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Short-rotation Willow Biomass Plantations Irrigated and Fertilised with Wastewaters 2 Field experiments2.1 Experimental procedures2.1.1 Experimental sites and climatic conditionsExperimental fields were established in Sweden, France, Northern Ireland (UK) and Greece. The experimental sites were located in the vicinity of wastewater treatment plants at Roma on the isle of Gotland (Sweden), at Orchies near to Lille (France), at Culmore near to Londonderry (UK), and at Larissa 300 km north of Athens (Greece), see Fig. 1.
Figure 1. Climatic conditions varied between the four experimental sites (Table 1) and could be characterised as:
Table 1. Due to the different needs of water supply during the growing season wastewater irrigation regimes were adapted for the specific sites. The water requirement to fill the gap up to a theoretical potential evaporation was calculated by using available normal climate variables from the sites. The irrigation requirement figures are given as daily average values for the actual months of growth (Table 2). The range of irrigation in mm/day during the growing season varies considerably between the sites showing an interesting range of extremes. Similarly, plant requirements of macronutrients (N, P, and K) were calculated from empirical data from the Swedish willow projects (Table 3). The average annual production level was estimated from practical and experimental cultivation. Table 2.
Willows were planted on an area of up to 5 hectares at each experimental site. Within the experimental field, sub-plots were placed with different treatments randomised in each of three replicates. The sub-plots were planted with the Svalöf Weibull AB variety "Jorr" (Salix viminalis). Guard rows, to give at least a 20 m buffer zone between each plot, were planted at the same density with the same clone (Sweden and France) or a mixture of S. viminalis clones (Culmore). Guard areas in Greece were left in grass. Three replicates resulted in 12 to 18 plots in total per site. The plot size was 16 m x 25 m = 400 m2 resulting in plot width of 7 double-rows (7 x 2.25 m = 15.75 m) and the length equivalent to a row of some 40 plants. The total number of plants was approximately 600 per plot. The treatment set-up for each site based on a randomised block design is shown in Table 4. The experimental fields at Roma, Culmore and Larissa were all irrigated with municipal wastewater. The wastewater used at Roma was biologically treated in oxidation ponds after primary settling and then stored. At Culmore and Larissa the wastewater was primary effluent taken from the outflow of primary clarifiers. The wastewater at Orchies was a stored industrial effluent from a chicory processing plant. The willow plantations were supplied with wastewater via drip irrigation systems at Larissa, Roma and Orchies and with a low-level sprinkler system at Culmore. The human urine mixture (urine + flush water), used at Roma, was mixed with water before application to the crop. The soil type can be described as follows:
Table 4.
PE = Potential evapotranspiration, i.e. site-specific rate applied according
to historical data on monthly basis. Thus, monthly irrigation rates are equivalent to a
multiple (1, 2 or 3 times) of the corresponding calculated potential evapotranspiration
rate. 2.1.3.1 Water Samples of wastewater and human urine for chemical analyses were taken once a month during the irrigation period for chemical analyses, i.e. 5-9 analyses per year depending on local conditions. Analyses were carried out concerning pH, COD, BOD, Total N, NH4-N, NO3-N, Total P, PO4-P, K, Cu, Zn, Pb, Cd and Cl-. Chemical analyses of pure water and rainwater were performed where applicable. A plastic groundwater pipe with slits was installed centrally in each plot with the bottom end placed below the known or assumed lowest groundwater table. The upper 0.5 m around the pipes was sealed with bentonite (impermeable volcanic clay) or common clay to prevent short-circuit flow of wastewater or rainwater along the pipes. Sampling of superficial groundwater from these pipes was carried out year round bimonthly (BOD, NH4-N, NO3-N, PO4-P and Cl-) or three times per year (pH, COD, Total N, Total P, K, Cu, Zn, Pb, and Cd). Groundwater levels were measured with a plumb bob twice a month during the irrigation periods and once a month rest of the year. 2.1.3.2 Soil Soil sampling, primarily for analyses of nutrients and heavy metals, was carried out at the start (in winter 1998/99) and at the end of the experiment (in winter 2001/02). The objective of this sampling was twofold. First it enabled description and characterisation of soil properties at the four locations and secondly it allowed detection of changes in soil properties due to the various treatments. In each plot 10 soil cores were taken from topsoil (0-30 cm) and subsoil (30-90 cm), respectively. The soil samples were mixed to form pooled samples from each plot and level. The soil was classified according to the FAO-system.
Test pit for soil characterisation at the experimental site at Roma. 2.1.3.3 Vegetation Biomass estimations were based on non-destructive measurements each year during 1999-2001 and a destructive weighing after a final harvest during the final winter, 2001/2002. For non-destructive biomass measurements, the mean weight of 10 randomly selected living stools (plants) in each net plot was estimated. The net plot was allocated as the centre of each plot and had a width of 3 double-rows (6.75 m) and a length of 10 m (15-17 plants), or in total about 100 plants. On each randomly selected stool, the diameter of all living shoots was measured to the nearest tenth of a millimetre using a calliper. The diameter was measured at the 55 cm height of the shoot, i.e. 55 cm along a straight line from the ground at the point where the shoot is attached to the stump, distance "A" in Fig. 2. Dead shoots and shoots shorter than 55 cm were not included. If the shoot was branching below 55 cm, the main shoot that showed apical dominance was measured. Plant shoots shorter than 55 cm were not measured, but still regarded as living. For calculation of the relation between shoot diameter and shoot dry weight an exponential function describing the relationship was established for each location and each year. For this purpose 20-25 shoots were destructively sampled. The diameter was measured at the 55 cm point and the stem cut at 10 cm above the ground surface, corresponding to the cutting height of mechanised harvesters, distance "B" in Fig. 2. The fresh weight of each shoot was determined separately to the nearest tenth of a gram (0.1 g), dried at 85 °C until constant weight occurred (48 to 72 hours), and the dry weight measured. The best data for calculation of the relationship between dry weight and diameter were obtained by (selectively) collecting shoots of all diameter sizes represented in the stand. During the final winter (2001/2002), a final harvest was performed in each plot. All stools in the net plots were included in the harvest and weighed in the field. A representative sample of 20-30 shoots was also weighed separately, oven dried to a constant weight (105 °C, 48 h), and weighed again. The dry matter percentage was calculated and used for calculating the dry matter content of the total harvested biomass. Stems were also sampled for chemical analyses at the final harvest. Chemical analyses of stem biomass were carried out concerning N, P, K, Cu, Zn, Pb, Cd and chloride. Samples were weighed fresh, air-dried a day or two, milled/finely divided, dried in the oven at 85 °C for 3 days. After decomposition with acid solution, the sample solutions were analysed.
