Combined transcriptome and metabolome analysis reveals the toxic effects of antimony on the earthworm.

Antimony (Sb) poses a significant ecological threat. This study combines biochemical, pathological, transcriptome, and metabolome analyses to assess the short-term (14-day) toxic impact of two Sb levels (25 mg/kg and 125 mg/kg) on earthworms (Eisenia fetida). Higher Sb concentration caused severe intestinal damage, elevated metallothionein (MT) levels, and reduced antioxidant capacity. Metabolome analysis identifies 404 and 1698 significantly differential metabolites in the two groups. Metabolites such as S(-)-cathinone, N-phenyl-1-naphthylamine, serotonin, 4-hydroxymandelonitrile, and 5-fluoropentylindole contributed to the metabolic responses to Sb stress. Transcriptome analysis shows increased chitin synthesis as a protective response, impacting amino sugar and nucleotide sugar metabolism for cell wall synthesis and damage repair. Integrated analysis indicated that 5 metabolite-gene pairs were found in two Sb levels and 11 enriched pathways were related to signal transduction, carbohydrate metabolism, immune system, amino acid metabolism, digestive system, and nervous system. Therefore, the integration of multiomics approaches enhanced our comprehension of the molecular mechanisms underlying the toxicity of Sb in E. fetida.

Toxicological effects induced by two carbamates on earthworms (Eisenia fetida): Acute toxicity, arrested regeneration and underlying mechanisms.

Eisenia fetida is recognised as advantageous model species in ecotoxicological and regeneration investigations. The intensive utilization of carbamate pesticides (CARs) imposes heavy residue burdens and grave hazards on edaphic environments as well as soil fauna therein. However, precise mechanisms whereby the specific CAR exerted toxic effects on earthworms remain largely elusive, notably from regenerative perspective. Herein, acute responses and regenerative toxicity of two carbamates (metolcarb, MEB and fenoxycarb, FEB) against E. fetida were dissected using biochemical, histological as well as molecular approaches following OECD guidelines at the cellular, tissue and organismal level. The acute toxicity data implied that MEB/FEB were very toxic/medium to extremely toxic, respectively in filter paper contact test and low to medium toxic/low toxic, respectively in artificial soil test. Chronic exposure to MEB and FEB at sublethal concentrations significantly mitigated the soluble protein content, protein abundance while enhanced the protein carbonylation level. Moreover, severely retarded posterior renewal of amputated earthworms was noticed in MEB and FEB treatments relative to the control group, with pronouncedly compromised morphology, dwindling segments and elevated cell apoptosis of blastema tissues, which were mediated by the rising Sox2 and decreasing TCTP levels. Taken together, these findings not only presented baseline toxicity cues for MEB and FEB exposure against earthworms, but also yielded mechanistic insights into regenerative toxicity upon CAR exposure, further contributing to the environmental risk assessment and benchmark formulation of agrochemical pollution in terrestrial ecosystem.

Toxicokinetics of selenate in earthworm sub-tissues and potential bio-accessibility assessment of earthworm-derived selenium.

Selenium (Se) pollution is mainly caused by anthropogenic activities, and the resulting biosecurity concerns have garnered significant attention in recent years. Using one-compartmental toxicokinetic (TK) modelling, this study explored the kinetic absorption, sub-tissue distribution, and elimination processes of the main Se species (selenate, Se(VI)) in the cultivated aerobic soil of the earthworm Eisenia fetida. The bio-accessibility of earthworm-derived Se was assessed using an in vitro simulated gastrointestinal digestion test to evaluate its potential trophic risk. The results demonstrated that Se accumulated in the pre-clitellum (PC) and total tissues (TT) of earthworms in a time- and dose-dependent manner. The highest Se levels in the PC, post-clitellum (PoC), and TT were 70.54, 57.93, and 64.26 mg/kg during the uptake phase, respectively. The kinetic Se contents in the earthworms PC and TT were consistent with the TK model but not with PoC. The earthworm TT exhibited a faster uptake (Kus = 0.83-1.02 mg/kg/day) and elimination rate of Se (Kee = 0.044-0.049 mg/kg/day), as well as a shorter half-life time (LT1/2 = 15.88-14.22 days) than PC at low soil Se levels (≤5 mg/kg). Conversely, the opposite trend was observed with higher Se concentrations (10 and 20 mg/kg). These results are likely attributable to the tissue specificity and concentration of the toxicant. Earthworms PC and TT exhibited a higher kinetic Se accumulation factor (BAFk) than steady-state BAF (BAFss), with values ranging from 8 to 24 and 3-13, respectively. Furthermore, the bio-accessibility of earthworm-derived Se to poultry ranged from 66.25 % to 84.35 %. As earthworms are at the bottom of the terrestrial food chain, the high bio-accessibility of earthworm-derived Se poses a potential risk to predators. This study offers data support and a theoretical foundation for understanding the biological footprint of soil Se and its toxicological impacts and ecological hazards.

