That search has now produced an intriguing result. A team of South African researchers, publishing in the journal MicrobiologyOpen, used DNA sequencing to map the fungal communities living in the rhizosphere of soybean plants grown under different organic fertilization regimes. The rhizosphere, the narrow zone of soil hugging the plant’s roots, is one of the most biologically active environments on Earth, teeming with bacteria, fungi, and other microorganisms that feed on sugars and other compounds exuded by the roots. Because more than 95 percent of soil microbes cannot be grown in a laboratory dish, the team turned to amplicon sequencing of the internal transcribed spacer region, a genetic barcode widely used to identify fungi, to catalogue organisms that traditional culturing would have missed entirely.

The experiment took place at the North-West University farm in Molelwane, Mafikeng, in a region receiving about 540 millimeters of rain per year. The researchers laid out a randomized complete block design with three treatments applied to soybean plots: soil amended with cattle dung manure, soil amended with poultry waste, and an untreated control, alongside bulk soil collected from open inter-row areas with no root contact. Organic amendments were incorporated before planting at a rate of 20 tons per hectare, and rhizosphere samples were harvested at the flowering stage, seven weeks after sowing, when interactions between roots and microbes reach their peak. Physicochemical analysis showed that poultry waste enriched the soil with phosphorus and calcium, while cattle dung boosted phosphorus, potassium, and magnesium.

Sequencing on an Illumina NovaSeq 6000 system produced remarkably clean data, with each sample yielding roughly 165,000 to 172,000 non-chimeric reads and quality scores exceeding standard thresholds. After processing with the DADA2 denoising algorithm within QIIME 2 and classifying sequences against the UNITE reference database, the researchers could compare the fungal profiles of each treatment with unprecedented resolution. The results revealed that the type of organic matter added to the soil, not merely its presence, exerts a selective force on which fungi thrive around soybean roots.

At the phylum level, cattle dung-amended soils showed higher relative abundances of Chytridiomycota, Aphelidiomycota, and Rozellomycota, groups often associated with saprotrophic living and the breakdown of tough plant polymers such as cellulose. Poultry waste, by contrast, favored Mortierellomycota and Ascomycota, lineages that include fast-growing copiotrophs suited to nutrient-rich conditions. Mortierellomycetes, in particular, are known as active nitrogen mineralizers and phosphate solubilizers, suggesting that the nutrient-cycling potential of poultry-amended soil differs meaningfully from that of cattle dung. The untreated control and bulk soils were dominated by Olpidiomycota, Zoopagomycota, and Kickxellomycota, a less diverse but more stress-tolerant community consistent with their lower nutrient status.

The genus-level patterns were equally striking. Cattle dung soils supported filamentous Ascomycete genera such as Chaetomium, Neurospora, Achaetomium, and Humicola, organisms thought to play important roles in cellulose degradation and organic matter turnover. Poultry waste favored Trichoderma and Penicillium, two genera celebrated for promoting plant growth, suppressing pathogens through biocontrol mechanisms, and enhancing nutrient flow to crops, along with a marked dominance of Keratinophyton, a fitting match for the keratin-rich feathers and litter that characterize poultry waste. Meanwhile, the control and bulk soils were dominated by Fusarium, Curvularia, and Stagonosporopsis, taxa that include notorious plant pathogens. The authors interpret this as evidence that organic amendments can suppress potentially harmful taxa by shifting competitive hierarchies in the soil toward beneficial saprotrophs.

Diversity metrics added the clearest statistical evidence of treatment effects. Bulk soil exhibited the highest fungal richness, with a Chao1 index of 1603.33 and 1557 observed features, followed by cattle dung at 1456.64, while poultry waste showed the lowest richness at 957.93. Shannon diversity was highest in cattle dung soils at 6.561, closely followed by bulk soil at 6.528, with poultry waste trailing at 5.525 and showing the lowest evenness. Good’s coverage remained at 1.000 across all treatments, confirming that the sequencing captured the communities thoroughly. The researchers suggest that poultry waste imposes nutrient-driven ecological filtering that favors fast-growing copiotrophs at the expense of rarer taxa, whereas the diverse organic substrates in cattle manure support a more balanced and heterogeneous fungal community.

