Harnessing Endophytic Fungi for Sustainable Agriculture: Interactions with Soil Microbiome and Soil Health in Arable Ecosystems

Abstract

Sustainable food production for a growing population requires farming practices that reduce chemical inputs while maintaining soil as a living, renewable foundation for productivity. This review synthesizes current advances in understanding how endophytic fungi (EFs) interact with the soil microbiome and contribute to the physicochemical and biological dimensions of soil health in arable ecosystems. We examine evidence showing that EFs enhance plant nutrition through phosphate solubilization, siderophore-mediated micronutrient acquisition, and improved nitrogen use efficiency while also modulating plant hormones and stress-responsive pathways. EFs further increase crop resilience to drought, salinity, and heat; suppress pathogens; and influence key soil properties including aggregation, organic matter turnover, and microbial network stability. Recent integration of multi-omics, metabolomics, and community-level analyses has shifted the field from descriptive surveys toward mechanistic insight, revealing how EFs regulate nutrient cycling and remodel rhizosphere communities toward disease-suppressive and nutrient-efficient states. A central contribution of this review is the linkage of EF-mediated plant functions with soil microbiome dynamics and soil structural processes framed within a translational pipeline encompassing strain selection, formulation, delivery, and field scale monitoring. We also highlight current challenges, including context-dependent performance, competition with native microbiota, and formulation and deployment constraints that limit consistent outcomes under field conditions. By bridging microbial ecology with agronomy, this review positions EFs as biocontrol agents, biofertilizers, and ecosystem engineers with strong potential for resilient, low-input, and climate-adaptive cropping systems.

1. Introduction

Feeding the growing global population while reducing reliance on agrochemical input requires farming systems that treat soil as a living and resilient resource. Soil serves as a dynamic ecosystem that supports plants, animals, and humans and depends largely on the diversity and activity of microorganisms, which drive nutrient cycling, organic matter turnover, and soil structure formation. Therefore, microbial diversity is both a feature of healthy soil and a key driver linking farm management to productivity, environmental quality, and climate resilience [1,2]. Endophytic fungi (EFs), which belong to both clavicipitaceous and non-clavicipitaceous groups, are a diverse group of fungi that inhabit internal plant tissues for at least part of their life cycle without causing visible disease symptoms. Originally defined solely by their location within plant tissues, EFs are now recognized as functionally versatile symbionts whose interactions with host plants range from mutualistic to context-dependent latent pathogenicity [3,4]. Functionally, EFs contribute to plant nutrition, phytohormone modulation, abiotic stress tolerance, pathogen suppression, and metabolic reprogramming of host tissues, positioning them as key biological agents in sustainable agroecosystems [5,6].

Modern research reframes plants as meta-organisms whose growth and resilience rely on diverse microbial partners across the rhizosphere, rhizoplane, and endosphere. These microbial communities, which are shaped by plant traits and environmental factors, influence the nutrient uptake, immunity, and stress tolerance of plants. EFs are key players in this network that exchange carbon for nutrients, trigger defense responses, and modify root growth to support plant and soil health [7,8]. Beyond direct plant effects, EFs influence the composition and function of surrounding soil microbiomes, thereby shaping nutrient fluxes, microbial network stability, and ecosystem multifunctionality. Similarly, arbuscular mycorrhizal fungi (AMF) enhance nutrient acquisition, reduce nutrient losses, and improve soil structure, porosity, and water retention, which are critical functions that sustain soil fertility and ecosystem stability [9]. Because AMF are obligate root endophytes and frequently co-occur with non-mycorrhizal EFs, both groups are considered together in this review to capture their complementary and sometimes synergistic contributions to soil health and plant performance.

Soil structure and stability rely on aggregation, which resists breakdown and protects organic carbon. AMF strengthen these aggregates through hyphal networks and glomalin-related soil proteins (GRSPs), which improve water infiltration, aeration, and erosion control in arable soils [10]. Non-mycorrhizal EFs contribute more indirectly by reshaping rhizosphere microbial communities, altering carbon inputs, and producing extracellular polymeric substances and organic acids that promote microaggregate formation and nutrient retention [11,12]. In addition to their structural benefits, EFs also enhance plant stress tolerance. Studies have shown that they can transfer resistance to heat, salinity, and drought by modulating antioxidant activity and water use in plants, leading to improved survival and yield stability under climate stress [13]. Additionally, endophytes suppress pathogens through competition and by producing antimicrobial compounds and inducing systemic resistance. When integrated with crop rotation, organic amendments, and reduced chemical inputs, EF-mediated biocontrol offers a scalable pathway toward environmentally sustainable disease management [6,14,15].

Incorporating these mechanisms into real-world farming requires aligning the host plant, endophytes, management practices, and the environment. The structure and stability of the soil microbiome are highly sensitive to factors such as tillage, fertilization, moisture, and temperature. Therefore, endophyte inoculation or “banking” should be integrated into sustainable practices such as reduced tillage, residue retention, cover cropping, and organic amendments that create favorable conditions for fungal growth and persistence. Recent studies have emphasized the design of cropping systems that support and stabilize diverse beneficial microbial communities rather than depending on single-strain inoculants alone [16].

Despite growing recognition of their potential, the widespread application of EFs in agriculture remains constrained by several challenges. EF performance is often context-dependent, varying with soil type, resident microbiota, host genotype, and climate, underscoring the need for ecological matching and site-adapted inoculation strategies. Mechanistic understanding is also incomplete as it remains difficult to disentangle direct EF effects from those mediated by associated bacteria or soil physicochemical factors. Addressing these gaps will require integrative multi-omics approaches, long-term field trials, and the use of sensitive soil health indicators, such as aggregate stability, particulate organic carbon, and nutrient use efficiency. The novelty of this review lies in its integrative perspective that explicitly links EF-mediated plant functions with soil microbiome dynamics and soil structural processes in arable systems. Unlike previous reviews that focus primarily on plant growth or stress tolerance, this work synthesizes evidence across ecological, physiological, and soil biophysical scales and proposes a functional framework connecting EF traits to ecosystem-level outcomes and field-scale management strategies. By integrating recent findings from the fields of microbial ecology, plant physiology, and soil science, this review highlights how EFs can enhance nutrient cycling, stress tolerance, and soil structural stability, ultimately supporting resilient and eco-friendly crop systems.

2. Materials and Methods

This review was conducted using a structured literature search across major academic databases, including Web of Science, Scopus, PubMed, and Google Scholar, to capture international studies on endophytic fungi (EFs), soil microbiomes, nutrient cycling, and soil health in arable systems. Publications from 2000 to March 2025 (except for a foundational historical reference of EFs in 1866) were identified using Boolean combinations of keywords including “endophytic fungi”, “soil microbiome”, “rhizosphere”, “nutrient cycling”, “arable land”, and “sustainable agriculture”. Peer-reviewed research articles and reviews were prioritized, with the gray literature consulted selectively to support applied and management perspectives. Studies were screened for relevance to plant–soil–microbe interactions and ecosystem functions, and the evidence was qualitatively synthesized to identify recurring mechanisms, emerging trends, and knowledge gaps across diverse agroclimatic regions.

