concentration increased 40 and 30% in STW and TTW, respectively, over the three years of the experiment (Table 3) without showing visible symptoms. In spite of the high boron levels observed in the irrigation water and the soil of both wastewater

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irrigation treatments, no significant increase in the foliar concentration was seen during the experiment (Table 3). 3.4. Yield and fruit quality In 2005 hail impact caused discoloured scars on fruit, unevenly affected on the different plots, resulting in a significant loss of production. Therefore there was no record. The total yield harvested in 2006 was 57.4 and 50.5 t ha−1 in TTW and STW respectively; while in 2007 the crop production was 56.5 and 49.5 t ha−1 in TTW and STW. This difference of 13% between treatments was statistically significant. No significant differences were observed between quality indexes of lemon fruits obtained from STW and TTW in 2006 (Table 4). However, in 2007, lemon fruits from STW showed higher TA and SSC than lemon fruits obtained from TTW (Table 4). In addition, fruits from trees irrigated with reclaimed water in STW had a higher SSC:TA ratio and therefore, they reached maturity standards earlier than fruits from trees irrigated in TTW (Table 4). The microbial load of total aerobic bacteria, yeast and moulds was very similar for both wastewater irrigation treatments in all lemons fruits, and no risk of thermotolerant coliforms was observed as E. coli spp. counts were undetected for all the samples (Table 4). 4. Discussion In both sources of irrigation water the average sodium concentrations observed over the three years were higher than the threshold of restriction on use (<9 meq l−1 ) recommended in the FAO-bulletin on water quality for agriculture (Ayers and Westcot, 1989). However, the corresponding concentrations of calcium and magnesium were high enough to maintain the sodium adsorption ratio “SAR” within the range of 0–10 (Table 1). This indicates the presence of a reduced sodification power in both treatments (Rhoades, 1982). The Cl− concentrations in both treatments were higher than the threshold of restriction on use (<4.28 meq l−1 ) determined in Australia for citrus trees by Cole (1985) (Table 1). This author estimated a yield decrease of about 20% for each increase of 1 meq l−1 of Cl− concentration in the irrigation water above the threshold value, but this general behaviour is not always observed. It has been demonstrated the rootstock selection plays an important role in the intensive agriculture developed in the semi-arid and arid areas, and the rootstock used in this experiment (Macrophyla) is more salt tolerant than other citrus rootstocks (Nieves et al., 1991; Fernández-Ballester et al., 2003). In numerous studies it has been demonstrated that excess B reduces tree growth and productivity, and contributes to defoliation and leaf injury (Chapman, 1968; Walker et al., 1982; Ayers and Westcot, 1989; Parsons and Wheaton, 1996). In our case, although B concentration in STW soil exceeded the range of 18.5 meq l−1 recommended for citrus crops by Asano et al. (2007), the leaf boron concentration was always far below the limit of toxicity for citrus leaves (>260 ppm) (Embleton et al., 1973). This result is probably due, on the one hand, to water and soil pH, since B is assimilated with difficulty in an alkaline medium (Hu and Brown, 1997) and, on the other hand, to the use of high frequency fertigation system which improve N and P fertilization and therefore mitigate B toxicity (Levy and Syvertsen, 2004). Furthermore, the lemon trees of this research are grafted on Macrophyla rootstock which is considered among the most tolerant rootstocks to high soil B concentration (Levy et al., 1993). The foliar nitrogen levels were always in the optimum range considered for citrus trees development (2.4–2.7%) (Parsons and Wheaton, 1996), but an increase in soil nitrate concentration was observed during the experimental period (Table 2), indicating

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F. Pedrero et al. / Agricultural Water Management 109 (2012) 54–60

Table 1 Inorganic analysis, physical–chemical and microbiological characteristics of irrigation water used in secondary treated wastewater (STW) and tertiary treated wastewater (TTW). Each data represents the annual mean of 36 values ± the standard deviation measured on water samples collected during 2005, 2006 and 2007. Irrigation water