Ecotoxicity of five veterinary antibiotics on indicator organisms and water and soil communities.

This study explores the environmental effects of five common veterinary antibiotics widely detected in the environment, (chlortetracycline,CTC; oxytetracycline,OTC; florfenicol,FF; neomycin, NMC; and sulfadiazine, SDZ) on four bioindicators: Daphnia magna, Vibrio fischeri, Eisenia fetida, and Allium cepa, representing aquatic and soil environments. Additionally, microbial communities characterized through 16 S rRNA gene sequencing from a river and natural soil were exposed to the antibiotics to assess changes in population growth and metabolic profiles using Biolog EcoPlates™. Tetracyclines are harmful to Vibrio fisheri (LC50 ranges of 15-25 µg/mL), and the other three antibiotics seem to only affect D. magna, especially, SDZ. None of the antibiotics produced mortality in E. fetida at concentrations below 1000 mg/kg. NMC and CTC had the highest phytotoxicities in A. cepa (LC50 = 97-174 µg/mL, respectively). Antibiotics significantly reduced bacterial metabolism at 0.1-10 µg/mL. From the highest to the lowest toxicity on aquatic communities: OTC > FF > SDZ ≈ CTC > NMC and on edaphic communities: CTC ≈ OTC > FF > SDZ > NMC. In river communities, OTC and FF caused substantial decreases in bacterial metabolism at low concentrations (0.1 µg/mL), impacting carbohydrates, amino acids (OTC), and polymers (FF). At 10 µg/mL and above, OTC, CTC, and FF significantly decreased metabolizing all tested metabolites. In soil communities, a more pronounced decrease in metabolizing ability, detectable at 0.1 µg/mL, particularly affected amines/amides and carboxylic and ketonic acids (p < 0.05). These new ecotoxicity findings underscore that the concentrations of these antibiotics in the environment can significantly impact both aquatic and terrestrial ecosystems.

Effects of pyroligneous acid on acute, chronic, and cyto-genotoxicity to earthworms (Eisenia fetida).

The pyroligneous acid (PA), or wood vinegar, is a byproduct of wood carbonization during the slow pyrolysis process. PA is recognized globally as a safe compound for agriculture due to its various beneficial properties, such as antioxidant, antibacterial, antifungal, and termiticidal properties. However, the impact of different PA concentrations on beneficial soil organisms, such as earthworms has not been investigated. The present study aims to understand the effects of different PA concentrations on earthworm Eisenia fetida. The earthworms were exposed to nine different concentrations of PA in soils, including their control. The acute toxicity assay was performed after 14 days of exposure, and the chronic toxicity assay was performed up to 8 weeks after exposure. The results from the acute toxicity assay demonstrated no significant effect on earthworm mortality. The chronic toxicity assay showed that lower PA concentrations (0.01-0.2% of weight/weight PA in soil) promoted cocoon and juvenile production in soils, whereas higher PA concentrations (0.5 and 1%) had a negative effect. These findings highlight the potential of PA to enhance soil fertility at lower concentrations, up to 0.2%, by stimulating worm activity and subsequent manure production. The outcomes of this study have significant implications for the careful management of PA concentrations within agricultural operations.

Assessment of growth, reproduction, and vermi-remediation potentials of Eisenia fetida on heavy metal exposure.