Venn diagram analysis of amplicon sequence variants revealed a core microbiome of 300 ASVs shared across all treatments, likely reflecting the filtering influence of soybean root exudates, alongside large treatment-specific fractions: bulk soil harbored 756 unique ASVs and cattle dung 655, compared with 330 for poultry waste and 476 for the control. Beta diversity analysis showed the greatest compositional dissimilarity between poultry waste and the control, and between cattle dung and poultry waste, indicating that the two organic amendments reshape the fungal community in distinctly different ways. Ordination by principal component analysis and principal coordinates analysis based on UniFrac distances reinforced this picture, with cattle dung and poultry waste samples forming distinct clusters while control and bulk soils overlapped more and spread heterogeneously. Notably, the weighted UniFrac metric showed cattle dung most strongly influenced abundant taxa, while the unweighted metric revealed shifts in rare taxa across all treatments, with possible long-term consequences for soil resilience.

Functional prediction using the FUNGuild database translated these taxonomic shifts into ecological meaning. Cattle dung soils were strongly enriched for dung saprotrophs, animal pathogens, and wood saprotrophs, consistent with their cellulose-degrading Ascomycete residents. Poultry waste showed distinct enrichment of endophyte and plant pathogen guilds, likely reflecting its nitrogen- and keratin-rich character, while bulk soils clustered separately with wood saprotrophs and ectomycorrhizal fungi. Crucially, 32 functional guilds were shared across all treatments, indicating a stable core of ecological functions underpinning soil health and soybean growth. Linear discriminant analysis effect size identified Hypocreales, a group rich in biocontrol agents, as a biomarker for cattle dung, and keratin-degrading Onygenales as a biomarker for poultry waste. Predicted metabolic pathways confirmed that carbohydrate and amino acid metabolism dominated across all samples, underscoring the central role of carbon and nitrogen turnover in rhizosphere function.

The authors are careful to frame their conclusions with appropriate statistical caution. With only three field replicates per treatment, the ANOSIM and MRPP tests did not reach conventional significance despite moderate effect sizes, most notably the R-value of 0.74 separating cattle dung from poultry waste. Differences in underlying soil chemistry, including extractable phosphorus, calcium, potassium, magnesium, sodium, and pH, may also partly confound the interpretation of amendment effects. The alpha diversity findings, however, represent the most statistically robust conclusions of the study. Taken together, the work delivers a clear message for sustainable agriculture: the choice between organic fertilizers is not interchangeable. Cattle dung appears to cultivate diverse, functionally rich saprotrophic communities likely to benefit soil health, while poultry waste fosters specialized, nutrient-responsive assemblages. As farmers worldwide seek to rebuild degraded soils, understanding these amendment-specific microbial signatures could help tailor fertilization strategies that harness the hidden fungal workforce beneath every soybean field.

Subject of Research: Fungal diversity and functional prediction of the soybean rhizosphere microbiome under different organic fertilization regimes

Article Title: Amplicon Characterization of Fungal Diversity and Functional Prediction of Glycine max L. Rhizosphere Microbiome Under Different Organic Fertilization

Article References: Osuji, I. E., Akanmu, A. O., & Babalola, O. O. (2026). Amplicon Characterization of Fungal Diversity and Functional Prediction of Glycine max L. Rhizosphere Microbiome Under Different Organic Fertilization. MicrobiologyOpen, 15(5), Article e70437. https://doi.org/10.1002/mbo3.70437

Image Credits: AI Generated

DOI: 10.1002/mbo3.70437

Keywords: soybean, rhizosphere, fungal diversity, organic fertilizer, cattle dung, poultry waste, ITS amplicon sequencing, FUNGuild, soil microbiome, sustainable agriculture, alpha diversity, soil health

Neonicotinoids are systemic insecticides applied as coatings on crop seeds before planting. As the seed germinates and grows, the chemical is taken up into plant tissues, protecting the young seedling from chewing and sucking insects. But only a fraction of the active ingredient ends up inside the plant. The rest can persist in the soil, where it encounters an enormous community of microorganisms that drive nutrient cycling, decompose organic matter, and form partnerships with plant roots. While the impacts of neonicotinoids on insects and aquatic invertebrates have been studied extensively, their effects on soil fungal communities have remained surprisingly underexplored, a gap the new research set out to fill.

