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Effect of stocking density and food quality on the growth and fecundity of an epigeic earthworm


Environmentalist (2008) 28:483–488 DOI 10.1007/s10669-008-9173-5

Effect of stocking density and food quality on the growth and fecundity of an epigeic earthworm (Eisenia fetida) during vermicomposting
V. K. Garg ? Priya Kaushik ? Y. K. Yadav

Published online: 7 May 2008 ? Springer Science+Business Media, LLC 2008

Abstract Physico-chemical characteristics of the feed and optimum worm density are important parameters for the ef?cient working of a vermicomposting system. Overcrowding of worms can affect the ef?ciency of a vermicomposting system even if all other parameters have been optimized. This article reports the effect of stocking density and feed quality on the growth and fecundity of Eisenia fetida under laboratory conditions. The feed mixtures contained cow dung and textile mill wastewater sludge in different ratios. Three feed mixtures and ?ve stocking rates (1, 2, 4, 8, and 12) were tested for 12 weeks. The results showed that E. fetida growth rate was faster at higher stocking densities; however, biomass gain per worm was faster at lower stocking densities. Sexual maturity was attained earlier at higher stocking densities. Growth rate was highest in 100% cow dung at all the stocking densities when compared to textile mill wastewater sludge containing feed mixtures. A worm population of 27–53 worms per kg of feed was found to be the most favorable stocking density. Keywords Vermicomposting ? Eisenia fetida ? Feed mixture ? Stocking density ? Growth rate ? Fecundity ? Cow dung ? Textile mill wastewater sludge

1 Introduction It has been well established that epigeic earthworms can hasten the composting process signi?cantly with

V. K. Garg (&) ? P. Kaushik ? Y. K. Yadav Department of Environmental Science and Engineering, Guru Jambheshwar University of Science and Technology, Hisar, Haryana 125001, India e-mail: vinodkgarg@yahoo.com

production of a better quality of compost when compared to compost prepared through traditional methods (Suthar 2006). Use of earthworms for waste management, organic matter stabilization, soil detoxi?cation, and vermicompost production has been well documented in literature (Bansal and Kapoor 2000; Kaushik and Garg 2003; Garg and Kaushik 2005; Suthar 2006). There are several reports regarding the potential utilization of epigeic earthworms for successful degradation of organic wastes, generated from different industries such as paper and pulp industry (Elvira et al. 1997, 1998); dairy industry (Gratelly et al. 1996); winery and distillery industry (Nogales et al. 2005); wood and chips industry (Maboeta and van Rensburg 2003); and textile mill (Kaushik and Garg 2004; Garg and Kaushik 2005). The results of these studies have shown that environmental conditions and earthworm population density affect earthworm growth and reproduction. Several studies have been made on demographic parameters of the earthworms in relation to different physico-chemical conditions (Reinecke and Kriel 1981; Kaplan et al. 1980). Dominguez and Edwards (1997) have reported that individual worms grew more and faster at the lowest stocking density; the total biomass production was more at the higher population density. Ndegwa et al. (2000) have reported that a stocking rate of 1.60 kg-worms/m2 and a feeding rate of 1.25 kg-feed/kg-worm/day resulted in the highest bioconversion of the substrate into worm biomass. Kaushik and Garg (2003, 2004), Garg et al. (2006) have reported the vermicomposting of textile mill wastewater sludge (STMS) under different conditions. The results of these studies have showed that STMS can be vermicomposted if it is mixed up to 30% with other organic wastes like cow dung (CD), agricultural residues, poultry droppings, and biogas plant slurry. As stated earlier, growth and reproduction in developing groups of Eisenia fetida at

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Environmentalist (2008) 28:483–488

different worm densities is one of the important parameters in the optimization of a vermicomposting process. This article reports the effect of E. fetida stocking densities on three different feed mixtures of STMS and CD. It was hypothesized that different worm density would affect growth and reproduction of E. fetida.

