3.1 Characterizations of vermicompost and organic manure
Vermicompost and organic manure's physical, chemical, and biological characteristics were determined and shown in Table 2. MC and pH of vermicompost and organic manure ranged from 17.2 to 19.3%,7.9 ± 0.2 to 8.1 ± 0.7, respectively. TC, TN, and SOC of vermicompost and organic manure varied from 3.6 ± 0.7 to 7.8 ± 0.6%, 2.7 ± 0.4 to 5.7 ± 0.8%, 3.2 ± 0.4 to 7.2 ± 0.7%, respectively. The concentration of P and S of vermicompost and organic manure were 20.0 ± 2.7 to 39.1 ± 2.3 g kg− 1, 29.5 ± 2.6 to 68.6 ± 2.2 g kg− 1, respectively. The concentration of K and Ca of vermicompost and organic manure were found 29 ± 2.1 to 60 ± 5.4 mg g− 1, and 230.0 ± 5.8 to 432.7 ± 6.4 mg g− 1, respectively. The total culturable bacteria (CFU) of vermicompost and organic manure was 1.95x105 to 6.75x107g− 1 sample. These results revealed that the chemical (SOC, P, K+, S, TC, TN, Ca2+) and biological (CFU) properties were significantly higher (p < 0.05) in earthworms with fresh cow-dung vermicompost as compared with other vermicompost and organic manure.
Table 2
Characterizations of vermicompost and organic manure
Sample name
|
Moisture content (%)
|
pH
|
TC %
|
TN %
|
SOC %
|
P
(gkg-1)
|
K
(mgg-1)
|
S
(gkg-1)
|
Ca
(mgg-1)
|
CFU
(Isolated bacteria)
|
V1
|
18.4
|
7.9 ± 0.2a
|
6.2 ± 0.4b
|
4.6 ± 0.3b
|
5.9 ± 0.4b
|
27.0 ± 1.4c
|
44 ± 2.2c
|
59.4 ± 2.b
|
374.5 ± 5.5b
|
3.85x106b
|
V2
|
17.2
|
7.9 ± 0.4a
|
4.1 ± 0.4c
|
3.1 ± 0.4d
|
3.5 ± 0.4c
|
25.0 ± 1.1d
|
39 ± 1.1c
|
49.9 ± 1.2c
|
270.7 ± 5.5c
|
2.55x105c
|
V3
|
18.7
|
8.1 ± 0.5a
|
7.8 ± 0.6a
|
5.7 ± 0.8a
|
7.2 ± 0.7a
|
39.1 ± 2.3a
|
52 ± 4.3b
|
68.6 ± 2.2a
|
432.7 ± 6.4a
|
6.75x107a
|
T1
|
19.2
|
8.1 ± 0.7a
|
5.8 ± 0.6b
|
3.7 ± 0.4c
|
5.6 ± 0.5b
|
32.1 ± 3.4b
|
60 ± 5.4a
|
56.7 ± 3.0b
|
372.0 ± 8.7b
|
4.55x106b
|
M1
|
19.3
|
7.9 ± 0.5a
|
3.6 ± 0.7c
|
2.7 ± 0.4d
|
3.2 ± 0.4c
|
20.0 ± 2.7e
|
29 ± 2.1d
|
29.5 ± 2.6d
|
230.0 ± 5.8d
|
1.95x105c
|
V1 = raw material earthworm plus cow-dung that was stored more than 1 years, V2 = raw materials earthworm plus cow-dung with waste materials, V3 = raw materials earthworm plus fresh cow-dung that was stored in 40 days, T1 = raw materials waste materials with Trichoderma, M1 = raw materials cow-dung with waste materials. a, b, c, d, and e indicate significant difference at 5% level. MC- Moisture content, TC- Total carbon, TN- Total nitrogen, SOC- Soil organic carbon, P- Phosphorus, K- Potassium, S- Sulphur, Ca- Calcium, and CFU- Colony forming Unit. |
3.2 Determination of NNRs and NPs in vermicompost and organic manure
The NNRs in vermicompost and organic manure were quantified and fluctuated from 5.14 ± 0.82to 21.98 ± 0.23mg kg− 1 h− 1. Similarly, the NPs in vermicompost and organic manure were from 3.07 ± 0.16 mg kg− 1 h− 1to 18.70 ± 0.60 mg kg− 1 h− 1(Fig. 1). NNR was highest in the fresh cow-dung vermicompost and lowest in the cow-dung manure, demonstrating that earthworms in fresh cow-dung affect the NNR and NP. According to a Pearson correlation study, as shown in Table S2, the chemical characteristics SOC, P, Ca, S, TC, and TN were positively linked (p < 0.001) with NPs and NNRs. The Pearson correlation analysis further showed that the chemical characteristics (MC, pH, and K) did not correlate with NNR and NP. The scatter plot matrix analysis in Fig. 2(a) shows that NNR was linearly linked with SOC, P, Ca, S, TC, and TN in vermicompost and organic manure. Similarly, as shown in Fig. 2(b), NP had linear relationships with SOC, P, TC, and TN. The chemical characteristics TN, TC, SOC, and P displayed linear correlations with one another according to the dependent variables NNRs and NPs (Fig. 2).