2005

−1

Na (meq l ) Cl (meq l−1 ) B (ppm) Ca (meq l−1 ) Mg (meq l−1 ) SAR NO3 (ppm) H2 PO4 /HPO4 (ppm) SO4 (meq l−1 ) EC (dS m−1 ) TDS (mg l−1 ) Thermotolerant coliforms (cfu 100 ml−1 ) E. coli (cfu 100 ml−1 ) Helmints (eggs 10 l−1 )

2006

STW

TTW

16.1 ± 0.4 16.3 ± 3.1 0.7 ± 0.1 6.1 ± 0.6 4.1 ± 0.1 8.1 ± 0.5 4.2 ± 1.2 3.8 ± 0.3 3.4 ± 1.5 3.3 ± 0.5 2060 ± 235 3800 ± 120 820 ± 43 <10

14.5 ± 0.5 13.5 ± 0.6 0.3 ± 0.1 6.1 ± 1.2 4.2 ± 0.4 6.4 ± 0.7 4.8 ± 1.3 4.2 ± 1.2 5.1 ± 0.2 2.2 ± 0.2 754 ± 41 280 ± 21 92 ± 11 <10

* * *

ns ns ns ns ns ns * ** ** **

ns

2007

STW

TTW

14.5 ± 0.4 16.2 ± 0.3 1.1 ± 0.1 5.3 ± 1.3 4.1 ± 0.8 6.7 ± 0.4 3.8 ± 1.3 3.1 ± 0.6 3.6 ± 1.9 2.8 ± 0.2 1589 ± 362 4320 ± 125 1265 ± 150 <10

13.5 ± 0.2 7.6 ± 0.3 0.7 ± 0.1 3.4 ± 1.1 3.3 ± 0.7 7.4 ± 0.8 5.9 ± 1.2 3.0 ± 0.1 5.5 ± 1.7 2.1 ± 0.1 945 ± 54 234 ± 32 78 ± 21 <10

* ** *

ns ns ns ns ns ns * * ** **

ns

STW

TTW

15.9 ± 0.4 15.1 ± 0.5 1.1 ± 0.1 5.1 ± 0.6 3.2 ± 0.1 7.8 ± 0.3 7.5 ± 2.1 4.8 ± 2.1 2.6 ± 0.5 2.8 ± 0.3 1510 ± 254 2240 ± 86 760 ± 70 <10

14.6 ± 0.1 10.7 ± 1.5 0.5 ± 0.1 5.8 ± 0.6 3.2 ± 0.2 6.9 ± 0.5 4.2 ± 1.5 5.5 ± 2.1 4.6 ± 1.9 2.1 ± 0.2 883 ± 110 478 ± 56 45 ± 8 <10

* ** **

ns ns ns ns ns ns * * ** **

ns

Mean content (n = 36). * Statistically significant at P < 0.05 level of significance. ** Statistically significant at P < 0.01 level of significance. Table 2 Chemical ion concentration in the soil of secondary treated wastewater (STW) and tertiary treated wastewater (TTW). The data contain average values derived from all samples collected during 2005, 2006 and 2007. Values represent the mean (n = 48) ± the standard deviation. Soil chemical analysis

Na (meq l−1 ) K (meq l−1 ) Ca (meq l−1 ) Mg (meq l−1 ) B (meq l−1 ) Chlorides (meq l−1 ) Nitrates (meq l−1 ) Sulfates (meq l−1 ) ECe (dS m−1 )

STW

TTW

2005

2006

2007

41.4 ± 7.9 11.1 ± 1.1 36.5 ± 2.3 40.1 ± 2.4 19.2 ± 1.3 58.1 ± 3.1 6.4 ± 0.5 4.3 ± 0.5 1.7 ± 0.1

39.1 ± 9.7 9.1 ± 0.8 35.4 ± 2.5 36.5 ± 3.9 23.9 ± 0.1 62.9 ± 4.7 6.8 ± 0.6 4.6 ± 0.7 1.9 ± 0.1

35.6 ± 5.8 9.7 ± 0.6 34.1 ± 2.4 38.4 ± 2.8 24.9 ± 1.1 71.1 ± 3.5 7.8 ± 0.3 4.2 ± 0.5 2.2 ± 0.2

ns ns ns ns * * *

ns *

2005

2006

2007

29.2 ± 2.7 10.5 ± 2.3 36.4 ± 1.9 34.5 ± 2.2 5.4 ± 0.3 38.8 ± 2.5 5.3 ± 0.4 4.6 ± 1.3 1.1 ± 0.1