3. Fungal Endophytes: Definition, Categorization, Distribution, and Ecological Niches

3.1. Definition

The term “endophyte” (from the Greek endon meaning “within” and phyton meaning “plant”) was first introduced by De Bary [17] to describe organisms living inside plant tissues as opposed to epiphytes that exist on plant surfaces. Initially, it referred only to location rather than function [18,19]. Later, Petrini [3] (see also Petrini 1986) broadened the concept, defining endophytes as organisms that inhabit internal plant tissues without causing visible disease. This inclusive view emphasized the symbiotic and symptom-free nature of these associations and laid the foundation for the modern understanding of endophytes [3].

Carroll (1988) proposed a narrower definition of endophytes, focusing mainly on foliar fungi that inhabit healthy leaves while excluding known pathogens and mycorrhizal mutualists [20]. Although this clarified the study of foliar endophytes, it restricted the broader perspective offered by Petrini. Later, Wilson [21] refined the concept, suggesting that endophytes should simply be defined as organisms living inside plant tissues without producing symptoms, a view that brought much needed precision. The concept evolved further with Schulz & Boyle [4], who introduced the “endophytic continuum”, recognizing that fungal interactions can shift from mutualistic to pathogenic depending on environmental and host conditions. Building on this, Rodriguez et al. [5] highlighted the ecological functions of endophytes, such as stress tolerance and growth promotion, and advanced classification schemes based on life history and transmission modes, integrating both functional and ecological perspectives into modern definitions.

With advances in microbiome research, Hardoim et al. [22] redefined endophytes within the plant holobiont concept as internal members of plant microbiota that live within the plant without causing disease. Their interactions range from mutualistic to neutral, depending on environmental and host conditions. Building on this view, the modern consensus defines fungal endophytes as fungi that asymptomatically inhabit healthy plant tissues, with relationships that shift along a context-dependent continuum. They are now characterized not only by their location within the plant but also by their ecological roles, life cycles, and transmission modes.

Importantly, this broad definition encompasses classical (non-mycorrhizal) endophytic fungi and arbuscular mycorrhizal fungi (AMF), although these groups differ in key biological traits. Classical endophytes are taxonomically diverse, include facultative symbionts, and often colonize roots, stems, leaves, or reproductive tissues without forming specialized exchange structures. In contrast, AMF are obligate biotrophs that form highly structured symbioses within root cortical cells and are traditionally treated as a distinct functional group due to their specialized role in phosphorus acquisition and carbon exchange. In this review, both groups are considered together because they converge functionally at the root–soil interface and exert complementary influences on rhizosphere processes and soil health. AMF contribute directly to soil aggregation, nutrient capture, and hyphal network formation, while non-mycorrhizal endophytic fungi influence soil structure and nutrient cycling more indirectly by reshaping rhizosphere microbial communities, modifying root exudation patterns, and altering carbon inputs to the soil. In arable systems, AMF and other endophytic fungi frequently co-occur within roots and interact with overlapping microbial networks, jointly affecting nutrient fluxes, pathogen suppression, and soil physical stability.

Accordingly, this review adopts an integrative perspective that treats endophytic fungi, including AMF, as functionally connected components of the plant–soil continuum. By revisiting the definition of EFs through the lens of soil architecture and rhizosphere microbiome restructuring, we aim to highlight how distinct endophytic strategies collectively contribute to soil health and ecosystem resilience in arable agroecosystems.

3.2. Classification

EFs can be classified based on several criteria, as shown in

Table 1. Summary of major EF classes based on taxonomy, transmission mode, host tissue preference and ecological function. The table highlights functional distinctions among clavicipitaceous, non-clavicipitaceous, and mycorrhiza-like endophytes and their roles in nutrient acquisition, stress tolerance and soil health.

Class of Endophytes	Representative Fungal Genera	Common Host/Colonized Tissues	Transmission Mode	Ecological/Functional Roles	Key References
Class 1 Clavicipitaceous (C-endophytes)	Epichloë, Neotyphodium	Grasses (Poaceae); mainly leaf sheaths, stems, and seeds	Vertical (seed-borne); occasionally mixed	Systemic colonization, drought tolerance, herbivore resistance via alkaloid production, and stress resilience	[5,25,26]
Class 2 Non-clavicipitaceous root-associated endophytes	Trichoderma, Clonostachys, Fusarium, Penicillium	Roots of cereals, legumes, and vegetables	Horizontal (soil- and rhizosphere-acquired)	Nutrient mobilization, abiotic stress tolerance, induced systemic resistance (ISR) induction, and VOC-mediated pathogen suppression	[5,27,28]
Class 3 Non-clavicipitaceous foliar/stem endophytes	Aspergillus, Alternaria, Colletotrichum, Cladosporium	Leaves and stems of various crops (e.g., maize, rice, and wheat)	Horizontal (airborne or phyllosphere sources)	VOCs and secondary metabolites for pathogen defense, stress mitigation, and sometimes latent pathogens	[22,29,30]
Class 4 Endophytic mycorrhiza-like (Dark septate fungi, AMF-related)	Phialocephala, Cadophora, Rhizophagus, Glomus	Root cortical tissues of cereals, pulses, and oilseeds	Horizontal (soil propagules and hyphal contact)	Nutrient acquisition (especially N and P), improved water uptake, modulation of rhizosphere microbiome, and soil aggregation	[5,31,32]
Seed-associated and reproductive endophytes	Epichloë, Trichoderma, Fusarium spp.	Seeds and reproductive organs of crops	Vertical or mixed (seed + environment)	Early plant protection, enhanced germination, seedling vigor, and microbiome assembly	[33,34]

: (a) Taxonomically, most belong to the phyla Ascomycota and Basidiomycota, with genera such as Trichoderma, Clonostachys, Fusarium, Aspergillus, and Penicillium, which are commonly found in agricultural systems [5,23]. (b) Functionally, they include clavicipitaceous endophytes (C-endophytes), which are mainly found in grasses and vertically transmitted through seeds, and non-clavicipitaceous endophytes (NC-endophytes), which are found in diverse plants and horizontally acquired from the environment. (c) Ecologically, EFs are grouped as root, foliar, or reproductive endophytes enhancing nutrient uptake, stress tolerance, and pathogen defense [24]. (d) Transmission occurs either vertically (seed-borne, e.g., Neotyphodium) or horizontally (from soil, litter, or air), with the latter being the most common in crop systems.