3.3 PTEs quantification and contamination levels in vermicompost and organic manure
The PTE contents in vermicompost and organic manure are shown in Table 3. The respective PTE contents in vermicompost and organic manure were 21.5 ± 0.9 and 318.5 ± 12.6 mg kg-1 f for Pb2+, 12.3 ± 0.4, and 41.4 ± 0.8 mg kg-1 for Cu2+, 2.9 ± 0.7 and 116.0 ± 4.4 mg kg-1 for Zn2+, 1.3 ± 0.2 and 10.5 ± 0.2 mg kg-1 for Cd2+, and 436.8 ± 5.6 and 22533.9 ± 897.8 mg kg-1 for Fe3+. Cow-dung manure had the greatest quantities of Cu2+, Zn2+, Pb2+, and Cd2+, when compared to other vermicompost and organic manure. While the concentrations of Fe2+ were the highest in stored cow-dung vermicompost as compared to other vermicompost and organic manure see details in Table 3. According to PI analysis, stored cow-dung vermicompost, cow-dung, and waste vermicompost, fresh cow-dung vermicompost showed moderate Cu2+ contamination whereas cow-dung manure showed slight pollution. Fe2+waste materials with Trichoderma and cow-dung manure show very slight contamination whereas stored cow-dung vermicompost showed severe contamination. For Pb2+, stored cow-dung vermicompost showed slight contamination, whereas waste materials with Trichoderma and cow-dung manure showed moderate pollution. For Cd2+, Zn2+ stored cow-dung vermicompost showed slight pollution/very slight pollution, whereas cow-dung manure showed very severe pollution/very severe contamination. NNR and NP were positively linked (p < 0.001) with Fe2+ in vermicompost and organic manure, according to a Pearson correlation study. Table S3 shows that Cu2+, Pb2+, Cd2+, and Zn2+ were negatively linked with NNR and NP. As demonstrated in Fig. 2c, the Scatter plot matrix study showed that NNR were linearly linked with Fe2+ in vermicompost and organic manure. Similarly, in vermicompost and organic manure, NP was linearly linked with Fe2+ (Fig. 2d). Scatter plot matrix study also showed that the interactions of all PTEs had a linear association between Cu2+, Pb2+, Cd2+, and Zn2+.
Table 3
PTEs and contamination level in vermicompost and organic manure
Heavy metals
|
Sample
|
Concentration (mg kg− 1)
|
Target value (mg kg− 1)
|
Contamination factor (CF)
|
Contamination
|
Cu2+
|
V1
|
12.3 ± 0.4e
|
36
|
0.34
|
Moderate contamination
|
V2
|
17.4 ± 0.9c
|
0.48
|
Moderate contamination
|
V3
|
15.4 ± 0.9d
|
0.42
|
Moderate contamination
|
T1
|
34.5 ± 1.4b
|
0.96
|
Very severe contamination
|
M1
|
41.4 ± 0.8a
|
1.15
|
Slight pollution
|
Fe2+
|
V1
|
22533.9 ± 97.7a
|
38000
|
0.59
|
Severe contamination
|
V2
|
11535.9 ± 3.3c
|
0.30
|
Moderate contamination
|
V3
|
16103.5 ± 52.3b
|
0.42
|
Moderate contamination
|
T1
|
464.1 ± 3.3d
|
0.01
|
Very slight contamination
|
M1
|
436.8 ± 5.6e
|
0.01
|
Very slight contamination
|
Pb2+
|
V1
|
21.5 ± 0.9e
|
85
|
0.25
|
Slight contamination
|
V2
|
26.1 ± 1.6d
|
0.31
|
Moderate contamination
|
V3
|
30.5 ± 1.8c
|
0.36
|
Moderate contamination
|
T1
|
186.6 ± 4.5b
|
2.20
|
Moderate pollution
|
M1
|
318.5 ± 12.6a
|
3.75
|
Moderate pollution
|
Cd2+
|
V1
|
1.3 ± 0.2e
|
0.8
|
1.67
|
Slight pollution
|
V2
|
3.4 ± 0.2c
|
4.27
|
Severe pollution
|
V3
|
2.5 ± 0.1d
|
3.09
|
Moderate pollution
|
T1
|
9.9 ± 0.3b
|
12.34
|
Very severe pollution
|
M1
|
10.5 ± 0.1a
|
13.09
|
Very severe pollution
|
Zn2+
|
V1
|
2.9 ± 0.7e
|
140
|
0.02
|
Very slight contamination
|
V2
|
4.5 ± 0.9c
|
0.03
|