Heavy metal pollution is a significant environmental concern with detrimental effects on ecosystems and human health, and traditional remediation methods may be costly, energy-intensive, or have limited effectiveness. The current study aims were to investigate the impact of heavy metal toxicity in Eisenia fetida, the growth, reproductive outcomes, and their role in soil remediation. Various concentrations (ranging from 0 to 640 mg per kg of soil) of each heavy metal were incorporated into artificially prepared soil, and vermi-remediation was conducted over a period of 60 days. The study examined the effects of heavy metals on the growth and reproductive capabilities of E. fetida, as well as their impact on the organism through techniques such as FTIR, histology, and comet assay. Atomic absorption spectrometry demonstrated a significant (P < 0.000) reduction in heavy metal concentrations in the soil as a result of E. fetida activity. The order of heavy metal accumulation by E. fetida was found to be Cr > Cd > Pb. Histological analysis revealed a consistent decline in the organism's body condition with increasing concentrations of heavy metals. However, comet assay results indicated that the tested levels of heavy metals did not induce DNA damage in E. fetida. FTIR analysis revealed various functional group peaks, including N-H and O-H groups, CH2 asymmetric stretching, amide I and amide II, C-H bend, carboxylate group, C-H stretch, C-O stretching of sulfoxides, carbohydrates/polysaccharides, disulfide groups, and nitro compounds, with minor shifts indicating the binding or accumulation of heavy metals within E. fetida. Despite heavy metal exposure, no significant detrimental effects were observed, highlighting the potential of E. fetida for sustainable soil remediation. Vermi-remediation with E. fetida represents a novel, sustainable, and cutting-edge technology in environmental cleanup. This study found that E. fetida can serve as a natural and sustainable method for remediating heavy metal-contaminated soils, promising a healthier future for soil.

Evaluation of anthelminthic and antiprotozoal activity of myrrh (Commiphora myrrha) methanolic extract.

To treat and control parasitic infections, traditional medical remedies using plant products are utilized as antiparasitic agents rather than standard synthetic chemicals due to drug resistance. Myrrh, a resinous exudate of Commiphora myrrha (Burseraceae), is a powerful antioxidant with a variety of medicinal uses. This study aimed to investigate the effect of the myrrh methanolic extract (MyE) of three concentrations (100, 50, and 25 mg/ml) on the sporulation of oocysts and as an anthelminthic effector via in vitro study. Characterization of the plant was done by Fourier-transform infrared spectroscopy (FT-IR). The earthworm, Eisenia fetida, is used as a model worm to evaluate the anthelminthic activity of MyE. Eimeria labbeana-like oocysts are used as a model protozoan parasite in anticoccidial assays. The sporulation and inhibition (%) of E. labbeana-like were assessed by MyE compared to other chemical substances. FT-IR revealed the presence of twelve active compounds. Our results showed that paralysis and death of earthworms at MyE (100 mg/ml) were 7.88 ± 0.37 and 9.24 ± 0.60 min, respectively, which is more potency when compared to mebendazole (reference drug). In all treated worms, microscopic examinations revealed obvious surface architecture abnormality. This study shows that MyE affects oocysts sporulation in a dose-dependent manner. At 24 and 36 hr, a high concentration of MyE (100 mg/ml) inhibits sporulation by 90.95 and 87.17 %. At 36 hr, other concentrations of MyE (50 and 25 mg/ml), as well as amprolium, DettolTM, and phenol inhibits oocyst sporulation by 40.17 %, 29.34 %, 45.09 %, 85.11 %, and 61.58 %, respectively. According to our research, the MyE extract had powerful anthelmintic and anticoccidial properties.

Production and characterization of enriched vermicompost from banana leaf biomass waste activated by biochar integration.

