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Influence of biogas digestate on density, biomass and community composition of earthworms
Industrial Crops and Products 66 (2015) 206–209
Contents lists available at ScienceDirect
Industrial Crops and Products journal homepage: www.elsevier.com/locate/indcrop
Influence of biogas digestate on density, biomass and community composition of earthworms Barbara Koblenz a,∗ , Sabine Tischer b , Jan Rücknagel a , Olaf Christen a a
University of Halle-Wittenberg, Institute of Agricultural and Nutritional Sciences, Agronomy and Organic Farming, Betty-Heimann-Straße 5, 06120 Halle (Saale), Germany University of Halle-Wittenberg, Institute of Agricultural and Nutritional Sciences, Soil Biogeochemistry, Germany
b
a r t i c l e
i n f o
Article history: Received 10 September 2014 Received in revised form 20 November 2014 Accepted 13 December 2014 Available online 8 January 2015 Keywords: Biogas digestate Slurry Earthworms Community composition Biogas plants
a b s t r a c t In recent years, the increasing number of biogas plants in operation has also led to a considerable rise in fermentative substrates, which are now widely used as agricultural fertilizers. The impact on earthworm fauna of using biogas digestate as a fertilizer has yet to be sufficiently researched. At two different tests sites, the short-term (four months after fertilization) and longer-term (three-year test period) influence of using fermented residues as a fertilizer was examined on earthworm density, biomass and community composition compared to using traditional fertilizers (cattle and pig slurry, chemical fertilizers as well as an unfertilized control). The crop grown was maize (Zea mays L.). Applying biogas digestate and slurry had a positive overall impact at both sites on earthworm density and biomass. Observing different fertilization regimes in the short term, the significantly highest earthworm density was seen where slurry had been applied. In the treatments with digestate and conventional slurry, earthworm biomass differed significantly in comparison with chemical fertilization and the untreated variant. After three years, earthworm biomass in the variants fertilized with conventional slurry and digestate tended to be higher than in the chemical fertilizer and untreated variants. Community composition was strongly influenced by the application of digestate. A decrease in the species Aporrectodea rosea was accompanied by an increase in Aporrectodea caliginosa. The earthworm population was supported equally positively at both sites by the variants with conventional slurry and digestate. © 2014 Elsevier B.V. All rights reserved.
1. Introduction In recent years, the promotion of renewable energy generation has resulted in a significant increase in the importance of agricultural biogas production (Weiland, 2010). In addition to biogas, digestate is a by-product of microbial anaerobic digestion (Bauer et al., 2009). The considerable amounts of digestate that accumulate are now being used more widely as secondary fertilizers in agriculture. So far it has not been possible to make any sufficient statements about the influence of using digestate as a fertilizer on soil quality. The properties of this substance are decisively influenced on the one hand by the anaerobic, microbial fermentation process and on the other by the actual substrates used. Apart from a reduction in the amounts of dry matter caused by the ´ et al., decomposition of easily convertible organic matter (Voca 2005), major changes to its properties include a higher NH4 –N
∗ Corresponding author. Tel.: +49 345 5522603. E-mail address: [email protected] (B. Koblenz). http://dx.doi.org/10.1016/j.indcrop.2014.12.024 0926-6690/© 2014 Elsevier B.V. All rights reserved.
concentration (Möller, 2009). In terms of its properties, therefore, this type of fertilizer is considerably different from conventional cattle and pig slurry. As representatives of macrofauna, earthworms respond very sensitively to various forms of land cultivation, which is why they are used as an important bio-indicator when evaluating soil quality (Paoletti, 1999). Organic as well as chemical fertilizers serve as a food source for earthworms, either directly or indirectly by increasing crop and root residues. Fertilization is thus, also highly important for earthworm activity in addition to tillage, pH value, soil moisture and a site’s weather conditions. In many respects, earthworms have important functions in the agricultural ecosystem. For the most part this involves decomposing organic matter as well as forming stable clay-humus complexes and the establishment of a consistent macro-pore system. Due to the rapid growth of biogas plants there is still a considerable lack of underlying data concerning the influence on earthworm populations of fertilizing soil using digestate. So far there have only been rudimentary attempts at comprehensively studying the impact on earthworms – as a bio-indicator – of fertilizing soil using
B. Koblenz et al. / Industrial Crops and Products 66 (2015) 206–209 Table 1 Main characteristics of cattle slurry and digestate used in the experiment in Cunnersdorf on a dry weight basis for cattle slurry and digestate respectively (idm = in dry matter). Cattle slurry Applied amount[m3 ] Dry matter[%] Total nitrogen[%] NH4 –N[%]
Digestate 86 9.00 0.38 idm 0.21 idm
Applied amount[m3 ] Dry matter[%] Total nitrogen[%] NH4 –N[%]
207
Table 2 Main characteristics of pig slurry manure and digestate used in the experiment in Pfahlheim on a fresh weight basis for pig slurry and digestate respectively (ifm = in fresh matter). Pig slurry
70 4.90 0.42 idm 0.25 idm
digestate (Ernst et al., 2008; Bermejo Domínguez, 2012). In particular, it is still not possible to draw any conclusions about the influence of fertilization with digestate on earthworms after a couple of years. Based on two field trials, this study aims to assess the impact, both in the short term and over a three-year period, on earthworms of applying fermentation residues to soil. The study will also investigate and evaluate the influence of using digestate as a fertilizer compared with using traditional slurry, chemical fertilizers and an untreated control. 