eDNA Soil Biodiversity Analysis — 2024 Baseline

Project: PROJ05706 · NatureMetrics Terrestrial Soil eDNA
Samples received: July 2024
Sites: 8 sampling locations
Assays: Soil Fungi · Soil Bacteria · Soil Invertebrates
Analysis date: April 2026

1. Introduction and Overview

This report provides a detailed interpretation of the environmental DNA (eDNA) results from eight soil samples collected in 2024 and analysed by NatureMetrics. The three assays — fungi, bacteria, and soil invertebrates — together offer a multi-kingdom snapshot of what lives in your soils, and what that biological community tells us about soil health, function, and condition.

eDNA metabarcoding detects organisms by extracting and sequencing DNA directly from soil, without needing to see or culture individual organisms. The results are expressed as OTUs (Operational Taxonomic Units)¹ — groups of DNA sequences that roughly correspond to species or species-groups. The number of OTUs detected is a proxy for species richness, while their relative abundance (expressed as percentage of DNA reads) gives an indication of dominance, though it should be interpreted with care as eDNA read counts don't translate directly to organism counts.

A note on sample types

Before describing the sites, an important distinction: most of the eight samples are mix probes — composite samples taken across a broader area containing multiple cultures or plant communities. Two are single-stripe probes, taken from a specific strip within a field representing a single culture:

• E1.1 Market Garden 70 is a stripe within the market garden where a spade test (Spatenprobe) scored a perfect 5 out of 5 on soil structure — the best possible result. Its eDNA data therefore represents the biology of the highest-quality, most structurally advanced soil on the project.
• E3.1 Spannweid 14 is a stripe within the Spannweid field planted with a deliberate nettle (Urtica dioica) culture established in autumn 2022 — a distinct single-species environment with its own specific organic matter chemistry and root dynamics.

This distinction matters for interpretation: the stripe samples represent a narrower, more specific ecological situation than the mix probes, which capture a broader and more varied community. Differences between E3 Spannweid Mix and E3.1 Spannweid 14, or between E1 Market Garden Mix and E1.1 Market Garden 70, should be understood in this light.

The eight sampling sites

All sites are managed organically with no fertiliser or manure inputs of any kind. The key differentiator between sites is how long they have been under active regenerative management, and their soil type.

A-sites (loamy soils):

• A1 Forsthaus (ForsthausUnten) — the lowest biodiversity across all three assays. The most extreme starting point in the dataset: liquid manure (Gülle) was applied by the previous land manager shortly before handover; the field was then essentially open, bare soil through spring 2024; and at the time of sampling in July 2024, a pumpkin and clover understory culture had only just begun establishing and was only partially green. The 2024 data captures a true before-picture — bare, recently disturbed, manure-affected soil with almost no plant cover. Mix probe.
• A2 Wiese Mix (ForsthausOben / Adlisberg Wiese) — a deliberately established Swiss Kunstwiese (sown meadow), established in spring 2020. By July 2024 this sward was approximately four years old. A Kunstwiese typically contains a diverse mix of grasses and legumes (clover species) — grass roots support a rich nematode community, and legumes support nitrogen-fixing bacterial associations. Mix probe.
• A3 Alter Obstgarten Mix — old orchard area, newly brought into current management but with a cover crop mix established since autumn 2023. Loamy soil. Despite being newly managed, the cover crop has already had a measurable effect on bacterial diversity. Mix probe.

E-sites (longer under management):

• E1 Market Garden Mix — the longest-managed site. Consistently the most biodiverse across all three kingdoms. Loamy soil. Mix probe.
• E1.1 Market Garden 70 — a single-stripe probe within the market garden, representing one specific culture strip. This stripe scored a perfect 5/5 on the spade test (Spatenprobe) for soil aggregate structure — the best possible physical result. Its eDNA data represents the biology of the most structurally advanced soil on the project, and provides the clearest direct link between biological richness and measurable soil health.
• E2 Haus Huegel Mix — several years under management. Peat and loamy soil. Strong invertebrate diversity, particularly earthworm relatives and mites. Mix probe.
• E3 Spannweid Mix — several years under management. Contains both peat and loamy soil within a single field. The highest fungal functional diversity and the most unique fungal OTUs — partly reflecting the two distinct soil communities present. Mix probe.
• E3.1 Spannweid 14 — a single-stripe probe within the Spannweid field. A deliberate nettle (Urtica dioica) culture established autumn 2022, giving approximately 1.5–2 years of establishment by July 2024. Nettles are deep-rooted, nitrogen-accumulating perennials whose biomass decomposes rapidly — a chemically and structurally specific soil environment, distinct from the broader Spannweid mix.

The management timeline is crucial context: the diversity gradient from the species-rich Market Garden down to the bare Forsthaus likely reflects years of organic, no-input management building soil biological complexity over time — with the 5/5 spade test at Market Garden 70 as direct physical proof of what that process delivers.

Summary of detections

¹ OTU (Operational Taxonomic Unit): When DNA is sequenced from soil, many organisms have never been formally named by scientists, so results cannot always be reported as species. Instead, DNA sequences that are ≥97% identical to each other are grouped into a single "bucket" called an OTU — one OTU approximates one species, named or unnamed. Think of it as a way of counting distinct biological entities even when their names aren't yet known.

All samples passed quality control. DNA amplification, sequencing, and target OTU detection were successful across all eight sites and all three assays. The fungi assay returned 100% target reads at all sites. The bacteria assay showed some non-target reads (2.7%–12.1%), which is typical and does not affect data quality.

2. Soil Fungi — A Deep Dive

2.1 Overall fungal diversity

A total of 358 fungal OTUs were detected across the eight sites, of which 41 could be identified to species level. The community is overwhelmingly dominated by Ascomycota (318 OTUs, 89%), followed by Basidiomycota (27), Zygomycota (8), Chytridiomycota (2), and a single representative of the Glomeromycota (the arbuscular mycorrhizal fungi). This dominance of Ascomycota is entirely typical for agricultural and managed soils — these are the workhorses of organic matter decomposition.

An important distinction here: E1 Market Garden has the highest raw OTU count (174), but E3 Spannweid has the highest functional diversity (2.08) and evolutionary diversity (34.67). This means Spannweid doesn't just have many fungi — it has fungi that are doing a wider range of different things in the soil, drawn from a broader evolutionary lineage. This is a sign of a particularly functionally rich soil ecosystem.

2.2 Fungal community composition

All sites are dominated by Sordariomycetes (a class of Ascomycota), which is typical for temperate agricultural soils. The dominant orders are Hypocreales (decomposers and biocontrol agents) and Sordariales (dung and decaying organic matter specialists), followed by Pleosporales (diverse plant-associated fungi).

The key classes detected and their significance:

Sordariomycetes (dominant at all sites, 62–102 OTUs per site): This class includes many of the most functionally important soil fungi — decomposers that break down cellulose, lignin, and other recalcitrant plant material; biocontrol fungi that attack insects and other fungi; and plant endophytes. Their dominance is a good sign for organic matter cycling.