3.3. Global Distribution and Ecological Niches

EFs occur in all major biogeographic regions and display distinct distribution patterns, reflecting both plant diversity and environmental gradients. Tropical regions are particularly rich in EF diversity, dominated largely by non-clavicipitaceous taxa associated with forest trees and perennial crops. Temperate and arid regions tend to have lower taxonomic diversity but often exhibit strong functional specialization; for example, endophytes adapted to extreme environments or grass-dominated ecosystems [29,35]. In arable landscapes, agricultural practices such as tillage intensity, residue retention, crop rotation, and fertilizer inputs shape the EF community structure and abundance, often selecting taxa that can persist under disturbance regimes. Importantly, EF colonization is not random as shown in Figure 1; many strains exhibit tissue-specific preferences (e.g., roots vs. leaves vs. reproductive organs) and occupy distinct ecological niches that enable long-term persistence and coevolution with host plants [36,37].

This niche differentiation has direct implications for application in crop–soil systems. Root-associated endophytes, which dominate in arable soils, are particularly relevant targets for seed coatings, in-furrow inoculation, or soil amendments aimed at improving nutrient acquisition, pathogen suppression, and soil aggregation. In contrast, foliar or reproductive endophytes may be more effectively deployed through seed-borne transmission or targeted foliar applications to enhance early plant vigor or stress tolerance. Management practices that reduce disturbance, such as conservation tillage, residue retention, and diversified rotations, can further promote the establishment and persistence of beneficial EF guilds by maintaining favorable soil habitats and inoculum reservoirs [38,39].

Overall, the structured distribution and niche specialization of EFs underscore their ecological integration within plant holobionts and highlight opportunities for strategic, context-specific deployment in arable systems [22]. Recognizing how EF niches align with crop traits, soil conditions, and management practices is essential for translating their ecological potential into reliable agronomic benefits.

4. Diversity and Ecological Roles of EFs in Arable Land

EFs are widespread across crop species and can be detected in most plant compartments, yet their apparent diversity in arable systems is often underestimated when only culture-dependent methods are used. High-throughput sequencing (HTS) approaches, particularly ITS amplicon profiling and shotgun metagenomics, have consistently revealed a much larger “hidden” endophytic pool, including taxa that are rare, unculturable, or transient colonizers. In agricultural landscapes, EF communities are shaped by a combination of host genotype, soil physicochemical conditions (pH, texture, organic carbon, and nutrient status), and climate and management intensity with strong spatial (field-to-field and within-field) and seasonal turnover patterns. These dynamics align with broader soil biodiversity syntheses emphasizing that agricultural practices can simplify or restructure microbial diversity and functions, thereby altering nutrient cycling capacity and resilience of agroecosystems [40].

4.1. Diversity Patterns and Drivers in Arable Systems

EF assemblages in cropland are taxonomically dominated by Ascomycota, commonly including genera such as Fusarium, Penicillium, Trichoderma, Cladosporium, and Alternaria, with contributions from Basidiomycota and early diverging fungal lineages depending on soil and crop type [41]. Importantly, diversity patterns in arable land are not random: EF communities show tissue filtering (root vs. stem vs. leaf endosphere), host selection, and strong sensitivity to disturbances that modify residue inputs and soil habitat structure. Long-term field studies using HTS show that tillage intensity, fertilization regime, and crop rotation history measurably alter fungal community structure [42,43,44] and guild composition (including endophytic-associated taxa), supporting the idea that arable EF diversity is partly “management-selected”. Management-driven structuring is particularly clear in long-term rotation and tillage experiments. For example, ITS-based community profiling in replicated crop rotation trials has shown that conservation tillage vs. conventional tillage, nitrogen inputs, and preceding crops shift fungal community composition and functional guild distribution over time. Such findings reinforce that EF pools available for colonization in crops are not only determined by plant genetics but also by the “soil fungal reservoir” shaped by farming practices [45].

4.2. Ecological Roles in Arable Land: From Plant Traits to Soil Functions

While EFs are often discussed in relation to plant growth promotion, their ecological roles in cropland extend to soil processes and microbiome organization. In arable systems, EFs can contribute to nutrient cycling through several pathways: (i) mobilization of phosphorus and micronutrients via secretion of organic acids, phosphatases, and chelators; (ii) siderophore-mediated iron mobilization that can indirectly influence competitive outcomes among rhizosphere microbes; and (iii) facilitation of nutrient acquisition through interaction networks with bacteria and other fungi in the rhizosphere. A growing body of experimental and synthesis literature supports EF contributions to phosphate solubilization and nutrient mobilization mechanisms relevant to crop nutrition and fertilizer use efficiency [46]. EFs also influence disease regulation and community stability in arable soils. Many EF taxa produce bioactive secondary metabolites and/or enzymes that inhibit pathogens, compete for space and nutrients, and prime host defenses. These interactions can reduce pathogen establishment and help maintain a more functionally balanced microbiome, particularly when integrated with diversified crop rotations and reduced disturbance practices. Reviews focused on fungal functional groups in annual cropping systems emphasize that these ecological roles are strongly context-dependent and influenced by soil organic matter, residue management, and the crop sequence that regulates inoculum carryover and habitat continuity [47].

4.3. How Arable Management Reshapes EF Diversity and Function

EF diversity in arable land is especially responsive to management because croplands experience recurring disturbance (tillage, planting, and harvest), rapid nutrient pulses (fertilizers), and crop-specific selection pressures. Evidence from long-term experimental sites shows that tillage regime and fertilization intensity are strong predictors of fungal community shifts, which can cascade into changes in organic matter dynamics and soil structure formation through altered fungal biomass, hyphal networks, and decomposition pathways [45]. Crop rotation is another major driver (rotation alters residue chemistry, root exudate profiles, and soil habitat continuity) which collectively restructures fungal reservoirs and may influence endophyte recruitment into crop tissues. Global syntheses and meta-analyses demonstrate that rotation generally supports higher microbial diversity and can improve system stability though responses that can differ across microbial groups and environmental contexts [48].

4.4. Methodological Advances and Functional Validation

The study of EF diversity and ecological function in agricultural systems has advanced rapidly through ITS rRNA gene amplicon sequencing, metagenomics, and multi-omics integration, enabling deeper resolution of community composition and functional potential. However, because sequence-based surveys alone cannot confirm ecological function, complementary culture-based isolation remains essential for trait screening (e.g., nutrient solubilization, stress tolerance, and antagonism), mechanism testing, and eventual formulation as bioinoculants. Integrating HTS discovery with functional assays provides a translational pipeline from “who is there” to “what they do,” supporting the identification of EF candidates that are both ecologically persistent in arable soils and agronomically relevant [49].

5. Mechanisms of Plant Growth Promotion by EFs

EFs promote plant growth through a combination of direct physiological modulation of the host and indirect restructuring of the rhizosphere environment leading to improved nutrient acquisition, root development and stress-buffering capacity, as illustrated in Figure 2. The diverse mechanistic pathways underlying EF-mediated growth promotion are summarized in

Table 2. Summary of EF’s mechanisms in plant growth promotion. This table illustrate the compilation of representative EF taxa, host crops, dominant growth promoting mechanisms and observed agronomic benefits. The table links specific functional traits (e.g., hormone production, phosphate solubilization, antagonism) with plant performance outcomes.