Vermicomposting uses less energy and requires fewer infrastructures, and it is capable of restoring soil nutrition and carbon. Banana cultivation produces lots of trash in a single crop season, with 30 tonnes of waste generated per acre. The biodegradable fraction of banana leaf waste is thrown out in large quantities from temples, markets place wedding halls, hotels, and residential areas. Vermicomposting can be used for recovering lignin, cellulose, pectin, and hemicellulose from banana leaves. Earthworm digests organic materials with the enzymes produced in gut microflora. Biochar adds bulk to vermicomposting, increases its value as fertilizer. The goal of this study was to amend biochar (0, 2, 4 and 6%) with banana leaf waste (BLW) + cow dung (CD) in three different combinations (1:1, 2:1 and 3:1) using Eisenia fetida to produce enriched vermicompost. In the vermicompost with biochar groups, there were higher levels of physicochemical parameters, as well as macro- and micronutrient contents. The growth and reproduction of earthworms were higher in groups with biochar. A maximum of 1.82, 1.18 and 1.67% of total nitrogen, total phosphorus and total potassium was found in the final vermicompost recovered from BLW + CD (1:1) amended with 4% biochar; while the other treatments showed lower levels of nutrients. A lower C/N ratio of 18.14 was observed in BLW + CD (1:1) + 4% biochar followed by BLW + CD (1:1) + 2% biochar amendment (19.92). The FTIR and humification index studies show that degradation of organic matter has occurred in the final vermicompost and the substrates with 4% biochar in 1:1 combination showed better degradation and this combination can be used for nutrient rich vermicompost production.

Study on the life maintenance mechanism of Eisenia fetida under low-density polyethylene stress: Based on path analysis and canonical correlation analysis.

The widespread use but low recovery rate of agricultural films has led to microplastic accumulation in farmlands, which poses a serious threat to the health of the soil ecosystem. There is an urgent need for early warning and monitoring of soil microplastics pollution, as well as the performance of bioremediation research. In this study, earthworms were used as test organisms to carry out toxicological tests under low-density polyethylene (LDPE) stress. A canonical correlation analysis model (CCA) was established to analyze the relationship between oxidative stress and microbial community. A path analysis model (PA) was also constructed to examine the detoxification mechanism of earthworms under LDPE stress. The results showed that low concentrations (100 and 500 mg/kg) of LDPE did not cause oxidative damage to earthworms but stimulated their physiological metabolism. Meanwhile, 1000 mg/kg LDPE concentrations caused oxidative damage to earthworms and altered their internal microbial community structure. Furthermore, at 1500 mg/kg LDPE concentrations, the oxidative stress to the earthworms is aggravated, and their physiological responses work in conjunction with the microbial community to cope with the adverse condition. Lastly, treatment with 2000 mg/kg LDPE induced the appearance of LDPE tolerant populations in the microbial community in vivo. Taken together, our results provide a theoretical basis for revealing the physiological response of earthworms when challenged in a polluted environment and provide a model for pollution remediation and ecological security monitoring of soil ecosystems.

Assessing the chronic toxicity of spreading organic amendments on agricultural soil: Tests on earthworms and plants.

Recycling organic wastes on agricultural soils improves the soil quality, but the environmental and health impact of these organic amendments closely depends on their origins, their bio-physicochemical characteristics and the considered organisms potentially affected. The aim of this study was to assess the potential chronic ecotoxicity of spreading organic amendments on agricultural soils. To do this, we characterized three different organic amendments: sewage sludge from an urban wastewater treatment plant, cow manure and liquid dairy manure. Their chronic ecotoxicity was studied through assays exposing earthworms of the species Eisenia fetida and two plants: Medicago sativa and Sinapis alba. Of the three amendments, the sewage sludge presented the highest concentrations of micropollutants and a considerable fraction of available and biodegradable organic matter. The cow manure and liquid dairy manure had lower chemical contamination and similar characteristics with lower biodegradable fractions of their organic matter. No chronic phytotoxicity was evidenced: on the contrary, particularly with sewage sludge, the germination rate and aerial and root biomass of the two plants increased. Considering earthworms, their biomass increased considerably during the reproduction assays in soil amended with sewage sludge, which contained the more bioavailable organic matter. Nonetheless, the earthworms presented an inhibition close to 78% of the production of juveniles when exposed to sewage sludge exceeding 20 g.kg-1 DW (that means 2 times the agronomic dose). This reprotoxic effect was also observed in the presence of liquid dairy manure, but not with cow manure. At the end of the assays, the glycogen and protein reserves in earthworms exposed to sewage sludge were inferior to that of control earthworms, respectively around 50% and 30%. For the earthworms exposed to liquid dairy manure, protein and lipid reserves increased. In the case of liquid dairy manure, this reprotoxic effect did not appear to be linked to the presence of micropollutants. In conclusion, our results confirm the need to use several ecotoxicity assays at different biological levels and with different biological models to assess the ecotoxic impacts of soil amendments. Indeed, although certain organic wastes present a strong nutritional potential for both plants and earthworms, a not inconsiderable risk was apparent for the reproduction of the latter. An integrated ecotoxicity criterion that takes into account a weighted sum of the different results would guide the utilization of organic amendments while ensuring the good health of agricultural ecosystems.

