3.1. Concentratons of heavy metals in final bioremediated susbstrates
The vermiremediation using E. fetida significantly detoxified the substrates in different reactors as depicted by the changes in heavy metal concentrations (Cd, Cr, Cu, Ni, Pb & Zn) in Table 1. The difference among the reactors was statistically significant for Cd (ANOVA; F = 12.67, P ˂ 0.0001), Cr (ANOVA; F = 69.36, P ˂ 0.0001), Cu (ANOVA; F = 257.47, P ˂ 0.0001), Ni (ANOVA; F = 83.56, P ˂ 0.0001), Pb (ANOVA; F = 166.03, P ˂ 0.0001) and Zn (ANOVA; F = 45.77, P ˂ 0.0001). At the end of bioremediation process, Cd ranged from 4.93 ± 0.003 mg.kg-1 (R1) to 7.79 ± 0.001 mg.kg-1 (R3), Cr ranged from 31.8 ± 0.9 mg.kg-1 (R2) to 57.5 ± 0.6 mg.kg-1 (R1), Cu ranged from 27.68 ± 0.43 mg.kg-1 (R3) to 38.73 ± 0.48 mg.kg-1 (R6), Ni ranged from 2.98 ± 0.001 mg.kg-1 (R2) to 3.45 ± 0.024 mg.kg-1 (R1), Pb ranged from 12.26 ± 0.3 mg.kg-1 (R3) to 13.52 ± 0.5 mg.kg-1 (R5) and Zn ranged from 57.54 ± 0.4 mg.kg-1 (R6) to 91.60 ± 0.45 mg.kg-1 (R3). R3 (PMS: CD = 1:1) was the most feasible reactor in terms of reduction in heavy metal concentrations with average removal efficiency of 74.45%. The heavy metal removal order was: R3 (74.45%) > R2 (72.98%) > R4 (72.93%) > R6 (68.22%) > R1 (67.25%) > R5 (66.63%). Wang et al. (2013a) stated that there are a variety of factors in the vermiremoval process such as pH, equilibrium between absorbing and accumulating forms of metals, metal speciation, chemical-interactions between various ions, physico-chemical characteristics of wastes, etc. among others that work to detoxify the substrates. The efficiency of removal of heavy metals (ƞr) for different metals ranged as: Cd (37.2–58.2%), Cr (57.0-74.3%), Cu (67.3–79.8%), Ni (74.7–81.9%), Pb (78.8–83.4%) and Zn (71.2–77.4%). Removal efficiencies of reactors for individual heavy metals were also obtained: Cd [R3 > R1 > R4 > R2 > R6 > R5], Cr [R4 > R2 > R3 > R5 > R6 > R1], Cu [R3 > R2 > R4 > R5 > R6 > R1], Ni [R3 > R2 > R4 > R6 > R5 > R1], Pb [R3 > R4 > R2 > R6 > R5 > R1] and Zn [R6 > R2 > R5 > R4 > R1 > R3]. Figure 1 shows decrease in concentration of heavy metals in different reactors during the course of this study. The results obtained were significant (P ˂ 0.05) and can be correlated with the findings of Suthar et al. (2014) whose study concluded that the increase in proportion of PMS resulted in effective removal of heavy metals, especially Cu and Pb. Yuvaraj et al. (2020) made use of two epigeic earthworm species E. eugeniae and P. excavates to bio-stabilize textile mill wastewater sludge and reported that the treatment combination (PMS: CD = 1:1) removed heavy metals up to a significant level. Gupta et al. (2007) stabilized water hyacinth using cow dung and significantly observed reduction of Pb (42.7–72.4%), Cd (20.8–58.1%) and Cu (26.9–49.1%) in the final vermicompost. The organic matter breakdown results in release of soluble fractions of heavy metals from substrates (Suthar and Singh, 2008). In this study, the rate of heavy metal removal was of the order: Pb > Ni > Zn > Cu > Cr > Cd. This reduction in heavy metal concentrations is also dependent on the assimilation/ accumulation and excretion of heavy metals through the gut of earthworms. The relationship of reduction of heavy metal concentrations with load of heavy metals in earthworm tissues was also examined by linear regression analysis (Table 3). The reduction of metals and genotoxicity in the final vermistabilized product is indicative of the capability of earthworms and the overall process of vermitechnology in cleaning up of industrial wastes.