Figure 2. 2.2 Results and discussion2.2.1 Operational and technical aspectsSome problems with weeds in the plantations were apparent. Weeds were controlled with herbicides, mechanically or by hand before plantation and during the establishing year. After the first year it was assessed that all field trials were well established. At the start of the second irrigation season at Culmore some difficulty was experienced with problems caused by solid particles in the irrigation system. The amount of suspended solids in the wastewater led on some occasions to clogging of the sprinkler heads. To help overcome this, a filter was installed in line after the pump in the irrigation system. The problems decreased but the spray units still needed to be checked frequently. The following season, a finer steel mesh cylinder was placed around the outside of the filter cylinder. With this system it was only necessary to clean the filter once a week. A water meter was installed in order to monitor the actual irrigation rates. At Larissa no significant difficulties were experienced during the project. However, from December 2000 onwards, an unexpectedly deep water table (below the well bottom) prohibited sampling of the ground water. At Roma problems arose with clogged filters due to sediments and growth of algae. As a result of this, it was decided to invest in a sand filter. Water gauges were installed in order to measure more precisely the water flow. The injection of human urine worked properly during the irrigation season except for some minor problems of mechanical nature. At Orchies parts of the willow plantations suffered from drought and leaf beetle attacks. Occasionally there were also problems with heavy precipitation incidents making the ground in the lower parts of the plantation wet and flooded. The soil was very compact, which decreased the transportation of air and water into the soil horizon. Most of the problems that occurred were normal disturbances for these kind of studies and acceptable within the frame of project. However, the rather poor trial conditions at Orchies, which were not obvious before the project started, limited the delivery of test results from this site. 2.2.2 Application of wastewater and water requirementThe amount of water used for irrigation was initially calculated using the normal climate values from adequate nearby meteorological stations. In order to control the irrigation regimes the evapotranspiration data was continuously updated during the trial period using most recent measured data. Irrigation rates and precipitation at the four sites are shown in Table 5. The plots were only irrigated during the growing season and when the weather was appropriate, i.e. the irrigation was stopped on major rain occasions. The application of wastewater at Orchies was reduced due to limited supply of wastewater and also sometimes due to clogged emitters. For instance, irrigation during 2001 amounted to approximately half the calculated values. Table 5.
Some studies have indicated that the total water loss to the atmosphere (evaporation, interception and transpiration) from a willow biomass plantation could exceed the potential evapotranspiration according to the Penman formula, which refers to a water saturated and mowed grass area (Penman, 1956). For instance, Persson and Lindroth (1994) compared actual water losses from irrigated and well-established willow stands with Penman evapotranspiration. They reported ratios of 0.7-1.0 in the beginning, 1.2-1.6 in the middle and ca 2.0 at the end of the growing season, indicating the importance of water access when the leaf canopy is fully developed. Lindroth and Båth (1999) estimated the difference between actual and potential evapotranspiration to be ca 1 mm/d, resulting in 200 mm per season in southern Sweden. Calculated differences between precipitation and potential evapotranspiration according to Penman gave water surpluses in the western part of south Sweden of ca 200 mm and deficits of about 50 mm for the southeastern part of Sweden. A conclusion was that for optimum biomass growth it could be assumed that a general water deficit, by and large, prevails in normal years in the southern parts of Sweden. Hence, irrigation of plantations would most likely enhance biomass production in that area. It is obvious that the plant water requirement was reached at the sites already after 1 PE wastewater application (except, possibly, for Orchies during 2001) indicating that water availability was probably not a growth limiting factor. 2.2.3 Biomass productionBy the non-destructive method, dry weight of biomass in tonnes per hectare was calculated each year (Fig. 3). According to these estimations the urine treatment (Roma) gave the highest biomass production. At Culmore and Larissa the productivity increased with increased load of wastewater. However, this was not the case at Roma and Orchies where the highest productivity after wastewater irrigation occurred when the load was 2 PE WW. In December 2001 net plot areas were harvested. According to the results from the final harvest at Culmore (Fig. 4), there were fewer differences between treatments compared with the non-destructive estimations (Fig. 3). The 3 PE WW treatment and the sludge treatment produced the largest amounts of biomass, ca 9 t DM/ha/year. These values were similar to those calculated for the estimated biomass production for these treatments. The 1 PE WW treatment and the control treatment resulted in higher values in harvested biomass than estimated by the non-destructive method, while the 2 PE WW treatment and the 1 PE PW treatment showed lower values. In general, however, the number of stools measured becomes much larger at harvest, and these results are likely to be more robust than those derived from the non-destructive method. Similar to the non-destructive estimates, the differences between treatments in biomass were not statistically significant. In general, however, the results indicated that the wastewater and sludge treatments did not confer any significant production advantage to the coppice. The high rainfall values at this field trial indicate that irrigation treatments were unlikely to give plants any advantage in terms of water availability. Any advantage in terms of nutrient availability might not be apparent yet because of the fertile soil at this site. Another harvest cycle with irrigation treatments would help ascertain if treatment differences due to nutrient availability would emerge. On the other hand, it is possible to conclude that the wastewater and sludge treatments have not been detrimental to coppice growth.