Toxicity of benzyl paraben on aquatic as well as terrestrial life.

Parabens are derivatives of alkyl esters of p-hydroxybenzoic acid and come in different classes. These compounds are primarily used as antimicrobial preservative agents in many commercial products, including cosmetics and pharmaceuticals. Accordingly, Benzyl paraben (BeP) is known to be a potential endocrine disruptor. The aim of this study was to determine the toxicity of benzyl paraben (BeP) on aquatic and terrestrial organisms, specifically Scenedesmus sp., Moina macrocopa, and Eisenia fetida. All the organisms were treated with different concentrations of BeP (0.025 mg/L and 1000 mg/L), and LC25, LC50, and LC90 values were used to measure the toxicity levels. Results showed the LC values of BeP for M. macrocopa (3.3 mg/L, 4.7 mg/L, 7.3 mg/L) and E. fetida (173.2 mg/L, 479.8 mg/L, 1062 mg/L), respectively. Toxicity tests on green algae (Scenedesmus sp.) were conducted, the green algae were subjected to various BeP concentration. At 50 mg/L of BeP, cell viability was reduced to 56.2% and the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay indicated 35.4% viable cells. The chlorophyll value and the biochemical parameters of the algal cells were corroborative with the cell viability test. Lethal indices (LC50) for M. macrocopa and E. fetida were evaluated for their toxicity on biochemical properties and were found to be catalase (0.111 mg/L, 0.5 mg/L), lipid peroxidation (0.072 mg/L, 0.056 mg/L), and total protein (0.309 mg/L, 0.314 mg/L), respectively. Overall, this study demonstrated the toxic impact of BeP on non-target aquatic as well as terrestrial species.

Evaluation of the treatment performance and reuse potential in agriculture of organized industrial zone (OIZ) wastewater through an innovative vermifiltration approach.

Vermifiltration (VF) is a natural and sustainable biofilter that has many advantages, including being energy-free, cost-effective, and allowing ease of application and maintenance. In this study, the effectiveness of a lab-scale VF system was assessed by the removal efficiency of total suspended solids, electrical conductivity, chemical oxygen demand, total nitrogen, total phosphorus, fecal coliform, and heavy metals in organized industrial zones (OIZ) and domestic wastewater (DW) for the first time. Additionally, the reuse suitability of the treated wastewater was determined by comparing different countries' and global irrigational criteria. The lab systems were built with four layers: one worm-bed and three varying filtering materials, and operated at an optimum hydraulic loading rate of 1.8-2 m3/m2/day for 45 days with Eisenia fetida as the earthworm species. The results demonstrated that removal efficiencies of total suspended solids and chemical oxygen demand were found to be 95% and 80% in OIZ wastewater and 90% and 88% in DW, respectively. Total nitrogen and total phosphorus were removed at rates of 69% and 67% in OIZ wastewater, respectively, and 84% and 74% in DW. Besides, the VF system has shown satisfactory removal performance for heavy metals ranging from 51% to 77% in OIZ wastewater that has met Turkish national wastewater discharge limits. Although the final characterization of treated wastewater was suitable, heavy metal and fecal coliform levels have not met many countries' irrigation water quality criteria. To meet global irrigation standards and to enhance the VF performance, further experimental studies should be carried out, including parameters such as bed material type in the reactor, worm type, and different operating conditions.