The team designed a controlled microcosm experiment in which soil was treated with three commonly applied neonicotinoids: imidacloprid, thiamethoxam, and clothianidin. These compounds represent the backbone of modern seed-treatment agriculture, coating millions of tonnes of seed annually across the world’s major cropping systems. By working in microcosms, the researchers could isolate the effect of each compound on the fungal community without the confounding noise of weather, cropping history, or management variation that complicates field studies. Soil samples were then collected at multiple time points after exposure, allowing the team to track not just whether the fungi responded, but when.

To profile the fungal communities, the researchers turned to amplicon sequencing of the internal transcribed spacer, or ITS, region of fungal DNA. The ITS region is the standard molecular barcode for fungi, allowing scientists to identify which fungal taxa are present in a soil sample even when those organisms cannot be cultured in the laboratory. This technique revealed hundreds of fungal taxa across the samples, spanning dominant phyla such as Ascomycota and Basidiomycota as well as rarer, more enigmatic lineages. Sequencing-based approaches like this have transformed microbial ecology in recent years, making it possible to detect subtle community shifts that would be invisible under a microscope.

One of the study’s most striking findings concerns timing. Overall diversity indices remained largely stable across the treatments when averaged over the whole experiment, which might initially suggest the fungicide exposure had little effect. But post-hoc comparisons told a different story: on day 10, fungal diversity dropped significantly under both imidacloprid and thiamethoxam exposure. This kind of delayed, transient response is exactly the sort of signal that single end-point sampling would miss. The lesson, the authors suggest, is that the ecological footprint of a pesticide cannot be judged from a single snapshot in time.

Beta diversity analysis, which measures how community composition differs between samples, reinforced this temporal picture. The researchers found significant effects of sampling day and, critically, a treatment-by-time interaction, meaning the fungal communities under different insecticide treatments did not simply follow the same trajectory. Instead, each compound appeared to push the community along its own path as the experiment unfolded. Such temporally variable responses complicate risk assessment, because a pesticide that looks benign in one week of a field season may produce measurable disruption in another.

Not all fungi responded equally. The dominant phyla, Ascomycota and Basidiomycota, which include many decomposers and plant-associated species, remained relatively stable throughout the experiment. But several less abundant phyla declined over time, including Mortierellomycota, Rozellomycota, and Olpidiomycota. These obscure-sounding groups are far from ecologically trivial. Mortierellomycota species are important decomposers and plant growth promoters, Rozellomycota comprises widespread intracellular parasites of other microorganisms, and Olpidiomycota includes fungi that can vector plant viruses. A decline in these rarer lineages may signal subtle erosion of functions that only become apparent when soil health degrades.

Perhaps the most consequential result came from functional guild analysis, which classifies fungi by their ecological roles rather than their taxonomy. Here the researchers found that clothianidin exposure drove an increase in saprotroph abundance, the fungi that break down dead organic matter, alongside a decrease in symbiotroph abundance, the fungi that live in mutually beneficial partnerships with plants. The most important symbiotrophs in agricultural soil are the arbuscular mycorrhizal fungi, which colonize crop roots and exchange soil nutrients for plant sugars. A shift away from symbiotrophs and toward saprotrophs could alter how nutrients flow through the soil food web, potentially affecting crop nutrition in ways that standard pesticide evaluations never measure.

Differential abundance analysis, a statistical technique for identifying which taxa increase or decrease under specific conditions, revealed that each compound left its own fingerprint on the community. Imidacloprid was associated exclusively with suppressed fungal biomarkers, meaning every taxon linked to this compound was depleted rather than enriched. Thiamethoxam induced both enriched and depleted taxa, suggesting a more mixed restructuring of the community. Clothianidin stood out as the most disruptive of the three, associated with the greatest number of discriminatory fungal biomarkers and accompanied by the increase in saprotrophs and reduction in symbiotrophs. The compound-specific nature of these responses suggests that treating all neonicotinoids as a single ecological hazard may obscure important differences among them.

