Abstract Beneficial plant–microbe associations (BMAs) offer a valuable opportunity to reduce dependence on synthetic inputs. However, traditional breeding has rarely targeted traits that enhance beneficial interactions, and conventional agricultural practices have often degraded soil health and microbial diversity. We present a framework that combines breeding for traits that facilitate BMA with soil management practices that enrich BMA. This approach integrates advanced breeding technologies with strategies for precise production and inoculation of microbes. Strengthening these complementary plant- and microbe-centered approaches and encouraging their adoption by farmers can foster more resilient and productive agricultural systems with reduced dependence on pesticides and fertilizers.

Societal Impact Statement Field inoculation with arbuscular mycorrhizal fungi (AMF) offers a promising route to improve sustainable agriculture and food production, yet results are often variable. We investigated the mycorrhizal growth response of maize and wheat varieties recommended for farmers and found substantial variation under greenhouse conditions. The mycorrhizal growth response varied from +7% to -12% for maize and from +6% to -25% for wheat and varied by breeding company. Selecting suitable varieties is crucial to fully harness AMF benefits. Field trials under diverse climatic and biotic conditions are required to support AMF application to ultimately promote sustainable agriculture and food security.Summary Arbuscular mycorrhizal fungi (AMF) enhance plant nutrient uptake and can reduce reliance on external inputs. Yet, it is unresolved whether plant breeding influences the ability to benefit from AMF and whether certain modern varieties benefit more from AMF associations compared to older varieties. We tested 15 maize (Zea mays) and 15 wheat (Triticum aestivum) varieties recommended for farmers, as well as three older wheat cultivars under greenhouse conditions. The plants were inoculated with an AMF (Rhizoglomus irregulare), and effects on mycorrhizal root colonization and mycorrhizal growth response (MGR) were assessed. To explore whether genetic background influences AMF responsiveness, we assessed whether response patterns varied across breeding companies and differed when comparing modern and old plant varieties. For maize, most modern varieties showed positive or neutral MGR, ranging from +7% to -12%. In contrast, the majority of modern wheat varieties exhibited neutral or negative responses, with MGR ranging from +6% to -25%. Crucially, modern varieties did not respond uniformly; maize varieties from the same breeder tended to cluster together in terms of MGR, suggesting a strong influence of genetic background. Three old wheat varieties generally showed more positive responsiveness than modern ones, but variation remained within both groups. Our study demonstrates that modern crop varieties differ markedly in their responsiveness to AMF, and that genetic background is a key determinant. These findings suggest that breeding programs and variety testing should integrate beneficial microbial interactions as selection criteria. Such an approach would maximize crop benefits from soil microorganisms and advance sustainable agriculture.

As the human population grows, so does the demand for higher agricultural yields. As a result, agricultural intensification practices are increasing while soil health is often declining. Integrating the benefits of microorganisms into agricultural management systems can reduce the need for external resource inputs. One particular group of plant symbionts that can help plants to acquire additional nutrients and promote plant growth are arbuscular mycorrhizal fungi (AMF). The application of AMF in agricultural practice has been hampered by the variability in the success of mycorrhizal inoculation and the lack of consistency in different fields. Here, we tested whether it is possible to predict mycorrhizal inoculation success based on soil health and productivity. We hypothesized higher inoculation success on fields with poor soil health because in such fields, mycorrhiza can improve nutrient uptake and biotic resistance to pathogens. We calculated a soil health index by aggregating six biotic and abiotic variables from 54 maize fields and tested its correlation with the mycorrhizal growth response (MGR). The MGR was linked to soil health and significantly higher in less healthy soils and less productive fields. This implies that soil inoculation with AMF has most potential in fields with poor soil health and low productivity. Based on these findings, we propose a soil health framework that highlights the potential benefits of AMF field inoculation.