Table 1
Parameters | Cd (mg.kg− 1) | Cr (mg.kg− 1) | Cu (mg.kg− 1) | Ni (mg.kg− 1) | Pb (mg.kg− 1) | Zn (mg.kg− 1) |
Initial Substrate | | | | | | |
R1 [CD (100%)] | 10.53 ± 0.02 | 133.7 ± 0.4 | 117.8 ± 0.3 | 13.66 ± 0.04 | 63.6 ± 0.3 | 215.92 ± 0.6 |
R2 [PMS:CD(1:2)] | 14.76 ± 0.04 | 119.4 ± 0.5 | 125.6 ± 0.4 | 15.73 ± 0.08 | 71.5 ± 0.7 | 267.56 ± 0.3 |
R3 [PMS:CD(1:1)] | 18.64 ± 0.03 | 141.8 ± 0.3 | 137.2 ± 0.6 | 17.56 ± 0.04 | 74.2 ± 0.4 | 317.83 ± 0.6 |
R4 [PMS:TW:CD(1:1:1)] | 16.38 ± 0.02 | 139.5 ± 0.2 | 133.4 ± 0.4 | 16.88 ± 0.06 | 72.3 ± 0.5 | 292.67 ± 0.4 |
R5 [PMS:TW:CD(1:2:1)] | 11.97 ± 0.03 | 134.5 ± 0.3 | 120.2 ± 0.5 | 13.98 ± 0.02 | 65.8 ± 0.4 | 239.53 ± 0.8 |
R6 [PMS:TW:CD(2:1:1)] | 13.53 ± 0.04 | 136.8 ± 0.6 | 122.6 ± 0.3 | 14.43 ± 0.05 | 66.5 ± 0.8 | 255.43 ± 0.5 |
Final Substrate | | | | | | |
R1 [CD (100%)] | 4.93 ± 0.003 | 57.5 ± 0.6 | 38.53 ± 0.2 | 3.45 ± 0.024 | 13.43 ± 0.4 | 59.28 ± 0.4 |
R2 [PMS:CD(1:2)] | 7.65 ± 0.002 | 31.8 ± 0.9 | 29.13 ± 0.1 | 2.98 ± 0.001 | 13.08 ± 0.2 | 61.85 ± 0.6 |
R3 [PMS:CD(1:1)] | 7.79 ± 0.001 | 39.4 ± 0.4 | 27.68 ± 0.4 | 3.18 ± 0.002 | 12.26 ± 0.3 | 91.60 ± 0.4 |
R4 [PMS:TW:CD(1:1:1)] | 7.93 ± 0.005 | 35.8 ± 0.5 | 32.49 ± 0.3 | 3.26 ± 0.003 | 12.73 ± 0.2 | 78.54 ± 0.5 |
R5 [PMS:TW:CD(1:2:1)] | 7.51 ± 0.001 | 48.4 ± 0.9 | 36.45 ± 0.2 | 3.34 ± 0.005 | 13.52 ± 0.5 | 63.45 ± 0.3 |
R6 [PMS:TW:CD(2:1:1)] | 7.44 ± 0.002 | 54.8 ± 0.4 | 38.73 ± 0.4 | 3.28 ± 0.003 | 12.48 ± 0.2 | 57.54 ± 0.4 |
Changes in heavy metal concentrations in different reactors during bioremediation of substrates (Mean ± SD; n = 3). SD = Standard deviation, mean values indicate that difference between reactors is statistically different (ANOVA; Tukey’s t-test, p ˂ 0.05). |
Table 2
Heavy metal concentrations in earthworm tissues |
Reactors | Cd (mg.kg− 1) | Cr (mg.kg− 1) | Cu (mg.kg− 1) | Ni (mg.kg− 1) | Pb (mg.kg− 1) | Zn (mg.kg− 1) |
R1 [CD (100%)] | 2.92 ± 0.01 | 23.54 ± 0.4 | 9.75 ± 0.2 | 3.76 ± 0.03 | 9.43 ± 0.02 | 25.78 ± 0.8 |
R2 [PMS:CD(1:2)] | 2.79 ± 0.02 | 24.77 ± 0.8 | 10.68 ± 0.8 | 3.81 ± 0.02 | 8.96 ± 0.02 | 27.35 ± 0.5 |
R3 [PMS:CD(1:1)] | 3.24 ± 0.04 | 28.76 ± 0.7 | 11.25 ± 0.3 | 3.99 ± 0.05 | 8.92 ± 0.04 | 28.81 ± 0.4 |
R4 [PMS:TW:CD(1:1:1)] | 3.08 ± 0.05 | 27.58 ± 0.3 | 11.56 ± 0.3 | 3.92 ± 0.05 | 8.76 ± 0.01 | 29.72 ± 0.4 |
R5 [PMS:TW:CD(1:2:1)] | 2.94 ± 0.03 | 24.32 ± 0.4 | 9.72 ± 0.5 | 3.61 ± 0.03 | 8.83 ± 0.03 | 23.12 ± 0.3 |
R6 [PMS:TW:CD(2:1:1)] | 2.86 ± 0.01 | 23.92 ± 0.5 | 9.53 ± 0.6 | 3.52 ± 0.08 | 8.79 ± 0.04 | 26.53 ± 0.5 |
Bioaccumulation Factors (BAFs) for metals |
Reactors | BAFCd | BAFCr | BAFCu | BAFNi | BAFPb | BAFZn |
R1 [CD (100%)] | 0.27 ± 0.002 | 0.17 ± 0.004 | 0.083 ± 0.001 | 0.27 ± 0.001 | 0.15 ± 0.001 | 0.119 ± 0.002 |