The study, which was funded by the Australian National University and conducted with colleagues Julia F. Jasonsmith, Nilantha R. Hulugalle, and Craig L. Strong, carries implications well beyond the laboratory. As regulators and farmers weigh the costs and benefits of neonicotinoid seed treatments, fungal communities have rarely featured in the calculus, even though these organisms underpin soil fertility, carbon storage, and crop resilience. The authors argue that fungal community responses deserve a place in evaluations of the broader ecological effects of neonicotinoids, and they call for long-term, functionally oriented studies in real agroecosystem contexts. A ten-day microcosm can reveal the shape of a disturbance, but only sustained field research can determine whether these shifts persist, compound across seasons, or ultimately translate into measurable consequences for the crops that depend on the hidden life below ground.

Subject of Research: Effects of neonicotinoid seed treatments on soil fungal community diversity, composition, and function over time

Article Title: Fungal Responses to Neonicotinoid Seed Treatments in Soil: Temporal Shifts in Community Diversity and Composition

Article References: Akter, S., Jasonsmith, J. F., Hulugalle, N. R., & Strong, C. L. (2026). Fungal Responses to Neonicotinoid Seed Treatments in Soil: Temporal Shifts in Community Diversity and Composition. Microbial Ecology. https://doi.org/10.1007/s00248-026-02892-2

Image Credits: AI Generated

DOI: 10.1007/s00248-026-02892-2

Keywords: neonicotinoids, soil fungi, fungal diversity, imidacloprid, thiamethoxam, clothianidin, ITS amplicon sequencing, soil microbiome, temporal dynamics, saprotrophs, symbiotrophs, agroecosystems

The study, published in Advanced Biotechnology, arrives at a moment of growing anxiety over the ecological footprint of engineered nanomaterials. Layered double hydroxides carry the general formula [M2+1−xM3+x(OH)2]An−x/n·yH2O, and their high surface area, anion exchange capacity, and thermal stability have made them favored adsorbents for heavy metals, dyes, and other pollutants. China’s chemical industry standard HG/T 5549–2019 even recognizes LDHs as an environmentally friendly adsorbent material. But as production volumes climb, so do emissions into the environment, and soil is the compartment where these particles tend to accumulate. Because LDHs can interact directly with cell surfaces, dissolve toxic elements, and generate reactive oxygen species, the team argued that understanding their fate in terrestrial ecosystems is no longer optional.

The researchers synthesized the two-dimensional NiFe-LDHs hydrothermally from ferric chloride and nickel nitrate with urea, and converted part of that product into the three-dimensional NiFeS-LDHs by reacting it with thioacetamide in ethanol. X-ray diffraction and electron microscopy confirmed the structures. Ryegrass, a fast-growing annual grass with high biomass and documented tolerance to heavy metals and nanomaterials, served as the model plant. Pots received 2 kilograms of sieved farmland soil each, amended with either material at concentrations ranging from 200 to 800 milligrams per kilogram, alongside untreated controls, and the plants grew for 50 days before harvest.

The growth results split cleanly along structural lines. Two-dimensional NiFe-LDHs promoted ryegrass growth at specific concentrations: chlorophyll content rose significantly at amendment levels between 200 and 500 milligrams per kilogram compared with untreated soil, and the moderate suppression of fresh weight at 200 milligrams per kilogram gave way to a rebound before declining again at the highest doses. The three-dimensional NiFeS-LDHs told a darker story, significantly reducing fresh weight at every concentration tested and pushing chlorophyll levels below control values at 650 and 800 milligrams per kilogram. Hydrogen peroxide measurements revealed the mechanism at work: both materials triggered oxidative stress, but the 3D particles produced more pronounced accumulation of reactive oxygen species, forcing the plants into a stronger antioxidant response involving catalase, superoxide dismutase, and peroxidase.

Metal uptake analysis helped explain why. Nickel and iron accumulated in ryegrass roots far more than in leaves, and root nickel content was consistently higher in the three-dimensional treatment group than in the two-dimensional group across all concentrations. Nickel is a micronutrient in small doses, supporting urease activity, photosynthesis, and nutrient absorption, but excess nickel disrupts chloroplast function, auxin transport, and iron uptake. At low and moderate concentrations the benefits of trace nickel and iron likely stimulated chlorophyll synthesis, while at high doses metal accumulation overwhelmed those gains and suppressed photosynthesis, a pattern consistent with the hormetic responses seen for other metal-based nanomaterials.