Endophytic Fungus	Host Plant/Crop	Mechanism of Plant Growth Promotion	Observed Benefits	References
Trichoderma harzianum	Tomato, avocado, maize, and rice	Production of IAA, phosphate solubilization, siderophore release, and antagonism against soil pathogens	Enhanced germination, root growth, and disease resistance	[50,51,52]
Piriformospora indica (Serendipita indica)	Barley, maize, and rice	Symbiotic association enhancing nutrient uptake (P, N, and Zn), hormone modulation, and stress tolerance	Improved biomass, drought, and salt tolerance	[53,54]
Fusarium oxysporum (non-pathogenic strains)	Tomato and cucumber	Secretion of growth hormones (IAA and gibberellins), enhanced antioxidant enzyme activity, and induced systemic resistance	Increased yield and tolerance to abiotic stress	[55,56]
Penicillium chrysogenum	Wheat, lettuce, cabbage, croccoli, and orchid	Phosphate solubilization, siderophore production, and ACC deaminase activity	Improved nutrient uptake and shoot biomass	[57,58]
Aspergillus niger	Mung bean, cassava, and forage grass	Organic acid secretion, phosphate solubilization, and enzyme production	Enhanced P availability and root elongation	[59,60]
Chaetomium globosum	Maize, cotton, and wheat	Antagonistic metabolites, cellulase/chitinase production, and induced systemic resistance	Protection against root pathogens and improved vigor	[61,62]
Clonostachys rosea	Tomato, wheat, and soybean	Mycoparasitism, production of antifungal compounds, and phytohormones	Disease suppression and growth enhancement	[63,64]
Colletotrichum truncatum	Pepper, snakeweed, and cucumber	Modulation of host metabolism and nitrogen uptake and IAA synthesis	Enhanced photosynthesis and biomass accumulation	[65,66]
Talaromyces pinophilus	Wheat, rice, and maize	Production of siderophores, phosphate solubilization, and secretion of stress-protective enzymes	Improved nutrient efficiency and drought resilience	[67,68]
Purpureocillium lilacinum	Soybean, cotton, and cucumber	Root colonization, phytohormone production, and nematode antagonism	Improved root growth, yield, and nematode resistance	[69,70]
Beauveria bassiana	Maize and corn	Endophytic colonization, secondary metabolite production, insect deterrence, and growth promotion	Enhanced biomass and biocontrol potential	[71,72,73]
Alternaria alternata	Wheat, rice, and tomato	IAA synthesis, phosphate solubilization, and antioxidant modulation	Improved germination, nutrient uptake, and stress tolerance	[74,75]

, encompassing nutrient mobilization, phytohormone regulation, pathogen suppression and microbiome recruitment.

Quantitative evidence supporting these effects across crop systems is described in Section 8.3, which compiles reported effect sizes for plant biomass, nutrient-use efficiency and soil biological responses. These growth promoting effects are often most pronounced under low-input or stress-prone conditions, where microbial functions compensate for limited nutrient availability or constrained plant metabolism. As EF-host interactions are context dependent, plant growth promotion typically reflects the integration of multiple mechanisms rather than a single dominant pathway [5,22,76].

5.1. Hormone Modulation and Root System Reprogramming

Many EFs influence plant growth by producing or regulating phytohormones, particularly IAA, gibberellins (GAs), and cytokinins, which reshape root architecture by stimulating lateral root formation, root hair development, and overall root biomass (Figure 2). Enhanced root systems improve soil exploration and nutrient uptake efficiency. Several EF strains also modulate ABA signaling and stomatal behavior, which can improve growth under fluctuating water availability by balancing carbon assimilation and transpiration. These hormone-linked effects are widely reported across crop systems, including maize, wheat, rice, and horticultural crops [77,78,79].

5.2. Ethylene Stress Reduction via ACC Deaminase and Stress Signaling Dampening

Stress conditions often elevate ethylene, which suppresses root elongation and reduces growth. EF-associated mechanisms that lower ethylene sensitivity, including ACC deaminase activity (reported more frequently for bacteria but also for some fungal associated endosymbionts and EF consortia), can reduce stress-induced growth inhibition and maintain root development. In parallel, many EFs enhance antioxidant and osmoprotectant systems that limit oxidative damage, indirectly stabilizing growth and enabling continued nutrient uptake under stress. Even when ACC deaminase is not the primary EF trait, EF-driven reductions in stress ethylene responses and ROS damage are repeatedly observed in plant–endophyte studies (Figure 2) [13,78,80].

5.3. Nutrient Mobilization: P Solubilization, Micronutrient Acquisition, and N-Use Efficiency

A major growth promotion pathway is nutrient mobilization, especially in nutrient-limited soils. EFs can solubilize inorganic phosphates through the secretion of organic acids and phosphatases, increasing plant available P and improving P use efficiency, as highlighted in Figure 2. They also enhance acquisition of micronutrients, such as Fe and Zn, often through chelation-related processes (including siderophore-like activity in fungi and cooperative interactions with siderophore-producing bacteria). Additionally, EFs can improve nitrogen use efficiency by enhancing root uptake capacity and coordinating microbial transformations in the rhizosphere. These mechanisms are directly linked to reduced fertilizer dependence, which is a core goal in sustainable arable agriculture [46,76,78,81].

5.4. Indirect Growth Promotion via Microbiome Recruitment and Rhizosphere Engineering

As presented in Figure 2, EFs alter the quantity and composition of root exudates (sugars, amino acids, organic acids, and phenolics), which can recruit beneficial bacteria and fungi, suppress opportunistic pathogens, and shift the rhizosphere networks toward higher nutrient cycling capacity. This indirect mechanism is increasingly supported by amplicon sequencing and metagenomic studies, showing EF-driven changes in community composition and functional genes related to N and P cycling. EF-mediated recruitment of beneficial microbiota can also enhance mycorrhizal establishment, particularly when EFs and AMF occupy complementary niches in the root system. These “microbiome engineering” effects help explain why EF inoculation can produce sustained growth benefits beyond short-lived hormone effects [7,8,82,83,84].

5.5. Bioactive Metabolites, VOCs and Immune Priming That Support Growth

Some EFs promote growth by releasing secondary metabolites and volatile organic compounds (VOCs) that modulate plant signaling, inhibit pathogens, or influence neighboring microbes. VOC-mediated effects include stimulation of root growth, modulation of defense pathways, and suppression of pathogen activity at a distance. At the same time, EF-induced immune priming (ISR-like responses) can lower disease burden and reduce the resource cost of chronic defense activation, indirectly improving growth and yield stability. Although VOC outcomes can be concentration dependent and variable across soils, controlled studies repeatedly demonstrate their potential contribution to plant vigor and rhizosphere balance (Figure 2) [6,85,86].

