Impact of earthworms on suppressing dissemination of antibiotic resistance genes during vermicomposting treatment of excess sludge.

Earthworms play a crucial role in suppressing the dissemination of antibiotic resistance genes (ARGs) during vermicomposting. However, there is still a lack of how earthworms influence the spread of ARGs. To address this gap, a microcosm experiment was conducted, incorporating earthworms and utilizing metagenomics and quantitative PCR to assess the impact of earthworms on microbial interactions and the removal of plasmid-induced ARGs. The findings revealed that vermicomposting led to a reduction in the relative abundance of ARGs by altering microbial communities and interactions. Significantly, vermicomposting demonstrated an impressive capability, reducing 92% of ARGs donor bacteria and impeding the transmission of 94% of the RP4 plasmid. Furthermore, through structural equation model analysis, it was determined that mobile genetic elements and environmental variables were the primary influencers of ARG reduction. Overall, this study offers a fresh perspective on the effects of vermicomposting and its potential to mitigate the spread of ARGs.

Elimination of antibiotic-resistant bacteria and resistance genes by earthworms during vermifiltration treatment of excess sludge.

Vermifiltration (VF) and a conventional biofilter (BF, no earthworm) were investigated by metagenomics to evaluate the removal rates of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs), and class 1 integron-integrase (intI1), as well as the impact mechanism in combination with the microbial community. According to the findings of qPCR and metagenomics, the VF facilitated greater removal rates of ARGs (78.83% ± 17.37%) and ARB (48.23% ± 2.69%) than the BF (56.33% ± 14.93%, 20.21% ± 6.27%). Compared to the control, the higher biological activity of the VF induced an increase of over 60% in the inhibitory effect of earthworm coelomic fluid on ARB. The removal rates of ARGs by earthworm guts also reached over 22%. In addition, earthworms enhanced the decomposition of refractory organics, toxic, and harmful organics, which led to a lower selective pressure on ARGs and ARB. It provides a strategy for reducing resistant pollution in sewage treatment plants and recognizing the harmless stability of sludge.

Effects of Hydrophobic Biochar-Modified Landfill Soil Cover on Methane Oxidation.

Landfill cover soils play an important role in mitigating landfill methane (CH4) emissions. Incorporating biochar into the soil has proven effective in reducing CH4 emissions. However, the role of hydrophobic biochar in this context remains underexplored. This study investigated the CH4 removal efficiency of a biochar-modified landfill soil cover column (RB) and hydrophobic biochar-modified landfill soil cover column (RH) under varying CH4 influx gas concentrations (25 and 35%), simulated CH4 inflow rates (10, 15, and 20 ml/min), and temperatures (20, 25, 30, 35, and 40 °C). RH consistently outperformed RB in terms of CH4 removal efficiency under these experimental conditions. The optimal conditions for CH4 degradation by both RB and RH were observed at a CH4 influx gas concentration of 35%, a simulated CH4 inflow rate of 10 ml/min, and a temperature of ~30 °C. RH achieved a CH4 removal rate of up to 99.96%. In summary, the addition of hydrophobic biochar enhanced the air permeability and hydrophobicity of landfill cover soils, providing a promising alternative to conventional cover soils for reducing CH4 emissions from landfills.

Electric field applications enhance the electron transfer capacity of dissolved organic matter in sludge compost.

Dissolved organic matter (DOM) plays an important role in heavy metal passivation and organic pollutant degradation owing to its redox ability. The structure and composition of DOM are determinants of redox ability changes during composting. Electric field-assisted aerobic composting (EAAC) has been shown to promote the degradation and humification of organic matter in compost. However, how EAAC affects the redox ability of DOM remains unclear. Hence, electron transfer capacity (ETC) of DOM extracted from EAAC was studied using the electrochemical method. Various spectral methods, such as excitation-emission matrix and ultraviolet and visible spectrophotometry were used to study the relationship of ETC with the compositional and structural changes of DOM. Results indicated that EAAC enhanced ETC of DOM at the later stage of composting, and ETC of DOM extracted from the final EAAC product was 10.4% higher than that of the control group. Spectral and correlation analyses showed that EAAC resulted in structural and compositional changes of DOM, and humification degree, aromatic compounds, molecular weight, and fulvic- and humic-like substance contents were improved in EAAC. This conversion increased ETC of DOM. Results of this study will contribute to the understanding of the redox of DOM and in expanding the application of EAAC products.