R2 [PMS:CD(1:2)] | 0.19 ± 0.001 | 0.21 ± 0.003 | 0.085 ± 0.002 | 0.24 ± 0.003 | 0.12 ± 0.001 | 0.102 ± 0.003 |
R3 [PMS:CD(1:1)] | 0.17 ± 0.001 | 0.20 ± 0.001 | 0.081 ± 0.004 | 0.22 ± 0.004 | 0.12 ± 0.005 | 0.090 ± 0.001 |
R4 [PMS:TW:CD(1:1:1)] | 0.18 ± 0.004 | 0.19 ± 0.005 | 0.086 ± 0.003 | 0.23 ± 0.001 | 0.12 ± 0.004 | 0.101 ± 0.002 |
R5 [PMS:TW:CD(1:2:1)] | 0.24 ± 0.006 | 0.18 ± 0.003 | 0.080 ± 0.006 | 0.25 ± 0.003 | 0.13 ± 0.002 | 0.096 ± 0.004 |
R6 [PMS:TW:CD(2:1:1)] | 0.21 ± 0.003 | 0.17 ± 0.004 | 0.077 ± 0.005 | 0.24 ± 0.005 | 0.13 ± 0.003 | 0.103 ± 0.006 |
Bioavailability of heavy metal in tissues of earthworms in different reactors and bioaccumulation factor (BAF) (Mean ± SD, n = 3). SD = Standard deviation, mean values indicate that difference between reactors is statistically different (ANOVA; Tukey’s t-test, p ˂ 0.05). |
Table 3
Linear regression analysis | R2 | P-value |
Loss of heavy metals and load of heavy metals in earthworm tissue |
Cd removal = -24.7642 + 10.7204 Cd earthworm | 0.58 | ** |
Cr removal = -36.4782 + 4.9504 Cr earthworm | 0.92 | *** |
Cu removal = -38.0901 + 12.5193 Cu earthworm | 0.84 | *** |
Ni removal = -17.0926 + 7.7534 Ni earthworm | 0.66 | ** |
Pb removal = +139.4968–9.3235 Pb earthworm | 0.25 | ** |
Zn removal = -30.7809 + 8.4394 Zn earthworm | 0.59 | ** |
Loss of heavy metals and BAF |
Cd removal = +17.3067–48.6351 Cd BAF | 0.67 | ** |
Cr removal = 13.46 + 408.25 Cr BAF | 0.36 | ** |
Cu removal = -18.9142 + 1356.25 Cu BAF | 0.15 | ** |
Ni removal = 33.6876–89.2247 Ni BAF | 0.81 | *** |
Pb removal = 100.2446–344.2439 Pb BAF | 0.76 | *** |
Zn removal = 396.9294–1972.0071 Zn BAF | 0.55 | ** |
The relationship of loss of heavy metal concentrations with both bioavailability of heavy metals in earthworm tissues and BAF, obtained by linear regression analysis. |
**Significant (P ˂ 0.05). |
***Significant (P ˂ 0.001). |
On the contrary, increase in heavy metal concentrations were also reported by previous studies. Vig et al. (2011) used Eisenia fetida to bioremediate tannery sludge, but, reported 2.6–13.4% and 4.6–17.3% increase in concentrations of Mn and Zn respectively. This increase was due to increase in loss of carbon content due to mineralization. Li et al. (2009) studied the transition changes in Cu and Zn contents by passing pig manure through the gut of earthworms and observed 1.2–3.4 folds increase in Cu content and 1.3–2.5 folds increase in Zn in the end-product. The study signified this increase to the affinity of Cu and Zn to Fe and Mn oxides, which may not have significantly changed after transit through earthworm gut. Bioavailability of metals, therefore, depend upon different mechanisms such as bioaccumulation (accumulation of heavy metals in inoculated worms) (Suthar and Singh, 2008), leaching of heavy metals from sludge during bioremediation and adsorption of heavy metals on waste surface fractions (Wang et al., 2013a).
