Copper toxicity on Eisenia fetida in a vineyard soil: a combined study with standard tests, genotoxicity assessment and gut metagenomic analysis.

Copper (Cu) toxicity is a pressing concern for several soils, especially in organic viticulture. The objective of this work was to assess Cu toxicity on the non-target organism Eisenia fetida, employing both traditional and novel tools for early identification of Cu-induced damages. In addition to traditional tests like avoidance and reproductive toxicity experiments, other tests such as the single cell gel electrophoresis (SCGE) and gut microbiome analysis were evaluated to identify early and more sensitive pollution biomarkers. Four sub-lethal Cu concentrations were studied, and the results showed strong dose-dependent responses by the earthworm avoidance test and the exceeding of habitat threshold limit at the higher Cu doses. An inverse proportionality was observed between reproductive output and soil Cu concentration. Bioaccumulation was not detected in earthworms; soil concentrations of potentially bioavailable Cu were not affected by E. fetida presence or by time. On the contrary, the SCGE test revealed dose-dependent genotoxicity for the 'tail length' parameter already at the second day of Cu exposition. Gut microbiome analysis a modulation of microbial composition, with the most aboundant families being Pectobateriaceae, Comamonadaceae and Microscillaceae. Bacillaceae increased over time and showed adaptability to copper up to 165 mg/kg, while at the highest dose even the sensitive Acetobacteriaceae family was affected. The research provided new insights into the ecotoxicity of Cu sub-lethal doses highlighting both alterations at earthworms' cellular level and changes in their gut microbiota.

Agronomical valorization of eluates from the industrial production of microorganisms: Chemical, microbiological, and ecotoxicological assessment of a novel putative biostimulant.

Plant Biostimulants (BSs) are a valid supplement to be considered for the integration of conventional fertilization practices. Research in the BS field keeps providing alternative products of various origin, which can be employed in organic and conventional agriculture. In this study, we investigated the biostimulant activity of the eluate obtained as a by-product from the industrial production of lactic acid bacteria on bare agricultural soil. Eluates utilization is in line with the circular economy principle, creating economical value for an industrial waste product. The research focused on the study of physical, chemical, biochemical, and microbiological changes occurring in agricultural soil treated with the biowaste eluate, applied at three different dosages. The final aim was to demonstrate if, and to what extent, the application of the eluate improved soil quality parameters and enhanced the presence of beneficial soil-borne microbial communities. Results indicate that a single application at the two lower dosages does not have a pronounced effect on the soil chemical parameters tested, and neither on the biochemical proprieties. Only the higher dosage applied reported an improvement in the enzymatic activities of ß-glucosidase and urease and in the chemical composition, showing a higher content of total, nitric and ammonia N, total K, and higher humification rate. On the other hand, microbial communities were strongly influenced at all dosages, showing a decrease in the bacterial biodiversity and an increase in the fungal biodiversity. Bioinformatic analysis revealed that some Operative Taxonomic Units (OTUs) promoted by the eluate application, belong to known plant growth promoting microbes. Some other OTUs, negatively influenced were attributed to known plant pathogens, mainly Fusarium spp. Finally, the ecotoxicological parameters were also determined and allowed to establish that no toxic effect occurred upon eluate applications onto soil.

The extracellular DNA can baffle the assessment of soil bacterial community, but the effect varies with microscale spatial distribution.

Environmental DNA is made-up of intracellular (iDNA) and extracellular (eDNA) pools. In soils, eDNA can be present up to 40% and could distort the assessment of living microorganisms. Distribution of microbial community is inconsistent among different size-aggregates, and the persistence and turnover of eDNA are thus uneven. Uneven persistence and distribution of eDNA could lead to heterogeneity in community analysis biases that arise due to eDNA sequences at micro-scale distribution. Here, we investigated the diversity and structure of eDNA and iDNA bacterial communities in bulk soil and different size-aggregates. Significant differences were observed between eDNA and iDNA bacterial diversity and composition. Changes in community composition are more important than the amount of eDNA to assess the biases caused by eDNA in community analysis. Furthermore, variations were also observed in aggregates-levels for eDNA and iDNA community which indicates that colonization pattern of iDNA community and protection of eDNA through absorbance on particle surface within soil-matrix is heterogeneous. Our work provides empirical evidence that eDNA presence could mask the detection of aggregates-level spatial dynamics in soil microbial community and have potential to qualitatively baffle observed live effects of given treatment by adequately muting the actual response dynamics of the soil microbiome.