2. Materials and methods 2.1. Experimental site 2.1.1. Site 1 Cunnersdorf The field trial in Cunnersdorf (Saxony, 12◦ 13’E, 51◦ 24’N) is located in central Germany at 130–140 m AMSL. The predominant soil texture in the topsoil is a silt loam (90 g kg−1 clay, 330 g kg−1 sand). In terms of soil typology, the test site can be classified as a Stagnic Luvisol. Cunnersdorf has a mean annual temperature of 8.9 ◦ C (1977–2011) and mean annual precipitation of 619 mm (1977–2011). The experimental setup was a one-year, single-factor strip design with four replicates. The crop grown was maize, and the preceding crop was oat. In addition to an unfertilized control variant and a test element fertilized with mineral nitrogen only, the trial investigated raw slurry and fermentation residues. Before the experiment was set up, the land was ploughed to a depth of approximately 25 cm. The slurry was spread on individual test elements on 19.03.2009 using a special method to inject it at a soil depth of approximately 10 cm. As the capacity of the tractor’s slurry tank was not large enough for an entire test element, each test element was driven over twice in immediate succession. It was possible to place the second portion in precisely the area that remained after application of the first portion. Approximately 86 m3 of cattle slurry and 70 m3 of digestate were spread in order to meet nitrogen requirements of about 160 kg ha−1 NH4 –N. The contents of the two agricultural fertilizers are shown in Table 1. The digestate originated from a co-fermentation plant with a secondary fermenter. In addition to cattle slurry, the biogas plant mainly ferments maize. Earthworms were caught on 09.04.2009. The maize was drilled five weeks after spreading the slurry (22.04.2009). 2.1.2. Site 2 Pfahlheim The test site Pfahlheim is located 484 m AMSL in southern Germany in the foothills of the Swabian Jura. The soil type is a Luvisol. Based on the particle size composition, the soil texture in the Ap horizon can be described as a silt loam (244 g kg−1 clay, 146 g kg−1 sand). On average this site has 840 mm of precipitation each year as well as an annual average temperature of 7.7 ◦ C. The experiment was set up back in 2007 using a fully randomized, single-factor block design with four replications, and from the very beginning conservation tillage was employed down to a soil depth of 15 cm. The raw slurry and digestate were spread using drag hoses on 23.04.2009. With 30 m3 each of digestate and pig slurry, 130 kg N were spread per hectare. Next a tooth cultivator was used
Applied amount[m3 ] pH value Total nitrogen[%] NH4 –N[%]
Digestate 30 8.3 0.7 ifm 0.5 ifm
Applied amount[m3 ] pH value Total nitrogen[%] NH4 –N[%]
30 8.6 0.8 ifm 0.5 ifm
to carefully incorporate the organic fertilizers into the ground. The amount of chemical fertilizer used was equivalent to 165 kg N per hectare; it was applied by adding two doses of calcium ammonium nitrate (65/100 kg ha−1 N). Using the experimental design of the study presented here, the following test elements were examined on 28.05.2009 during the third year of the experiment: ‘untreated control variant’, ‘chemical fertilizer (calcium ammonium nitrate)’, ‘pig slurry’ and ‘digestate’. The digestate used in the study came from a co-fermentation plant. There was no secondary fermenter at the time of sampling. The substrates used in this biogas plant include renewable raw materials such as maize and grass silage as well as cereal grain. Poultry dung is also fed into the biogas plant to be used as an additional fermentation substrate. More information about the properties of the organic fertilizers is listed in Table 2. 2.2. Sampling and analysis Earthworms were caught approximately four weeks after fertilization (ISO 23611-1 (2006)) using a combination of hand-sorting to a depth of 30 cm and a subsequent extraction of deeper living earthworms by formaldehyde solution. By combining the ethological, active method of formalin extraction with the passive method of hand-sorting, it is possible to significantly increase the effectiveness of catching worms on arable land (Terhivuo, 1982). The total sampling area covered 1 m2 ; a metal ring 0.125 m2 in size was used to demarcate the border of the sampling area. Then a block of soil approximately a spade deep was removed from the demarcated area and searched for earthworms. Afterwards approximately 2 l of a 0.2% formalin solution were poured into the hole in the ground. Any earthworms that emerged were collected, washed in water for approximately 20 min and preserved in jars of ethanol. The test was performed with eight replicates per variant. The sampling areas were selected in such a way that they proportionately contained different aspects of one plot (row of maize and the space between two rows of maize). Species identification followed Sims and Gerard (1985). For each variant the parameters ‘abundance’ [individuals m−2 ], ‘biomass’ [g m−2 ] and ‘species dominance’ [%] were defined. 2.3. Statistical analyses The earthworm abundance and biomass results between earthworm populations in each variant were statistically verified using the distribution-free Mann-Whitney test (U-test) (Kasuya, 2001). Significant values (p < 0.05) are represented by different letters. 3. Results 3.1. Site 1 Cunnersdorf No significant differences in earthworm abundance and biomass were observed in the variants with raw slurry and digestate at the Cunnersdorf site (Table 3). In these two organic fertilizers, significantly higher biomasses were identified in comparison with the remaining variants. Overall abundance (adult and juvenile specimens) was also higher in the test elements with raw slurry and
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B. Koblenz et al. / Industrial Crops and Products 66 (2015) 206–209
Table 3 Influence of different treatments on earthworm numbers m−2 and biomass (g m−2 ) in Cunnersdorf, sampled on 09.04.2009. Different letters indicate significant differences between the treatments according to Mann–Whitney test (P < 0.05). Treatment
Control
Mineral fertilizer
Cattle slurry
Digestate
Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total
Abundance [indiv. fresh weight m−2 ]
Earthworm biomass [g m−2 ]
35 11 46 b 32 6 38 b 72 14 86 a 62 14 76 ab
12.76 7.87 20.63 b 7.69 4.37 12.06 b 21.69 15.42 37.11 a 18.05 11.22 29.27 a
ical fertilizer and untreated variants (Table 4). At 46.1 g m−2 , the test element with raw slurry displayed the greatest biomass. The unfertilized variant returned the highest abundance, at 77 ind m−2 . This is attributable to the high number of A. rosea. The variants with raw slurry and digestate had 57 and 60 ind m−2 , respectively. Analyzing the ecological groups, it can be seen that each variant contained mostly endogeic earthworms as well as one anecic species in the form of L. terrestris. The absence of epigeic life forms can be explained by the lack of leaf litter on arable land. Essentially the species observed (A. caliginosa, Aporrectodea nocturna, A. rosea, A. chlorotica and L. terrestris) are typical of arable land. The range of species was the same in all of the variants investigated. However, there were considerable differences in terms of dominance structure (Table 5). There tended to be less individuals of the species A. rosea observed in the variants with raw slurry and digestate. In contrast, there were increased levels of A. caliginosa.