3.2. Bioavailability of heavy metals in tissues of earthworms and BAFs
The reduction of heavy metal concentrations in different reactors clearly suggests the role and bioaccumulating ability of E. fetida in the bioconversion process. The concentrations of heavy metals in tissues of earthworms were statistically significant (P ˂ 0.05) for Cd (ANOVA; F = 128.61, P < 0.0001), Cr (ANOVA; F = 177.26, P < 0.0001), Cu (ANOVA; F = 132.37, P < 0.05), Ni (ANOVA; F = 229.68, P ˂ 0.0001), Pb (ANOVA; F = 323.24, P < 0.0001) and Zn (ANOVA; F = 221.24, P < 0.0001). The concentrations of heavy metals in tissues of earthworms ranged as: Cd [2.79 ± 0.02 (R2) to 3.24 ± 0.04 mg.kg-1 (R3)], Cr [23.54 ± 0.4 (R1) to 28.76 ± 0.8 mg.kg-1 (R3)], Cu [9.53 ± 0.6 (R6) to 11.56 ± 0.3 mg.kg-1 (R4)], Ni [3.52 ± 0.08 (R6) to 3.99 ± 0.05 mg.kg-1 (R3)], Pb [8.76 ± 0.01 (R4) to 9.43 ± 0.02 mg.kg-1 (R1)] and Zn [23.12 ± 0.3 (R5) to 29.72 ± 0.4 mg.kg-1 (R4)], as shown in Table 2. The difference among the reactors for concentration values of heavy metals in tissues of earthworms may have been due to changes in physico-chemical characteristics of substrates. The bioaccumulation of heavy metals in the tissues of earthworms for different reactors can be arranged in descending order as: Cd [R3 > R4 > R5 > R1 > R6 > R2], Cr [R3 > R4 > R2 > R5 > R6 > R1], Cu [R4 > R3 > R2 > R1 > R5 > R6], Ni [R3 > R4 > R2 > R1 > R5 > R6], Pb [R1 > R2 > R3 > R5 > R6 > R4] and Zn [R4 > R3 > R2 > R6 > R1 > R5]. Heavy metals such as Co, Cu, Cd and Zn have high affinity to metal binding protein called metallothionein, which renders them biologically inactive (Samal et al., 2019). The rate of accumulation of heavy metals in tissues of earthworms largely depends on the metal pollution level of the environment into which the earthworms are fed. If the metal pollution level would be high, then earthworms would not be able to accumulate the heavy metals in their gut and will excrete them out at a faster rate. This is solely done to maintain equilibrium conditions in their physiological metabolism. The defence mechanism of Eisenia species is prolific at cellular levels, as a result of which they are strong bioaccumulators of toxic metals (Suleiman et al., 2017) and by which the bioremediation of metals can be achieved sustainably through vermicomposting. The bioaccumulation of heavy metals in the tissues of earthworms has also been in well documented in published literature (Suthar and Singh, 2008; Azizi et al., 2013; Srivastava et al., 2005).