2 Materials and methods 2.1 Cow dung (CD), textile mill wastewater sludge (STMS), and earthworms Fresh CD was procured from the Devi Bhawan cowshed, Hisar, India. The main characteristics of CD were: total solids: 434 g/kg, pH (1:10 ratio, CD: water, w/v) 7.62, total organic carbon (TOC): 416 g/kg, total Kjeldhal nitrogen (TKN): 6.5 g/kg, total phosphorus (TP): 6.3 g/kg, and C:N ratio: 64.0. Fresh STMS was obtained from the wastewater treatment plant of a textile factory (H.P. Cotton Mill Ltd.) located near Hisar, India. The main characteristics of STMS were: total solids: 197 g/kg, pH (1:10 ratio, CD: water, w/v): 8.3, TOC: 142 g/kg, TKN: 0.74 g/kg, and C:N ratio: 199. The sludge was dried in shade prior to use for vermicomposting. All the organic waste quantities were used on dry weight basis that was obtained by oven drying known quantities of material at 110°C in hot air oven to constant mass. Non-clitellated hatchlings of E. fetida earthworm were randomly picked for use in the experiments from several stock cultures containing 500–2000 earthworms in each, maintained in the laboratory with cow dung as culturing material. Each hatchling weighed between 0.100 and 0.250 g. The pH was determined by pH system 361 systronics, using double-distilled water suspension of each mixture in the ratio of 1:10 (w/v). Total organic carbon (TOC) was measured using the method of Nelson and Sommers (1982); Total Kjeldhal nitrogen (TKN) was determined by digesting the samples with conc. H2SO4 and HClO4 (9:1, v/v) by Bremner and Mulvaney (1982) procedure. Total available phosphate was analyzed by using the spectrophotometric method with molybdenum in sulfuric acid (Kaushik and Garg 2004). 2.2 Experimental design There were three feed treatments consisting of one with CD only (100%CD) and two mixtures containing 80%CD + 20%STMS and 70%CD + 30%STMS. Cylindrical plastic containers were ?lled with 150 g of each feed mixture (on dry weight basis). All containers were kept in darkness at room temperature (22–26°C). The

moisture content of the feed in each container was maintained at 60–80%, throughout the study period by sprinkling adequate quantities of water on the surface of the feed. These mixtures were turned over manually every day for 15 days to eliminate the potential accumulation of volatile toxic substances. For each feed, ?ve stocking densities (1, 2, 4, 8, and 12 worms per container) were maintained by introducing non-clitellated earthworms after 15 days. There were three replicates for each feed mixture. No additional food was added at any stage during the study period. The duration of the experiment was 12 weeks. 2.3 Growth and fecundity study Biomass gain and cocoon production were recorded every 4th day for 12 weeks. The feed in the container was turned out, and earthworms and cocoons were separated from the feed by hand sorting, after which they were counted and weighed after washing with water and drying them with paper towels. The worms were weighed individually. The worms were weighed without purging their gut content. Then all earthworms, cocoons, and the feed were returned to their respective container.

3 Results and discussion The initial physico-chemical characteristics of CD and STMS were determined. The pH values of CD and STMS were in alkaline range (7.62 and 8.3). The TOC content of CD was much higher than that of STMS. The TKN content of cow dung and STMS were 6.5 and 0.74 g/kg, respectively. The TKN content of feed mixture decreased as STMS ratio in the feed mixture was increased. The C:N ratio of cow dung was 64 and that for STMS was 199. Earlier studies have reported that STMS alone (Kaushik and Garg 2003, 2004) cannot be used as a raw material for vermicomposting and suitable additives were required in the feed mixture for proper growth and reproduction of E. fetida. No mortality was observed at any stocking density in all the feed mixtures. The in?uence of stocking rates in different feed mixtures on the growth of individual worm has been represented in Figs. 1–3. The maximum worm biomass was observed in 100%CD (1480 ± 214 mg/ earthworm) at the stocking density of one worm per container. The minimum biomass was 610 ± 136 mg/ earthworm at the stocking density of 12. The maximum biomass was similar at 830 ± 184 mg/earthworm for stocking densities 4 and 8 worms per container. Similar trends were recorded with respect to biomass gain Figs. 2 and 3) in 80%CD + 20%STMS and 70%CD + 30%STMS feed mixtures. The net increase in biomass by an earthworm