Very slight contamination
|
V3
|
3.7 ± 0.2d
|
0.03
|
Very slight contamination
|
T1
|
76.8 ± 5.9b
|
0.55
|
Very slight contamination
|
M1
|
115.9 ± 4.4a
|
0.83
|
Very severe contamination
|
a, b, c, d, and e indicate significant difference at 5% level. V1 = raw material earthworm plus cow-dung that was stored more than 1 years, V2 = raw materials earthworm plus cow-dung with waste materials, V3 = raw materials earthworm plus fresh cow-dung that was stored in 40 days, T1 = raw materials waste materials with Trichoderma, M1 = raw materials cow-dung with waste materials. |
3.4 Abundances of comammox N. inopinata in vermicompost and organic manure and relations with AOB and NOB
The abundances of comammox N. Inopinata bacteria fluctuated from 3.76E + 05 ± 6.73E + 04 to 5.29E + 06 ± 3.42E + 05 copies number per g vermicompost and organic manure. This result showed that the abundances of comammox N. inopinata bacteria were the highest in vermicompost and the lowest in cow-dung manure, as shown in Fig. 3(a). Pearson correlation study revealed that comammox N. inopinata bacteria was positively linked (p < 0.001) with NNR and NP see details in Table S4. The range of copy number of comammox N. inopinata, AOB, and NOB was from 3–5%, 59–81%, and 13–36% in vermicompost and organic manure, respectively (Fig. 3b).
3.5 Abundance of the total bacterial community (16S rDNA) and AOA, AOB, NOB, and relative abundances in vermicompost and organic manure
Abundances of 16S rDNA fluctuated from 3.61E + 11 ± 8.23E + 10 to 1.13E + 12 ± 6.86E + 11 (not shown in Fig. 4), and AOA, AOB, and NOB were observed at 3.44E + 07 ± 4.83E + 06 to 2.55E + 08 ± 9.49E + 06, 8.20E + 06 ± 4.22E + 05 to 7.90E + 07 ± 4.11E + 06, and 3.30E + 06 ± 5.07E + 05 to 1.27E + 07 ± 5.95E + 06 copies number g− 1vermicompost and organic manure, respectively as shown in Fig. 4A. These results confirmed that the abundances of 16S rDNA copies were higher than those of AOA, AOB, and NOB. The abundances of AOA, AOB, and NOB significantly differed between organic manure and vermicompost samples. AOA, AOB, and NOB relative abundances varied from 9.53E-05 ± 9.40E-06 to 2.26E-04 ± 1.01E-05, 1.38E-05 ± 1.40E-06 to7.02E-05 ± 3.65E-06, 9.15E-06 ± 5.88E-07 to 1.13E-05 ± 5.28E-06 copies number g− 1vermicompost and manure, respectively (Fig. 4. B). Pearson correlation among NNR and NP with an abundance of CFU, 16S rDNA, AOA, AOB, and NOB were represented in Table S4. According to these findings, NNR and NP were strongly positively linked with CFU, 16S rDNA, AOA, AOB, and NOB. In vermicompost and organic manure, a scatter plot matrix study discovered that NNR were linearly linked with AOB, NOB, and AOA as shown in Fig. 5A. In vermicompost and organic manure, NP had a linear relationship with AOB, NOB, and AOA (Fig. 5B). The scatter plot matrix study also discovered a linear association among the interactions of all bacteria and archaea in AOA, AOB, comammox N. inopinata, and 16S rDNA. Based on PCA the chemical properties, of NR, NP, PTEs, CFU, 16S rDNA, AOB, NOB, AOA, comammox N. inopinata, of vermicompost and manure were shown in Fig. 6. According to the PCA results, cow dung manure was unique and distinct from other vermicompost and manure. This outcome also demonstrated that waste materials with Trichoderma manure were distinct and separate from other vermicompost and organic manure.