The presence and distribution of mycorrhizal symbionts can influence plant distribution through specific host-mycorrhiza symbiosis interactions. However, generalist hosts also exist, such as dual-mycorrhizal plants that form symbiotic associations with both ectomycorrhizal fungi (EM) and arbuscular mycorrhizal fungi (AM). Little is known about the effect of dual mycorrhization status on the hosts' global distribution and acclimation to specific environments. This study investigates the potential advantage of dual associations of more than 400 woody genera spread at a global scale. We found that dual-host woody species occupy a broader geographical range and environmental niche space compared to those associating exclusively with either AM or EM. We show that the increased geographic range and expanded environmental niche space are independent of the phylogenetic architecture and evolutionary history of the woody genera. Our results highlight the advantage of generalist host-microbe symbioses between woody species and fungi to expand their range, and their potential role in colonising dry climates.

Tree species differ in their carbon (C) allocation strategies during environmental change. Disentangling species-specific strategies and contribution to the C balance of mixed forests requires observations at the individual tree level. We measured a complete set of C pools and fluxes at the tree level in five tree species, conifers and broadleaves, co-existing in a mature evergreen mixed Mediterranean forest. Our study period included a drought year followed by an above-average wet year, offering an opportunity to test the effect of water availability on tree C allocation. We found that in comparison to the wet year, C uptake was lower in the dry year, C use was the same, and allocation to belowground sinks was higher. Among the five major C sinks, respiration was the largest (ca. 60%), while root exudation (ca. 10%) and reproduction (ca. 2%) were those that increased the most in the dry year. Most trees relied on stored starch for maintaining a stable soluble sugars balance, but no significant differences were detected in aboveground storage between dry and wet years. The detailed tree-level analysis of nonstructural carbohydrates and δ13 C dynamics suggest interspecific differences in C allocation among fluxes and tissues, specifically in response to the varying water availability. Overall, our findings shed light on mixed forest physiological responses to drought, an increasing phenomenon under the ongoing climate change.

We used five mature Picea abies continuously labeled with 13C-depleted CO2 in a broadleaf-dominated Swiss forest to assess the spatial extent and lag time of carbon fluxes to ectomycorrhizal fungi differing in hyphal development and host association. We traced labeled carbon into ectomycorrhizal sporocarps collected for two seasons at different distances from labeled Picea. Picea-derived photosynthate reached conifer-specific sporocarps up to 6-12 m away and reached other sporocarps only 0-6 m away. At 0-6 m, genera of lesser hyphal development acquired more Picea-derived photosynthate than those of greater hyphal development, presumably from preferential fungal colonization of inner root zones by the former genera. Correlations of sporocarp 613C with daily solar radiation integrated for different periods indicated that carbon fluxes from Picea to sporocarps peaked 17-21 days after photosynthesis. Thus, these results provided rough estimates of the spatial extent and temporal lags of carbon transfer from Picea to ectomycorrhizal fungi.

Disentangling species-specific strategies and contribution to the C balance of mixed forests, requires observations at the individual tree-level. We measured a complete set of C pools and fluxes at the tree-level in five tree species, conifers and broadleaves, co-existing in a mature evergreen mixed Mediterranean forest. Field measurements of mature trees were conducted in 1 ha forest research plot at the Judean foothills in Israel (Yishi forest, 31º 43´N 34º 57´E, 320 m elevation) (Klein et al. 2013, Lapidot et al. 2019, Rog et al. 2021b). The climate is Mediterranean with annual precipitation of 510 mm falling between Sep and May, rendering May-Sep as a prolonged drought. Mean diurnal temperatures are 16.0°C in February and 25.3°C in August. During two consecutive hydrological years (Oct 2017 to Sep 2019) we assessed monthly C mass pool sizes and fluxes and up-scaled them to the compartment and tree levels. For this purpose, we combined direct and indirect measurements and data from previous onsite studies

The mutualistic interaction between trees and ectomycorrhizal fungi (EMF) can have a major effect on forest dynamics and specifically on seedling establishment. Here, we compared the EMF community composition associated with the roots of young saplings and mature trees of two co-habiting Pinaceae: Pinus halepensis and Cedrus deodara growing together in a post-fire forest plot, using fungal ITS metabarcoding. We found that the differences in the EMF community between the two sapling groups were mostly attributed to changes in the relative abundance of specific fungal species, with little species turnover. Specifically, Tomentella showed high abundance on pine roots, while Tuber, Russula and Sebacina were more common on the roots of cedars. The physical proximity to a specific host species was correlated with the EMF community composition of young saplings. Specifically, regardless of the sapling's own identity, the roots of saplings growing next to mature cedars had higher abundance of Tuber species, while Tomentella coerulea (H & ouml;hn. & Litsch), Russula densifolia (Secr. ex Gillet) and Tuber nitidum (Vittadini) dominated saplings next to mature pines. Cedar saplings' shoot structure was correlated with a specific EMF species. Overall, these results suggest that when germinating next to mature trees, the EMF community of saplings could be determined by extrinsic factors such as the small-scale distribution of mature trees in the forest.