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Environmentalist (2008) 28:483–488 Fig. 1 Mean individual growth of E. fetida in 100%CD at ?ve different stocking densities

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1 worm
1.6 1.4 1.2

2 worms

4 worms

8 worms

12 worms

Mean Individual Biomass (g)

1 0.8 0.6 0.4 0.2 0 0 8 16 24 32 40 48 56 64 72 80 88 96

Time (days)

Fig. 2 Mean individual growth of E. fetida in 80%CD + 20%STMS feed mixture at ?ve different stocking densities

1 worm

2 worms

4 worms

8 worms

12 worms

1.4

1.2

Mean Individual Biomass (g)

1

0.8

0.6

0.4

0.2

0 0 8 16 24 32 40 48 56 64 72 80 88 96

Time (days)

decreased as the stocking density increased (Table 1). The results showed that earthworms grew more at the lowest population density. Similar observations have been reported by Dominguez and Edwards (1997) for Eisenia andrei in pig manure. The total biomass gain attained by earthworms was highest at maximum stocking density (12 worms per container). Total biomass gained by earthworms was 7.32 ± 0.84, 8.88 ± 0.63, and 7.44 ± 0.65 g per container at the stocking density of 12 in 100%CD, 80%CD + 20%STMS, 70%CD + 30% STMS, respectively (Table 2).

The average growth rates of the earthworms varied between 16.81 ± 1.19 and 10.0 ± 1.47 mg/earthworm/day in 100%CD. The average growth rate of the earthworms varied in the range of 13.15 ± 1.61 to 7.77 ± 1.56 mg/ earthworm/day in 80%CD + 20%STMS. In 70%CD + 30%STMS feed mixture it varied from 17.08 ± 2.75 to 6.25 ± 1.78 mg/earthworm/day (Table 3). As the population density increased the time for maximal biomass decreased (Figs. 1–3); therefore, growth rates were faster at higher worm densities. The net biomass produced

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486 Fig. 3 Mean individual growth of E. fetida in 70%CD + 30%STMS feed mixture at ?ve different stocking densities

Environmentalist (2008) 28:483–488

1 worm
1.6

2 worms

4 worms

8 worms

12 worms

1.4

1.2

Mean Individual Biomass (g)

1

0.8

0.6

0.4

0.2

0 0 8 16 24 32 40 48 56 64 72 80 88 96

TIme (days)

Table 1 Mean net worm biomass for an individual earthworm (mg/earthworm) in different feed mixtures at different stocking densities

Stocking density

Feed mixture 100%CD 80%CD + 20%STMS 1000 ± 245 780 ± 233 640 ± 234 590 ± 103 560 ± 78 70%CD + 30%STMS 1230 ± 279 880 ± 247 620 ± 195 520 ± 132 436 ± 154

1 2 4 8 12

1210 ± 214 880 ± 198 590 ± 173 572 ± 197 440 ± 136

Table 2 Total earthworm biomass attained (g) in different feed mixture at different stocking densities

Stocking density

Feed mixture 100%CD 80%CD + 20%STMS 1.48 ± 0.39 2.50 ± 0.53 3.28 ± 0.5 6.00 ± 0.7 7.44 ± 0.65 70%CD + 30%STMS 1.22 ± 0.43 1.70 ± 0.38 83.00 ± 0.36 66.08 ± 0.59 8.88 ± 0.63

1 2 4 8 12

1.48 ± 0.31 2.24 ± 0.47 3.32 ± 0.49 6.64 ± 0.73 7.32 ± 0.84

was 8.07 ± 1.31, 6.66 ± 0.93, and 8.20 ± 1.38 mg/g at the stocking density of one in 100%CD, 80%CD + 20% STMS, and 70%CD + 30%STMS feed mixtures, respectively. Whereas, at the stocking density of 12, the net biomass produced was 35.2 ± 2.25, 33.71 ± 3.15,

and 29.58 ± 1.47 in 100%CD, 80%CD + 20%STMS, and 70%CD + 30%STMS feed mixtures, respectively (Table 4). The worms were sexually mature at a younger age at higher stocking rates although cocoon production did not