Figure 3.
Figure 4. After the first year of growth in Larissa, all the treatments using wastewater for irrigation resulted in significantly higher biomass production than the treatment using pure water for irrigation (Fig. 3). The 2 PE WW treatment was the most productive one and there were statistically significant differences between this treatment and the 1 PE WW and 3 PE WW, respectively. There was no significant difference between the 1 PE WW and the 3 PE WW treatments.
Figure 5. Also after the second year it was found that all wastewater treatments gave biomass production significantly higher than the treatment using pure water for irrigation. The 2 PE WW and 3 PE WW treatments were the most productive ones with statistically significant differences between these treatments and the 1 PE WW. There was no significant difference between the 2 PE WW and 3 PE WW treatments. Also during the third growth period, all wastewater treatments produced significantly higher biomass than the 1 PE PW treatment. The 2 PE WW and 3 PE WW treatments were the most productive ones. There was no significant difference between these treatments. The yield per hectare, based on harvested three-year-old stools, showed that the estimation of dry matter per hectare based on the diameter measurements, slightly overestimated the production of 1 PE WW but underestimated the production of 3 PE WW (Fig. 5). Statistical analysis on the yield the final year, demonstrates that 3 PE WW yielded most (almost 10 t DM/ha/y as annual mean) and 1 PE WW yielded least (ca 5 t DM/ha/y as annual average). No significant difference was found between 1 PE WW and 1 PE PW. The estimated biomass production the first year at Orchies averaged 3.5 t DM/ha. The survival rate was 91.2 %. Due to different factors the biomass production was limited to a rather modest level in spite of the contributions made by irrigation: 7.7 t DM/ha/y was the annual average of all treatments, see Fig. 3. The low biomass production could be the result of poor soil conditions or the documented leaf beetle attacks. The final harvest of the willow plantation was not completed due to difficulties caused by very wet soil conditions. Two different machines were tested but the bearing capacity of the wet soil was unfortunately not sufficient. At Roma the estimated shoot growth during the growing season 2001 was high and exceeded the growth in 2000. On average during 2001, the growth was 11.4 t DM/ha with the lowest growth, 7.5 t DM/ha, in the control treatment and the highest growth, 16.7 t DM/ha, found after the urine treatment, see Fig. 3. The destructive measurement during the final harvest gave similar results to the non-destructive estimations. The urine treatment resulted in an annual production of almost 10 t DM/ha, where as growth after wastewater irrigation gave ca 7 t DM/ha as annual averages (Fig. 6). Control and PW plots produced ca 6 t DM/ha/y.
Figure 6.
Combined harvesting and chipping of a willow plantation in Sweden. The results from the experimental sites clearly indicate that wastewater, human urine and sewage sludge can stimulate willow production substantially. Application of human urine mixture mixed with water to reach expected plant water requirement resulted in the highest willow growth when non-destructive estimation and weighing after harvest are evaluated together. In general, growth results recorded in this study were at higher levels than those of other production results from commercial plantations in Sweden. During the winters 1995/96 and 1996/97, some 1600 hectares of willow plantations in southern Sweden were harvested. The stem growth of the 4-year stands varied between 2 and 15 t DM/ha/y with an average of 4.8 t DM/ha/y (Larsson, 1998). Jonsson (1997) reported an average growth amounting to 6.8 and 7.2 t DM/ha/y during the first rotation period (5 years) and the second rotation period (3 years), respectively, based on data collected from about hundred commercial plantations in the southern parts of Sweden. Jonsson (1997) concluded that the greatest restrictions on growth potential were 1/ frost damage, 2/ water shortage, and 3/ insufficient fertilisation and/or weed control. 2.2.4 Biological attacks in vegetationLeaf damage estimations were carried out in 1999, 2000 and 2001, in the same way and by the same observers, on leaf samples from all four sites. In addition, aphids on stems were scored during year 2000 at the Culmore site. At this site bird activity and flora in the plantations were investigated as well. There were differences between the trials in the four countries in terms of leaf damages and which organisms/conditions causing the damage. Larissa differed from the other sites in that symptoms due to the leaf roll gall midge (Dasineura marginemtorquens) were never observed there. On the other hand, the Larissa plantation had symptoms of abiotic stress, which were not observed in the other trials. The willow leaves at the Larissa site were curling, particularly in 1999, and starting from the margins, the leaves first became chlorotic, then necrotic. Furthermore, leaf size was just half of that at the other sites in 2001. Yet the stem wood production at Larissa was quite high. However, it is unclear if the stress, presumably due to the hot climate/high solar irradiation, might threaten the sustainability of the plantation. The trial at Orchies had extremely high levels of attack by leaf beetles (chrysomelids). At Culmore populations of these beetles also increased to high levels over the three years of study. Rust (Melampsora epitea var. epitea) disease levels were fairly low at all sites. The variety Jorr used in the plantations is partially resistant to leaf rust. The plantations at Larissa and Roma had very low levels of rust. The trial at Roma also had very little of mechanical damage to leaf blades caused by insects and wind.