Dothideomycetes (16–31 OTUs per site): The second most diverse class. These fungi interact with plants in several ways: some decompose dead plant material (leaves, stems, woody debris), others live as endophytes — quietly inside living plant tissue without causing obvious harm, in a relationship that may be neutral or subtly beneficial — and a smaller number are pathogens that cause leaf spots, blights, or stem cankers. The vast majority detected in your soils are likely decomposers or endophytes; Boeremia exigua is the only confirmed pathogen named in the results, and its low-level presence in soil does not indicate active disease. Their higher count at E1 Market Garden and E3 Spannweid (31 OTUs each) likely reflects the greater variety of plant species and plant material at those sites — more plants means more substrates and more hosts for this diverse class.

Mortierellomycetes (3–7 OTUs per site): All Mortierellales — saprotrophic fungi that are excellent indicators of soil organic matter quality. They tend to be more abundant in soils with high-quality organic inputs (fresh plant material, compost) rather than recalcitrant, low-quality residues. Their consistent presence is positive.

2.3 Named fungal species of interest

Several identified species tell us something specific about your soils:

Claroideoglomus claroideum (Glomeromycota) — Detected only at E3 Spannweid Mix. This is an arbuscular mycorrhizal fungus (AMF) that forms symbiotic partnerships with plant roots, extending the root's reach for phosphorus and other nutrients. AMF are widely considered among the most important organisms in soil for plant nutrition. The fact that it was detected at only one site may be significant — AMF are sensitive to soil disturbance, high phosphorus levels, and the absence of host plants. Its presence at Spannweid and absence elsewhere is interesting to interpret in light of the soil heterogeneity there: E3 Spannweid contains both peat and loamy soil within a single field. The AMF signal may be coming specifically from the loamy zone — where phosphorus availability, root density, and soil structure may be more favourable for mycorrhizal establishment — rather than from the peat zone, which tends to be more acidic and less hospitable for some AMF species. Alternatively, the peat zone's naturally low available phosphorus could be driving strong mycorrhizal demand from plants growing there. In either case, several years of no-input management combined with this soil diversity appears to have created conditions where AMF can persist — something not yet visible at the other sites. It is also important to note that eDNA metabarcoding tends to underdetect AMF because the standard ITS primers used for fungi do not capture Glomeromycota well. The true AMF community in your soils is almost certainly richer than this single detection suggests. If mycorrhizal health is a priority, a dedicated AMF-specific assay (targeting the SSU rRNA gene) would give a much more complete picture.

Trichoderma reesei (Hypocreales) — Found at A3 Alter Obstgarten and E1 Market Garden. Trichoderma species are among the best-known beneficial soil fungi. They are powerful decomposers of cellulose, but also act as biocontrol agents — they parasitise and suppress pathogenic fungi, and can stimulate plant immune responses. T. reesei in particular is famous for its cellulase production. Its presence at the old orchard and market garden sites makes ecological sense: these are sites with diverse organic inputs (fallen fruit, vegetable residues, composts) that support cellulolytic communities.

Clonostachys intermedia (Hypocreales) — Found at A1 Forsthaus, E3 and E3.1 Spannweid. A beneficial biocontrol fungus that suppresses plant pathogens including Botrytis and, notably, Fusarium — one of the most damaging soil-borne pathogens in agriculture, responsible for root rots, crown rots, wilts, and mycotoxin contamination across a wide range of crops. Clonostachys suppresses Fusarium through several simultaneous mechanisms: it physically parasitises Fusarium hyphae (mycoparasitism), outcompetes it for space and nutrients, produces antifungal compounds that inhibit spore germination, and can prime plant root defences — much like its close relative Trichoderma. Critically, it thrives under exactly the conditions of your management system: diverse organic matter inputs, minimal soil disturbance, and no fungicide use. In conventionally managed soils with regular tillage and chemical inputs, it typically disappears. Its natural presence at three of your sites — without any inoculation — is a direct result of the organic approach building the conditions it needs³. Its strongest signal at E3 Spannweid (0.40% of reads) suggests that site currently has the most active biological Fusarium suppression.

Mortierella alpina (Mortierellales) — Detected only at E3 Spannweid. A saprotrophic fungus that thrives in soils with good organic matter inputs. It is notable for producing unusually large quantities of arachidonic acid (an omega-6 polyunsaturated fatty acid), which is rare in the plant kingdom but common in fungi and animals. In the soil food web, this matters: invertebrates that feed on Mortierella acquire arachidonic acid and use it to regulate reproduction, immune responses, and stress tolerance — meaning this fungus may be actively supporting the health of the invertebrate community above it in the food web. There is also emerging evidence that fungal arachidonic acid in the root zone can prime plant immune responses, potentially increasing disease resistance². Its presence at Spannweid is a positive sign of both organic matter quality and food web connectivity at that site.

² For a fuller explanation of arachidonic acid and its ecological roles in soil, see the supplementary notes: Species Notes & Expanded Explanations

Podospora appendiculata (Sordariales) — Strikingly dominant at A1 Forsthaus (3.69% of reads, the highest percentage for any named species at any site). Podospora species are coprophilous — they specialise in decomposing animal dung. Forsthaus received applications of liquid manure (Gülle) from the previous land manager immediately before the site came under current management. The dominant Podospora signal is almost certainly a direct legacy of that: the fungal community is still processing residual organic matter from the Gülle input at the time of sampling. This is a textbook transition signal — the previous management's biological footprint is still visible in the fungal community a year or more later. It also tells us that the Forsthaus baseline captured here is not a neutral starting point but a soil in the process of transitioning away from a high-nutrient, manure-driven biology toward the more plant-residue-driven community that the current no-input management will eventually establish.

Paracremonium variiforme (Hypocreales) — The only named species detected at all eight sites, and notably dominant at A1 Forsthaus (2.84%). This is a relatively poorly-studied soil fungus, but its ubiquity suggests it is a core member of your soil community across all land-use types.

Humicola udagawae (Sordariales) — Detected at seven of eight sites (absent only from A1 Forsthaus), with a particularly strong signal at E3 Spannweid (1.64%). Humicola species are thermotolerant decomposers that break down cellulose and hemicellulose. Their wide distribution confirms active organic matter cycling.

Solicoccozyma aeria and S. terricola (Tremellomycetes/Basidiomycota) — Yeasts found at five to six sites, strongest at A1 Forsthaus. Basidiomycete yeasts are common in surface soils and are involved in simple sugar metabolism and nutrient cycling. Their relative prominence at Forsthaus, combined with the Podospora and Keratinophyton signals, paints a coherent picture of a soil in active transition: the fungal community is still processing the readily available organic matter left behind by the previous Gülle applications — sugars, simple nitrogen compounds, and animal-derived keratin — rather than the complex, diverse plant residues that characterise the longer-managed sites. This is not a failure; it is simply where the soil is in its journey. As Forsthaus develops under current management and plant diversity builds, we would expect to see the dominant organisms at this site shift progressively toward plant-residue decomposers, biocontrol fungi, and eventually mycorrhizal associations. Monitoring this transition through future surveys will be one of the most ecologically instructive aspects of the whole project.

Rhizophlyctis rosea (Chytridiomycota) — Found at A1 Forsthaus and E3.1 Spannweid. Chytrids are aquatic or semi-aquatic fungi, and Rhizophlyctis is a cellulose decomposer found in moist soils. Its presence may indicate wetter soil conditions at these sites.