Beneath the surface, the two materials reshaped soil geochemistry in opposite directions. The three-dimensional NiFeS-LDHs significantly lowered soil pH while raising electrical conductivity by as much as 271 percent at 800 milligrams per kilogram. The authors attribute the acidification to sulfide oxidation in the aerated soil, which generates sulfate and hydrogen ions, compounded by hydrolysis of released nickel and iron and by a self-amplifying cycle in which iron-oxidizing microbes regenerate ferric iron that attacks remaining sulfides. The flat sheets behaved more gently, gradually reducing electrical conductivity, likely by adsorbing ions, and stabilizing total carbon and phosphorus. Both materials significantly boosted soil organic carbon, by up to roughly 49 percent for the 2D particles and 56 percent for the 3D particles, while both cut total nitrogen sharply, by 16 to 36 percent depending on dose and material.

Soil enzyme activity, a sensitive barometer of nutrient cycling, diverged along the same structural fault line. The two-dimensional particles suppressed sucrase activity by up to 53 percent at high doses but enhanced urease activity by 25 to 54 percent, leaving catalase and neutral phosphatase largely untouched. The three-dimensional particles did nearly the opposite, significantly inhibiting catalase by 13 to 30 percent and urease by 7 to 29 percent, while boosting neutral phosphatase by roughly 50 to 64 percent at low to moderate doses. These enzymatic fingerprints imply that the two architectures push soil metabolism toward different functional states, with the flat sheets tilting the system toward nitrogen transformation and the structured particles toward phosphorus mobilization at the expense of oxidative and urea-processing capacity.

High-throughput sequencing of 16S rDNA and fungal ITS regions showed that both materials altered the composition of rhizosphere bacterial and fungal communities, but the three-dimensional particles hit harder. At elevated concentrations they significantly depressed fungal diversity indices, a concerning signal because greater fungal diversity has been linked to healthier soils and more sustainable crop production. Moderate concentrations of the 2D particles enriched taxa such as Bryobacter, Flavisolibacter, Rhodoplanes, Cryptococcus, and Fusarium, organisms associated with nitrate reduction, denitrification, plant growth promotion, and stress resistance. Correlation analyses tied these community shifts to soil pH, electrical conductivity, ammonium, and iron levels, and structural equation modeling delivered the study’s headline number: the total negative path coefficient of the 3D material on ryegrass growth through the soil ecosystem, −1.071, was nearly three times that of the 2D material, −0.368, with soil acidification, salt stress, and nickel toxicity acting as the dominant downward forces while the microbial community itself continued to support plant growth.

The findings do not mean that nickel-iron LDHs are unsuitable for environmental work, but they do mean that their ecological risk cannot be assessed by chemistry alone. Two materials with identical elemental composition behaved like entirely different pollutants once their geometry changed, altering enzyme stoichiometry, reshaping microbial assemblages, and shifting the balance of nitrogen and phosphorus cycling. The authors caution that field-scale toxicity will require multi-season trials, ionic and bulk controls, and molecular studies to confirm the causal pathways they observed in pots. Still, as engineered nanomaterials flood into remediation schemes, fertilizers, and wastewater treatment plants, the lesson is blunt and timely: in the soil beneath our feet, nanostructure is destiny, and regulators evaluating the safety of layered double hydroxides would do well to demand the blueprint of every particle before it touches the ground.

Subject of Research: The differential biological effects of 2D and 3D NiFe-based layered double hydroxide nanomaterials on the ryegrass-soil ecosystem.

Article Title: Bio-effects of engineering nanomaterials NiFe-based LDHs on ryegrass-soil system

Article References: Xu, H., Jiang, X., He, C., Peng, Y., Xin, G., & Li, X. (2026). Bio-effects of engineering nanomaterials NiFe-based LDHs on ryegrass-soil system. Advanced Biotechnology, 4(3), Article 24. https://doi.org/10.1007/s44307-026-00114-x

Image Credits: AI Generated

DOI: 10.1007/s44307-026-00114-x

Keywords: engineered nanomaterials, layered double hydroxides, NiFe-LDHs, ryegrass, soil health, soil enzymes, soil microbiome, oxidative stress, soil acidification, fungal diversity, nanotoxicology, soil remediation