Henry W. G. Birt

Mycorrhizal fungi form essential symbiotic relationships with plant roots, facilitating nutrient exchange and promoting plant health. Understanding their interactions can benefit from advanced imaging techniques capable of visualising nutrient exchange and structural colonisation at subcellular resolutions across large sample sizes. This review explores novel imaging approaches that are revolutionising our understanding of root-mycorrhizal fungal symbioses. Several techniques can now visualise and characterise mycorrhizal fungi and associated root structures non-destructively and in three dimensions, for example, X-ray computed tomography (micro-CT), X-ray fluorescence (XRF) and X-ray absorption near edge structure (XANES) spectroscopy. Metabolic processes and nutrient exchange can be tracked through positron emission tomography (PET), fluorescent nanoparticles (FNPs), and the monitoring of electrical signalling. AI-powered image processing software is enabling high-throughput analysis of complex images generated from a range of sources. Mycorrhiza systems are also able to be tracked in-field at multiple scales: hyperspectral imaging can detect mycorrhizal associations at the kilometre scale, while portable MRI imagers can detect changes at the tissue scale. These converging technologies enable the direct, continuous measurement of structural and metabolic root-mycorrhizal fungi interactions, paving the way for a mechanistic understanding of these vital symbiotic partnerships and their impact on plant health and ecosystem functioning.

Societal Impact Statement: Our study tests how soil and plant biodiversity can enhance sustainability of crop production in Kenya. We tested whether mixtures of maize varieties performed better than monocultures and tested their response to arbuscular mycorrhizal fungi. Mycorrhizal responsiveness differed significantly by maize variety, and genetic mixtures outperformed monocultures. These findings demonstrate the benefits of using naturally‐occurring soil microorganisms in combination with genetic mixtures of crops to enhance food security. Summary: Plant genetic diversity is a key component of biodiversity but one that is often overlooked when considering the adoption of sustainable strategies to enhance crop production, such as inoculation with arbuscular mycorrhizal (AM) fungi. Here, we tested the hypothesis that the biomass of the most responsive maize genotypes to AM fungal inoculation when grown in genetic monoculture would be enhanced when grown in mixtures comprising a mix of genotypes (polyculture). In a first experiment, maize varieties were inoculated with Rhizophagus irregularis or Funneliformis mosseae or left uninoculated. We measured key growth parameters, AM fungal colonisation and phosphorus uptake and calculated mycorrhizal responsiveness. A second experiment evaluated the effect of crop genetic diversity on productivity by growing the four most responsive varieties as either monocultures or polyculture, with and without AM fungal inoculation. Mycorrhizal responsiveness differed significantly by maize variety, with some varieties demonstrating greater benefits from AM fungi. Polycultures outperformed monocultures in terms of AM fungal colonisation, biomass and phosphorus capture, which were driven by a combination of complementarity and facilitation effects resulting from biodiversity. Our findings demonstrate the critical role played by AM fungi in shaping crop genotype performance and the potential benefits of moving away from cropping systems that rely on genetic monocultures. The use of AM fungal inoculum in combination with targeted locally adapted crop genotype mixtures maximises plant nutrient efficiency and productivity and provides a sustainable approach to maize production in sub‐Saharan agroecological systems.

Summary: There is uncertainty in whether there is a common pattern of nestedness and modularity in plant–arbuscular mycorrhizal (AM) fungi associations, partly because of limitations arising from the use of null models that randomly rewire the observed connections to test for non‐random patterns in the network. Here, we overcome these limitations by generating null association matrices using maximum entropy network modelling, and specifically the bipartite binary configuration model (BiCM) with degree distributions as soft constraints. This was used to test the hypothesis that nestedness and modularity are prevalent in plant–AM fungi associations. In contrast to past findings, we found most plant–AM fungi associations were anti‐nested and modular. This pattern was almost universal, being consistent across habitat types, multiple spatial scales, and multiple levels of plant node aggregation, from communities and species to populations. Anti‐nestedness can easily emerge from modularity when network patterns are determined by the identity of the plant and AM fungal nodes. Our findings emphasize the need for experiments that test the factors that cause the observed network structure and how that structure determines the function and stability of plant–AM fungi association networks.