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Environmentalist (2008) 28:483–488 Table 3 Average growth rate of E. fetida (mg/worm/day) in different feed mixture at different stocking densities

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Stocking density

Feed mixture 100%CD 80%CD + 20%STMS 13.15 ± 1.61 (76) 10.23 ± 1.82 (52) 11.07 ± 2.01 (56) 9.83 ± 2.15 (60) 7.77 ± 1.56 (72) 70%CD + 30%STMS 17.08 ± 2.75 (72) 12.94 ± 2.18 (68) 9.64 ± 1.03 (56) 9.28 ± 2.17 (56) 6.25 ± 1.78 (72)

1 2 4
a

16.81 ± 1.19 (72)a 16.92 ± 1.32 (52) 11.34 ± 1.87 (52) 13.63 ± 1.16 (44) 10.0 ± 1.47 (44)

Figures in parentheses is the day on which maximum biomass was attained

8 12

Table 4 Net biomass gain per unit feed mixture (mg/g) by E. fetida in different feed mixtures at different stocking densities

Stocking density

Feed mixture 100%CD 80%CD + 20%STMS 6.66 ± 0.93 11.40 ± 0.83 14.45 ± 1.02 31.43 ± 0.90 33.71 ± 3.15 70%CD + 30%STMS 8.20 ± 1.38 11.73 ± 1.34 13.86 ± 1.25 27.74 ± 1.42 29.58 ± 1.47

1 2 4 8 12

8.07 ± 1.31 11.73 ± 1.28 15.74 ± 1.45 32.00 ± 1.36 35.2 ± 2.25

Table 5 The day cocoons appeared in the different treatments Stocking density Feed mixture 100%CD Day 52 Day 40 Day 36 Day 36 Day 32 80%CD + 20%STMS Day 68 Day 48 Day 40 Day 36 Day 32 70%CD + 30%STMS Day 72 Day 52 Day 40 Day 40 Day 36

Table 6 Total number of cocoons produced by E. fetida in different treatments at the different stocking densities Stocking density Feed mixture 100%CD 13 ± 5 21 ± 8 62 ± 13 82 ± 17 113 ± 28 80%CD + 20%STMS 8±3 20 ± 9 39 ± 21 41 ± 27 72 ± 38 70%CD + 30%STMS 5±3 18 ± 8 30 ± 19 44 ± 23 54 ± 26

1 2 4 8 12

1 2 4 8 12

occur before day 32 at any of the stocking densities regardless of the feed mixtures (Table 5). The results also indicated that although cocoon production was started late at lower stocking densities, it continued for longer periods of time than at higher stocking densities. The maximum cocoon production occurred between day 40 and 60 at the stocking densities of 4, 8, and 12 in all the feed mixtures. At all the stocking densities, the worms reach sexual maturity, although the time to reach sexually maturity was inversely related to stocking density. A similar observation was reported by Dominguez and Edwards (1997). The total number of cocoon produced per container was directly related to the stocking density (Table 6). The highest number of cocoons was counted (113 ± 28) in 100%CD at the stocking density of 12, whereas fewest were counted (05 ± 03) in 70%CD + 30%STMS at the stocking density of 1. Higher cocoon production is predictable at higher stocking densities. The production of cocoons at the stocking densities of 1 was strange but other co-workers have also reported production of cocoons at the stocking

density of 1. The rate of cocoon production per earthworm was maximum at the stocking densities of 4 in 100%CD, 2 in 80%CD + 20%STMS and in 70%CD + 30%STMS feed mixtures (Table 7). The results indicate that loading densities between 27 and 53 worms per kg of substrate is optimal.