The mycorrhizal symbiosis between fungi and plants is among the oldest, ubiquitous and most important interactions in terrestrial life on Earth. Carbon (C) transfer across a common mycorrhizal network (CMN) was demonstrated over half a century ago in the lab ( Reid & Woods, 1969), and later in the field ( Simard et al., 1997a). Recent years have seen ample progress in this research direction, including evidence for ecological significance of carbon transfer ( Klein et al., 2016). Furthermore, specific cases where the architecture of mycorrhizal networks have been mapped ( Beiler et al., 2015) and CMN-C transfer from mature trees to seedlings has been demonstrated ( Orrego, 2018) have suggested that trees in forests are more connected than once thought ( Simard, 2021). In a recent Perspective, Karst et al. (2023) offered a valuable critical review warning of over-interpretation and positive citation bias in CMN research. It concluded that while there is evidence for C movement among plants, the importance of CMNs remains unclear, as noted by others too ( Henriksson et al., 2023). Here we argue that while some of these claims are justified, factual evidence about belowground C transfer across CMNs is solid and accumulating.

The mycorrhizal symbiosis between fungi and plants is among the oldest, ubiquitous and most important interactions in terrestrial life on Earth. Carbon (C) transfer across a common mycorrhizal network (CMN) was demonstrated over half a century ago in the lab (Reid and Woods 1969), and later in the field (Simard et al. 1997). Recent years have seen ample progress in this research direction, including evidence for ecological significance of carbon transfer (Klein et al. 2016). Furthermore, specific cases where the architecture of mycorrhizal networks have been mapped (Beiler et al. 2015) and CMN-C transfer from mature trees to seedlings has been demonstrated (Orrego 2018) have suggested that trees in forests are more connected than once thought (Simard 2021). In a recent Perspective, Karst et al. (2023) offered a valuable critical review warning of over-interpretation and positive citation bias in CMN research. It concluded that while there is evidence for C movement among plants, the importance of CMNs remains unclear, as noted by others too (Henriksson et al. 2023). Here we argue that while some of these claims are justified, factual evidence about belowground C transfer across CMNs is solid and accumulating.

Survival and growth of woody species in the Mediterranean are mainly restricted by water availability. We tested the hypothesis that Mediterranean species acclimate their xylem vulnerability and osmotic potential along a precipitation gradient. We studied five predominant co-occurring Mediterranean species; Quercus calliprinos, Pistacia palaestina, Pistacia lentiscus, Rhamnus lycioides and Phillyrea latifolia, over two summers at three sites. The driest of the sites is the distribution edge for all the five species. We measured key hydraulic and osmotic traits related to drought resistance, including resistance to embolism (Psi(50)) and the seasonal dynamics of water and osmotic potentials. The leaf water potentials (Psi(l)) of all species declined significantly along the summer, reaching significantly lower Psi(l) at the end of summer in the drier sites. Surprisingly, we did not find plasticity along the drought gradient in Psi(50) or osmotic potentials. This resulted in much narrower hydraulic safety margins (HSMs) in the drier sites, where some species experienced significant embolism. Our analysis indicates that reduction in HSM to null values put Mediterranean species in embolism risk as they approach their hydraulic limit near the geographical dry edge of their distribution.