Curled willow leaves from the experimental site at Larissa. Very few effects of the wastewater/urine/sludge treatments on leaf damages were found. The urine treatment was more attacked by leafhoppers in 2000 and by rust in 2001 than the other treatments at the Roma site. Also, damages to leaf edges were more frequent in the urine treatment in 2001. It is likely that the fertilisation effect of urine is favourable for organisms like herbivorous insects and for a biotrophic fungus, such as leaf rust, since nitrogen often is a limiting factor for their growth. However, since the urine fertilisation is encouraging willow growth as well, there is no reason to avoid urine treatment in commercial applications. For unknown reasons willows on the lowest dose of wastewater had more galling on leaf margins at Orchies in 1999 and more lost leaf area at Culmore in 1999 and 2000. There is no clear explanation why the rust was most frequent on the middle dose of wastewater at Orchies in 2001, compared to other treatments at the site. The result was probably due to the influence of other environmental factors at the site, such as high humidity caused by wet soil conditions. Stress symptoms occurred on all treatments at Larissa, but the symptoms were less severe in the PW treatment. Mites were also less frequent on those leaves. In an additional study at Culmore in 2000, where stem dwelling aphids (Pterocomma salicis) were quantified, it was found that irrigated plots had fewer aphids. In this case it is likely that the sprinklers used at this site had a disturbing effect on the aphids. 2.2.5 Application and plant uptake of nutrients and metals2.2.5.1 Nutrient and metal application with wastewater and urine Concentrations in applied wastewater and urine of main nutrients (nitrogen (N), phosphorus (P) and potassium (K)) and easy biodegradable organic substances, measured as BOD, were calculated from mean concentrations during each irrigation season (Table 6). The values fluctuated to some extent from one year to another. The wastewater quality at Larissa was fairly normal considering a primary effluent with low impact of storm water. The wastewater at Culmore was also a primary effluent but probably more diluted by rainwater from the existing combined sewer system. The relatively high P content is more difficult to explain. It may be due to a specific industrial effluent connected to the municipal sewer system. The industrial wastewater at Orchies showed high concentrations compared to municipal wastewater quality in general, especially considering K and BOD. The high BOD content probably explains the clogging problems with the irrigation equipment. Table 6.
ND=No data
High-strength effluent from the chicory processing plant at Orchies. The extended biologically treated wastewater at Roma showed a low content of measured wastewater components. A willow plantation filter in this case will serve more as a general polishing step rather than specific wastewater treatment. The chemical composition of urine differed from the other wastewaters to a large extent, particularly with regard to the nutrient content. In order to reach the water requirement, the urine mixture was mixed with pure water and applied in practice at a ratio of about 1:100 – 3:100 (1 – 3 % urine mixture). Due to the different chemical composition of the wastewaters with respect to their origin and/or pre-treatment level and the various water requirements of the willow depending primarily on the climate, the applied amounts of nutrients varied greatly between the experimental sites (Fig. 7 and Table 7). At Larissa the nutrient application with wastewater was higher than the need of the willow stems even at a level of 1 PE WW (cf. the following chapter). At Roma the application of N and P was close to the stem uptake while the K supplied was 2-3 times above the need. The urine treatment resulted in a fairly balanced nutrient application, with the exception of the P amount, which was low. At Culmore, the P application was almost 10 times above the plant requirement while N and K fertilisation were at suitable levels. At Orchies all measured nutrients were above the stem uptake levels. The main part of applied nitrogen was in plant available forms (NO3- an NH4+). A small proportion of the nitrogen was bound to organic material (especially at Larissa). Most of the organic nitrogen should be released and available through mineralisation in coming years (if accumulation in the soil has occurred). In the primary effluent at Culmore only a small part of the phosphorus was in the plant available phosphate form (15-20 %). Potassium exists almost always in dissolved forms in wastewater and thus is readily available to plants. At Orchies and Roma the measured macronutrients in the wastewater and the urine were in readily plant absorbable forms. Figure 7. Table 7.
ND=No data Other required macronutrients, i.e. calcium (Ca), magnesium (Mg) and sulphur (S), exist in municipal wastewater and were in some cases analysed. However, these nutrients are rarely limiting in normal soils and, hence, are not applied as conventional fertilisers. In Table 7 results are shown of all treatments with wastewater and urine regarding measured macronutrients and heavy metals. The wastewaters contained sufficient amounts of nutrients at 1 PE and in most cases the amounts were above the plant requirement. At Roma, however, application of nitrogen with wastewater and phosphorus with the urine mixture seemed lower than the plant requirement (cf. Table 7 and Table 8). Due to a fairly high content of metals, the application rate of 1 PE WW is recommended for long-term use in a plantation. In a sustainable crop system, the nutrient and metal load should not exceed the need and uptake in harvested crop parts. Although willows in general have the ability to extract metals to a large extent, the metal load in these field trials was high, in general higher than the stem uptake (see the following chapter). 2.2.5.2 Nutrient and metal uptake in stem wood Assimilation of N in willow stems varied largely between the sites, from 18 to 73 kg N/ha as annual averages (Table 8). Table 8.