Influence of vermifiltered wastewater on the morphological, physiological and biochemical developments of Zea mays, Triticum aestivum and Sorghum bicolor seed germination.

BACKGROUND: Reclaimed wastewater is a potential source for irrigation, although its effect on seed germination and plant growth has not been widely investigated. The present study focused on treating wastewater by vermifiltration and investigating the effect on seed germination and seedling development of Zea mays, Triticum aestivum and Sorghum bicolor and their morphological, physiological and biochemical developments. A filter bed was constructed using indigenous and improved materials. The constituents used to develop the filter bed consist of matrix of garden soil, lateritic hardpan gravels and coconut coir inoculated with Eisenia fetida. RESULTS: The wastewater was obtained from a public septic tank toilet facility and contained 35-40% dry matter with high concentrations of total dissolve solids (TDS), total suspended solids (TSS), chemical oxygen demand (COD), biochemical oxygen demand (BOD), total nitrogen (Ntot ) and phosphorus (Pavail ). Portable water was used as the control. Vermifiltration of the wastewater was shown to be effective for the removal of 52-65% TDS, 56-62% TSS, 34-40% COD, 48-52% BOD, 56-62% Ntot and 53-58% Pavail . The results showed that the morphological, physiological and biochemical processes in Z. mays, T. aestivum and S. bicolor improved when seeds were subjected to treated wastewater. CONCLUSION: The present study demonstrated that a potential reuse of vermifiltered wastewater as a source of irrigation water with continuous monitoring of the water quality is productive and sustainable. © 2023 Society of Chemical Industry.

Arsenic toxicity to earthworms in soils of historical As mining sites: an assessment based on various endpoints and chemical extractions.

Eisenia fetida is an earthworm species often used to assess the toxicity of contaminants in soils. Several studies indicated that its response can be unpredictable because it depends both on total concentrations of contaminants and also on their forms that differ in susceptibility to be released from soil solid phase. The issue is complex because two various uptake routes are concurrently involved, dermal and ingestion in guts, where the bioavailability of contaminants can considerably change. The aim of this study was to analyze the toxicity of arsenic (As) in various strongly contaminated meadow and forest soils, representative for former As mining and processing area, to earthworms E. fetida and its accumulation in their bodies. An attempt was made to find relationships between the response of earthworms and chemical extractability of As. In the bioassay, carried out according to the standard ISO protocol, different endpoints were applied: earthworm survival, fecundity measured by the numbers of juveniles and cocoons, earthworm weight and As accumulation in the bodies. The results proved that E. fetida can tolerate extremely high total As concentrations in soils, such as 8000 mg/kg, however, the individual endpoints were not correlated and showed different patterns. The most sensitive one was the number of juveniles. No particular soil factor was identified that would indicate an exceptionally high As susceptibility to the release from one of soils, however, we have demonstrated that the sum of non-specifically and specifically bound As (i.e. fractions F1 + F2 in sequential extraction according to Wenzel) could be a good chemical indicator of arsenic toxicity to soil invertebrates.

Effects of soybean oil exposure on the survival, reproduction, biochemical responses, and gut microbiome of the earthworm Eisenia fetida.

With increasing production of kitchen waste, cooking oil gradually enters the soil, where it can negatively affect soil fauna. In this study, we explored the effects of soybean oil on the survival, growth, reproduction, tissue structure, biochemical responses, mRNA expression, and gut microbiome of earthworms (Eisenia fetida). The median lethal concentration of soybean oil was found to be 15.59%. Earthworm growth and reproduction were significantly inhibited following exposure to a sublethal concentration of soybean oil (1/3 LC50, 5.2%). The activity of the antioxidant enzymes total superoxide dismutase (T-SOD), peroxidase (POD), and catalase (CAT) were affected under soybean oil exposure. The glutathione (GSH) content decreased significantly, whereas that of the lipid peroxide malondialdehyde (MDA) increased significantly after soybean oil exposure. mRNA expression levels of the SOD, metallothionein (MT), lysenin and lysozyme were significantly upregulated. The abundance of Bacteroides species, which are related to mineral oil repair, and Muribaculaceae species, which are related to immune regulation, increased within the earthworm intestine. These results indicate that soybean oil waste is toxic to earthworms. Thus, earthworms deployed defense mechanisms involving antioxidant system and gut microbiota for protection against soybean oil exposure-induced stress.