Letters a and b indicate significant differences between the treatments.
4. Discussion Table 4 Influence of different treatments on earthworm numbers m−2 and biomass (g m−2 ) in Pfahlheim, sampled on 28.05.2009. Different letters indicate significant differences between the treatments according to Mann–Whitney test (P < 0.05). Treatment
Control
Mineral fertilizer
Pig slurry
Digestate
Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total
Abundance [indiv. fresh weight m−2 ]
Earthworm biomass [g m−2 ]
45 32 77 a 21 22 43 a 26 31 57 a 27 33 60 a
17.26 23.69 40.95 a 7.50 23.50 31.00 a 9.45 36.69 46.14 a 12.57 31.56 44.13 a
digestate. The variant with raw slurry (86 ind m−2 ) was the only one to differ significantly from the unfertilized and chemical variants (46 ind m−2 and 38 ind m−2 respectively). Compared with digestate, there was a tendency for fertilization with unfermented slurry to yield comparatively higher earthworm abundance and biomass. The determination of age ranges resulted in far more juvenile earthworms than adults per square meter in all of the variants examined. A total of four species were recorded (Aporrectodea caliginosa, Aporrectodea rosea, Allolobophora chlorotica and Lumbricus terrestris) from three genera. The species A. caliginosa and L. terrestris were present in all of the fertilization variants. Accounting for more than 50% in each variant, the most dominant species was the mineral soil dweller A. caliginosa. As regards community composition (Table 5), spreading digestate points to a decrease in the endogeic species A. rosea. Furthermore, the proportions of each ecological group in the total population change. Evidence of the species A. chlorotica is only present in the raw slurry variant. The species A. caliginosa and L. terrestris are the only ones present in the digestate variant. 3.2. Site 2 Pfahlheim Earthworm abundance and biomass were examined in the variants ‘unfertilized’, ‘chemical’, ‘raw slurry’ and ‘digestate’. Overall there are no statistically reliable differences between these variants. At the Pfahlheim site, earthworm biomass tended to be higher in the test elements with raw slurry and digestate than in the chem-
In the field trials, the differentiation that occurred among earthworm populations from various fertilization variants largely confirms previous results from the literature, which show that an organic fertilizer has a far more positive impact on earthworms than a chemical fertilizer or an untreated control can (Edwards and Lofty, 1982; Whalen et al., 1998; Timmerman et al., 2006; Ulrich et al., 2010). Organic fertilizers provide earthworms with an immediate, large supply of organic matter (Unwin and Lewis, 1986; Timmerman et al., 2006), while chemical fertilizers indirectly supply organic matter more slowly in the form of crop and root residues. At the Cunnersdorf site as well as Pfahlheim, there were no statistically reliable differences between conventional slurry and digestate when comparing earthworm abundance. Both organic fertilizers had an equally positive influence on earthworm population. In Pfahlheim, three years after the study began the earthworm population tended to have been increased by both organic fertilizers, although there are different. The raw slurry included pig manure and the digestate based on poultry dung, maize, grass silage and cereal corn which have an effect on several properties of the organic fertilizers such as dry matter content and nutritional values. At the Cunnersdorf site, significant differences can be observed between fertilization with slurry or digestate, on the one hand, and using a chemical fertilizer or the untreated control on the other. Our results are in agreement with the work of Leroy et al. (2007), who also observed a larger earthworm population shortly after applying organic matter. Despite all this, it is surprising that in Cunnersdorf significant differences appeared between the different fertilization treatments just four weeks after fertilization. Investigations by Ernst et al. (2008) do however indicate that fertilization using a conventional slurry results in increased earthworm biomass compared with fermented slurry. During the biogas production process, the substrates’ readily soluble carbon bonds are mostly broken down. Even so, in both experiments the digestate did apparently provide improved nutritional quality and availability for earthworms than was the case with chemical fertilizers and the unfertilized control. The reasons for this seem to be related to biogas plant technology. Especially in stirrer tank digesters, which is where the digestate used in the field trials in Cunnersdorf and Pfahlheim came from, the mixing process causes part of the fresh substrate to end up in the outlet. This in turn provides soil fauna with a food source in the form of readily degradable carbon compounds. As digestate continues to be removed from the fermenter, ´ parts of the active biomass also continue to be discharged (Voca et al., 2005). While these anaerobic organisms do die immediately after being deposited in the fermentation residue storage container, they nevertheless offer earthworms an additional food source in
B. Koblenz et al. / Industrial Crops and Products 66 (2015) 206–209