The bioaccumulation factors (BAFs) of heavy metals are shown in Table 2. The values of BAF obtained were statistically significant for Cd (ANOVA; F = 154.98, P < 0.0001), Cr (ANOVA; F = 34.21, P < 0.0001), Cu (ANOVA; F = 325.54, P < 0.05), Ni (ANOVA; F = 255.78, P ˂ 0.0001), Pb (ANOVA; F = 128.87, P < 0.0001) and Zn (ANOVA; F = 344.79, P < 0.0001). The BAF for different heavy metals ranged as: Cd [0.17 ± 0.001 (R3) to 0.27 ± 0.002 mg.kg-1 (R1)], Cr [0.17 ± 0.004 (R1, R6) to 0.21 ± 0.003 mg.kg-1 (R2)], Cu [0.077 ± 0.001 (R6) to 0.086 ± 0.004 mg.kg-1 (R4)], Ni [0.22 ± 0.004 (R3) to 0.27 ± 0.001 mg.kg-1 (R1)], Pb [0.12 ± 0.001 (R2) to 0.15 ± 0.001 mg.kg-1 (R1)] and Zn [0.090 ± 0.001 (R3) to 0.119 ± 0.002 mg.kg-1 (R1)]. The BAF for heavy metals can be arranged in the following order: Ni > Cd > Cr > Pb > Zn > Cu. The relationship of reduction of heavy metal concentrations with bioaccumulation factors (BAFs) was also determined by linear regression analysis (Table 3). The results obtained are similar to that obtained by Suthar et al. (2014), thus, support his hypothesis which suggested that leaching is the main mechanism of loss of heavy metals in this study rather than absorption by worms. Previous studies have also used BAF in tissues of earthworms for evaluation of toxicity in bioremediation of substrates. (Mountouris et al., 2002; Wang et al., 2013a; Yuvaraj et al., 2018a).
3.3. Changes in pH, EC, TOC, TN, TP, TK level and C/N ratio during vermiremediation of substrate
Vermistabilization of PMS and TW resulted in increase of nutrient contents beneficial for the soil and growth of plants. The difference among the reactors was statistically significant: pH(ANOVA; F = 13.43, P ˂ 0.0001), EC (ANOVA; F = 263.56, P ˂ 0.0001), TOC (ANOVA; F = 344.71 P ˂ 0.0001), TN (ANOVA; F = 113.89, P ˂ 0.0001), TP (ANOVA; F = 166.26, P ˂ 0.0001), TK (ANOVA; F = 49.54, P ˂ 0.0001) and C/N ratio (ANOVA; F = 534.75, P ˂ 0.0001). The physico-chemical characteristics of both initial and final substrate are given in Table 4 and ranged as: pH[7.32 ± 0.50 (R1) to 7.65 ± 0.50 (R6)], EC [3.79 ± 0.20 (R1) to 5.25 ± 0.10 (R6)], TOC [38.79 ± 0.75 (R3) to 42.63 ± 0.50 (R6)], TN [1.24 ± 0.03 (R6) to 1.87 ± 0.01 (R3)], TP [0.68 ± 0.01 (R6) to 1.08 ± 0.01 (R3)], TK [0.28 ± 0.01 (R2) to 0.49 ± 0.02 (R6)] and C/N ratio [20.74 ± 0.90 (R3) to 34.37 ± 0.33 (R6)]. Previous researchers have also documented the enrichment of nutrients by the process of vermistabilization (Yuvaraj et al., 2020; Kaur et al., 2010). Formation of organic acids and other intermediate metabolic products in the gut of earthworms with the help of mucus and enzymic activities would have accelerated the degradation of organic matter (TOC) and resulted in simultaneous decrease of pH and increase of EC (Badhwar et al., 2020; Lim et al., 2011). Maximum reduction of TOC content and increase of earthworm population in R3 (PMS: CD = 1:1) indicates the efficacy of good vermistabilization process. The earthworms have the ability to eliminate the nitrogenous substances from the substrate which would have resulted increase of TN content (Suthar et al., 2017). The increase in TP content is due to the ability of earthworms to convert the insoluble phosphorus to soluble phosphorus in the process of vermitechnology (Fu et al., 2015). High TK content in R5 is attributed to the TK content of tea waste. C/N ratio of all the reactor substrates decreased significantly, which indicates efficient recovery of wastes and promotes the sustainable cycle of transformation of waste to resource.