Many applications of network theory to plant-mycorrhizal associations have used a bipartite description, in which one set of nodes is the plants, and the other set is the fungi. Most applications have relied on null models from algorithms that randomly rewire the observed connections to test for non-random patterns in the network. We used existing plant-arbuscular mycorrhizal (AM) fungal datasets to apply a new, well validated generation of network models relaxing the very limiting assumptions of traditional null models. We focused on nestedness and modularity, which have been related to the functioning and stability of communities. Given the existent literature, we expected nestedness and modularity to be prevalent. We modelled plant-AM fungal associations using maximum entropy networks with a degree sequence, soft constraint to generate null distributions for nestedness and modularity. Most plant-AM fungal associations were anti-nested and modular. This pattern was consistent across habitat types and multiple spatial scales. Anti-nestedness can easily emerge from modularity when network patterns are determined by the identity of the plant and AM fungal nodes. Future studies will have to test how the observed patterns determine the ability of the associations to adapt to environmental changes.

Conventional dryland cropping systems are characterised by low crop diversity and frequent fallows. This has significant impacts on soil microbes that underpin soil function. Diversifying crop rotations can potentially counter these effects; however, limited data exists on the impacts of diversified crop rotations on soil microbes in drylands. Using phylogenetic marker gene sequencing, we characterised soil microbial diversity in conventional and diversified dryland crop rotations in subtropical Australia. This included winter and summer dominant rotations. Conventional systems were cereal-dominant with a crop-fallow rotation. Diversified systems included greater crop diversity, double crops, cover crops, and a multi-year ley pasture. In summer rotations with increased crop diversity and cover crops, bacterial and fungal richness increased, and distinct communities were formed compared to fallow land. Often, these community shifts were associated with greater soil organic carbon (SOC) and nitrogen. All winter rotations had distinct fungal communities and ley pasture resulted in greater fungal diversity compared to other rotations. No effects of the winter rotations were evident on bacterial communities. Our results show that diversification of dryland crop rotations leads to significant shifts in soil microbial communities in both winter and summer cropping systems. Both summer and winter rotations incorporating cover crops and ley pasture had greater soil respiration and nitrogen, indicating increases in soil fertility. These rotations may offer an alternative to conventional crop-fallow rotations to counter ongoing declines in soil health.

Many Australian native plants from regions with ancient, highly weathered soils have specialised adaptations for acquiring phosphorus (P) and can exhibit negative effects of excess P supply on growth and survival. Despite this, fertiliser (including P) is routinely applied in post-mining and other restoration schemes. In this study we investigated the effect of a range of applied P on the growth and tissue P concentrations for six woody species from the Great Western Woodlands (GWW) of Western Australia – a region that it not only biodiverse, but that has experienced significant levels of mining related activities. Our data from a pot-based experiment show that all six species exhibited greater growth with increased P application up to 15 mg kg sand–1. However, at P concentrations in excess of 15 mg kg–1, dry mass accumulation did not increase further for three of the species tested. For the other three species, dry mass accumulation declined as the P concentration increased above 15 mg kg–1. For all of the study species, root and shoot P concentrations increased as the concentration of applied P increased. The internal shoot P concentration, at which dry matter accumulation either plateaued or started to decline, was in the range 1.95 to 3.2 mg P g–1 dry matter. This was ~2–4 times the concentration found in natural vegetation. These data suggest that in a restoration context, there is a potential risk that, excess P application may decrease plant growth rates for some species. Consequently, the addition of fertiliser to restored sites may have unpredictable impacts on the plant community by directly reducing the growth of some species but increasing the growth of others. We suggest that careful consideration should be given to designing appropriate fertiliser regimes for land restoration schemes in ancient P deplete landscapes to avoid the risk that fertiliser addition has the unwanted outcome of decreasing growth and survival of the target native species and increasing the abundance of unwanted weeds or aggressive pioneer species.