Boeremia exigua (Pleosporales) — Detected at A2 Wiese, E1, and E1.1 Market Garden. This is a known plant pathogen (causes leaf spots and stem lesions on a wide range of hosts). Its presence at low levels is not alarming — many low-level plant pathogens persist in soil without causing disease — but it is worth noting as part of the baseline.

Keratinophyton wagneri (Onygenales) — Found at five sites, strongest at A1 Forsthaus (0.28%) and A3 Alter Obstgarten (0.22%). Keratinophyton species decompose keratin (hair, feathers, horn). Liquid manure (Gülle) contains significant amounts of keratin-rich material — fragments of hair, skin cells, and other animal-derived organic matter from the livestock it originates from. The strong Keratinophyton signal at Forsthaus is consistent with the Gülle legacy interpretation, and together these two species paint a coherent picture of a soil whose decomposer community is still oriented around the previous management's inputs. Their also being present at A3 Alter Obstgarten at lower levels may reflect historic inputs at that site as well, or simply wildlife activity around the old orchard.

2.4 What's notably absent

More Glomeromycota (AMF) — Only a single AMF species was detected, and at only one site. As noted, this is likely an artefact of the ITS primers used, but it is still a gap in the picture. If plant nutrition and soil structure are priorities, a targeted AMF survey would be valuable.

Ectomycorrhizal fungi — No ectomycorrhizal species (e.g. Cortinarius, Russula, Amanita, Boletus) were detected. This is expected if the sites are grassland, agricultural, or orchard — ectomycorrhizal fungi associate primarily with forest trees (oaks, beeches, pines). If A1 Forsthaus includes or borders woodland, you might expect some, and their absence could suggest the soil sampling point was away from tree roots.

Entomopathogenic fungi — Species like Beauveria bassiana or Metarhizium anisopliae (which naturally control insect pest populations in soil) were not detected. Tyrannicordyceps fratricida (a mycoparasitic fungus, found only at E3 Spannweid) is the closest to an entomopathogen detected. The absence of classical entomopathogens is not necessarily a problem — they can be present at very low levels that fall below the eDNA detection threshold — but their presence would have been a positive sign for integrated pest management.

2.5 Site comparison — Fungi

E3 Spannweid Mix stands out as the fungal diversity champion. It has the highest functional diversity (2.08), the highest evolutionary diversity (34.67), the second-highest OTU count (169), and the most unique OTUs found only at this site (31). It is the only site with AMF (Claroideoglomus claroideum) and Mortierella alpina, and hosts the strongest Clonostachys (biocontrol) and Humicola (cellulose decomposition) signals. An important factor here: E3 Spannweid contains two distinct soil types within a single field — peat and loam — and the sample is a mix across both. This means the eDNA is capturing two ecologically distinct fungal communities simultaneously, which partly explains the elevated diversity metrics and high number of unique OTUs. Species adapted to the acidic, moisture-retaining peat zone and species more typical of loamy mineral soil are both present, and their overlap produces a richer combined picture than either soil type alone would. The AMF detection, for instance, may be coming specifically from the loamy zone where phosphorus dynamics differ from the peat. This doesn't diminish the site's richness — both communities are genuinely present and functional — but it is worth bearing in mind when comparing E3 to sites with a single, uniform soil type.

E1 Market Garden Mix has the highest raw OTU richness (174) and the most named species (20). It hosts Trichoderma, Stachylidium, and strong Musicillium presence — all associated with active decomposition of diverse organic substrates. The market garden's high input diversity (vegetable waste, different crops, likely composting) is clearly reflected in a correspondingly diverse fungal community.

A1 Forsthaus is an interesting outlier. While its OTU count (141) is mid-range, its community character is distinctive — dominated by Podospora (dung fungi), Solicoccozyma (yeasts), and Keratinophyton (keratin decomposers). This suggests a soil environment shaped by animal inputs rather than diverse plant residues. It also has 20 unique OTUs, suggesting a genuinely different habitat.

A3 Alter Obstgarten has the lowest fungal OTU count (112) and functional diversity (1.71). This site has only recently come into management, though a cover crop mix was established from autumn 2023. Given that timeline, the fungal community is still in an early stage of development. The presence of Trichoderma and a range of Sordariales suggests the decomposer community is already functional and responding to the new plant inputs from the cover crop — a promising sign for a site so early in its journey.

2.6 Assessment

Your soils support a diverse and functionally active fungal community. The dominance of decomposer guilds (Hypocreales, Sordariales) across all sites indicates effective organic matter cycling. The presence of biocontrol fungi (Trichoderma, Clonostachys) at multiple sites is a positive sign for natural disease suppression. The main gap is the very limited AMF detection — addressing this with a dedicated survey would strengthen the picture considerably, especially for understanding nutrient availability to plants.

3. Soil Bacteria — A Deep Dive

3.1 Overall bacterial diversity

The bacterial community is by far the richest of the three assays, with 628 OTUs detected across the eight sites. Bacterial species richness per site ranges from 315 (E2 Haus Huegel) to 389 (E1 Market Garden). However, only 21 OTUs could be identified to species level — this is normal for bacteria, where the vast majority of soil species remain unnamed and are known only from their DNA sequences.

Notable: A3 Alter Obstgarten and E1 Market Garden share the highest bacterial functional diversity (3.27). These two sites appear to offer the most complex bacterial metabolic environment — likely reflecting diverse carbon sources and nutrient conditions.

A shared core of 160 OTUs (25% of total) was found at all eight sites. An additional 95 OTUs were found at 6–7 sites. This means the bacterial communities share a strong common foundation, with differentiation happening at the margins — 133 OTUs were unique to a single site. Interestingly, A3 Alter Obstgarten had the most unique bacterial OTUs (47), more than any other site, suggesting its soil chemistry or microhabitat conditions support distinctive bacterial lineages that don't occur elsewhere.

3.2 Phylum-level composition and what it means

The bacterial phylum profile is a remarkably powerful diagnostic tool for soil condition. Here is what your soils show:

Actinobacteriota (128 OTUs, dominant at all sites with 76–93 OTUs each): The single largest phylum. Actinobacteria are gram-positive bacteria that are the backbone of soil organic matter decomposition. They break down tough compounds like chitin, cellulose, and humic substances. Many produce antibiotics that help structure the microbial community and suppress pathogens. Their dominance is a hallmark of well-aerated, reasonably fertile agricultural soils with moderate to neutral pH. If Actinobacteria were absent or rare, it would be a serious concern — their strong presence across all your sites is reassuring.

Proteobacteria (107 OTUs, 54–69 per site): The second most diverse phylum, and the most metabolically versatile group of bacteria on Earth. They include nitrogen fixers, nitrifiers, denitrifiers, sulphur oxidisers, and many plant-growth-promoting bacteria. E1 Market Garden has the highest Proteobacteria count (69), which likely reflects the diverse nutrient conditions in an actively managed market garden soil. Proteobacteria tend to thrive where nutrients are available and organic inputs are frequent — they are often characterised as "r-strategists" (fast-growing, nutrient-responsive).

Planctomycetota (81 OTUs, 31–46 per site): A fascinating phylum that is increasingly recognised as important in soil. Planctomycetes are involved in the breakdown of complex organic molecules and may play roles in anaerobic ammonium oxidation (anammox). Their relatively high diversity across your sites is a positive indicator of functional complexity. A2 Wiese Mix has the highest Planctomycetota count (46), which may reflect wetter or more variable moisture conditions in the meadow soil.