Table 4
Parameters | pH | EC (mS.cm− 1) | TOC (%) | TN (%) | TP (%) | TK (%) | C/N ratio |
Initial Substrate | | | | | | | |
R1 [CD (100%)] | 8.13 ± 0.50 | 1.48 ± 0.05 | 43.26 ± 0.60 | 1.17 ± 0.01 | 0.62 ± 0.02 | 0.29 ± 0.01 | 36.97 ± 0.20 |
R2 [PMS:CD(1:2)] | 8.46 ± 0.30 | 1.67 ± 0.05 | 44.85 ± 0.25 | 1.28 ± 0.03 | 0.58 ± 0.01 | 0.17 ± 0.01 | 35.03 ± 0.55 |
R3 [PMS:CD(1:1)] | 8.63 ± 0.50 | 1.96 ± 0.10 | 46.65 ± 0.45 | 1.41 ± 0.01 | 0.65 ± 0.04 | 0.15 ± 0.01 | 33.08 ± 0.30 |
R4 [PMS:TW:CD(1:1:1)] | 8.75 ± 0.50 | 2.08 ± 0.05 | 47.90 ± 0.20 | 1.33 ± 0.01 | 0.51 ± 0.03 | 0.21 ± 0.03 | 36.01 ± 0.64 |
R5 [PMS:TW:CD(1:2:1)] | 8.83 ± 0.10 | 2.19 ± 0.15 | 48.72 ± 0.15 | 0.99 ± 0.02 | 0.43 ± 0.01 | 0.25 ± 0.01 | 49.21 ± 0.36 |
R6 [PMS:TW:CD(2:1:1)] | 8.92 ± 0.20 | 2.27 ± 0.10 | 49.51 ± 0.10 | 0.94 ± 0.01 | 0.39 ± 0.02 | 0.31 ± 0.02 | 52.67 ± 0.27 |
Final Substrate | | | | | | | |
R1 [CD (100%)] | 7.32 ± 0.50 | 3.79 ± 0.20 | 39.96 ± 0.40 | 1.65 ± 0.02 | 0.82 ± 0.02 | 0.41 ± 0.03 | 24.21 ± 0.35 |
R2 [PMS:CD(1:2)] | 7.42 ± 0.20 | 4.23 ± 0.15 | 39.12 ± 0.55 | 1.79 ± 0.03 | 0.95 ± 0.02 | 0.28 ± 0.01 | 21.85 ± 0.60 |
R3 [PMS:CD(1:1)] | 7.48 ± 0.40 | 4.38 ± 0.05 | 38.79 ± 0.75 | 1.87 ± 0.01 | 1.08 ± 0.01 | 0.34 ± 0.01 | 20.74 ± 0.90 |
R4 [PMS:TW:CD(1:1:1)] | 7.53 ± 0.30 | 4.65 ± 0.10 | 39.43 ± 0.30 | 1.68 ± 0.05 | 0.89 ± 0.04 | 0.30 ± 0.04 | 23.47 ± 0.45 |
R5 [PMS:TW:CD(1:2:1)] | 7.62 ± 0.50 | 5.12 ± 0.05 | 40.84 ± 0.15 | 1.38 ± 0.02 | 0.75 ± 0.02 | 0.49 ± 0.01 | 29.59 ± 0.25 |
R6 [PMS:TW:CD(2:1:1)] | 7.65 ± 0.50 | 5.25 ± 0.10 | 42.63 ± 0.50 | 1.24 ± 0.03 | 0.68 ± 0.01 | 0.46 ± 0.02 | 34.37 ± 0.33 |
Changes in physico-chemical parameters of reactors during bioremediation process (Mean ± SD, n = 3). SD = Standard deviation, mean values indicate that difference between reactors is statistically different (ANOVA; Tukey’s t-test, p ˂ 0.05). |
3.4. Survival of earthworms, biomass changes, cocoon production and mortality rate
Earthworms are known to improve the quality of soil by adding organically bound nutrients (C, N, P, K, S, etc.) and detoxify heavy metals in the soil environment. Bioremediation occurs due to unique bioaccumulation mechanism of earthworms (Maity et al., 2009) and their strong metabolic system where diverse intestinal microflora, enzymic activities and chloragocyte cells function accordingly in the remediation of industrial sludges (Srivastava et al., 2005). The changes in the biomass of earthworms, reproduction rate and rate of mortality can be directly correlated with the