Influence of earthworms on the nitrogen transfer of sewage sludge in the vermifilter process.

A 6-year laboratory study was conducted to explore the performance of the vermifilter (VF) on reducing sewage sludge. Sewage sludge was found to be reduced significantly in the VF and exhibited a better performance of sludge reduction as compared with the conventional biofilter (BF), which could be traced through the nitrogen-rich organic matter. The nitrogen stable isotope technology was applied to study the matter flow of sewage sludge in the VF process and the influence of earthworms Eisenia fetida on sewage sludge reduction. Results showed that (1) the protein material could be consumed more than the polysaccharide and lipid materials, respectively, in the VF; (2) the presence of earthworms could enhance the consumption capacity of the VF on the protein-rich material of the sewage sludge; (3) earthworms played a leading role on the nitrogen lifting of the sewage sludge, leading to the performance difference of the effluent sludge or biofilm sludge between the VF and BF; (4) in the VF, nitrogen-isotope accumulation in either biofilms or earthworms happened more significantly in the lower layer than in the upper one, while nitrogen transfer of sewage sludge happened more significantly in the upper layer than in the lower one; (5) earthworms improved the feeding environment of organisms (microorganisms in biofilms, moth fly larvae, Limacidaes, and Lymnaeidaes); (6) biofilms, Limacidaes, and moth fly larvae performed better than leeches, Lymnaeidaes, and moth fly adults at nitrogen transfer of sewage sludge.

Study on the biomass and size spectra of bio-particles in vermifilter biofilms.

In biological processes of sludge treatment, the sludge yield is closely related to the energy dissipation of entire microbial system. The vermifilter (VF), a novel biofilter, works efficiently due to the introduction of earthworms, which modifies the energy flow pathway through the variations of microbial size structure. For a deep insight into the sludge reduction in the VF, the biomass size spectrum (BSS) was employed to map the energy dissipation in the VF. The results indicated that bio-particles in the size class of [31, 63] µm were reduced most in the excess sludge after the VF treatment. In biofilms, bio-particles in the size class of [31, 63] µm varied most with the filter depth and earthworm density. Eight biomass and size spectra (BSS) were established for all beds of the VF and BF (the control of the VF, without earthworms). The normalized BSS were all linear both in the VF and BF, and their linear regression parameters, the slopes (k) and intercepts (b), varied with the filter depth and the earthworm density. The k and b of the VF were both significantly different from those of the BF. According to the k, the productivity level of largest bio-particles was higher in the VF than in the BF. According to the b, bio-particles at the bottom of size structure could be taken faster in the VF than in the BF. At last, some improvement approaches with some tries were proposed to enhance the sludge treatment capacity of the VF.

Exploring the effects of earthworms on bacterial profiles during vermicomposting process of sewage sludge and cattle dung with high-throughput sequencing.

This study aims to reveal the effects of earthworms (Eisenia fetida) on bacterial profiles during the vermicomposting process of sewage sludge and cattle dung with the high-throughput sequencing technology. The earthworms could accelerate organic degradation and improve the stabilization process. Moreover, the addition of earthworms not only affected the bacterial numbers, but also increased the bacterial community diversity. The activity of earthworms had significant effects on the bacterial community structure as the bacterial community was clearly different between the vermicomposting and the control treatment. Furthermore, the earthworms affected the physical and chemical properties of substrates, thus promoting the growth of some microorganisms, such as Flavobacteria, Acidbacteria, and Planctomycetes. Earthworms largely inhibited the growth of various human pathogenic bacteria. In summary, earthworms significantly affected the bacterial community in vermicomposting and it could be applied as an authentically effective technique for the stabilization of organic wastes.

Optimizing vermifilter depth by process performance collaborated with the evolutions of microbial characteristics during sewage sludge treatment.

The present study focuses on optimizing filter depth on sludge reduction in a four-stage vermifiltration during the course of treating excess sludge continuously. The results indicated that when the filter depth exceeded 75 cm, though the fourth stage can further advance the sludge reduction, its contribution for the total sludge reduction was lower than 10%, while the aerobic bacteria, especially the dominant bacteria (Proteobacteria and Bacteroidetes), kept a high similarity as the filter depth varied. Furthermore, earthworm activities attributed to aerobic bacteria being preferentially selected in the system, positively supporting the organic decomposition. As far as economic cost and process performance are concerned, a 75-cm vermifilter was recommended to efficiently and economically achieve the required standard for sewage sludge reduction and stabilization.

