Dataset DOI: 10.5061/dryad.qfttdz0vk

Description of the data and file structure

This dataset originates from a greenhouse experiment investigating how arbuscular mycorrhizal fungi (AMF) influence the phyllosphere microbial community of alfalfa and modulate plant defenses against pea aphid and pathogen attack.

The data include plant growth measurements, disease assessments, AMF colonization, enzyme activities, plant hormone levels, protease inhibitor levels, and microbial alpha diversity indices. Additional files contain pyllosphere microbiome rawdata for statistical analysis.

• Treatment groups:
  • NM: No AMF, no aphid, no pathogen
  • NMA: No AMF + aphid
  • NMP: No AMF + pathogen
  • NMAP: No AMF + aphid + pathogen
  • AM: AMF only
  • AMA: AMF + aphid
  • AMP: AMF + pathogen
  • AMAP: AMF + aphid + pathogen
• Plant growth and disease:
  • Shoot dry weight (g/pot): Aboveground biomass
  • Root dry weight (g/pot): Belowground biomass
  • Disease incidence (%): Percentage of diseased plants
  • Disease index: Composite index of disease severity
  • Mycorrhizal colonization (%): Proportion of roots colonized by AMF
  • Root length: Total root length (cm)
• Defense enzymes and hormones:
  • POD, SOD, PPO: Antioxidant and defense-related enzyme activities
  • JA: Jasmonic acid content
  • SA: Salicylic acid content
  • TI: Trypsin inhibitor activity
• Microbial community indices:
  • Fungi/Bacteria Chao1: Richness estimator
  • Fungi/Bacteria Shannon: Diversity index
  • Fungi/Bacteria Simpson: Dominance index
  • Fungi/Bacteria Pielou: Evenness index

Analysis workflow

1. Alpha diversity
   • Calculation of Chao1, Shannon, Simpson, Pielou, and Good’s coverage using vegan.
2. Beta diversity
   • Bray–Curtis distance matrix and PCoA visualization.
   • Group comparison via PERMANOVA (adonis2).
3. Differential abundance (DESeq2)
   • Normalization and statistical testing of OTU counts.
   • Identification of significantly enriched taxa.
4. Co-occurrence network
   • Spearman correlation among OTUs (psych, Hmisc).
   • Network construction with igraph (GraphML export).
5. Random forest analysis
   • Classification of treatments using OTU features.
   • Ranking of important OTUs (%IncMSE).
6. Structural Equation Modeling (SEM)
   • Piecewise SEM analysis linking AMF, aphids, pathogen, plant hormones, enzymes, and microbial diversity (piecewiseSEM).

The following R packages are required:

• vegan (alpha and beta diversity, PERMANOVA)
• ggplot2 (visualization)
• DESeq2 (differential abundance analysis)
• igraph (co-occurrence network)
• psych, Hmisc (correlation analysis)
• randomForest, rfPermute (random forest analysis)
• piecewiseSEM (structural equation modeling)

Files and variables

File: code.txt

Description: R script containing the code used for statistical analyses.

File: Functional_ecology-data.xlsx

Description: Excel spreadsheet containing the experimental dataset. Each row corresponds to an individual replicate, and each column represents a measured variable, including plant growth traits (shoot/root dry weight, root length, mycorrhizal colonization), disease indices (incidence, index), plant defense parameters (POD, SOD, PPO, JA, SA, TI), and microbial diversity indices (Chao1, Shannon, Simpson, Pielou for both fungi and bacteria).

Code/software

R (version ≥ 4.0.0)

Access information

Other publicly accessible locations of the data:

• NCBI Sequence Read Archive under accession number PRJNA1055717 and PRJNA1303415.