Acidobacteriota (55 OTUs, 33–41 per site): Acidobacteria are slow-growing "K-strategists" that dominate in acidic, low-nutrient soils. They are specialists in degrading complex polymers under nutrient-limited conditions. The fact that they maintain a consistent presence (but don't dominate) suggests your soils are moderately fertile with some pH variation. E3 Spannweid and E3.1 Spannweid 14 have the highest Acidobacteriota counts (40–41), which could indicate slightly more acidic or lower-nutrient conditions at these sites compared to the Market Garden.

Verrucomicrobiota (44 OTUs, 22–32 per site): Verrucomicrobia are common in soils but were historically underappreciated. They include some methane oxidisers and are generally considered positive indicators of soil health — they tend to be more abundant in undisturbed, biologically active soils. A3 Alter Obstgarten has the highest count (32), which fits with a long-established, relatively undisturbed site.

Chloroflexota (38 OTUs): Green non-sulphur bacteria, often found in soils with complex carbon cycling. Their presence contributes to overall metabolic diversity.

Gemmatimonadota (27 OTUs): Gemmatimonads are adapted to low-moisture conditions and are often found in drier soils. Their presence adds resilience to the community — they continue functioning when wetter-adapted bacteria slow down during dry spells.

Nitrospirota (4 OTUs): Small in number but disproportionately important — see below.

Thermoproteota (8 OTUs): Formerly classified as Thaumarchaeota, these are actually archaea, not bacteria. Their detection here is significant — see below.

3.3 Named bacterial species and their significance

Nitrososphaera viennensis (Thermoproteota/Nitrososphaeraceae) — Detected at all eight sites, and the single most abundant named organism in the entire bacterial dataset (2.14% at Forsthaus, 2.16% at Spannweid 14). Despite being classified here under "bacteria," Nitrososphaera is actually an ammonia-oxidising archaeon (AOA). It performs the first step of nitrification: converting ammonia (NH₃) to nitrite (NO₂⁻). This is critical for nitrogen cycling — without ammonia oxidisers, nitrogen from organic matter decomposition and fertilisers cannot be converted into forms that plants can use. Its high abundance at Forsthaus and Spannweid 14, and lower abundance at Market Garden (0.37%), is noteworthy. Since none of these sites receive any fertiliser or manure, all nitrogen must come from atmospheric deposition, nitrogen fixation, and the mineralisation of organic matter. The pattern makes ecological sense: at the longer-managed Market Garden, years of diverse plant growth and organic matter accumulation have likely built a more complex nitrogen cycling community where many organisms participate and no single species dominates. At the recently-started Forsthaus and at Spannweid 14 (with its peat soil, which locks nitrogen in organic forms), the community may be more reliant on this single archaeal species to drive ammonia oxidation. The Spannweid peat soils in particular are characteristically nitrogen-poor in available forms despite containing large amounts of total organic nitrogen — exactly the environment where a specialist ammonia oxidiser would be critically important.

Nitrospira japonica (Nitrospirota/Nitrospiraceae) — Found at all eight sites (0.24%–0.52%). Nitrospira completes the second step of nitrification: oxidising nitrite (NO₂⁻) to nitrate (NO₃⁻). Together with Nitrososphaera, it forms a complete nitrification pathway in your soils. Nitrospira species are now known to be the dominant nitrite oxidisers in most soils worldwide, having replaced the previously-assumed Nitrobacter. The consistent presence of Nitrospira across all sites is an excellent sign — it means your soils have a fully functioning nitrogen cycle. Recent research has also shown that some Nitrospira can perform "comammox" — complete ammonia oxidation (ammonia all the way to nitrate in a single organism). This makes their presence even more significant for plant nutrition.

Nitrospira moscoviensis — A second Nitrospira species, found only at E1 and E1.1 Market Garden. Having two Nitrospira species at the Market Garden sites adds functional redundancy to nitrogen cycling — if one species is stressed (e.g. by pH changes or drought), the other can maintain nitrite oxidation. This is a sign of a robust nitrogen cycling system.

Nocardioides islandensis (Actinobacteriota) — Found at all eight sites (0.49%–0.67%). Nocardioides are versatile decomposers that can break down a wide range of organic compounds, including some pollutants and pesticide residues. Their ubiquity suggests they are a fundamental part of the soil's decomposer workforce.

Microlunatus panaciterrae (Actinobacteriota) — Found at all eight sites, strongest at E2 Haus Huegel (0.71%). Microlunatus is a polyphosphate-accumulating bacterium — it stores phosphorus. In a soil context, organisms like this contribute to phosphorus cycling by accumulating and then releasing phosphorus over time. Its consistent presence adds to the picture of functional nutrient cycling.

Cellvibrio fulvus and C. gandavensis (Proteobacteria) — Cellvibrio species are cellulose decomposers, found at three to five sites, strongest at A3 Alter Obstgarten and E1 Market Garden. Cellulose is the most abundant organic polymer on Earth, and effective cellulose decomposition is fundamental to soil health. Combined with the cellulolytic fungi (Trichoderma, Humicola), these bacteria indicate a well-functioning cellulose decomposition system.

Flavisolibacter ginsengiterrae (Bacteroidota) — Found at seven sites. Flavisolibacter species are involved in the decomposition of complex organic polymers and are typically found in soils with good organic matter content.

Nakamurella flavida (Actinobacteriota) — Found at seven sites. A polysaccharide-accumulating bacterium that contributes to soil aggregate formation — one of the most important physical processes in soil health. Soil aggregates are clusters of mineral particles glued together by biological secretions and fungal hyphae, creating the crumb structure that allows water infiltration, root penetration, and long-term carbon storage. Nakamurella is one of several detected organisms contributing to this process, and the full picture of soil aggregation across your sites is discussed in detail in the supplementary notes⁴.

Singulisphaera acidiphila (Planctomycetota) — Found at six sites. As its name suggests, it prefers slightly acidic conditions. Its widespread presence adds to the functional complexity of the community.

Verrucomicrobium spinosum (Verrucomicrobiota) — Found at A3 Alter Obstgarten, E3 Spannweid, and E3.1 Spannweid 14. A well-studied soil bacterium that degrades complex polysaccharides. Its presence at these three sites reinforces their characterisation as soils with active, complex organic matter decomposition.

Sphingomonas (Proteobacteria) — Multiple OTUs found across sites. Sphingomonas species are metabolically versatile and can degrade a wide range of organic compounds, including some environmental contaminants. They also produce plant-growth-promoting substances. Their presence adds both nutrient cycling and potential bioremediation capacity.

Gemmatimonas aurantiaca (Gemmatimonadota) — Found at A1 Forsthaus, A3 Alter Obstgarten, and E2 Haus Huegel. A photoheterotroph (can use light energy) that is adapted to dry conditions. Its presence at these three sites may indicate periods of lower soil moisture.