29.6 ± 1.6 9.7 ± 1.2 37.8 ± 1.5 35.2 ± 1.1 5.6 ± 0.5 39.4 ± 5.5 5.5 ± 0.2 5.3 ± 0.5 1.3 ± 0.3

31.1 ± 0.5 9.6 ± 1.1 38.7 ± 4.1 36.3 ± 3.2 6.5 ± 0.2 52 ± 6.1 6.9 ± 0.4 5.7 ± 1.1 1.2 ± 0.2

ns ns ns ns * * *

ns ns

Mean content (n = 48). * Statistically significant at P < 0.05 level of significance.

that reclaimed water is a potential source of additional nutrients (Ahmed et al., 2009). This result is in accordance with other researchers who claim that reclaimed water is an important source of nitrogen for citrus trees (Zekri and Koo, 1994; Jimenez-Cisneros, 1995). The characteristic features of irrigation in the Spanish southeast are the predominance of smallholdings, the wide variety of crops grown in one single irrigation zone, and the presence of irrigation channels and drainage ditches where the reclaimed water is mixed with resources from other sources (Iglesias and de Miguel, 2008). In this experiment, it has been shown that the practice

of blending saline reclaimed water with underground water of better quality (TTW) assured a suitable irrigation water which considerably decreased pathogenic hazards. Therefore, the thermotolerant coliforms concentration in STW exceeded the limits of use restriction recommended by the World Health Organization (>1000 cfu 100 ml−1 ) (Cairncross and Mara, 1989) and the U.S. Environmental Protection Agency (>14 cfu 100 ml−1 ) (Asano and Cortuvo, 1998), while the concentration in TTW was always below these limits (Table 1). The thermotolerant coliforms are a subset of coliform bacteria often used to estimate the concentration of E. coli in general, and the presence of E. coli O157:H7 in particular

Table 3 Leaf mineral analysis of secondary treated wastewater (STW) and tertiary treated wastewater (TTW). The data contain average values derived from all samples collected during 2005, 2006 and 2007. Values represent the mean (n = 48) ± the standard deviation. Leaf chemical analysis

STW

TTW

2005 −1

N (mmol kg ) Na (ppm) K (mmol kg−1 ) Ca (mmol kg−1 ) Mg (mmol kg−1 ) B (ppm) Chlorides (mmol kg−1 ) N (mmol kg−1 ) Na (ppm)

1235 6.5 201 2875 354 3.9 18.5 1235 6.5

2006 ± ± ± ± ± ± ± ± ±

72 1.6 46 45 36 1.1 1.2 72 1.6

1217 5.4 230 2810 365 5.1 24.3 1217 5.4

Mean content (n = 48). * Statistically significant at P < 0.05 level of significance.

2007 ± ± ± ± ± ± ± ± ±

92 2.5 40 76 40 1.4 3.1 92 2.5

1226 6.3 225 2910 375 4.9 25.9 1226 6.3

2005 ± ± ± ± ± ± ± ± ±

35 2.3 25 36 20 1.2 1.8 35 2.3

ns ns ns ns ns ns *

ns ns

1935 6.4 212 2765 304 3.7 19.2 1935 6.4

2006 ± ± ± ± ± ± ± ± ±

70 1.4 67 31 33 0.5 1.3 70 1.4

2000 5.3 226 2720 295 4.3 24.4 2000 5.3

2007 ± ± ± ± ± ± ± ± ±

90 0.5 45 70 42 0.5 2.3 90 0.5

2014 6.8 215 2820 325 4.6 24.8 2014 6.8

± ± ± ± ± ± ± ± ±

30 1.3 23 33 28 0.6 1.8 30 1.3

ns ns ns ns ns ns *

ns ns

F. Pedrero et al. / Agricultural Water Management 109 (2012) 54–60

59

Table 4 Quality indexes [titratable acidity (TA), weight, soluble solid content (SSC) and maturity index (SSC/TA ratio)] and microbial quality of lemon fruits obtained from secondary treated wastewater (STW) and tertiary treated wastewater (TTW) in 2006 and 2007. Quality index