Interaction of earthworms-microbe facilitating biofilm dewaterability performance during wasted activated sludge reduction and stabilization.

Sludge dewaterability was chemically and morphologically explored during sewage sludge treatment by vermifiltration. The results, with a conventional biofilter (BF, no earthworms) as a control, demonstrated that the capillary suction time(CST) and specific resistance of filtration(SRF) of vermifilter (VF, with earthworms) treated sludge were 64.9±1.7s and (23.1±1.3)×1012m/kg,16.8% and 36.0% lower than that of the BF, respectively. Additionally, the VF could efficiently decompose loosely bound extracellular polymeric substances (LB-EPS), releasing more water trapped inside biofilm. Furthermore, the VF enable to reduce the electrostatic repulsive forces between particles, verified by 9.61±0.19mV of the absolute value of zeta potential, 19.6% lower than that of the BF. Notably, based on scanning electron microscopy (SEM) analysis, the fractal dimension (Df) of sludge floc structure stated that more small and loose sludge flocs tended to aggregate into bigger inorganic particles. Therefore sludge flocs with highly compacted structure and smooth surface can transform part of vicinal water and water of hydration into easier-removed interstitial water, improving the dewaterability.

Influence analysis for the behavior of dewaterability of excess sludge in a two-stage vermifilter.

To improve excess sludge dewaterability, a two-stage vermifilter was developed to qualitatively and quantitatively analyze sludge physico-chemical properties (fractal dimension, zeta potential, extracellular polymeric substances (EPS), particle size distribution, etc.) and to correlate them with sludge dewatering characteristics (specific resistance to filtration (SRF) and capillary suction time (CST)). Results demonstrated that sludge dewatering performance was significantly improved after the primary vermifilter VF1 and the second-stage vermifilter VF2. In addition, the further VF2 treatment exhibited higher effects on sludge dewatering performance. The particle boundary of sludge after VF2 treatment was clearer and smoother than VF1 sludge (VF1S), apart from the fact that sludge morphological structure got denser and more compact. Comparing with VF1S, the fractal dimension D1 calculated within 1D topological space was closer to 1 after VF2 treatment, and the fractal dimension D2 within 2D topological space closer to 2, indicating a better dewatering performance after VF2 treatment. Additionally, the changes of sludge floc surface properties (such as zeta potential and EPS) resulted in small particles agglomerating into larger ones and then the increase of particle diameter. In summary, the two-stage vermifilter got a better sludge dewatering performance, and thus beneficial for subsequent processing of sludge.

Highlighting earthworm contribution in uplifting biochemical response for organic matter decomposition during vermifiltration processing sewage sludge: Insights from proteomics.

A vermifilter (VF) was steadily operated to explore the mechanism of lower microbial biomass and higher enzymatic activities due to the presence of earthworms, with a conventional biofilter (BF) as a control. The analysis of 2-DE indicated that 432 spots and 488 spots were clearly detected in the VF and BF biofilm. Furthermore, MALDI-TOF/TOF MS revealed that six differential up-regulated proteins, namely Aldehyde Dehydrogenase, Molecular chaperone GroEL, ATP synthase subunit alpha, Flagellin, Chaperone protein HtpG and ATP synthase subunit beta, changed progressively. Based on Gene Ontology annotation, these differential proteins mainly performed 71.38% ATP binding and 16.23% response to stress functions. Taken the VF process performance merits into considerations, it was addressed that earthworm activities biochemically strengthened energy releasing of the microbial metabolism in an uncoupled manner.

Investigation on the difference between biofilm morphologies of the vermifilter and conventional biofilter with the flow cytometer.

With the demand of new sludge reduction processes, a vermifilter (VF) was studied based on a conventional biofilter (BF). The biofilm morphology was investigated using a new technique, the flow cytometer (FCM), to find a way to optimize VF structure. VF was inoculated with Eisenia fetida, packed with ceramsites, and operated stably at the organic load of 1.2kg-VSSm(-3)d(-1) with BF as the control. Compared with BF, VF had about 13% more removal efficiency of excess sludge and 45% shorter biofilm update period. FCM profile showed the morphology of microbial cells in VF biofilms was significantly different from that in BF in upper layers, with decreases of average refractive index (about 72%) and size (about 22%), and suggested it was better to keep earthworms there to remove rod-shaped microorganisms with other filter media in lower layers to remove spherical ones combining the findings in SEM images and extracellular polymeric substances.