209
Table 5 Community composition in numbers (%) in different fertilizer treatments at the study site Cunnersdorf (09.04.2009) and Pfahlheim (28.05.2009). Species
Lumbricus terrestris Aporrectodea caliginosa Aporrectodea rosea Allolobophora chlorotica Aporrectodea noctorna
Cunnersdorf
Pfahlheim
Control (%)
Mineral fertilizer (%)
Cattle slurry (%)
Digestate (%)
9.1
33.3
14.3
7.1
81.8
50.0
57.1
9.1
16.7
– –
Mineral fertilizer (%)
Pig slurry (%)
Digestate (%)
6.3
18.2
16.1
12.1
92.9
25.0
22.7
64.5
63.6
14.3
–
53.1
22.7
3.2
3.0
–
14.3
–
3.1
13.6
9.7
6.1
–
–
–
12.5
22.8
6.5
15.2
the form of microbial protein. The relatively high number of earthworms in the unfertilized control in Pfahlheim seems to be that earthworms have been negatively affected through the application of both organic fertilizers. Some studies show these results of slurry and digestate on earthworms (Timmerman et al., 2006; Ernst et al., 2008; Bermejo Domínguez, 2012). The negative effects of slurry are related to its high salt concentration and the occurrence of substances that might be toxic to earthworms (Curry, 1976). In addition, the ammonia content may adversely affect soil dwelling earthworms, especially in the digestate treatment. The high abundances and biomass levels at the Cunnersdorf site might be the result of earthworms having migrated. The experimental setup allowed them to migrate to variants with a more attractive food supply. Hoogerkamp et al. (1983) declare a natural spread rate of between 2 and 15 m per year when earthworms colonise habitats with more favorable living conditions. The earthworm population in Cunnersdorf was characterised by a high number of juvenile specimens. This might indicate a higher reproduction rate. In terms of their dominance structure, both of the sites examined are typical representatives of arable land (Tischer, 2008). At Cunnersdorf, applying digestate resulted in a less varied range of species. Abundance of the endogeic earthworm species A. rosea declines. This contradicts research by Ernst et al. (2008), where something of a decrease in A caliginosa biomass was observed after treatment with digestate. In the variant with digestate, the most common species were A. caliginosa and L. terrestris. As a primary decomposer, the anecic earthworm L. terrestris finds sufficient food in this fertilization variant. The mineral soil dweller A. caliginosa benefits from the higher feeding activity of L. terrestris (Ernst et al., 2009). Overall, the differentiation that occurred in the field trials among earthworm populations from various fertilization variants rather confirms previous results from the literature, which show that an organic fertilizer has a far more positive impact on earthworms than a chemical fertilizer or an untreated control can (Edwards and Lofty, 1982; Whalen et al., 1998; Timmerman et al., 2006; Ulrich et al., 2010). Acknowledgements We thank Prof. Niclas, Dr. Schuster, Dr. Kreuter and M. Fuchs from SKW Stickstoffwerke Piesteritz GmbH as well as Dr. M. Mokry and M. Müller from LTZ Augustenberg for allowing us to use their study area in order to investigate this problem. A.-K. Schmitt, T. Leithold and E. Koblenz are acknowledged for their support during soil fauna sampling.
Control (%)
References Bauer, A., Mayr, H., Hopfner-Sixt, K., Amon, T., 2009. Detailed monitoring of two biogas plants and mechanical solid–liquid separation of fermentation residues. J. Biotechnol. 142, 56–63. Bermejo Domínguez, G., 2012. Agro-ecological aspects when applying the remaining products from agricultural biogas processes as fertilizer in crop production. In: Dissertation. Humboldt-Universität, Berlin. Curry, J.P., 1976. Some effects of animal manures on earthworms in grassland. Pedobiologia 16, 425–438. Edwards, C.A., Lofty, J.R., 1982. Nitrogenous fertilizers and earthworm population in agricultural soils. Soil Biol. Biochem. 14, 515–521. Ernst, G., Müller, A., Göhler, H., Emmerling, C., 2008. C and N turnover of fermented residues from biogas plants in soil in the presence of three different earthworm species (Lumbricus terrestris, Aporrectodea longa, Aporrectodea caliginosa). Soil Boil. Biochem. 40, 1413–1420. Ernst, G., Henseler, I., Felten, D., Emmerling, C., 2009. Decomposition and mineralization of energy crop residues governed by earthworms. Soil Biol. Biochem. 41, 1548–1554. Hoogerkamp, M., Rogaar, H., Eijsackers, H.J.P., 1983. Effect of earthworms on grassland on recently reclaimed polder soils in the Netherlands. In: Satchell, J.E. (Ed.), Earthworm Ecology: from Darwin to Vermiculture. Chapman and Hall, London, pp. 85–105. ISO23611-1, Soil quality - Sampling of soil invertebrates - Part 1 Hand-sorting and formalin extraction of earthworms, 2006. Kasuya, E., 2001. Mann–Whitney U test when variances are unequal. Anim. Behav. 61, 1247–1249. Leroy, B.L.M., Van den Bossche, A., De Neve, S., Reheul, D., Moens, M., 2007. The quality of exogenous organic matter: short-term influence on earthworm abundance. Eur. J. Soil Biol. 43, 196–200. Möller, K., 2009. Influence of different manuring systems with and without biogas digestion on soil organic matter and nitrogen inputs, flows and budgets in organic cropping systems. Nutr. Cycl. Agroecosyst. 