*Below detection level, 4 mg/kgTS (uptake < ~0.040 kg/ha/y) For Roma the N content in stems were relatively low and for Larissa both low and high. For Culmore more typical N levels were found, i.e. ca 50 kg N/ha/y. N concentrations in stem wood were higher after wastewater application than for other treatments at Culmore. At Larissa the 2 PE WW treatment resulted in the highest and 1 PE PW treatment in the lowest N concentrations in stems compared with the other treatments at this site. The low N uptake in harvested willow stems at Roma (ca 25 kg N/ha/y) could possibly be a result of the low content of nitrogen in applied wastewater. Plants supplied with the urine mixture grew best and contained higher amounts of nitrogen (ca 35 kg N/ha/y) than plants from the wastewater irrigation treatments, which could be a result of the high nitrogen content in the urine mixture compared with the biologically pre-treated wastewater. Large variations in the nitrogen content in the Salix crop has been reported earlier. Alriksson (1997), for instance, reported that the distribution of nitrogen varies throughout the plant; of the whole N pool in the plant, 9-57 % was found in the foliage and 13-47 % was found in the root system. This indicates that nitrogen fertilisation should to some extent exceed the stem uptake even when account is taken of part of the N in leaf litter being recycled within the stand. Mean uptake of phosphorus and potassium was in the range of 3-9 kg P/ha/y and 6-27 kg K/ha/y, respectively. Ericsson (1981) reported ratios of P/N = 0.13 and K/N = 0.65 for optimum cell growth for various Salix clones. Average P/N- and K/N-ratios for wastewater-irrigated willows found here amounted to 0.08 and 0.15 (Larissa), 0.14 and 0.36 (Culmore), and 0.21 and 0.65 (Roma), respectively. These results may indicate some imbalances in terms of nutrition. At Larissa, for instance, both the P and the K content in stem biomass appeared low, even though the nitrogen uptake for some treatments was high (cf. Table 8). Even after irrigation at the lowest rate (1 PE WW), application of P and K appeared to be sufficient, indicating that nutrients should have been available in the soil. The rates of uptake of heavy metals in stem biomass differed substantially between the sites. For lead, the uptake rates were about 50 times higher at Culmore (500-700 g Pb/ha/y) than at Larissa (7-14 g Pb/ha/y), irrespective of treatment. The mobility of Pb in soil is generally low and normal extraction of Pb in willows found elsewhere amounts to 1-30 g/ha/y (Aronsson and Perttu, 1994; Hasselgren, 1999a; Hasselgren, 1999b). There seems to be no rational explanation to this difference other than a possible error in the chemical analysis. With regard to zinc, the Larissa and Roma sites showed values within 200-600 g Zn/ha/y, being below those in Culmore (700-900 g Zn/ha/y). In other investigations were reported uptake ranges in willow plantations of various ages of 300-1700 g Zn/ha/y (Aronsson and Perttu, 1994; Hasselgren, 1999a; Hasselgren, 1999b). For the other metals analysed, the levels were similar at the different sites. The content of Cu and Cd in stems varied within 35-110 g Cu/ha/y and 4-60 g Cd/ha/y, respectively, with values increasing with higher rates of wastewater application. Plants irrigated with PW and control plants averaged 20-50 g Cu/ha/y and 7-63 g Cd/ha/y, indicating that uptake of Cd takes place quite readily from the Cd supply in the soil irrespective of treatment. In other studies, assimilation rates of 4-77 g Cd/ha/y from willows of various shoot age have been reported (Aronsson and Perttu, 1994; Hasselgren, 1999a; Hasselgren, 1999b).
Willow plants irrigated with the urine mixture at the Roma site. No obvious correlations were found between plant uptake of metals and soil or wastewater content of metals. Application of metals with the wastewater to the system exceeded in general the metal uptake of plants after 1 PE WW treatment. For Cu the application was 6 times and twice the withdrawal by plants at Larissa and Roma, respectively, while at Culmore approximately the same amounts were taken up as applied. The application of Zn was 3 times higher than found in harvested stems at Larissa, while the plants took up twice the amount applied at Culmore. At Roma the application of Zn with the wastewater and the urine mixture was fairly well balanced with the plant assimilation of Zn. At Culmore the plants extracted about half of the Cd applied with wastewater. In a sustainable perspective, added nutrients and metals to willow biomass plantations should balance with nutrients and metals removed from the field by the crop, i.e. harvested stem wood after defoliation. Phosphorus, a finite resource necessary for all biological life, is probably the most important parameter to consider in a sustainable wastewater irrigation system. Irrigation with 1 PE WW resulted in P application rates of ca 10 times the plant requirement, according to the results obtained at Larissa and Culmore. At Roma the water need was balanced quite well with the P requirement. Thus, if the system design were to be based on a balanced phosphorus concept, i.e. input of P equivalent to stem biomass extraction of P, the conditions for sustainable metal balances over the soil-plant system would improve. 2.2.5.3 Impact on soil and groundwater 2.2.5.3.1 Soil The final soil analysis at Larissa showed that the NO3-concentration in the surface and deeper soil layers from the 3 PE WW treatment was lower than from the 1 PE WW and 2 PE WW treatments, possibly because anoxic conditions due to high irrigation rates prohibit the transformation of NH4 to NO3. The potassium concentration was slightly higher at the end of the experiment both in surface and deep soil. There was a slight decrease of the pH levels at the end of the experiment in comparison to the beginning. The soil at Orchies consisted predominantly of clay with an acid tendency and a low CaCO3 reserve. Throughout the experiment the content of NH4 increased, while the content of NO3 decreased. This evolution was probably due to the high organic load from the wastewater resulting in temporary oxygen deficiency in the soil. The stock of nitrogen decreased sharply with the depth, which means that nitrogen could have leached to the saturated zone. The total N content was stable throughout the trial period but the NO3 content was low in some cases (less than 1 mg/kg). This could have contributed to the relatively poor willow growth at Orchies. Over the period, the values of other major elements were rather homogeneous. The organic material content increased, but was low compared to the desirable value for this kind of soil. At Roma interesting observations were noted for some of the metals. Wastewater irrigation had little or no effect on the Cu content of the soil. However, control treatment and 1 PE PW treatment resulted in a decrease of the Cu content. A similar result was seen for the Zn content. The decrease of Zn was approximately 10-30 %. Pb generally decreased for all treatments. Still, the