Classification of target tissues of Eisenia fetida using sequential multimodal chemical analysis and machine learning.

Acquiring comprehensive knowledge about the uptake of pollutants, impact on tissue integrity and the effects at the molecular level in organisms is of increasing interest due to the environmental exposure to numerous contaminants. The analysis of tissues can be performed by histological examination, which is still time-consuming and restricted to target-specific staining methods. The histological approaches can be complemented with chemical imaging analysis. Chemical imaging of tissue sections is typically performed using a single imaging approach. However, for toxicological testing of environmental pollutants, a multimodal approach combined with improved data acquisition and evaluation is desirable, since it may allow for more rapid tissue characterization and give further information on ecotoxicological effects at the tissue level. Therefore, using the soil model organism Eisenia fetida as a model, we developed a sequential workflow combining Fourier transform infrared spectroscopy (FTIR) and matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) for chemical analysis of the same tissue sections. Data analysis of the FTIR spectra via random decision forest (RDF) classification enabled the rapid identification of target tissues (e.g., digestive tissue), which are relevant from an ecotoxicological point of view. MALDI imaging analysis provided specific lipid species which are sensitive to metabolic changes and environmental stressors. Taken together, our approach provides a fast and reproducible workflow for label-free histochemical tissue analyses in E. fetida, which can be applied to other model organisms as well.

Juxtaposing the quality of compost and vermicompost produced from organic wastes amended with cow dung.

Composting is a propitious technology to change bio-degradable solid waste into organic fertilizers. Considering this, five types of organic waste viz., leaf litter (Tectona grandis), water hyacinth (Eichhornia crassipes), cauliflower waste (Brassica oleracea var. botrytis), coir pith, and mushroom spent waste were composted with and without the use of earthworm (Eisenia fetida). The reaction (pH) and electrical conductivity of compost and vermicompost ranged from 6.98 to 7.45 and 6.97 to 7.36, 0.11 to 0.21 dSm-1, and 0.11 to 0.25 dSm-1, respectively. The chemical oxygen demand both the compost and vermicompost ranged from 687 to 1170 mg l-1 and 633-980 mg l-1 respectively. Cation exchange capacity (CEC) ranged from, 75 to 121 (c mol (p+) kg-1, and 80 to 127 (c mol (p+) kg-1, respectively. The C:N of compost and vermicompost varied from 16:1 to 33:1 and 12:1 to 19:1, respectively. The organic carbon content was decreased (18.3-38.7%), while secondary and micronutrient contents increased over the initial concentration. The NH4+ and NO3- content of compost and vermicompost ranged from 270 to 510 mg kg-1 and 230-430 mg kg-1, 560 to 105 mg kg-1, and 690-1100 mg kg-1, respectively. The nitrification index (NH4+/NO3-) ranged from 0.3 to 0.9 in composts and 0.3 to 0.6 in vermicomposts. The dehydrogenase and urease activity varied from 685 to 1696 µg g-1 hr-1 and 938-2549 µg TPF g-1 day-1 respectively. The bacteria, fungi and actinomycetes population were 2-3, 0.3-0.7 and 3-8 times more in vermicompost over the corresponding compost. This study confirmed that compared to compost, vermicompost showed better nutrients and microbial properties.

Evolution of the earthworm (Eisenia fetida) microbial community in vitro and in vivo under tetracycline stress.