5.6. Evidence from Field Trials and Translational Constraints

Field and “field-adjacent” trials show that EF inoculation can increase biomass and yield particularly under drought, salinity, reduced fertilizer inputs, or pathogen pressure. However, performance is inconsistent across environments due to differences in soil microbiomes, climate, inoculum survival, and host genotype compatibility. The gray literature and international assessments highlight that successful translation depends on formulation stability, delivery methods (seed coating, granules, and liquid inoculants) and post-deployment monitoring of colonization and function over seasons. Therefore, growth promotion mechanisms should be discussed alongside practical deployment conditions that determine whether these mechanisms operate in real soils [76,87,88,89].

6. EFs in Abiotic Stress Tolerance

Plants are continuously exposed to a range of abiotic stressors, including drought, heat, salinity, and nutrient deficiency, all of which negatively affect their growth and productivity, and food security. One promising avenue for enhancing plant resilience to such stressors is the use of EFs. As depicted in Figure 3, EFs play an important role in plants coping with abiotic and biotic stresses.

EFs have been widely reported to enhance plant tolerance to abiotic stressors via multiple interconnected mechanisms. Endophytes confer tolerance to drought, salinity, extreme temperature, and heavy metals through osmolyte accumulation, antioxidant defense activation, and hormonal modulation [88,90,91]. They also sequester or immobilize toxic metals, reducing phytotoxicity [51,92,93,94]. They promote growth by modulating phytohormones such as IAA, GAs, and cytokinins, and by producing ACC deaminase to lower ethylene levels during stressful conditions [95,96]. Additionally, they assist in nutrient solubilization and uptake, particularly in nutrient-limited soils, through the secretion of organic acids and siderophores and stimulation of soil enzymatic activities [97,98,99,100].

Importantly, EFs help plants develop antioxidant defenses by enhancing activities of enzymes such as catalase, superoxide dismutase, and peroxidase, thereby reducing oxidative stress caused by reactive oxygen species during drought or salinity stress [101]. They also assist in osmotic adjustment through the accumulation of compatible solutes, such as proline and trehalose, helping maintain water balance and cell turgor.

Moreover, EFs influence root architecture and exudation patterns, which not only improve nutrient uptake but also reshape the rhizosphere microbiome by recruiting beneficial microbes and enhancing mycorrhizal colonization. These microbiome shifts contribute to abiotic stress tolerance by improving access to water and nutrients under limiting conditions, enhancing soil aggregation and moisture retention, and stabilizing nutrient cycling during stress episodes. For example, enhanced mycorrhizal networks can increase the effective root surface area and facilitate phosphorus and water uptake during drought or salinity stress, while EF-driven recruitment of beneficial bacteria can support nitrogen mobilization and osmotic adjustment [102,103]. Omics-based studies support this indirect mechanism by showing that EF colonization restructures rhizosphere communities toward beneficial taxa and away from stress-exacerbating pathogens, thereby strengthening overall plant resilience and ecosystem stability [49,104,105].

Field and greenhouse experiments further supported these findings. For example, in barley and maize, EF inoculation increased biomass, root length, chlorophyll content, and stress tolerance under drought and saline conditions [106,107]. Omics technologies such as metagenomics, metabolomics, and stable isotope probing are increasingly used to link endophyte activity to nutrient flux, microbial diversity, and plant health [108,109,110].

Table 3. Quantitative effects of endophytic fungi on plant growth, nutrient acquisition, soil microbiome, and soil physical properties in arable ecosystems (synthesis of representative studies and meta-analyses).

Functional Category	Response Variable	Reported Effect Size (Range)	System/Context	Key Notes	Representative References
Plant growth	Shoot or total biomass	+10–35%	Cereals, legumes, and horticultural crops	Stronger under nutrient limitation and abiotic stress	[6,27,78]
	Root biomass/root length	+15–50%	Root endophytes and AMF	Increased absorptive surface area	[5,38]
Nutrient acquisition	Phosphorus uptake/P-use efficiency	+15–60%	AMF and non-AMF EFs	Organic acids + hyphal foraging	[10,81]
	Nitrogen use efficiency	+10–40%	EF-primed rhizosphere	Microbiome-mediated mineralization	[111,112]
	Iron availability	+20–70% (relative uptake)	Siderophore-mediated	Enhanced micronutrient capture	[46]
Soil microbiome	Microbial biomass C (MBC)	+10–45%	Conservation and low-input systems	Carbon-driven assembly	[36,112]
	Enzyme activities (C, N, and P cycling)	+15–50%	EF-inoculated soils	Enhanced turnover rates	[113,114]
	Pathogen abundance	−20–70%	IPM and EF-based systems	Competitive exclusion + antibiosis	[115,116]
Soil structure	Aggregate stability (WSA)	+5–30%	AMF-rich arable soils	GRSP-driven stabilization	[10,97]
	Soil organic carbon	+5–25% (relative increase)	Long-term EF presence	Protected within aggregates	[117,118]
Stress resilience	Drought/salinity tolerance indices	+15–45%	Abiotic stress conditions	Improved water and nutrient uptake	[102,103]

shows the quantitative effects of EFs in stress tolerance.

7. EFs in Biocontrol of Plant Pathogens

Over the past two decades, EFs have moved from “promising isolates” to credible components of integrated pest management (IPM) because they can suppress diverse pathogens (fungi, bacteria, and oomycetes) and some plant parasitic nematodes while simultaneously improving plant vigor and stress tolerance. This multi functionality contrasts with single-target agrochemicals and helps explain why EF-based solutions are increasingly positioned as sustainable inputs in arable and horticultural systems [14,22,99]. Biocontrol by EFs can be organized into four interacting “layers” that operate at different spatial scales (pathogen surface, rhizosphere, root tissues, and whole-plant systemic responses). In practice, most successful EF strains deploy multiple layers at once [111,113,114]. The mechanism of EFs in plant defense is illustrated in a diagram in Figure 4.

7.1. Direct Antagonism of Pathogens (Contact-Dependent)

Mycoparasitism and lytic enzymes: Many EFs directly attack fungal pathogens through hyphal contact, coiling, penetration, and secretion of cell-wall-degrading enzymes (chitinases, β-1,3-glucanases, and proteases). Trichoderma spp. are the best characterized example with coordinated mycoparasitism and antibiosis during Trichoderma–plant–pathogen interactions [119]. Clonostachys rosea is likewise a strong mycoparasite with documented suppression of multiple fungal diseases and additional activity against nematodes in some systems [120]. Antimicrobial secondary metabolites: EFs can inhibit pathogen growth or sporulation via diffusible metabolites (e.g., peptaibols and other specialized metabolites in Trichoderma). These compounds often act synergistically with enzymes, increasing membrane permeability and accelerating pathogen collapse [27,53,119,121,122].

7.2. Volatile-Mediated Inhibition and “Distance Effects” (Contact-Independent)

Fungal VOCs act as biofumigant-like signals. Many EFs emit VOC blends that suppress pathogens without physical contact and can also modulate plant signaling and root development. VOC effects are frequently dose dependent and environment dependent, which explains variable outcomes across soils and application methods. Mechanistically, VOCs can (a) inhibit pathogen growth/sporulation, (b) alter pathogen stress responses, and (c) shift microbial interactions in the rhizosphere. Trichoderma VOC studies provide clear experimental support for disease suppression plus plant response modulation [123,124,125,126].