84, 179–202. Paoletti, M.G., 1999. The role of earthworms for assessment of sustainability and as bioindicators. Agric. Ecosyst. Environ. 74, 137–155. Sims, W., Gerard, B.M., 1985. Earthworms – Keys and Notes for the Identification and Study of the Species. Brill & Backhuys, London, pp. 171S. Terhivuo, J., 1982. Relative efficiency of hand-sorting, formalin application and combination of both methods in extracting Lumbricidae from finnish soils. Pedobiologia 23, 175–188. Timmerman, A., Bos, D., Ouwehand, J., de Goede, R.G.M., 2006. Long-term effects of fertilisation regime on earthworm abundance in a semi-natural grassland area. Pedobiologia 50, 427–432. Tischer, S., 2008. Lumbricidae communities in soil monitoring sites differently managed and polluted with heavy metals. Pol. Ecol. 56 (4), 635–646. Unwin, R.J., Lewis, S., 1986. The effect upon earthworm population of very large applications of pig slurry to grassland. Agric. Waste 16, 67–73. Ulrich, S., Tischer, S., Hofmann, B., Christen, O., 2010. Biological soil properties in a long-term tillage trial in Germany. J. Plant Nutr. Soil Sci. 173, 483–489. ´ ´ N., Kriˇcka, T., Cosi ´ T., Rupic, ´ V., Jukic´ Zˇ , Kalambura, S., 2005. Digested Voca, c, residues as a fertilizer after the mesophilic process of anaerobic digestion. Plant Soil Environ. 51, 262–266. Weiland, P., 2010. Biogas production: current state and perspectives. Appl.Microbiol. Biotechnol. 85, 849–860. Whalen, J.K., Parmelee, R.W., Edwards, C.A., 1998. Population dynamics of earthworm communities in corn agroecosystems receiving organic or inorganic fertilizers amendments. Biol. Fertil. Soils 27, 400–407.
Contents lists available at ScienceDirect
Industrial Crops and Products journal homepage: www.elsevier.com/locate/indcrop
Influence of biogas digestate on density, biomass and community composition of earthworms Barbara Koblenz a,∗ , Sabine Tischer b , Jan Rücknagel a , Olaf Christen a a
University of Halle-Wittenberg, Institute of Agricultural and Nutritional Sciences, Agronomy and Organic Farming, Betty-Heimann-Straße 5, 06120 Halle (Saale), Germany University of Halle-Wittenberg, Institute of Agricultural and Nutritional Sciences, Soil Biogeochemistry, Germany
b
a r t i c l e
i n f o
Article history: Received 10 September 2014 Received in revised form 20 November 2014 Accepted 13 December 2014 Available online 8 January 2015 Keywords: Biogas digestate Slurry Earthworms Community composition Biogas plants
a b s t r a c t In recent years, the increasing number of biogas plants in operation has also led to a considerable rise in fermentative substrates, which are now widely used as agricultural fertilizers. The impact on earthworm fauna of using biogas digestate as a fertilizer has yet to be sufficiently researched. At two different tests sites, the short-term (four months after fertilization) and longer-term (three-year test period) influence of using fermented residues as a fertilizer was examined on earthworm density, biomass and community composition compared to using traditional fertilizers (cattle and pig slurry, chemical fertilizers as well as an unfertilized control). The crop grown was maize (Zea mays L.). Applying biogas digestate and slurry had a positive overall impact at both sites on earthworm density and biomass. Observing different fertilization regimes in the short term, the significantly highest earthworm density was seen where slurry had been applied. In the treatments with digestate and conventional slurry, earthworm biomass differed significantly in comparison with chemical fertilization and the untreated variant. After three years, earthworm biomass in the variants fertilized with conventional slurry and digestate tended to be higher than in the chemical fertilizer and untreated variants. Community composition was strongly influenced by the application of digestate. A decrease in the species Aporrectodea rosea was accompanied by an increase in Aporrectodea caliginosa. The earthworm population was supported equally positively at both sites by the variants with conventional slurry and digestate. © 2014 Elsevier B.V. All rights reserved.
1. Introduction In recent years, the promotion of renewable energy generation has resulted in a significant increase in the importance of agricultural biogas production (Weiland, 2010). In addition to biogas, digestate is a by-product of microbial anaerobic digestion (Bauer et al., 2009). The considerable amounts of digestate that accumulate are now being used more widely as secondary fertilizers in agriculture. So far it has not been possible to make any sufficient statements about the influence of using digestate as a fertilizer on soil quality. The properties of this substance are decisively influenced on the one hand by the anaerobic, microbial fermentation process and on the other by the actual substrates used. Apart from a reduction in the amounts of dry matter caused by the ´ et al., decomposition of easily convertible organic matter (Voca 2005), major changes to its properties include a higher NH4 –N
∗ Corresponding author. Tel.: +49 345 5522603. E-mail address: [email protected] (B. Koblenz). http://dx.doi.org/10.1016/j.indcrop.2014.12.024 0926-6690/© 2014 Elsevier B.V. All rights reserved.