Speciation and transformation of heavy metals during vermicomposting of animal manure.

This work was conducted to evaluate the effects of vermicomposting on the speciation and mobility of heavy metals (Zn, Pb, Cr, and Cu) in cattle dung (CD) and pig manure (PM) using tessier sequential extraction method. Results showed that the pH, total organic carbon and C/N ratio were reduced, while the electric conductivity and humic acid increased after 90days vermicomposting. Moreover, the addition of earthworm could accelerate organic stabilization in vermicomposting. The total heavy metals in final vermicompost from CD and PM were higher than the initial values and the control without worms. Sequential extraction indicated that vermicomposting decreased the migration and availability of heavy metals, and the earthworm could reduce the mobile fraction, while increase the stable fraction of heavy metals. Furthermore, these results indicated that vermicomposting played a positive role in stabilizing heavy metals in the treatment of animal manure.

Microbial enzyme and biomass responses: Deciphering the effects of earthworms and seasonal variation on treating excess sludge.

This paper reports on a seasonal pattern comparison of microbial enzymatic activities and biomass responses based on a conventional biofilter (BF, without earthworm) and a vermifilter (VF, with earthworm, Eisenia fetida) for excess sludge treatment. The volatile suspended solids (VSS) reduction, viable cell number and enzyme activities were assayed to probe what made the VF operate stably. The results indicated that the earthworm activities can polish the VSS reduction with 27.17% more than the BF. Though the VF had a lower level in the viable cell number compared with the BF, the earthworm strongly improved the microbial enzymatic activities such as INT-dehydrogenase, protease, ß-glucosidase and amylase, which can explain the excellent performance of VSS reduction. The correlation analysis documented that the VSS reduction was positively correlated with microbial enzyme activities. More importantly, the earthworm enabled the VF to avoid the detrimental influence of temperature, which guaranteed a stable performance during seasonal variations.

Addressing the role of earthworms in treating domestic wastewater by analyzing biofilm modification through chemical and spectroscopic methods.

Vermifiltration eco-friendly system is an alternative and low-cost artificial ecosystem for decentralized wastewater treatment and excess sludge reduction. The biofilm characteristics of a vermifilter (VF) with earthworms, Eisenia fetida, for domestic wastewater treatment were studied. A conventional biofilter (BF) without earthworms served as the control. Pore number in VF biofilm was significantly more than BF biofilm, and VF biofilm showed a better level-administrative structure through scanning electron microscope. VF biofilms had lower levels of protein and polysaccharide, but phosphoric acids and humic acid showed the opposite results. Furthermore, in the presence of earthworms, VF biofilms contained higher total organic carbon (TOC) percentage composition in the condition of less volatile suspended substances (VSS) contents. Dehydrogenase activity (DHA) and adenosine triphosphate (ATP) contents along VF showed better results than BF by increment of 12.84 ∼ 16.46 %. Overall findings indicated that the earthworms' presence remarkably decreases biofilm contests but increases enzyme activity and improves the community structure of VF biofilms, which is beneficial for the wastewater disposal.

Pyrosequencing reveals bacterial community differences in composting and vermicomposting on the stabilization of mixed sewage sludge and cattle dung.

This study aimed to compare the microbial community structures and compositions in composting and vermicomposting processes. We applied 454 high-throughput pyrosequencing to analyze the 16S rRNA gene of bacteria obtained from bio-stabilization of sewage sludge and cattle dung. Results demonstrated that vermicomposting process presented higher operational taxonomic units and bacterial diversity than the composting. Analysis using weighted UniFrac indicated that composting exhibited higher effects on shaping microbial community structure than the vermicomposting. The succession of dominant bacteria was also detected during composting. Firmicutes was the dominant bacteria in the thermophilic phase of composting and shifted to Actinomycetes in the maturing stage. By contrast, Proteobacteria accounted for the highest proportions in the whole process of the vermicomposting. Furthermore, vermicomposting contained more uncultured and unidentified bacteria at the taxonomy level of genus than the composting. In summary, the bacterial community during composting significantly differed from that during vermicomposting. These two techniques played different roles in changing the diversity and composition of microbial communities.