7.3. Competition and Niche Exclusion in the Rhizosphere/Root Interface

Space and resource competition: Effective EFs rapidly colonize root surfaces and internal tissues, occupying infection courts and competing for carbon, micronutrients, and other limiting resources. This can reduce pathogen establishment probability even before antagonism is expressed [112,115,127,128,129,130]. Microbiome engineering (community-level suppression): A growing body of meta-omics and community ecology work supports the notion that beneficial fungi can restructure rhizosphere community assembly, strengthen beneficial networks, and reduce pathogen-favoring consortia. Seed coating is a particularly relevant delivery route because it can “steer” early microbiome assembly trajectories [131].

7.4. Host-Mediated Resistance: Immune Priming and Systemic Protection ISR-like Responses and Priming

Beyond killing pathogens directly, EFs can trigger induced systemic resistance (ISR) or priming-like states that enhance the speed and intensity of plant defense upon pathogen challenge, often involving JA/ET and sometimes SA-linked signaling depending on the pathosystem. Trichoderma and Sebacinales/Serendipita (including Piriformospora/Serendipita indica) are widely cited examples [116]. This matters for yield stability because priming can reduce chronic defense costs while maintaining readiness for attack, offering a plausible mechanistic bridge between biocontrol and growth promotion [14,112,128,129].

8. Interactions with the Soil Microbiome and Nutrient Cycling

EFs reside inside plant tissues yet influence belowground community assembly and function by altering host metabolism, root exudation, and immune status. These plant-mediated changes cascade into the rhizosphere [132,133,134], shifting the diversity and activity of microbes that control carbon input, nitrogen transformation, phosphorus mobilization, and disease suppression. They enhance nutrient cycling through organic acid secretion, siderophore production, and stimulation of soil enzyme activities [135,136,137,138] and restructure microbial networks to favor beneficial taxa over pathogens [129,139,140]. Conceptual syntheses now frame endophytes as integral members of the plant holobiont with ecosystem-level consequences for agro-ecosystems [14,22,112].

8.1. Mechanisms by Which Endophytes Remodel the Soil Microbiome

EFs remodel the soil microbiome primarily through plant-mediated pathways, including immune priming, shifts in root exudation, and the release of bioactive metabolites and volatile organic compounds (VOCs). These mechanisms do not operate in isolation; rather, they interact to stabilize microbial networks, enhance nutrient acquisition, and buffer plants against abiotic stress. Collectively, they link microbial community assembly with improved soil structure, nutrient use efficiency, and plant resilience, as illustrated in Figure 5.

8.1.1. EFs Influence on Soil Aggregation

EFs can strongly influence soil aggregation by acting both as physical architects of the soil matrix and as biochemical glue producers in the rhizosphere. Like other soil fungi, many root-associated endophytes form fine hyphal networks that extend from the roots into surrounding soil, physically enmeshing mineral particles and microaggregates into larger, more stable macroaggregates. Classic soil physics work has shown that fungal hyphae, particularly vesicular/arbuscular mycorrhizal (VAM/AMF) hyphae, are key stabilizers of macroaggregates (>250 µm) alongside roots, whereas polysaccharides and other organic compounds stabilize microaggregates [141,142,143]. Because many AMF are functionally root endophytes, this places EFs directly in the pathway of aggregate formation at the root–soil interface. Apart from this, under natural conditions, AMF and endophytic fungi frequently co-infect plant roots, and their complementary biological traits often act synergistically to enhance ecological reconstruction and vegetation recovery following disturbance [144].

A central biochemical mechanism is the production of glomalin-related soil proteins (GRSPs), a class of hydrophobic glycoproteins secreted by arbuscular mycorrhizal endophytes into the soil. GRSPs have repeatedly been linked to improved water-stable aggregation and greater mean weight diameter of aggregates, often showing a stronger direct effect on aggregate stability than hyphal length alone [10]. Recent reviews highlight three key functions of GRSPs, such as long-term carbon sequestration, soil aggregation, and soil remediation/fertility, emphasizing that their hydrophobic nature and metal-binding capacity help cross-link organic and mineral fractions within aggregates [145]. By enhancing GRSP pools and sustaining dense extraradical hyphal networks, endophytic AMF effectively “glue” soil particles together and protect soil organic carbon inside stable aggregates.

Non-mycorrhizal endophytes also contribute more indirectly by reshaping the rhizosphere microbiome, root exudation patterns, and soil organic carbon inputs. Studies with tall fescue endophytes show that endophyte infection can alter soil microbial community structure, increase glomalin content, and enhance soil organic carbon storage, all factors that favor aggregate formation and stability [146]. Fungal endophytes and other root-associated microbes release extracellular polymeric substances (EPSs), organic acids, and other metabolites that act as binding agents and modify soil wettability and pore structure, further promoting microaggregate formation and macroaggregate stabilization [147,148].

8.1.2. Immune Priming, Exudation Shifts, and Community Assembly

EF colonization frequently primes plant immune responses, including ISR mediated by jasmonic acid and salicylic acid signaling. Immune-primed plants alter the quantity and composition of root exudates, including organic acids, amino acids, sugars, and phenolics. These exudation shifts selectively recruit beneficial microbial guilds such as AMF, Actinobacteria, and plant-growth-promoting rhizobacteria while disfavoring pathogens, thereby restructuring rhizosphere networks toward disease suppressive and functionally resilient states [94]. Functionally, these community shifts enhance nutrient mobilization, improve water uptake, and stabilize microbial activity under abiotic stress. Enhanced mycorrhizal recruitment increases effective root surface area and hydraulic connectivity, improving phosphorus and water uptake during drought or salinity stress. At the same time, recruited bacterial partners support nitrogen mineralization, osmotic adjustment, and redox balance, collectively enhancing plant stress tolerance. Experimental evidence including Trichoderma inoculation studies in cacao and other crops demonstrates that endophytic colonization beyond roots can also confer systemic protection and growth benefits [149].

8.1.3. Competitive Exclusion and Niche Preemption

Competitive exclusion is the principle by which two species competing for the same limited resources cannot coexist indefinitely; one will eventually outcompete the other and cause local extinction. Niche preemption is a mechanism of competitive exclusion in which one species is superior in acquiring a resource that claims the entire niche, leaving nothing for the other species. In situations of niche overlap, competing species may evolve to use resources differently (resource partitioning), or a superior competitor may preempt the niche, leading to competitive exclusion. By occupying endosphere niches and competing for nutrients at the root interface, EFs limit pathogen entry and indirectly stabilize rhizosphere communities (reduced pathogen bloom → more even, function-rich networks) [14,121].