concentration (Möller, 2009). In terms of its properties, therefore, this type of fertilizer is considerably different from conventional cattle and pig slurry. As representatives of macrofauna, earthworms respond very sensitively to various forms of land cultivation, which is why they are used as an important bio-indicator when evaluating soil quality (Paoletti, 1999). Organic as well as chemical fertilizers serve as a food source for earthworms, either directly or indirectly by increasing crop and root residues. Fertilization is thus, also highly important for earthworm activity in addition to tillage, pH value, soil moisture and a site’s weather conditions. In many respects, earthworms have important functions in the agricultural ecosystem. For the most part this involves decomposing organic matter as well as forming stable clay-humus complexes and the establishment of a consistent macro-pore system. Due to the rapid growth of biogas plants there is still a considerable lack of underlying data concerning the influence on earthworm populations of fertilizing soil using digestate. So far there have only been rudimentary attempts at comprehensively studying the impact on earthworms – as a bio-indicator – of fertilizing soil using
B. Koblenz et al. / Industrial Crops and Products 66 (2015) 206–209 Table 1 Main characteristics of cattle slurry and digestate used in the experiment in Cunnersdorf on a dry weight basis for cattle slurry and digestate respectively (idm = in dry matter). Cattle slurry Applied amount[m3 ] Dry matter[%] Total nitrogen[%] NH4 –N[%]
Digestate 86 9.00 0.38 idm 0.21 idm
Applied amount[m3 ] Dry matter[%] Total nitrogen[%] NH4 –N[%]
207
Table 2 Main characteristics of pig slurry manure and digestate used in the experiment in Pfahlheim on a fresh weight basis for pig slurry and digestate respectively (ifm = in fresh matter). Pig slurry
70 4.90 0.42 idm 0.25 idm
digestate (Ernst et al., 2008; Bermejo Domínguez, 2012). In particular, it is still not possible to draw any conclusions about the influence of fertilization with digestate on earthworms after a couple of years. Based on two field trials, this study aims to assess the impact, both in the short term and over a three-year period, on earthworms of applying fermentation residues to soil. The study will also investigate and evaluate the influence of using digestate as a fertilizer compared with using traditional slurry, chemical fertilizers and an untreated control. 2. Materials and methods 2.1. Experimental site 2.1.1. Site 1 Cunnersdorf The field trial in Cunnersdorf (Saxony, 12◦ 13’E, 51◦ 24’N) is located in central Germany at 130–140 m AMSL. The predominant soil texture in the topsoil is a silt loam (90 g kg−1 clay, 330 g kg−1 sand). In terms of soil typology, the test site can be classified as a Stagnic Luvisol. Cunnersdorf has a mean annual temperature of 8.9 ◦ C (1977–2011) and mean annual precipitation of 619 mm (1977–2011). The experimental setup was a one-year, single-factor strip design with four replicates. The crop grown was maize, and the preceding crop was oat. In addition to an unfertilized control variant and a test element fertilized with mineral nitrogen only, the trial investigated raw slurry and fermentation residues. Before the experiment was set up, the land was ploughed to a depth of approximately 25 cm. The slurry was spread on individual test elements on 19.03.2009 using a special method to inject it at a soil depth of approximately 10 cm. As the capacity of the tractor’s slurry tank was not large enough for an entire test element, each test element was driven over twice in immediate succession. It was possible to place the second portion in precisely the area that remained after application of the first portion. Approximately 86 m3 of cattle slurry and 70 m3 of digestate were spread in order to meet nitrogen requirements of about 160 kg ha−1 NH4 –N. The contents of the two agricultural fertilizers are shown in Table 1. The digestate originated from a co-fermentation plant with a secondary fermenter. In addition to cattle slurry, the biogas plant mainly ferments maize. Earthworms were caught on 09.04.2009. The maize was drilled five weeks after spreading the slurry (22.04.2009). 2.1.2. Site 2 Pfahlheim The test site Pfahlheim is located 484 m AMSL in southern Germany in the foothills of the Swabian Jura. The soil type is a Luvisol. Based on the particle size composition, the soil texture in the Ap horizon can be described as a silt loam (244 g kg−1 clay, 146 g kg−1 sand). On average this site has 840 mm of precipitation each year as well as an annual average temperature of 7.7 ◦ C. The experiment was set up back in 2007 using a fully randomized, single-factor block design with four replications, and from the very beginning conservation tillage was employed down to a soil depth of 15 cm. The raw slurry and digestate were spread using drag hoses on 23.04.2009. With 30 m3 each of digestate and pig slurry, 130 kg N were spread per hectare. Next a tooth cultivator was used
Applied amount[m3 ] pH value Total nitrogen[%] NH4 –N[%]
Digestate 30 8.3 0.7 ifm 0.5 ifm
Applied amount[m3 ] pH value Total nitrogen[%] NH4 –N[%]
30 8.6 0.8 ifm 0.5 ifm
to carefully incorporate the organic fertilizers into the ground. The amount of chemical fertilizer used was equivalent to 165 kg N per hectare; it was applied by adding two doses of calcium ammonium nitrate (65/100 kg ha−1 N). Using the experimental design of the study presented here, the following test elements were examined on 28.05.2009 during the third year of the experiment: ‘untreated control variant’, ‘chemical fertilizer (calcium ammonium nitrate)’, ‘pig slurry’ and ‘digestate’. The digestate used in the study came from a co-fermentation plant. There was no secondary fermenter at the time of sampling. The substrates used in this biogas plant include renewable raw materials such as maize and grass silage as well as cereal grain. Poultry dung is also fed into the biogas plant to be used as an additional fermentation substrate. More information about the properties of the organic fertilizers is listed in Table 2. 