feed materials (Table 5). The difference in the biomass of earthworms was statistically significant (P ˂ 0.05) in different reactors and was in the order: R3 (62.0%) > R2 (59.8%) > R4 (55.9%) > R5 (52.8%) > R1 (51.3%) > R6 (50.3%). Maximum mortality rate (19%) and minimum reproduction rate (6.1 ± 0.2 cocoon.worm-1) was seen in R6 [PMS: TW: CD (2:1:1)] which clearly indicates the unsuitability of higher proportions of TW in feed mixtures, although lesser proportions can be used for the degradation of substrates as seen in R5 and R4. Conversely, least mortality rate (2%) and maximum reproduction rate (7.8 ± 0.1 cocoon.worm-1) was seen in R3 [PMS: CD (1:1)] which showed the best earthworm survival rate among all the reactors and indicates its enhanced applicability in vermiremoval process. The other important factors in the rise of earthworm population in vermistabilized waste substrates include cocoon viability, hatchling success and the survival rate of juveniles, etc. The production of cocoons was highest in R3 followed by R4, R2, R5, R1 and R6 (Table 5). Suthar et al. (2014) stated that the production of cocoons depend upon various factors such as the toxicity of wastes, palatability of food, quality of waste mixtures and the overall environmental conditions required for the successful bioremediation of industrial sludges. The rate of cocoon production (cocoon.worm-1) and juvenile production (juvenile.worm-1) also determines the overall reproduction rate of earthworms in the process of vermitechnology.
Table 5
Reactors | Individual biomass of earthworms(g) | Biomass change (%) | Cocoon Production (n) | Reproduction rate a | Mortality rate (%) |
Initial | Final |
R1 [CD (100%)] | 9.56 ± 0.14 | 19.63 ± 0.85 | + 51.3 | 42.00 ± 1.00 | 6.8 ± 0.3 | 12 |
R2 [PMS:CD(1:2)] | 9.14 ± 0.44 | 22.78 ± 0.55 | + 59.8 | 49.00 ± 2.00 | 7.2 ± 0.1 | 6 |
R3 [PMS:CD(1:1)] | 9.22 ± 0.57 | 24.26 ± 0.36 | + 62.0 | 63.00 ± 1.50 | 7.8 ± 0.2 | 2 |
R4 [PMS:TW:CD(1:1:1)] | 9.41 ± 0.19 | 21.35 ± 0.25 | + 55.9 | 51.50 ± 1.50 | 7.0 ± 0.1 | 6 |
R5 [PMS:TW:CD(1:2:1)] | 9.05 ± 0.28 | 19.18 ± 0.15 | + 52.8 | 44.00 ± 3.00 | 6.4 ± 0.3 | 13 |
R6 [PMS:TW:CD(2:1:1)] | 9.32 ± 0.09 | 18.76 ± 0.28 | + 50.3 | 40.50 ± 1.50 | 6.1 ± 0.2 | 19 |
Biological properties of earthworms E.fetida in different reactors during bioremediation process (Mean ± SD, n = 3). SD = Standard deviation, mean values indicate that difference between reactors is statistically different (ANOVA; Tukey’s t-test, p ˂ 0.05). |
a(cocoon worm− 1) |