3.4 What's notably absent or underrepresented

Rhizobia and free-living nitrogen fixers — No classic rhizobial nitrogen-fixing genera (Rhizobium, Bradyrhizobium, Sinorhizobium) were named in the results. This doesn't mean they're absent — with only 21 of 628 OTUs named to species, many unnamed OTUs likely include nitrogen fixers. However, if legumes are grown or the sites include legume-rich meadows, a dedicated check for nitrogen-fixing community diversity would be worthwhile.

Bacillus and Pseudomonas — These two genera are among the most commercially important soil bacteria (many biocontrol and biofertiliser products are based on them). Neither was detected by name, though again, they may be present among the unnamed OTUs. Bacillus can be underrepresented in eDNA surveys because its spores are difficult to lyse during DNA extraction.

Cyanobacteria — Only 3 OTUs of cyanobacteria were detected. These photosynthetic bacteria can form biological soil crusts and fix nitrogen, but they are primarily surface organisms and may simply not have been at the soil depth sampled.

Firmicutes — Only 6 OTUs. In many agricultural soils, Firmicutes (especially Bacillus and Clostridium) are more prominent. Their low representation could reflect extraction bias or genuinely low abundance.

3.5 Site comparison — Bacteria

The bacterial communities are more uniform across sites than the fungi or invertebrates, with a shared core of 160 OTUs. The major differentiating factors are:

E1 Market Garden Mix and E1.1 Market Garden 70 have the highest bacterial richness (382 and 377 OTUs) and evolutionary diversity (20.30 and 20.44). They are the only sites with two Nitrospira species and have strong Proteobacteria representation. These soils appear to be the most metabolically active and functionally complete from a bacterial perspective.

A3 Alter Obstgarten matches Market Garden for functional diversity (3.27) and has the most unique bacterial OTUs (47 found only here). The old orchard's long establishment history and likely lower disturbance have allowed a distinctive bacterial community to develop. Its high Verrucomicrobiota count (32) supports this interpretation.

A1 Forsthaus and E2 Haus Huegel have the lowest bacterial richness (319 and 315) and functional diversity (2.75 and 2.71). For Forsthaus, this fits the pattern of a site only just brought into management — the bacterial community simply hasn't had time to diversify. For Haus Huegel, this is more surprising given its several years of management and strong invertebrate community; its peat/loamy soil mix may create different bacterial niches than the pure loam of the Market Garden.

E3 Spannweid sites sit in the middle for bacterial metrics but have the highest Acidobacteriota counts (40–41 OTUs). This is consistent with the peat component of the soil: peat is characteristically acidic, high in recalcitrant organic matter, and lower in available nutrients — precisely the conditions where Acidobacteriota thrive. It is also worth noting that E3 Spannweid Mix contains both peat and loamy soil within one field, so the bacterial community similarly reflects two distinct soil environments overlapping in a single sample. This is not a concern but a natural signature of the soil heterogeneity at this site.

3.6 Soil aggregation — what the data shows

Soil aggregation deserves specific attention because it underpins almost everything else: water infiltration and retention, drainage, aeration, root penetration, erosion resistance, and long-term carbon storage all depend on a well-aggregated soil structure. Aggregation is also the mechanism by which biologically active soil physically locks up carbon — organic matter trapped inside aggregates is protected from decomposition in a way that free organic matter is not. When you disturb a well-aggregated soil through tillage or compaction, you break open those structures and release that carbon. When you build aggregation, you build both fertility and carbon storage simultaneously.

Aggregates form at different scales, and different organisms are responsible for each — like a construction process with different trades working at different levels.

Layer 1 — Microaggregates: bacteria lay the foundations

The smallest aggregates (under 250 micrometres — invisible to the naked eye) are built primarily by bacteria secreting exopolysaccharides (EPS): sticky, glue-like polymers that coat mineral particles and bind them together. Think of it as biological mortar between the grains of soil. Multiple organisms detected across your sites contribute to this layer. Nakamurella flavida (7 sites) and Microlunatus panaciterrae (all 8 sites) are polysaccharide accumulators whose secretions contribute directly to particle binding. Sphingomonas (multiple sites) produces sphingan polysaccharides — high-molecular-weight polymers particularly effective at bridging clay particles. Gemmatimonas (all sites) continues this work even during dry spells, when many other bacteria become inactive, giving the aggregate structure resilience through dry periods. The Bacteroidota broadly (Flavisolibacter, Terrimonas, Flavobacterium — found across 5–7 sites each) coat soil particle surfaces with biofilm material that anchors and stabilises early aggregates.

The Actinobacteriota deserve special mention. Unlike most bacteria, which are single-celled, Actinobacteria grow in filamentous networks — physically threading through soil like a fine mycelium, stitching particles together in a way no single-celled organism can. With 76–93 Actinobacteriota OTUs at every site, this hyphal microaggregation capacity is the most consistent and well-supported biological process in your entire dataset. It is, in effect, the structural skeleton on which everything else builds.

Layer 2 — Macroaggregates: fungi stitch the structure together

Above the microscale, fungal hyphae become the primary architect. As fungi grow through soil, their hyphae — physically much larger and longer than bacterial filaments — reach across and bind microaggregates into larger macroaggregates (250 micrometres to several millimetres). This is the scale that gives soil its visible crumb structure, the texture you can feel and see when you pick up a handful of good soil. Your Sordariomycetes (dominant at all sites), Humicola (7 sites), and Mortierella (E3 Spannweid) all contribute hyphae to this layer. As fungal biomass builds with continued management and increasing plant diversity, this macroaggregation layer will strengthen progressively. It is already well-established at the longer-managed sites.

Layer 3 — Earthworm casts: the most stable structures of all

The largest and most stable macroaggregates in soil are earthworm casts. As earthworms pass soil through their gut, they mix mineral particles with mucus, gut bacteria, and partially digested organic matter. The resulting cast material is chemically and physically unlike the soil that went in — it is denser in organic matter, more microbially active, and far more stable as an aggregate. Cast aggregates resist breakdown by water (which is when aggregates are most vulnerable) far better than non-ingested soil. Earthworm burrows also create channels that improve drainage and aeration, and the lining of those burrows becomes a zone of especially intense biological activity.

Your sites clearly have an earthworm community. Lumbricidae DNA was detected at multiple sites, and the enchytraeid (potworm) community is particularly strong — especially the very large Marionina argentea signal at E1 Market Garden (11.92% of invertebrate reads). Enchytraeids are smaller than earthworms but work the same way, processing organic matter through their guts and producing stable, biologically enriched cast material in the upper soil layers. The combination of Lumbricidae and a dense Enchytraeidae population at Market Garden means this site is almost certainly producing substantial quantities of cast-derived macroaggregates — and the 5/5 spade test at Market Garden 70 is the physical confirmation of that.

Layer 4 — Glomalin: the waterproofing coat

Above and across all three layers sits the contribution of arbuscular mycorrhizal fungi (AMF), through a protein called glomalin. AMF coat their hyphae and nearby soil particles with glomalin — a sticky, hydrophobic (water-repelling) glycoprotein that acts as a waterproof sealant on aggregate surfaces. Crucially, glomalin remains stable and functional even when the soil gets wet, which is precisely when aggregates built only from bacterial EPS and fungal hyphae are at their most vulnerable to collapse. In soils with a well-established AMF community, glomalin can account for 20–30% of total aggregate stability, and represents 5–8% of total soil carbon. It is not a minor contribution — it is, in many healthy soils, the single most important compound holding the whole structure together.