major decrease occurred for the non-irrigated plots. However, Cd increased up to twice the initial value in all treatments, which indicates that the Cd analyses were less reliable in this case. The pH level increased by around 0.5 units in the upper 30 cm soil layer and 0.8 units in the lower 30-90 cm layer. After treatment both nitrogen and potassium decreased. This was true for all treatments and both depths. For the upper soil layer an increase in organic matter was observed after wastewater irrigation. The result of the preliminary soil analysis from Culmore showed an acid soil with a high content of organic matter and, probably related to this, a high level of total N, particularly in the upper 0-30 cm layer. This could be expected since the site was previously used as grass-land. Organic matter decreased, in general, according to the final soil analysis, regardless of irrigation treatment, while total N values increased, except under the 3 PE wastewater treatment. The levels of several of the metals (Cu, Mn, Fe and Co) measured rose over the course of the experiment and these increases did not appear to be influenced by the irrigation treatment. Zn values rose in the 3 PE WW treatment, but did not change much under the other treatments. Levels of ammonium lactate extractable K generally increased under wastewater treatments and decreased under the control and pure water irrigation treatments. Total P levels rose in the 0-30 cm soil horizon and decreased further down the soil profile, regardless of treatment. Ammonium lactate extractable P decreased, throughout the soil profile, in all treatments over the course of the experiment. The soil impact in general was low and more or less independent of applied wastewater rates. However, changes in soil chemistry could take place in a long-term perspective. The decrease of metals (Cu, Zn and Pb) at the test site at Roma shows that willow has a capacity to remove certain metals from the soil. Other investigations have clearly shown that willow plantations are effective for soil remediation and are also commercially used for this purpose (e.g. Bertholdsson, 2001, and Greger and Landberg, 1999). 2.2.5.3.2 Groundwater Sampling of superficial groundwater was, in general, carried out according to the description in the ‘Experimental procedure’ section. At Larissa, however, sampling was possible only during the first half of the 3-year test period due to extremely low groundwater levels (> 7 m below the ground surface and below the sampling well bottom) during the rest of the period. At Culmore, generally lower concentrations of BOD and COD in superficial groundwater samples were recorded during the wintertime, probably due to dilution with rainwater. A general pattern was that the groundwater quality was not specifically affected by the different treatments compared with the control (Table 9). However, the Cl--concentration increased with wastewater application. Throughout the experiment concentrations of metals were low. Also nutrient concentrations were low in treated plots and comparable to concentrations in control plots, even though a relatively high content of N, especially as nitrate N, was detected in the 1 PE WW treatment. This may have been a result of the lower N uptake (relatively low biomass yield) after this treatment. Also, the lower wastewater application may have facilitated favourable conditions for nitrification and less favourable conditions for denitrification. Further, in most soils nitrate ions in general have a higher mobility than ammonia ions (e.g. USEPA, 1981). Generally, the 1 PE PW treatment resulted in the lowest impact on groundwater and was comparable to the control. However, the groundwater concentrations after pure water irrigation were in general higher than the concentrations of applied pure water, indicating downwards transport of constituents remaining in the soil from previous activities. Table 9.
ND=No data At Larissa, the groundwater quality appeared to be unaffected by wastewater irrigation in terms of BOD and K (Table 10). The nitrogen and phosphorus fractions, as well as the content of COD, Zn and Cu, increased to a certain extent compared with the pure water treatment. The groundwater table fluctuated between 3 and 7 m below the soil surface, resulting in a fairly deep unsaturated zone with good prerequisites for removal of various wastewater constituents. Similar to the results at Culmore, the concentration of Total N decreased with increased wastewater irrigation, probably as a result of more favourable conditions for nitrification/denitrification. The relatively low chloride content in applied wastewater resulted in similar concentrations in superficial groundwater independent of treatment. The wastewater used for irrigation at Roma was well pre-treated and resulted in very low concentrations of all measured parameters in the groundwater zone (Table 11). However, the Cl-concentration increased to some extent with wastewater application rates. Despite the relatively high N application with urine, this treatment did not affect the groundwater quality. Metal concentrations in superficial groundwater beneath the urine treatment plots were lower compared to the wastewater treatments as well as control and pure water treatments. This is likely to be a result of low content of metals in applied urine mixture (cf. Table 8). 2.2.5.4 Calculation of wastewater treatment effects Eutrophying components (N and P) and oxygen demanding organic material (BOD) are problematic constituents if untreated wastewater is discharged to surface waters or ground water aquifers. Traditional wastewater treatment processes could to some extent be replaced by a soil-plant system exemplified in this study by wastewater irrigation of biomass willow plantations. A soil-plant system may be defined as a "natural" physical-biological-chemical reactor including the following main active parts and processes (Hasselgren, 1992): Table 11.
As indicated in the previous chapter, willow plantations have the potentials to treat pre-treated wastewater to a high quality. For a conventional "closed" wastewater treatment process or system, it is easy to analyse and evaluate the treatment or purification efficiency, since a well defined outlet, normally in terms of a pipe from the unit, collects and discharges the same wastewater volume as was put in. In an "open" wastewater irrigation system with a diffuse outlet following natural drainage it is necessary to measure and/or calculate the terms of the water budget in order to evaluate the treatment efficiency. Precipitation could be explicitly collected and measured according to normal procedures, but for evapotranspiration and percolation indirect measures are needed as a base for calculation of the treatment effects. Since chloride is not adsorbed or released by most soils and is taken up by plants only to a small extent, it is often used as a tracer for calculation of the leaching fraction of water in irrigation design (e.g. Salameh Al-Jamal et al., 1997; Pratt et al., 1978).