8.1.4. VOCs and Signaling at a Distance

EFs use VOCs for long-distance signaling to communicate with their host plants and other organisms, influencing processes such as growth, defense, and development [150]. These airborne molecules can promote plant growth, enhance pathogen resistance, or inhibit pathogens by directly acting as biocontrol agents without physical contact. The VOCs emitted by EFs are a diverse mix of compounds, such as terpenes and alcohols, and are being explored for sustainable agricultural applications, although challenges remain in controlling their effects owing to their high concentration dependence and dispersal [123,126,151,152].

8.2. Consequences for Nutrient Cycling in Arable Soils

EFs influence nutrient cycling in arable soils through a combination of direct biochemical activities and indirect plant- and microbiome-mediated processes (Table 3). They contribute to nutrient turnover by decomposing plant litter and producing extracellular enzymes that release bound nutrients, modifying litter quality through host metabolic regulation and reshaping soil microbial communities that govern nutrient transformations. These processes operate across multiple spatial scales, from the root endosphere to the surrounding rhizosphere and bulk soil, and collectively regulate the availability, retention, and efficiency of carbon (C), nitrogen (N), phosphorus (P), and micronutrients in cropping systems (Figure 5). By coupling plant carbon inputs with microbial energy flow and nutrient mobilization, EFs help stabilize nutrient cycling under fluctuating environmental conditions, thereby enhancing soil fertility, suppressing disease-promoting feedback and supporting ecosystem resilience in intensively managed agroecosystems [111,112,113,136].

8.3. Complementary Soil Management Approaches That Amplify EF-Mediated Benefits

Recent work highlights that EF-mediated processes can be further strengthened by complementary soil management approaches (organic amendments, integrated nutrient management, cover crops and crop rotation, reduced/no-till farming, and optimizing soil pH), particularly organic amendments such as biochar. Biochar improves soil structure, porosity, water holding capacity, and nutrient retention while providing habitat niches that support microbial persistence and functional diversity. When combined with EFs, biochar-amended soils show enhanced microbial network stability, increased nutrient use efficiency, and improved plant stress tolerance. Biochar surfaces can adsorb organic acids, enzymes, and signaling molecules, potentially prolonging EF-mediated effects and buffering microbial activity under stress conditions. Integrating EFs with organic amendments therefore represents a promising systems-level approach for enhancing soil structure, nutrient availability, and agroecosystem resilience, particularly under climate stress scenarios [117].

To integrate the functional roles of endophytic fungi described in Section 4, Section 5, Section 6 and Section 7 with the mechanistic pathways detailed here, it is important to recognize that plant growth promotion, abiotic stress tolerance, and pathogen biocontrol emerge from a shared set of soil- and microbiome-mediated processes. Specifically, EF-induced immune priming, shifts in root exudation, competitive niche preemption, and VOC-mediated signaling (Section 8.1) form the mechanistic basis through which EFs enhance nutrient acquisition (Section 5), buffer abiotic stress (Section 6), and suppress plant pathogens (Section 7). These mechanisms operate across the plant–soil continuum, linking endophytic colonization to rhizosphere community assembly, nutrient cycling, soil aggregation, and ecosystem resilience.

9. Translational Pipeline: Strain → Formulation → Delivery → Monitoring

The successful integration of EFs into sustainable agriculture requires a translational pipeline that bridges fundamental ecological insights with practical field applications. Figure 6 shows this pipeline, which consists of four interlinked stages, namely strain selection, formulation, delivery, and monitoring, each of which is crucial for achieving reproducible performance, compatibility with soil microbiomes, and measurable soil health outcomes in arable systems.

9.1. Strain Selection and Trait Stacking

The development of effective EF bioinoculants begins with the selection of strains that are biologically robust, environmentally adaptable, and multifunctional. Modern omics tools, such as metagenomics, transcriptomics, and metabolomics, enable researchers to identify strains with key traits linked to nutrient cycling, microbial diversity, and ecosystem stability [111,114,153,154]. Promising candidates such as Clonostachys rosea and Trichoderma spp. show strong biocontrol potential through mycoparasitism, antibiosis, and enzyme production [121,155]. Screening for strains that can trigger ISR via the jasmonic and salicylic acid pathways is equally important [112]. Additionally, evaluating VOC profiles helps identify fungi capable of pathogen suppression in variable soil environments [123,126,150]. Finally, omics-based host strain compatibility testing ensures that the selected fungi persist in specific crop–soil systems without disrupting native microbiomes [114].

9.2. Formulation and Production Technologies

The formulation determines how effectively EF strains survive, establish, and function in the soil–plant environment. Traditional inoculants often face challenges in terms of shelf stability, colonization efficiency, and compatibility with abiotic conditions. Modern formulation strategies address these through (1) carrier systems, such as solid carriers (e.g., peat, talc, lignite, and biochar) or liquid carriers with protective polymers that help maintain viability and facilitate root colonization; (2) encapsulation through alginate or biopolymer encapsulation, which enhances stress tolerance and controlled release of propagules; (3) co-formulation with beneficial microbes, where combining EF with rhizobacteria or AMF can create synergistic consortia for disease suppression and nutrient cycling; and (4) bioprocess optimization, where submerged and two-stage fermentation systems improve propagule yield and physiological quality, as shown for Clonostachys rosea and Trichoderma spp. Well-formulated EF inoculants must balance biological efficacy with ease of storage, handling, and application in agricultural settings [53,99].

9.3. Delivery Methods in Arable Systems

Efficient delivery strategies ensure effective colonization and interaction between plants and the soil microbiome. Different delivery approaches can be tailored to the crop type, farming system, and soil conditions. EFs are delivered to arable systems through artificial inoculation methods, such as seed coating, soil inoculation, root dipping, and foliar spraying [156]. The choice of the delivery method should align with both biological objectives (colonization and activity window) and farm logistics (scalability and cost-effectiveness).

9.4. Monitoring and Feedback Mechanisms

Post application monitoring is essential to assess EF establishment, persistence, and its impact on the soil microbiome and soil health indicators. Robust monitoring frameworks typically include (1) molecular tracking, including quantitative PCR (qPCR), amplicon sequencing, and metagenomics to confirm colonization and community shifts–, where qPCR-based methods have been successfully applied to track Clonostachys rosea and Trichoderma spp. in crop systems [157]; (2) functional indicators, such as enzyme activity (e.g., phosphatase and β-glucosidase), nutrient mineralization rates, and disease incidence, which is a practical field metric; (3) soil health indices, determined by monitoring changes in aggregation, carbon pools, microbial biomass, and pathogen load over time, which provides insight into ecosystem-level effects [10]; and (4) isotopic tracing (SIP), where stable isotope probing can be used to identify active microbial guilds involved in nutrient cycling under the influence of EFs [113]. Establishing feedback loops between monitoring data and formulation/delivery refinement enables adaptive optimization of EF-based interventions.