Tracking the composition and transformation of humic and fulvic acids during vermicomposting of sewage sludge by elemental analysis and fluorescence excitation-emission matrix.

Sewage sludge (T1) and the mixture of sewage sludge and cattle dung (T2) were vermicomposted with Eisenia fetida, respectively. The transformation of humic acid (HA) and fulvic acid (FA) extracted from these two treatments were evaluated by a series of chemical and spectroscopic methods. Results indicated that the vermicomposting decreased pH, TOC, and C/N ratio, and increased EC, total extractable C, and HA contents. The FA content in treatment T1 was increased significantly, and only slight increasing was observed in treatment T2. Moreover, vermicomposting decreased H content, C/N ratio, proteinaceous and carbohydrates components, and increased the N content, C/H ratio, aromatic compounds and polycondensation structures in HA and FA. In addition, fluorescence spectra and fluorescence regional integration indicated that protein-like groups were degraded and HA compounds were formed. Furthermore, the addition of cattle dung enhanced the humification process and improved the HA quality in spite of no significant effect on the FA.

Towards understanding the effects of additives on the vermicomposting of sewage sludge.

This work evaluated the effects of additives on the chemical properties of the final products (vermicompost) from vermicomposting of sewage sludge and the adaptable characteristics of Eisenia fetida during the process. An experimental design with different ratios of sewage sludge and the additives (cattle dung or pig manure) was conducted. The results showed that the vermicomposting reduced total organic carbon and the quotient of total organic carbon to total nitrogen (C/N ratio) of the initial mixtures and enhanced the stability and agronomical value of the final products. Notably, principal component analysis indicated that the additives had significant effects on the characteristics of the vermicomposts. Moreover, the vermibeds containing cattle dung displayed a better earthworm growth and reproduction than those with pig manure. Additionally, redundancy analysis demonstrated that electrical conductivity (EC), pH, and C/N ratio played crucial roles on earthworm growth and reproduction. In all, the additives with high C/N ratio, pH buffering capacity, and low EC are recommended to be used for vermicomposting of sewage sludge.

Towards understanding the stabilization process in vermicomposting using PARAFAC analysis of fluorescence spectra.

In this study, fluorescence excitation-emission matrix (EEM) combined with parallel factor analysis (PARAFAC) was employed to trace the behavior of water extractable organic matter and assess the stabilization process during vermicomposting of sewage sludge and cattle dung. Experiments using different mixing ratios of sewage sludge and cattle dung were conducted using Eisenia fetida. The results showed that vermicomposting reduced the DOC, DOC/DON ratio and ammonia, while increased the nitrate content. A three-component model containing two humic-like materials (components 1 and 2) and a protein-like material (component 3) was successfully developed using PARAFAC analysis. Moreover, the initial waste composition had a significant effect on the distribution of each component and the addition of cattle dung improved the stability of sewage sludge in vermicomposting. The PARAFAC results also indicated that protein-like materials were degraded and humic acid-like compounds were evolved during vermicomposting. Pearson correlation analysis showed that components 2 and 3 are more suitable to assess vermicompost maturity than component 1. In all, EEM-PARAFAC can be used to track organic transformation and assess biological stability during the vermicomposting process.

Feeding behavior and trophic relationship of earthworms and other predators in vermifiltration system for liquid-state sludge stabilization using fatty acid profiles.

The sludge reduction capability (VSS reduction) of vermifilter (VF) was 14.7% higher than that of conventional biofilter (BF) due to the fact that there was a net loss of biomass and energy when the food web in VF is extended. Therefore, feeding behavior and trophic relationship of earthworms and other predators (leeches, lymnaeidaes and limaxes) in VF were investigated using fatty acid (FA) profiles for the first time. Compared with BF biofilm, microbial community structure of VF biofilm got optimized by earthworms that the percentage of protozoa increased from 14.2% to 20.4%. Furthermore, analysis of specific microbial FAs composition in each predator suggested different trophic level of predators resulted from their selective ingestion of different microorganisms, and earthworms were at the second high trophic level in VF food web. Overall findings indicated earthworms modified microbial community and extended the food web of VF and thus enhanced the sludge reduction.