Weight (g) SSC (%) TA (%) Maturity index Microorganisms (log cfu/cm2 ) Mesophilic Faecal coliforms Escherichia coli Yeast and moulds

2006

2007

STW

TTW

148.3 ± 20.3 8.5 ± 0.3 7.1 ± 0.3 1.2 ± 0.1

148.2 ± 23.7 8.5 ± 0.2 7.0 ± 0.3 1.2 ± 0.1

ns ns ns ns

5.3 ± 0.1 ≤0.1 ≤0.1 2.2 ± 0.1

5.2 ± 0.4 ≤0.1 ≤0.1 2.2 ± 0.1

ns ns ns ns

STW

TTW

162.8 ± 10.4 8.0 ± 0.3 6.5 ± 0.2 1.23 ± 0.03

136.2 ± 11.2 6.4 ± 0.5 5.9 ± 0.2 1.08 ± 0.05

3.3 ± 0.3 ≤0.1 ≤0.1 2.4 ± 0.2

2.9 ± 0.4 ≤0.1 ≤0.1 2.0 ± 0.2

* * * *

ns ns ns ns

Values are the mean of 100 fruits ± SE. * Statistically significant at P < 0.05 level of significance.

(an enterohemorrhagic strain referenced by Park et al., 1999 and Selma et al., 2007). Regardless of the different microbial load of SWT and TTW irrigation water, the microbial count in soil was always below the recommended range (<1000 cfu 100 ml−1 ) (Cairncross and Mara, 1989) (data not shown). Taking into account that the microbial quality of irrigation water applied in STW and TTW was different, it could be expected that this could have an influence on the microbial quality of lemon fruits obtained from both treatments. However no microbial risk could be associated with the use of wastewater to irrigate lemon trees in any case (Table 4). In Murcia region, the total lemon production obtained using fresh water irrigation is around 55–60 t ha−1 (Perez-Perez et al., 2004; Segura et al., 2006; Domingo et al., 1996). In this way, the yield obtained in TTW treatment was very similar to the values reported in different lemon orchards of this area, using nonpolluted irrigation. However, it is known that citrus yield decreases in direct proportion to soil salinity increases above an EC threshold of 1.4 dS m−1 (Maas, 1993; Maas and Grattan, 1999), and for this reason, the yield obtained in STW was significantly reduced respect to TTW. It is still an open question as to whether yield reduction observed by saline effects is due to an osmotic effect (Cerdá et al., 1990), Cl toxicity (Cole, 1985) or both. In the present case, the increase over time of Cl concentrations observed in the soil was also reflected in the leaf mineral status (Table 3), but the measured leaf concentrations did not exceed the toxic level for citrus (>200 mmol kg−1 ) (Embleton et al., 1973). The absence of visible symptoms, and the leaf concentration of B and Cl below their corresponding toxic levels, would indicate that the yield decrease observed in STW respect to TTW could be attributable to osmotic stress rather than toxicity. The higher values of TA and SSC recorded in STW fruits compared to the values observed in TTW fruits (Table 4) are in accordance with the results of Ahmed et al. (2009). These authors deduced that salinity could increase SSC and TA of lemon fruits due to an increase in phenolic content and other organic acids. Many plants adapt to salt stress by accumulating secondary metabolites, such as sugars, organic acids and proteins in plant cells, which increase quality and marketability of the product. For example, salinity stress increases the sugar and dissolved solids content of tomatoes and melons; increases the content of beneficial antioxidant compounds in strawberries; and increases the oil and lesquerolic acid in lesquerella (Dobrowolski et al., 2008). Recently, studies on citrus trees demonstrated that it is necessary to carefully monitor the concentration of different ions in the plant if reclaimed water irrigation is used in the long term (Pereira et al., 2011). Our data suggest that long-term detrimental effects could be observed if secondary treated wastewater (STW) is used without implementing intermittent leaching practices to prevent the build up in the root zone, or without blending with less

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