Bioremediation of contaminated soil has received increasing attention, and the effects of antibiotic residues on the soil ecological environment are a current research hotspot. Earthworms are the first choice of soil organisms to indicate the degree of soil pollution, and their detoxification mechanism after antibiotic stress must be further explored. Taking Eisenia fetida as the research object, an antibiotic (tetracycline) stress test was carried out in sterile artificial soil. The stress concentrations were set at 0, 0.3, 3, 30, 300 and 600 mg/kg. The ECO method was used to cultivate microbes in earthworms and soil. The carbon source utilization intensity algorithm developed by our team was used for data statistics, and a factor analysis model was constructed to explore the succession process of microbes of earthworms in vivo and in vitro under tetracycline stress. The results showed that there were four processes in the evolution of microbes under short-term tetracycline stress: at 1-3 days, the microbes in worms played a leading role; at 4-5 days, the microbes in the worms and the soil microbes jointly resisted TET stress; after 6-8 days of stress, the microbes in worms still played the main role, but their role was weakened; and after 9-10 days, soil microbes played a leading role, and tolerant microbes appeared. Under long-term stress, the microbes of earthworms in vivo and in vitro were obvious different, and there may be no regulatory relationship. And the factor analysis model is suitable for the analyse of the changes in microbial communities in vivo and in vitro under TET stress. The research results provide a reference method and model basis for the bioremediation of antibiotic-contaminated soil and the study of earthworm detoxification mechanisms, and help agricultural development.

In vitro cultivation of primary intestinal cells from Eisenia fetida as basis for ecotoxicological studies.

The earthworm Eisenia fetida is a commonly used model organism for unspecific soil feeders in ecotoxicological studies. Its intestinal cells are the first to encounter possible pollutants co-ingested by the earthworm, which makes them prime candidates for studies of toxic effects of environmental pollutants on the cellular as compared to the organismic level. In this context, the aim of this study was to demonstrate the suitability of preparations of primary intestinal E. fetida cells for in vitro ecotoxicological studies. For this purpose, a suitable isolation and cultivation protocol was established. Cells were isolated directly from the intestine, maintaining >85% viability during subsequent cultivations (up to 144 h). Exposure to established pollutants and soil elutriates comprising silver nanoparticles and metal ions (Cu2+, Cd2+) induced a significant decrease in the metabolic activity of the cells. In case of microplastic particles (MP particles), namely 0.2, 0.5, 2.0, and 3.0 µm diameter polystyrene (PS) beads as well as 0.5 and 2.0 µm diameter polylactic acid (PLA) beads, no active uptake was observed. Slight positive as well as negative dose and size dependent effects on the metabolism were seen, which to some extent might correlate with effects on the organismic level.

Metsulfuron-methyl induced physiological, behavioural and biochemical changes in exotic (Eisenia fetida) and indigenous (Metaphire posthuma) earthworm species: Toxicity and molecular docking studies.

In modern agricultural practices, Metsulfuron-methyl (sulfonylurea herbicide) is widely employed to inhibit the weeds and grasses. The current study revealed that Metaphire posthuma was more sensitive than Eisenia fetida against Metsulfuron-methyl (MSM). The LC50 values for Eisenia fetida were 2884.08 mgkg-1 and 1871.18 mgkg-1after 7 and 14 days, respectively. Similarly, the LC50 values for Metaphire posthuma were 2449.34 mgkg-1 and 1673.10 mgkg-1for 7 and 14 days, respectively. Reproduction parameters were significantly decreased at 400 (T3), 800 (T4) and 1600 (T5) mgkg-1 MSM in E. fetida whereas at 200 (T2), 400 (T3), 800 (T4), 1600 (T5) mgkg-1 MSM in M. posthuma. EC50 of avoidance response for 20% MSM by E. fetida and M. posthuma was recorded 901.76 mgkg-1and 544.21 mgkg-1 respectively. Malondialdehyde (MDA) content along with guaiacol peroxidase (POD), catalase (CAT) and superoxide dismutase (SOD) activities were initially increased up to 21st day by MSM, inducing a slight oxidative stress in earthworms and recovered to control level on 28th day. The GST activities were continuously stimulated throughout the exposure period and enhance the detoxification effect thereby preventing the earthworms from toxins. Molecular docking studies indicated that hydrogen bonding and hydrophobic interactions are key forces in binding between MSM and SOD/CAT/POD/GST. As a result, this is the first study to be reported on physiological, behavioural and biochemical changes in two different earthworm species under the exposure of sulfonyl urea herbicide.