Sampling of superficial groundwater for chemical analysis. Consequently, the chloride ion was used in an attempt to compensate for the dilution effect of rainwater and possible influence of surrounding groundwater with regard to analysed superficial groundwater. Mass balance calculations over the soil-plant system concerning N, P and BOD were exemplified with data from the experiment at Culmore (Table 12). With knowledge of the total amounts applied to the plots (wastewater + precipitation) calculated evapotranspiration, calculated amounts of percolated wastewater in superficial groundwater beneath the plots, and assimilated amounts in Salix stems, rest terms in the mass balances are given concerning uptake in plant foliage and roots (N and P), accumulation or degradation in the soil (N, P and BOD), and denitrification of nitrogen from the plots (N losses in terms of N2 and N20). Correction was made for percolated amounts from control plots (non-irrigated and non-fertilised plots) to better reflect the impact of wastewater, as indicated with "corr" in the table. Thus, calculated percolated amounts in superficial groundwater from the control treatment were subtracted from the calculated amounts found in superficial groundwater samples from wastewater-irrigated treatments. Removal of BOD from wastewater in soil is generally a result of filtration and microbiological activity in the soil profile. In well-aerated and well-drained systems, the degradation is easily sustained via aerobic microbes. Irrigation activities give altered saturated and drained conditions creating possibilities for anaerobic and anoxic (denitrification) degradation of the organic material as well. However, at some of the sites irrigation with 2 and 3 PE seemed to cause more or less constant anaerobic conditions which limited the degradation of organic matter and transformation of NH4 to NO3. This can also be a result of a high soil content of silt and clay, which decreases the supply of oxygen. The organic matter in the upper soil horizon increased generally. Table 12.
Volatilization of NH3-N of up to 20 % of applied N amounts, from spray irrigation with alkaline (pH 7.5-8.5) wastewater, was reported by Pettygrove and Asano (1984). In another American investigation 10 % ammonia volatilization at pH 7.8 was reported (USEPA, 1981). Ammoniacal nitrogen evaporation is promoted from soils with low cation exchange capacities (CEC:s), mainly sandy or organic soils (USEPA, 1981). The soil profiles in this study were mainly silty/clayey so it could be assumed that the NH3 volatilization rate was limited from the experimental plots. Phosphate ions not taken up by plants could be sorbed to oxides or hydroxides of iron and aluminium in clay soils or precipitated with iron, aluminium and calcium in the soil to crystalline mineral forms. In soils unsaturated with P normally small percentages will leach from the system (Wittgren and Hasselgren, 1992). The results clearly indicate that purification of primary effluent in willow plantations could be substantial. A hydraulic load up to three times the evapotranspiration rate from the system did not influence wastewater treatment capacities, as exemplified with data from Culmore. Thus, managing a system with wastewater irrigation according to the water and nutrient requirements of a willow biomass plantation seems possible without negative environmental impacts with regard to oxygen demanding substances and eutrophying components. 2.2.5.5 Sanitary aspects – micro-organism analyses Conventional sewage system reduces the concentration of human enteric pathogens in the treated wastewater reaching a water course but does not exclude later risks. It functions as the first barrier in disease prevention, but does not safeguard populations exposed through direct contact, recreational activities or the one using the recipient as a source for drinking water production. Within this project some questions to be solved in relation to sanitary aspects and potential secondary transmission to humans were related to the site-specific characteristics. These in turn may function as a base for generalised guidelines to be applied when irrigation with wastewater or use of sludge as fertiliser is practised in willow plantations. These questions further apply to a) the optimal supply of wastewater under the prevailing local conditions, b) potential sanitary problems arising in relations to humans, c) environmental problems due to the presence of wildlife. For these assessments a number of faecal indicator organisms were used including the traditional coliforms indicators, supplemented with faecal enterococci, anaerobic sulphite reducing Clostridium perfringens, and the somatic coliphages (bacteria virus). Coliphages are often used as a model virus to indicate transport of human viruses down to the groundwater zone. In addition, analyses were made concerning the bacterial pathogens Salmonella and Campylobacter and the protozoan parasites Giardia and Cryptosporidium. All of these represent organisms that can infect both humans and animals. The potential transmission routes from wastewater for disease causing organisms are through groundwater, surface drainage and aerosols, via animals and direct contact. These possibilities were compared at Roma, Larissa and Culmore. 2.2.5.5.1 Removal in pre-treatment plants At Roma all selected indicator microorganisms and pathogens were analysed in inflow water to the pre-treatment plant. In the untreated wastewater the faecal coliforms and E. coli varied between 4*102 - 3*103 cfu/ml and were reduced, over the pond system, to below the detection limit, between <1 to <0.01 cfu/ml, giving a total reduction of >99.8 to >99.99 % of faecal coliforms and E. coli. The faecal enterococci likewise gave a reduction of >3 log10 (>99.9 %) while the spore forming bacteria (Clostridium perfringens) initially occurred in low values but were still detectable, 0.1-0.5 cfu/ml after treatment. The coliphages had influent concentrations of 1-4*102 cfu/ml and a reduction of 99.8-99.98 % (from below detection limit <0.1 pfu/ml to 0.3 pfu/ml). Salmonella and Campylobacter (analysed semi-quantitatively) were detected just on one occasion between the first and the second pond but never in the effluent used for irrigation. No Giardia or Cryptosporidium were detected in the untreated wastewater (detection limit <2.5/L) or in the treated wastewater used for irrigation (detection limit <0.3/L). In general the treated water used for irrigation had low or undetectable values of both indicator and pathogenic organisms although the wastewater is treated in open ponds, which are also a habitat for birds that could give an input of the organisms to the water. The presence of Salmonella and Campylobacter in the wastewater can be due to either their presence in the wastewater or as a result of the presence of birds. Analyses of untreated and treated wastewater were also carried out at Larissa and Culmore (Figure 8). At Larissa the treated wastewater used for irrigation contained concentrations of E. coli and faecal streptococci (60 to 80 cfu/ml and 10 to 20 cfu/ml, respectively). For coliphages the corresponding values were 150-160 pfu/ml with a treatment reduction efficiency of 99.2-99.8 %. The concentrations of E. coli in the wastewater used for irrigation in Culmore were 4*104 cfu/ml (average of four samplings) and the reduction capacity varied between 12 and 99.8 % over the treatment plant. Also the reduction capacity varied for the Clostridium perfringens, 44–99.1 %, and the coliphages, 75-98.6 %. For coliphages the concentrations varied dramatically between the different sampling occasions, from 30 pfu/ml to 1.9*104 pfu/ml in the pre-treated wastewater used for irrigation.