Climate change is expected to increase the frequency and severity of droughts in Mediterranean forests. Tree survival and storage of atmospheric CO2 in these forests depend on how individual tree species allocate their carbon (C). Here, we measured a complete set of above- and belowground C pools and fluxes in five coniferous and broadleaf species co-existing in a mature evergreen forest. Our study period included a drought year, followed by an above-average wet year, and the seasonal long dry period characterizes Mediterranean climate. To quantify the exact timing and spatial distribution of belowground C allocation, we additionally applied 13CO2 pulse labelling of one of the tree species (Quercus calliprinos). We found that during the dry versus wet year, photosynthetic C uptake decreased, C use in the C sinks remained unchanged and C allocation to belowground sinks increased. Among the five major C sinks, respiration was the main flux (~64%), while smaller fluxes like exudation (~9%) and reproduction (~1%) were those which increased the most in the dry year. To cope with seasonal drought, most trees relied on starch to maintain the C supply, but between years we found no significant differences in starch and sugars in aboveground tissues. Relative to the C storage dynamics, higher water use efficiency was found in conifers, while species-specific differences between dry and wet years were found among the broadleaves. Interestingly, in the wet season, after pulse labelling, C was allocated from the labeled leaves to the roots in two main time-lags: first after 3-5 days and second after 15-20 days. Labeled C reached fine roots at a distance of 0-6 m from the labeled tree. These detailed tree-level observations expose inter-annual and interspecific differences in C allocation among fluxes and tissues, specifically in response to varying water availability.

Summary Woody species employ various strategies to cope with drought stress. We investigated similarities and differences in response to chronic drought to understand resistance strategies in co-occurring Mediterranean species. We studied five predominant Mediterranean species; Quercus calliprinos, Pistacia palaestina, Pistacia lentiscus, Rhamnus lycioides , and Phillyrea latifolia over two summers at three sites with different aridities. We measured key hydraulic and osmotic traits related to drought resistance, including resistance to embolism (Ψ 50 ), carbon isotope signature (δ 13 C), pre-dawn (Ψ PD ) and mid-day (Ψ MD ) water potentials, and native (Ψ s ) and full turgor (П 0 ) osmotic potentials. Significant differences among species appeared in resistance to embolism. The species also showed differences in the water potential plastic response over the dry season. This interspecific variation increased at the end of the dry season and resulted in very narrow hydraulic safety margins (HSM). Consequently, predicted loss of hydraulic conductivity revealed species with significant native embolism. Two of the species also had seasonal changes in osmotic adjustment. Our detailed analysis indicates that co-existing Mediterranean woody species combine various drought resistance strategies to minimize mortality risk. However, all of them risk mortality as they approach their hydraulic limit near the dry margin of their distribution.

The mutualistic interaction between trees and ectomycorrhizal fungi (EMF) can have a major effect on forest dynamics and specifically in seedling establishment. Both intrinsic (i.e., identity of the sapling) and extrinsic (i.e., the identity of mature trees in the vicinity of the sapling) factors can affect the EMF community composition of young saplings. Here, we compared the EMF community composition associated with the roots of young saplings and mature trees of two co-habiting Pinaceae: Pinus halepensis and Cedrus deodara growing together in a planted forest plot, using fungal ITS metabarcoding. We found that the differences between the two sapling groups were mostly attributed to changes in the relative abundance of specific fungal species. Moreover, we found that physical proximity to a specific host species had a significant effect on the community composition of young saplings. However, while no significant differences in sapling size were apparent, the sapling shoot structure was affected by the identity of the nearest mature tree and its unique EMF community composition. Synthesis : These results suggest that the dynamics of the EMF community are greatly determined by extrinsic factors such as the small-scale distribution of mature trees in the forest, with possible cascading effects on the development of young trees.

Mixed forests are typically more productive and are faster to recover from drought compared to monospecific forests. Disentangling the contribution of each species to the overall success of the forest requires observations at the individual tree level. In this study, we measured a complete set of carbon (C) pools and fluxes at the tree-level in five tree species, two conifers and three broadleaf, co-existing in a mature evergreen mixed Mediterranean forest. Our study period included a drought year, followed by an above-average year. Across species, C sinks of 38-91 kg tree-1 year-1 were 16-32% larger than C source of 27-77 kg tree-1 year-1 in the dry year, with larger belowground C investment of the shallow-rooted species. Overall, respiration was the largest sink across species and years, accounting for 26-62% of all assimilated C, followed by growth (16%) and root exudation (19%). Non-structural carbohydrates accumulation was similar between the wet and the dry year. These detailed tree-level observations expose large interspecific differences in C allocation among fluxes and tissues and specifically in response to varying water availability. These insights become useful for forest management under ongoing change.