This is currently the clearest gap in your aggregation picture. Only one AMF species (Claroideoglomus claroideum) was detected, at a single site (E3 Spannweid). Glomalin production is therefore likely low across most of your sites right now — not because the management is wrong, but because AMF populations take years to fully establish, and several of your sites are still early in their journey. E3 Spannweid, with its peat soil and several years of management, is furthest along. As management time accumulates and plant diversity builds, AMF should spread and the glomalin layer will gradually close the gap. This is the area most likely to show the most significant improvement in aggregate stability over the coming years — and the reason the planned AMF survey is particularly worthwhile.

What you are already seeing

The 5/5 spade test at E1.1 Market Garden 70 is the physical proof that layers 1–3 are already working well at the most developed site. The biology is rich, the earthworms and enchytraeids are there in large numbers, and the result is measurable, best-in-class soil structure. The glomalin layer — layer 4 — is the next frontier, and the one that will take the longest to build. But the foundation for it is already forming.

3.7 Assessment

The bacterial picture is strong. A complete nitrification pathway (Nitrososphaera → Nitrospira) is present at every site. Cellulose decomposition, polyphosphate accumulation, and complex organic matter breakdown are all well-represented. The community is dominated by Actinobacteriota and Proteobacteria — the phylum profile of a healthy, well-aerated temperate soil. The main areas where more information would be valuable are nitrogen fixation capacity and the Bacillus/Pseudomonas community, both of which may be underrepresented by the current assay's detection approach.

4. Soil Invertebrates — A Deep Dive

4.1 Overall invertebrate diversity

The soil invertebrate community is the smallest of the three assays — 107 OTUs detected, with 17 to 45 per site. However, this is arguably the most ecologically informative dataset, because soil invertebrates occupy a wide range of trophic levels (from bacterial feeders to predators to plant parasites), and their community structure directly reflects soil food web complexity.

Twenty-four OTUs were identified to species level, and many more were identified to family or genus — sufficient for ecological interpretation.

Only a single OTU was shared across all eight sites (likely a ubiquitous nematode), and 59 OTUs (55% of the total) were unique to a single site. This high turnover between sites means the invertebrate community structure differs dramatically from place to place — far more so than bacteria or fungi.

4.2 The major groups detected

Nematoda (45 OTUs — 42% of all invertebrates): Nematodes are the most abundant multicellular animals in most soils. They occupy every trophic level in the soil food web and are among the best-studied bioindicators for soil condition. The nematode community here can be grouped by feeding strategy:

• Bacterial feeders (Rhabditida: Cephalobidae, Bunonematidae; also Plectidae): These nematodes eat bacteria and are enrichment indicators — their abundance increases when bacteria flourish, typically after fresh organic matter inputs or disturbance. Eucephalobus oxyuroides, E. striatus, and Bunonema reticulatum were all detected. They are present at four to six sites and are most diverse at E1 Market Garden and A3 Alter Obstgarten — sites with rich organic inputs.

• Fungal feeders (Aphelenchida: Aphelenchidae, Aphelenchoididae; some Tylenchidae): Aphelenchus avenae is the standout here — found at seven of eight sites (absent only from Forsthaus), making it one of the most widespread invertebrates detected. Aphelenchus is a specialist fungal feeder that controls fungal populations. Its near-ubiquity confirms an active fungal community available as food, and its absence from Forsthaus aligns with that site's generally depauperate invertebrate community. Aphelenchoides saprophilus, another fungal feeder, was found only at A3 Alter Obstgarten, where it accounted for 1.46% of reads — a strong signal.

• Plant parasites (Tylenchida: Hoplolaimidae, Tylenchidae): This is a group to monitor. Pratylenchus neglectus (root lesion nematode) was detected at A2 Wiese Mix. Pratylenchus is one of the most economically damaging nematode genera worldwide — it migrates through root tissue, creating lesions that allow secondary infections. At 0.09% of reads, this is a low-level detection, but it confirms the presence of the organism. Helicotylenchus minzi (spiral nematode) was also found at A2 Wiese. Helicotylenchus feeds on root surfaces and can reduce plant vigour, particularly in grasslands. Several Tylenchidae species (Filenchus, Basiria, Irantylenchus, Boleodorus) were found — these are mostly weak plant parasites or fungal feeders and are generally not considered damaging. Interestingly, the plant-parasitic nematodes are concentrated at the A-sites (especially A2 Wiese) rather than the E-sites. This may reflect different crop histories, grass dominance at A2, or more effective biological suppression at the E-sites.

• Predators (Dorylaimida: Mononchidae, Nygolaimidae): Clarkus papillatus (Mononchidae) was found at A3 Alter Obstgarten and E2 Haus Huegel. Mononchids are predatory nematodes that eat other nematodes — including plant parasites. Their presence is a sign of a mature, multi-layered soil food web. Nygolaimidae (found at two sites) are also predatory. The Dorylaimida order is well-represented overall (found across all sites), though mostly as unnamed OTUs. Dorylaimids are generally considered sensitive to disturbance and pollution — their persistence across sites is a good indicator.

• Omnivores and others (Dorylaimida: Qudsianematidae, Alaimidae): Alaimus OTUs were found at one to three sites — Alaimus is a non-feeding (or microbial-feeding) nematode associated with relatively undisturbed soils. The Qudsianematidae OTU was found at four sites and is likely an omnivore.

Arthropoda (39 OTUs — 36% of all invertebrates):

• Collembola (springtails) — 12 OTUs, found at all sites but most diverse at A1 Forsthaus (5), E1 Market Garden (3), and E2 Haus Huegel (5). Springtails are critically important soil organisms: they feed on fungi, bacteria, decaying plant material, and algae, and their grazing activity stimulates microbial turnover and nutrient cycling. They also contribute to soil structure through their burrowing. The five collembolan OTUs at Forsthaus are notable — it is the one metric where Forsthaus exceeds most other sites, suggesting that while the overall invertebrate community is depauperate, the microarthropod community may be relatively intact.

• Arachnida (mites) — 14 OTUs. Mites occupy multiple trophic levels. Oppiella nova (Oribatida, found at E3 Spannweid) is one of the most important soil health bioindicators. Oribatid mites are slow-reproducing, sensitive to disturbance, and feed on decomposing material and fungi — their presence indicates stable, mature soil conditions. Veigaia nemorensis (Mesostigmata, found at E1 Market Garden, E1.1, and E2 Haus Huegel) is a predatory mite that feeds on nematodes, springtails, and other small soil invertebrates. Predatory mites are top predators in the soil food web, and their presence indicates sufficient prey populations and food web complexity. Histiostoma feroniarum (Sarcoptiformes, found only at E2 Haus Huegel at a striking 5.32% of reads) is a mite associated with decomposing organic matter and manure — it feeds on nematodes and bacteria in nutrient-rich microhabitats.

• Insecta — 7 OTUs. Poecilus cupreus (Carabidae — a ground beetle) was detected at E2 Haus Huegel. Poecilus cupreus is a well-known beneficial predator in agricultural systems — it feeds on aphids, slugs, and other crop pests. Its detection via soil eDNA likely reflects DNA from larvae (which live in soil) or from adult beetles sheltering in or on the soil surface.