Table 7 Number of cocoons produced per earthworm in different feed mixture Stocking density Feed mixture 100%CD 13 ± 0.5 10.5 ± 0.4 15.5 ± 0.7 10.25 ± 0.9 9.4 ± 1.0 80%CD + 20%STMS 8 ± 0.2 10 ± 0.3 9.75 ± 0.5 5.12 ± 0.6 6.0 ± 0.8 70%CD + 30%STMS 5 ± 0.2 9 ± 0.2 7.5 ± 0.5 5.5 ± 0.7 4.5 ± 0.5

1 2 4 8 12

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Environmentalist (2008) 28:483–488 Garg VK, Kaushik P (2005) Vermistabilization of textile mill sludge spiked with poultry droppings by an epigeic earthworm Eisenia fetida. Bioresour Technol 96:1189–1193 Garg VK, Kaushik P, Dilbaghi N (2006) Vermiconversion of wastewater sludge from textile mill mixed with anaerobically digested biogas plant slurry employing Eisenia fetida. Ecotoxicol Environ Saf 63:412–419 Gratelly P, Benitez E, Elvira C, Polo A, Nogales R (1996) Stabilization of sludges from a dairy processing plant using vermicomposting. In: Rodriguez-Barrueco C (ed) Fertilizers and environment. Kluwer, The Netherlands, pp 341–343 Kaplan DL, Hartenstein R, Neuhauser EF, Malecki MR (1980) Physico-chemical requirements in the environment of the earthworm Eisenia fetida. Soil Biol Biochem 12:347–352 Kaushik P, Garg VK (2003) Vermicomposting of mixed solid textile mill sludge and cow dung with epigeic earthworm Eisenia fetida. Bioresour Technol 90:311–316 Kaushik P, Garg VK (2004) Dynamics of biological and chemical parameters during vermicomposting of solid textile mill sludge mixed with dung and agricultural residues. Bioresour Technol 94:203–209 Maboeta MS, Rensburg L (2003) Vermicomposting of industrially produced woodchips and sewage sludge utilizing Eisenia fetida. Ecotoxicol Environ Saf 56:265–270 Ndegwa PM, Thompson SA, Das KC (2000) Effects of stocking density and feeding rate on vermicomposting of biosolids. Bioresour Technol 71:5–12 Nelson DW, Sommers LE (1982) Total carbon and organic carbon and organic matter. In: Page AL, Miller RH, Keeney DR (eds) Method of soil analysis. American Society of Agronomy, Madison, pp 539–579 Nogales R, Celia C, Benitez E (2005) Vermicomposting of winery wastes: a laboratory study. J Environ Sci Health Part B 40:659– 673 Reinecke AJ, Kriel JR (1981) In?uences of temperature on the reproduction of the earthworm Eisenia fetida. S Afr J Zool 16:96–100 Suthar S (2006) Potential utilization of guar gum industrial waste in vermicompost production. Bioresour Technol 97:2474–2477

4 Conclusion Even when the physical conditions (temperature and moisture) and quality of waste (size, TOC, TKN, and TAP) are appropriate for vermicomposting, problems can develop due to overcrowding of earthworms. This study clearly showed that when E. fetida was allowed to grow at different stocking densities the worms grew slowly at higher stocking densities. The maximum body weight of earthworm was higher at lower stocking densities. Maturation rate was also affected by stocking rate. Worms attained sexual maturity earlier in crowded containers. Worms of same age developed clitellum at different times at different population densities. The results indicate that population of 27–53 worms per kg and 4–8 worms per 150 g/feed mixture is optimum.

References
Bansal S, Kapoor KK (2000) Vermicomposting of crop residues and cattle dung with Eisenia fetida. Bioresour Technol 73:95–98 Bremner JM, Mulvaney RG (1982) Nitrogen total. In: Page AL, Miller RH, Keeney DR (eds) Method of soil analysis. American Society of Agronomy, Madison, pp 575–624 Dominguez J, Edward CA (1997) Effect of stocking rates and moisture content on the growth and maturation of Eisenia andrei (Oligochaeta) in pig manure. Soil Biol Biochem 29:743–746 Elvira C, Sampedro L, Dominguez J, Mato S (1997) Vermicomposting of wastewater sludge from paper-pulp industry with nitrogen rich materials. Soil Biol Biochem 29:759–762 Elvira C, Sampedro L, Benitez E, Nogales R (1998) Vermicomposting of sludges from paper mill and dairy industries with Eisenia andrei: a pilot scale study. Bioresour Technol 63:205– 211

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