Abstract Background. The spatial extent and timing of carbon fluxes from mature trees to ectomycorrhizal fungi associated with different hosts is challenging to assess in natural forests but could provide insights into carbon dynamics of fungi differing in exploration capabilities. Methods. We analyzed carbon movement into ectomycorrhizal sporocarps at the Swiss Forest Free-Air CO2 Enrichment (FACE) site in 2010 and 2011 during continuous labeling of five mature Picea abies (L.) H. Karst. with 13C-depleted CO2. Sporocarps were collected 0-6 m, 6-12 m, 12-18 m, and > 18 m from 13C-labeled trees. We then used stepwise regressions on patterns of 13C:12C ratios (δ13C) in ectomycorrhizal sporocarps as a function of distance (zone), solar radiation, fungal genus, and association type. Results. 18%, 10%, 40%, and 32% of ectomycorrhizal sporocarps were associated with conifers, broad-leaved trees, both, or of unknown association, respectively. Conifer-associated sporocarps derived 58 ± 7%, 41 ± 10%, and 20 ± 7% of their carbon from labeled trees in the 0-6 m, 6-12 m and 12-18 m zones, respectively, whereas other sporocarps in the 0-6 m zone only derived 34 ± 4% of their carbon from labeled trees. Sporocarp δ13C correlated positively with solar radiation for the 16-19 days prior to sporocarp harvest. Conclusions. These patterns indicated that (1) carbon spread further through conifer-associated mycorrhizal networks than through other fungal networks, and (2) carbon accumulation, storage, and transfer dynamics for sporocarp formation can take several weeks. Mature conifers supplied carbon to mycorrhizal fungi up to ~15 m from host trees.

Mycorrhizal fungi can colonize multiple trees of a single or multiple taxa, facilitating bidirectional exchange of carbon between trees. Mycorrhiza-induced carbon transfer was shown in the forest, but it is unknown whether carbon is shared symmetrically among tree species, and if not, which tree species are better donors and which are better recipients. Here, we test this question by investigating carbon transfer dynamics among five Mediterranean tree species in a microcosm system, including both ectomycorrhizal (EM) and arbuscular (AM) plants. Trees were planted together in "community boxes" using natural soil from a mixed forest plot that serves as a habitat for all five tree species and their native mycorrhizal fungi. In each box, only the trees of a single species were pulse-labelled with 13 CO2 . We found that carbon transfer was asymmetric, with oak being a better donor, and pistacia and cypress better recipients. Shared mycorrhizal species may have facilitated carbon transfer, but their diversity did not affect the amount, nor timing, of the transfer. Overall, our findings in a microcosm system expose rich, but hidden, belowground interactions in a diverse population of trees and mycorrhizal fungi. The asymmetric carbon exchange among cohabiting tree species could potentially contribute to forest resilience in an uncertain future.

Mechanisms of host–microbe interactions and their direct impact on both parties have been extensively researched, however, much less is known on the effect of these interactions on the ecology of the host-community. Here we investigate tree-fungi mycorrhizal interactions, focusing on mycorrhizal-meditated resource sharing among trees, while examining the dynamics between specialist and generalist fungi and their implications on the forest ecology. Using genetic meta- barcoding, we identified the fungal community colonizing different trees in a mixed forest, and generated an extensive mapping connecting fungal sequences to their tree hosts. The mycorrhizal fungal community diverged between ectomycorrhizal and arbuscular host trees, but, unexpectedly, multiple ectomycorrhizal species colonized roots of non-ectomycorrhizal host trees. We complemented these findings by a novel computational framework, modeling competition between generalist and specialist mycorrhizal fungi, accounting for fungal-mediated resource sharing among neighboring trees. The analysis of the model revealed that generalist mycorrhizal networks may affect the entire tree community, and contribute to the maintenance of forest diversity in the long run. Furthermore, higher initial spatial mixing of trees can promote the evolution of generalist mycorrhizal species. These novel belowground interactions among trees and fungi may significantly impact forest biodiversity.### Competing Interest StatementThe authors have declared no competing interest.