• Diplopoda (millipedes) — 4 OTUs, found at E1 Market Garden, E1.1, and E2 Haus Huegel. Millipedes are important decomposers that fragment plant litter, making it accessible to smaller organisms and microbes. Their presence at the E-sites reflects available leaf litter and organic matter.

• Chilopoda (centipedes) — 1 OTU at E3.1 Spannweid 14. Centipedes are predators, contributing to pest regulation.

Annelida (6 OTUs — worms):

• Enchytraeidae (potworms) — Enchytraeus dichaetus (E2 Haus Huegel, E3 Spannweid) and Marionina argentea (A2 Wiese, A3 Alter Obstgarten, E1 Market Garden at a very high 11.92%, E2 Haus Huegel). Enchytraeids are small (1–3 cm) white worms that process organic matter in the upper soil layers. In many temperate grassland and agricultural soils, they process more organic matter than earthworms. Marionina argentea's extremely strong signal at E1 Market Garden (11.92% of reads — the highest single-species read percentage in the entire invertebrate dataset) indicates a massive population. This is consistent with a soil rich in organic matter and with active composting or mulching practices.

• Lumbricidae (earthworms) — Unnamed Lumbricidae OTUs were detected at several sites (Crassiclitellata order). The absence of named earthworm species is a limitation — earthworms are keystone soil organisms, and species-level identification (e.g. Lumbricus terrestris vs. Aporrectodea caliginosa) matters greatly for understanding soil function. However, the eDNA assay used targets 18S rRNA, which has limited resolution for earthworm species identification.

Rotifera (7 OTUs): Microscopic animals found mostly in water films in soil. Detected primarily at E1 Market Garden (6 OTUs, including bdelloid rotifers). Their presence indicates moist microhabitats with active microbial communities — bdelloids are bacterial feeders.

Tardigrada (3 OTUs): Water bears — microscopic, incredibly resistant animals found in soil water films. Detected at E1 Market Garden, E1.1, E3 Spannweid, and E3.1 Spannweid 14. Their presence is a positive sign of microhabitat complexity.

Gastrotricha (3 OTUs): Tiny worm-like animals that live in water films between soil particles. Found at A1 Forsthaus and E1 Market Garden. Rarely reported in soil biodiversity surveys — their detection here via eDNA demonstrates the technique's ability to capture groups that traditional surveys miss entirely.

Platyhelminthes (2 OTUs): Stenostomum arevaloi (a flatworm) found at E1 Market Garden. Soil flatworms are bacterial feeders and predators of other microscopic organisms.

Mollusca (1 OTU): Found at E1 Market Garden and E2 Haus Huegel. Likely a small slug or snail — a surface/soil interface organism.

4.3 What's notably absent

Named earthworm species — Earthworms are among the most important soil ecosystem engineers, yet no earthworm species were identified by name. Lumbricidae DNA was detected, so earthworms are present, but the 18S marker doesn't resolve earthworm species well. A dedicated earthworm survey (either physical sampling or a CO1-based eDNA approach) would fill this gap.

Larger predatory arthropods — Only one ground beetle (Poecilus cupreus) was detected. Other beneficial predators like rove beetles (Staphylinidae), spiders (Araneae), or centipedes (Chilopoda, only 1 OTU) were barely represented. This is partly a limitation of the soil eDNA approach — many of these organisms live on or above the soil surface and may leave little DNA in the soil matrix.

Isopoda (woodlice) — Important decomposers in surface litter, not detected. Again, likely a reflection of the soil sampling depth rather than true absence.

4.4 Site comparison — Invertebrates

The invertebrate community shows the starkest differences between sites:

E1 Market Garden Mix is overwhelmingly the most diverse (45 OTUs, evolutionary diversity 5.60). It has the most phyla represented (8 plus one unassigned), the strongest enchytraeid signal (Marionina argentea at 11.92%), rotifers, tardigrades, flatworms, and a slug. This is a soil teeming with life at every trophic level. The market garden's combination of diverse organic inputs, likely mulching and composting, varied root environments from different crops, and probably irrigation creates ideal conditions for a complex soil food web.

E2 Haus Huegel Mix has the second-most diverse arthropod community (15 OTUs) and a strong predator presence (Veigaia, Histiostoma, Poecilus). Its enchytraeid community is also strong. The strong Histiostoma signal (5.32%) suggests organic-matter-rich microsites (possibly manure or composting spots).

A1 Forsthaus is strikingly depauperate — only 17 OTUs total, with only 1 named species (Filenchus vulgaris, a weak plant parasite). The community is dominated by springtails (5 OTUs) with very few nematodes (only 4 OTUs). Knowing the full picture of this site, this is entirely expected: at the time of sampling the field had been open, bare soil for months following Gülle application by the previous manager, with a pumpkin and clover culture only just beginning to establish. There were effectively no established plant roots in the soil — and plant roots are the primary engine of the soil food web, providing exudates that feed bacteria and fungi, which in turn feed nematodes and larger invertebrates. The springtails' presence (5 OTUs) is actually a positive sign — microarthropods can colonise quickly from surrounding soil — but the nematode community, which depends on established microbial populations fed by plant roots, is barely present. This site will be the most dramatic to re-survey: as the plant cover establishes and diversifies over coming seasons, the below-ground response should be measurable and rapid.

A2 Wiese Mix has the most diverse nematode community (18 OTUs, including both plant-parasitic and bacterial-feeding species). This makes sense for a four-year-old Kunstwiese: four years of dense, mixed grass and legume root growth has built up exactly the kind of well-developed below-ground root network that supports a rich nematode community. The plant-parasitic nematodes (Pratylenchus, Helicotylenchus) are typical companions to established grass swards — grass roots are a prime host — and at these levels are not a concern. The legume component of the Kunstwiese mix may also be quietly supporting nitrogen-fixing bacterial associations, which, while not directly visible in the named OTU list, could be contributing to the site's relatively strong bacterial functional diversity.

E3 Spannweid sites have moderate invertebrate diversity but include the important bioindicator Oppiella nova (oribatid mite) and tardigrades — organisms that indicate stable, undisturbed soil conditions.

4.5 The soil food web — Putting it together

The invertebrate data allows us to sketch the soil food web at each site. A healthy, complex soil food web should include organisms at multiple trophic levels:

1. Primary decomposers (feeding on dead organic matter): Enchytraeids, earthworms, millipedes, oribatid mites → Present at most sites, strong at E1, E2
2. Microbial grazers (feeding on bacteria and fungi): Bacterial-feeding nematodes, fungal-feeding nematodes, springtails, rotifers → Present at most sites
3. Secondary consumers (feeding on other invertebrates): Predatory nematodes, predatory mites, ground beetles, centipedes → Present but patchy; strongest at E1, E2
4. Plant parasites (feeding on roots): Present at A2, A3, some E-sites at low levels

E1 Market Garden and E2 Haus Huegel have the most complete food webs, with organisms at all four levels. A1 Forsthaus has a truncated food web, with springtails but few nematodes and no detected predators — consistent with a site at the very beginning of its management journey, where the soil food web has not yet had time to build complexity.