Figure 8. Salmonella was detected in both the untreated and treated wastewater used for irrigation at both Culmore and Larissa. Campylobacter were not detected at any time. For Giardia and Cryptosporidium the concentrations varied between the different sampling occasions. At Larissa the incoming numbers of Giardia varied between 100/L (100 % DAPI+) to 6326/L (of these 1217/L were DAPI+) and with a reduction of 99.5-99.97 % giving concentrations of 0.5 to 1.7 cysts/L in the water used for irrigation. For Cryptosporidium the numbers were low: in the untreated wastewater <2 to 4.3/L and in the treated wastewater the concentrations were in the same range, indicating no significant reduction over the treatment plant. The corresponding concentrations for these pathogens at Culmore were for Giardia the highest of all treatment plants included in this study. In the untreated wastewater the cysts varied between 24 cysts/L (100 % DAPI+) to 15 000 cysts/L, while in treated wastewater the concentrations varied between <0.7-2805 (39 % DAPI+)/L. The reduction varied from a very low 7 % to 99.5 %. For Cryptosporidium the concentrations in the untreated wastewater were 4 oocysts/L to 104 oocysts/L while the pre-treated wastewater had concentrations of <0.7-42 oocysts/L, giving a reduction capacity of 50-60 %. At Roma, the urine mixture collected from the toilets and the urinal were sampled and analysed for indicator organisms on one occasion, soon after filling up the tank. In the urine mixture, E. coli, faecal streptococci, Clostridium perfringens and the coliphages, were detected in numbers of 90 cfu/ml, 655 cfu/ml, 1400 cfu/ml and 140 pfu/ml, respectively. Salmonella, Campylobacter, Giardia and Cryptosporidium (detection limit <10/L) were not detected in human urine. 2.2.5.5.2 Leakage to groundwater The results from the sampling of superficial groundwater at Roma showed that most of the organisms were below the detection limit, <0.1 cfu/ml (Table 13). The exception was total coliforms which were detected in concentrations of 3-25 cfu/ml. Total coliforms in low numbers can be found naturally in the environment. The low numbers were anticipated at Roma since the pre-treated wastewater was stored prior to irrigation. Due to the low number of organisms no differences could be seen between the different irrigation regimes. The numbers of indicator organisms found in the superficial groundwater at Culmore were consistently much higher than values found at Roma (Fig. 9). The total coliforms showed very high concentrations, indicating a clear groundwater contamination. No clear differences could be seen between the different treatments even though increased numbers of organisms were noticed in the 3 PE WW treatment compared with the 1 PE WW and 2 PE WW treatments. Coliphages are the smallest organisms and more likely to be transported than bacteria, helminth or protozoan parasites. Table 13.
Figure 9. The rate of leakage to the groundwater depends on the soil texture and the distance to the saturated zone. The infiltration capacity is normally higher in a sandy soil compared with clayey soil, which could be an explanation of the relatively low reduction rates at the Culmore site. Further, the unsaturated zone at Culmore was shallow and varied between 0.5 and 2.5 m during the course of the project. In a soil type with larger parts of clay particles cracks can occur in which the water can be transported rapidly. However, the risk of contamination varies with the occurrence of nearby wells. At Culmore sprinklers were used for irrigation whereby creation of aerosols could occur. Transport of aerosols is dependent on the prevailing wind conditions and direction. At the sites, the irrigated field was placed in an open area with no natural barriers against the transport of aerosols. However, the willow vegetation itself could create a barrier if the outer parts are not irrigated. When the plantation is irrigated with wastewater, the animals living in the willow coppice area are exposed to pathogens occurring in the wastewater, with a potential risk of direct infection and/or as carrier of infection to house pets, humans or water reservoirs. Results from analysis of faecal samples from wildlife did not indicate that this would be the case. Still, the observations were few and no final conclusions regarding this aspect could be drawn. In order to reduce or minimise the risk for direct contact with the wastewater or the crop irrigated with the wastewater, the irrigated area can be fenced to keep the public or larger animals out of the field. Another way of reducing the risk is to inform the public that the irrigated field is part of the wastewater treatment system. No public recreational areas should be located near the irrigated field, which also reduces the risk for humans and especially children to come into contact with the wastewater. The distance from the different fields to houses or farms in this study varied between 30 m to 800-1000 m. One way of reducing direct contact, and also aerosols, is to restrict the irrigation and exclude the outer parts of the plantation.
A mature willow stand at the Roma site in mid-summer the third year.
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