2.2. Sampling and analysis Earthworms were caught approximately four weeks after fertilization (ISO 23611-1 (2006)) using a combination of hand-sorting to a depth of 30 cm and a subsequent extraction of deeper living earthworms by formaldehyde solution. By combining the ethological, active method of formalin extraction with the passive method of hand-sorting, it is possible to significantly increase the effectiveness of catching worms on arable land (Terhivuo, 1982). The total sampling area covered 1 m2 ; a metal ring 0.125 m2 in size was used to demarcate the border of the sampling area. Then a block of soil approximately a spade deep was removed from the demarcated area and searched for earthworms. Afterwards approximately 2 l of a 0.2% formalin solution were poured into the hole in the ground. Any earthworms that emerged were collected, washed in water for approximately 20 min and preserved in jars of ethanol. The test was performed with eight replicates per variant. The sampling areas were selected in such a way that they proportionately contained different aspects of one plot (row of maize and the space between two rows of maize). Species identification followed Sims and Gerard (1985). For each variant the parameters ‘abundance’ [individuals m−2 ], ‘biomass’ [g m−2 ] and ‘species dominance’ [%] were defined. 2.3. Statistical analyses The earthworm abundance and biomass results between earthworm populations in each variant were statistically verified using the distribution-free Mann-Whitney test (U-test) (Kasuya, 2001). Significant values (p < 0.05) are represented by different letters. 3. Results 3.1. Site 1 Cunnersdorf No significant differences in earthworm abundance and biomass were observed in the variants with raw slurry and digestate at the Cunnersdorf site (Table 3). In these two organic fertilizers, significantly higher biomasses were identified in comparison with the remaining variants. Overall abundance (adult and juvenile specimens) was also higher in the test elements with raw slurry and
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Table 3 Influence of different treatments on earthworm numbers m−2 and biomass (g m−2 ) in Cunnersdorf, sampled on 09.04.2009. Different letters indicate significant differences between the treatments according to Mann–Whitney test (P < 0.05). Treatment
Control
Mineral fertilizer
Cattle slurry
Digestate
Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total
Abundance [indiv. fresh weight m−2 ]
Earthworm biomass [g m−2 ]
35 11 46 b 32 6 38 b 72 14 86 a 62 14 76 ab
12.76 7.87 20.63 b 7.69 4.37 12.06 b 21.69 15.42 37.11 a 18.05 11.22 29.27 a
ical fertilizer and untreated variants (Table 4). At 46.1 g m−2 , the test element with raw slurry displayed the greatest biomass. The unfertilized variant returned the highest abundance, at 77 ind m−2 . This is attributable to the high number of A. rosea. The variants with raw slurry and digestate had 57 and 60 ind m−2 , respectively. Analyzing the ecological groups, it can be seen that each variant contained mostly endogeic earthworms as well as one anecic species in the form of L. terrestris. The absence of epigeic life forms can be explained by the lack of leaf litter on arable land. Essentially the species observed (A. caliginosa, Aporrectodea nocturna, A. rosea, A. chlorotica and L. terrestris) are typical of arable land. The range of species was the same in all of the variants investigated. However, there were considerable differences in terms of dominance structure (Table 5). There tended to be less individuals of the species A. rosea observed in the variants with raw slurry and digestate. In contrast, there were increased levels of A. caliginosa.
Letters a and b indicate significant differences between the treatments.
4. Discussion Table 4 Influence of different treatments on earthworm numbers m−2 and biomass (g m−2 ) in Pfahlheim, sampled on 28.05.2009. Different letters indicate significant differences between the treatments according to Mann–Whitney test (P < 0.05). Treatment
Control
Mineral fertilizer
Pig slurry
Digestate
Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total Juvenile Adult Total
Abundance [indiv. fresh weight m−2 ]
Earthworm biomass [g m−2 ]
45 32 77 a 21 22 43 a 26 31 57 a 27 33 60 a
17.26 23.69 40.95 a 7.50 23.50 31.00 a 9.45 36.69 46.14 a 12.57 31.56 44.13 a
digestate. The variant with raw slurry (86 ind m−2 ) was the only one to differ significantly from the unfertilized and chemical variants (46 ind m−2 and 38 ind m−2 respectively). Compared with digestate, there was a tendency for fertilization with unfermented slurry to yield comparatively higher earthworm abundance and biomass. The determination of age ranges resulted in far more juvenile earthworms than adults per square meter in all of the variants examined. A total of four species were recorded (Aporrectodea caliginosa, Aporrectodea rosea, Allolobophora chlorotica and Lumbricus terrestris) from three genera. The species A. caliginosa and L. terrestris were present in all of the fertilization variants. Accounting for more than 50% in each variant, the most dominant species was the mineral soil dweller A. caliginosa. As regards community composition (Table 5), spreading digestate points to a decrease in the endogeic species A. rosea. Furthermore, the proportions of each ecological group in the total population change. Evidence of the species A. chlorotica is only present in the raw slurry variant. The species A. caliginosa and L. terrestris are the only ones present in the digestate variant. 3.2. Site 2 Pfahlheim Earthworm abundance and biomass were examined in the variants ‘unfertilized’, ‘chemical’, ‘raw slurry’ and ‘digestate’. Overall there are no statistically reliable differences between these variants. At the Pfahlheim site, earthworm biomass tended to be higher in the test elements with raw slurry and digestate than in the chem-
