Unraveling Fungal-Bacterial Carbon Exchange in the Hyphosphere via Quantitative SIP and Cross-Domain Networks

Soil is a complex and densely populated ecosystem where fungi and bacteria coexist, compete, cooperate, or otherwise interact via nutrient exchanges. Although co-occurrence network methods and omics profiling have long offered hypotheses about microbial interactions, linking those statistical correlations to actual carbon (C) exchange in situ remains a key bottleneck. The recent study offers a methodological advance by combining ¹³C-labeling quantitative stable isotope probing (qSIP) with network inference to more precisely pinpoint which bacteria and fungi are actively exchanging plant-derived carbon in the hyphosphere (i.e. the soil volume influenced by fungal hyphae) ^[1].

Related Service from Alfa Chemistry

• Stable Isotope Probing (SIP) Service
  Alfa Chemistry provides SIP service with following three types:
  • Phospholipid derived fatty acid- stable isotope probing (PLFA-SIP)
  • Deoxyribonucleic acid-stable isotope probing (DNA-SIP)
  • Ribonucleic acid-stable isotope probing (RNA-SIP)
• Isotope-labeled Lipids

Methodology: ¹³C qSIP and Enriched Networks

Researchers conducted a ¹³CO₂ pulse-labeling experiment in California grassland. Ingrowth bags filled with quartz sand allowed fungal hyphae, but not roots, to colonize. DNA from colonized bags was fractionated by density gradients, and qSIP identified taxa with significant ¹³C incorporation. Only these enriched taxa were used to build cross-domain co-occurrence networks, ensuring that inferred links reflected active carbon flow rather than background presence.

Key Findings: Which Microbial Players Exchange Carbon?

Taxonomic composition of ¹³C-enriched taxa

• Among fungi, the enriched OTUs spanned Ascomycota and Basidiomycota, including saprotrophic and biotrophic species (e.g. Aspergillus, Coprinus, Aureobasidium, Ijuhya).
• Among bacteria, enriched ASVs came from phyla such as Actinobacteriota, Bacteroidota, Bdellovibrionota, Proteobacteria, and Verrucomicrobiota.
• A notable pattern: about 70% of ¹³C-enriched bacterial ASVs were motile, consistent with the idea that they may follow fungal "highways" (i.e. move along hyphae) into substrates.

Network structure and fungal–bacterial link enrichment

• The ¹³C-enriched network (only active taxa) was more densely connected: among 56 nodes, there were 914 links, of which 137 were fungus–bacterium links (≈ 15% of edges).
• In contrast, the total community network had 233 nodes and 589 links, but only 4 cross-domain links (< 1% of edges).
• The enriched network had a far higher average degree (≈ 32.6 edges per node) than the total network (≈ 5.05).

Specific cross-domain interaction hypotheses

Several strong positive associations (R² ≥ 0.65) emerged in the ¹³C-enriched network:

• A fungal OTU of Alternaria strongly linked with bacterial ASVs of Bacteriovorax, Mucilaginibacter, and Flavobacterium.
• Podospora (fungus) showed strong correlations with Peredibacter, Oligoflexus, and Pedobacter ASVs.
• Notably, predatory bacteria from Bdellovibrionota (e.g. Bacteriovorax, Peredibacter, Oligoflexus) were often connected positively to fungi, suggesting that these predatory bacteria might assimilate fungal-derived carbon (either via predation on bacteria that consumed fungal C or direct grazing).
• Some taxa links observed in the total community network (e.g. Chaetomium with Massilia) differ from those in the enriched network, highlighting those enriched-network inference hones in on active C exchange relationships.

Interpretation: trophic cascades and microbial food webs

• The authors propose a multi-step cascade: fungi assimilate plant-fixed ¹³C and release exudates or hyphal compounds → bacteria (often motile) consume those fungal-derived carbon sources → predatory bacteria consume bacteria (or necromass) that have engaged in fungal–bacterial exchange.
• Indeed, as fungal abundance increases, associated carbon-consuming bacteria increase; then abundances of predatory bacteria increase, and some other non-fungal-C-consuming bacteria decline — consistent with expectation under food-web dynamics.
• The enriched-network filtering strategy allowed these patterns to emerge more cleanly, reducing noise from taxa that were present but not participating in that carbon pathway.

Reference

[1] Slanzon G. S., Yuan M., Estera-Molina K., et al. Quantitative stable isotope probing (qSIP) and cross-domain networks reveal bacterial-fungal interactions in the hyphosphere[J]. Microbiome, 2025, 13(1): 109.

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Unraveling Fungal-Bacterial Carbon Exchange in the Hyphosphere via Quantitative SIP and Cross-Domain Networks

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