4.6 Assessment

The invertebrate data reveals stark differences in soil food web complexity between your sites — and these differences map remarkably well onto the management timeline. E1 Market Garden (longest under management) has 45 OTUs spanning eight phyla and all trophic levels. A1 Forsthaus (just started) has only 17 OTUs and a truncated food web. The sites with several years of management (Spannweid, Haus Huegel) fall in between. This pattern strongly suggests that the organic, no-input management approach is building soil food web complexity over time — and that the process takes years, not months. The presence of plant-parasitic nematodes (Pratylenchus, Helicotylenchus) at A2 Wiese is notable but currently at low levels. The detection of key bioindicators — Oppiella nova, Veigaia nemorensis, Clarkus papillatus — at several E-sites confirms that years of organic management can build mature, multi-layered food webs with natural pest regulation capacity.

5. Cross-Kingdom Synthesis — What Your Soils Are Telling You

5.1 Site rankings

Combining all three assays and the management context, the sites tell a compelling story about how organic, no-input land management builds soil biological complexity over time:

Longest managed — the benchmark:
- E1 Market Garden Mix: Highest diversity in all three kingdoms. The most complete soil food web. Two Nitrospira species for nitrogen cycling. Highest fungal OTU count. Massive enchytraeid population. Years of diverse cropping without external inputs have built a soil ecosystem that generates its own fertility and pest regulation. This is your proof of concept.
- E3 Spannweid Mix: Highest fungal functional and evolutionary diversity. Only site with AMF. Highest Clonostachys (biocontrol). Good invertebrate diversity including Oppiella nova. This field contains both peat and loamy soil within a single sampling area, meaning the results capture two overlapping communities simultaneously — which partly explains the exceptional diversity metrics and the high number of unique OTUs (31). The biological richness is real, but some of it is structural: the two soil types together produce a wider range of ecological niches than either alone. This also makes E3 harder to compare directly with single-soil-type sites, and worth bearing in mind when interpreting future re-surveys.

Several years under management — building well:
- E2 Haus Huegel Mix: Strong arthropod diversity. Predatory mites and ground beetles present. Good enchytraeid community. The peat/loam soil supports a distinctive invertebrate community. Lower bacterial diversity than Market Garden but solid functional representation — possibly still catching up.
- E1.1 Market Garden 70: A single-stripe probe, and the one with a confirmed 5/5 spade test result for soil aggregation. Its biology is the most direct link in the entire dataset between eDNA richness and physical soil quality — strong Actinobacteriota, abundant enchytraeids, detected earthworms, rich fungi, all corresponding to the best possible physical structure.

Recently started — early signs of life:
- A3 Alter Obstgarten Mix: Despite being newly managed, the autumn 2023 cover crop appears to have had a rapid effect on bacteria — this site has the highest bacterial uniqueness (47 unique OTUs), highest Verrucomicrobiota, and shares the highest bacterial functional diversity with Market Garden (3.27). The cover crop may have introduced root exudates and organic matter that stimulated bacterial diversification ahead of the other newly-started sites. A promising early result.
- A2 Wiese Mix (ForsthausOben / Adlisberg Wiese): A four-year-old Swiss Kunstwiese (sown grass/legume meadow) — not newly established but a well-rooted, diverse sward. Good nematode diversity reflecting four years of established root growth. Plant-parasitic nematodes are typical companions to grass roots and not a concern at current levels. The legume component likely supports nitrogen-fixing bacterial associations not captured as named species but contributing to the functional community.

Just started — and carrying a legacy:
- A1 Forsthaus: Consistently lowest invertebrate diversity (17 OTUs, only 1 named species). Low bacterial richness. Fungal community still dominated by organisms processing the legacy of liquid manure (Gülle) applied by the previous land manager — dung-decomposing Podospora and keratin-decomposing Keratinophyton are both fingerprints of that input, still visible in the soil biology at the time of sampling. This makes Forsthaus not just a "newly started" site but a transitional one — the soil is actively shifting from a manure-driven biology toward the plant-residue-driven biology that no-input management will build. The 2024 data captures that transition in progress, and Forsthaus will be the most instructive site of all to re-survey: how quickly does the Gülle footprint fade? When do the first biocontrol fungi and mycorrhizal organisms appear? This site is a slow-motion before-and-after.
- E3.1 Spannweid 14: Lower overall diversity than the Spannweid Mix, which makes sense: it is a single-stripe probe from a deliberate nettle monoculture, not a mixed-community sample. Nettles produce distinctive, nitrogen-rich organic matter that decomposes rapidly — feeding a specific subset of decomposer fungi rather than the broad range supported by a diverse plant community. Its unique OTUs (20 found nowhere else) likely reflect the specific chemistry of nettle litter and root exudates, which select for a fungal community not found under other crops at your sites.

5.2 Soil health indicators summary

5.3 Recommendations

1. Re-survey in 2–3 years, especially Forsthaus and Alter Obstgarten — The most powerful use of this 2024 data is as a baseline. The recently-started sites (Forsthaus, Alter Obstgarten) should show measurable increases in biodiversity if the management approach is working. Forsthaus in particular will be the clearest test case: it starts from the lowest point, so any improvement will be most visible there. A3 Alter Obstgarten, where the cover crop already appears to have stimulated bacterial diversity, could show dramatic change in a relatively short time.

2. Dedicated AMF survey — planned — Arbuscular mycorrhizal fungi are arguably the most important group for both plant nutrition and soil aggregate stability in a no-input system, yet the standard ITS metabarcoding assay severely underdetects them. A targeted SSU rRNA survey for Glomeromycota will reveal whether the near-absence of AMF across most sites is real or an artefact. This is particularly relevant given the aggregation implications: AMF-produced glomalin can account for 20–30% of soil aggregate stability, and understanding how developed the AMF community is across your sites will give a much clearer picture of where your soils are structurally.

3. Monitor plant-parasitic nematodes at A2 Wiese — Pratylenchus neglectus and Helicotylenchus minzi are currently at low levels, typical for grassland. They could build up if susceptible crops are introduced. Worth watching in future surveys.

4. E1 Market Garden tells the story of your approach — With the highest diversity across all three kingdoms after the longest management period, E1 is living proof that organic, no-input management builds extraordinary soil biological richness over time. Documenting what has been done at this site — the specific practices, the timeline, the crop rotations — alongside this eDNA data creates a compelling evidence base.

5. Consider the A3 cover crop effect — The strikingly high bacterial uniqueness at A3 Alter Obstgarten (47 unique OTUs, more than any other site despite being newly managed) may be a direct result of the autumn 2023 cover crop mix. If this can be confirmed with a re-survey, it would demonstrate that cover crops are an effective tool for rapidly kickstarting bacterial diversity in newly managed soils — a valuable finding for other sites at the beginning of their management journey.

6. Differentiate loam vs. peat management expectations — The Spannweid and Haus Huegel peat/loam sites show characteristically different biological profiles (more Acidobacteriota, different AMF dynamics, different nutrient cycling patterns) than the loam sites. These differences are inherent to the soil type, not management failures. Setting separate benchmarks and expectations for peat vs. loam sites would make monitoring more meaningful.

This report was generated from NatureMetrics eDNA data files NM-QOC358 (Fungi), NM-SXP470 (Bacteria), and NM-JTS265 (Invertebrates), all from project PROJ05706. Analysis performed April 2026.