In the field trials, the differentiation that occurred among earthworm populations from various fertilization variants largely confirms previous results from the literature, which show that an organic fertilizer has a far more positive impact on earthworms than a chemical fertilizer or an untreated control can (Edwards and Lofty, 1982; Whalen et al., 1998; Timmerman et al., 2006; Ulrich et al., 2010). Organic fertilizers provide earthworms with an immediate, large supply of organic matter (Unwin and Lewis, 1986; Timmerman et al., 2006), while chemical fertilizers indirectly supply organic matter more slowly in the form of crop and root residues. At the Cunnersdorf site as well as Pfahlheim, there were no statistically reliable differences between conventional slurry and digestate when comparing earthworm abundance. Both organic fertilizers had an equally positive influence on earthworm population. In Pfahlheim, three years after the study began the earthworm population tended to have been increased by both organic fertilizers, although there are different. The raw slurry included pig manure and the digestate based on poultry dung, maize, grass silage and cereal corn which have an effect on several properties of the organic fertilizers such as dry matter content and nutritional values. At the Cunnersdorf site, significant differences can be observed between fertilization with slurry or digestate, on the one hand, and using a chemical fertilizer or the untreated control on the other. Our results are in agreement with the work of Leroy et al. (2007), who also observed a larger earthworm population shortly after applying organic matter. Despite all this, it is surprising that in Cunnersdorf significant differences appeared between the different fertilization treatments just four weeks after fertilization. Investigations by Ernst et al. (2008) do however indicate that fertilization using a conventional slurry results in increased earthworm biomass compared with fermented slurry. During the biogas production process, the substrates’ readily soluble carbon bonds are mostly broken down. Even so, in both experiments the digestate did apparently provide improved nutritional quality and availability for earthworms than was the case with chemical fertilizers and the unfertilized control. The reasons for this seem to be related to biogas plant technology. Especially in stirrer tank digesters, which is where the digestate used in the field trials in Cunnersdorf and Pfahlheim came from, the mixing process causes part of the fresh substrate to end up in the outlet. This in turn provides soil fauna with a food source in the form of readily degradable carbon compounds. As digestate continues to be removed from the fermenter, ´ parts of the active biomass also continue to be discharged (Voca et al., 2005). While these anaerobic organisms do die immediately after being deposited in the fermentation residue storage container, they nevertheless offer earthworms an additional food source in
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209
Table 5 Community composition in numbers (%) in different fertilizer treatments at the study site Cunnersdorf (09.04.2009) and Pfahlheim (28.05.2009). Species
Lumbricus terrestris Aporrectodea caliginosa Aporrectodea rosea Allolobophora chlorotica Aporrectodea noctorna
Cunnersdorf
Pfahlheim
Control (%)
Mineral fertilizer (%)
Cattle slurry (%)
Digestate (%)
9.1
33.3
14.3
7.1
81.8
50.0
57.1
9.1
16.7
– –
Mineral fertilizer (%)
Pig slurry (%)
Digestate (%)
6.3
18.2
16.1
12.1
92.9
25.0
22.7
64.5
63.6
14.3
–
53.1
22.7
3.2
3.0
–
14.3
–
3.1
13.6
9.7
6.1
–
–
–
12.5
22.8
6.5
15.2
the form of microbial protein. The relatively high number of earthworms in the unfertilized control in Pfahlheim seems to be that earthworms have been negatively affected through the application of both organic fertilizers. Some studies show these results of slurry and digestate on earthworms (Timmerman et al., 2006; Ernst et al., 2008; Bermejo Domínguez, 2012). The negative effects of slurry are related to its high salt concentration and the occurrence of substances that might be toxic to earthworms (Curry, 1976). In addition, the ammonia content may adversely affect soil dwelling earthworms, especially in the digestate treatment. The high abundances and biomass levels at the Cunnersdorf site might be the result of earthworms having migrated. The experimental setup allowed them to migrate to variants with a more attractive food supply. Hoogerkamp et al. (1983) declare a natural spread rate of between 2 and 15 m per year when earthworms colonise habitats with more favorable living conditions. The earthworm population in Cunnersdorf was characterised by a high number of juvenile specimens. This might indicate a higher reproduction rate. In terms of their dominance structure, both of the sites examined are typical representatives of arable land (Tischer, 2008). At Cunnersdorf, applying digestate resulted in a less varied range of species. Abundance of the endogeic earthworm species A. rosea declines. This contradicts research by Ernst et al. (2008), where something of a decrease in A caliginosa biomass was observed after treatment with digestate. In the variant with digestate, the most common species were A. caliginosa and L. terrestris. As a primary decomposer, the anecic earthworm L. terrestris finds sufficient food in this fertilization variant. The mineral soil dweller A. caliginosa benefits from the higher feeding activity of L. terrestris (Ernst et al., 2009). Overall, the differentiation that occurred in the field trials among earthworm populations from various fertilization variants rather confirms previous results from the literature, which show that an organic fertilizer has a far more positive impact on earthworms than a chemical fertilizer or an untreated control can (Edwards and Lofty, 1982; Whalen et al., 1998; Timmerman et al., 2006; Ulrich et al., 2010). Acknowledgements We thank Prof. Niclas, Dr. Schuster, Dr. Kreuter and M. Fuchs from SKW Stickstoffwerke Piesteritz GmbH as well as Dr. M. Mokry and M. Müller from LTZ Augustenberg for allowing us to use their study area in order to investigate this problem. A.-K. Schmitt, T. Leithold and E. Koblenz are acknowledged for their support during soil fauna sampling.
Control (%)
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