10. Integration with Sustainable Crop Production Practices

The integration of EFs into sustainable crop production involves the use of beneficial microbes to improve plant growth, nutrient uptake, and biotic and abiotic stress tolerance, thereby reducing the need for chemical fertilizers and pesticides. This can be achieved through advanced inoculation techniques, such as seed treatments, soil inoculations, and bioformulations, which can be combined with other sustainable practices, such as organic farming and conservation tillage, to enhance overall soil health and crop resilience. EFs can be aligned with IPM, conservation agriculture, and organic farming systems [158,159,160]. Delivery methods include seed coatings, granules, and liquid inoculants. However, their adoption depends on farmer awareness, performance consistency, and economic viability [161].

Seed coating is the most common and scalable method that allows for early niche preemption and ensures intimate contact between EF propagules and germinating roots. For example, Trichoderma seed coatings have been shown to restructure the rhizosphere communities and improve crop performance. For in-furrow or drench applications, inoculants are introduced directly into the rhizosphere to ensure broader soil coverage. Granular and slow-release formulations enable long-term persistence of EF propagules in soil aggregates. Consortium delivery involving EFs with AMF or phosphate-solubilizing bacteria can synergistically enhance nutrient mobilization and suppress disease.

10.1. Methods for Integrating EFs

Methods for integrating EFs include traditional techniques, such as seed treatment and soil inoculation, along with more advanced methods, such as foliar sprays, root drenches, and encapsulation, to protect the fungi. Innovative delivery systems and specialized media have also been used to promote EFs’ growth and colonization [156].

10.1.1. Traditional Methods

Traditional methods include (1) seed treatment, which involves coating seeds with endophyte formulations to ensure early colonization; (2) soil inoculation, which involves application of endophytes directly to the soil to enhance root colonization; (3) foliar sprays, which involves applying fungi as a spray on leaves; and (4) root drenching, which involves pouring endophyte formulations directly onto the root zone [156].

10.1.2. Advanced and Innovative Methods

Several advanced and innovative methods have been discovered: (1) encapsulation, which uses micro or nano-encapsulation to protect endophytes from environmental stress and allow for controlled release; (2) specialized media, where optimized culture media are used to improve fungal growth, such as potato dextrose agar for multiple species, or variations, such as potato dextrose agar with added plant extracts or lower nutrient concentrations to promote specific growth characteristics [162]; and (3) multi-omics approaches, such as metagenomics, transcriptomics, and metabolomics, to understand host–endophyte interactions at a molecular level and guide the development of more effective bioformulations [19,49,163].

11. Challenges and Limitations for EFs Application

The field performance of EF products is often inconsistent because their efficacy depends on crop genotype, soil type, climate, background microbiome, and even the application method/timing (e.g., pre-transplant vs. during transplant vs. post-transplant), which reshapes rhizosphere communities differently. These constraints include host specificity, environmental sensitivity, competition with native microbiota, production and formulation issues, and regulatory hurdles [4,28,164,165]. However, limited large-scale field validation remains a major challenge. Production and formulation bottlenecks that persist in solid-state systems can limit yields and quality, whereas implementing submerged/two stage fermentation improves propagule output but requires process optimization and QA/QC to achieve uniform, stable products [19,161]. After manufacturing, the shelf life and viability of seeds/coatings are recurrent constraints; maintaining live inocula through storage and handling is non-trivial and a known cause of variable outcomes in grower fields [166]. Ecologically, EF establishment can be host- and context-specific, with biotic and abiotic filters in the rhizosphere shaping whether the introduced strains persist or are out competed, underscoring the need for local matching and consortium design [167]. In practice, adoption is tempered by market and regulatory issues (quality standards, labeling, and registration), farmer confidence, and economic viability, all of which have been highlighted in recent inoculant market and technology reviews [19]. Finally, programs need post-application verification, which involves molecular tracking (e.g., qPCR) to confirm colonization and link their presence to their effects. Without this, null results are difficult to interpret, and iterative improvement is stalled.

12. Future Perspectives and Research Priorities of EFs

Future studies on EFs should leverage multi-omics, genome editing, and synthetic biology to optimize beneficial traits [168,169,170]. Climate-resilient endophyte selection, long-term field trials, regulatory reforms, and farmer engagement are critical for scaling adoption. EFs research in sustainable agriculture involves the development of integrated, data-driven strategies that couple ecological understanding with agronomic impacts. One key direction is the use of multi-omics platforms, such as metagenomics, transcriptomics, metabolomics, volatilomics, and stable isotope probing, to reveal the functional roles of EFs in nutrient cycling, plant immunity, and rhizosphere engineering at the ecosystem scale. By linking EF activity to carbon, nitrogen, and phosphorous fluxes, researchers can identify strains and functional guilds that provide the greatest benefits in specific soil–crop contexts [49,171].

The second priority is trait stacking and consortium development. Instead of relying on single-strain inoculants, EF consortia combining complementary functions, such as mycoparasitism, ISR induction, nutrient mobilization, and VOC-mediated pathogen suppression, may offer more robust performance across variable field environments. These consortia should be tailored to crop genotypes, soil microbiome structure, and management practices, such as conservation tillage or organic farming [123].

Third, scaling EF applications in arable systems requires optimized delivery platforms. Seed coating and in-furrow delivery are particularly promising because they are compatible with current agricultural practices and facilitate early root endophyte interactions. Developing stable formulations (e.g., encapsulated granules and liquid carriers with an extended shelf life) is crucial. Post deployment, molecular monitoring using qPCR, amplicon sequencing, and isotopic labeling are required to confirm colonization, persistence, and ecosystem effects [149,157].

Finally, research should place stronger emphasis on linking EF inoculation to soil health metrics, including aggregate stability, microbial biomass, enzyme activity, nutrient use efficiency, and agronomic outcomes, such as yield stability and stress resilience. Building iterative “monitor-refine-reapply” frameworks that combine field data with mechanistic insights will enhance reliability, scalability, and farmer adoption [172,173].

13. Conclusions

The increasing need for sustainable and low-input agriculture has brought endophytic fungi (EFs) to the forefront as integral components of plant–soil systems rather than passive members of the plant microbiome. This review highlights how EFs function as ecological engineers that enhance crop performance, suppress pathogens, and regulate nutrient cycling through coordinated interactions with the soil microbiome. By linking endophytic activity to phosphorus mobilization, nitrogen use efficiency, immune priming, and soil structural improvement, EFs emerge as key drivers connecting plant health with soil health in arable ecosystems. Recent advances in multi-omics, metabolomics, and stable isotope approaches have shifted the field from descriptive surveys toward mechanistic understanding, enabling direct links between EF activity and carbon, nitrogen, and phosphorus fluxes. However, EF performance remains context dependent, shaped by host genotype, soil properties, resident microbiota, and management practices. Addressing these constraints requires integrative frameworks that combine ecological matching, functional trait screening, robust formulation, and field-scale monitoring. Future progress will depend on embedding EFs within sustainable cropping systems, such as conservation tillage, crop diversification, and IPM, while developing shared data resources that link EF traits to soil health outcomes. By advancing mechanistic insight alongside translational strategies, EFs hold strong potential to support resilient, climate-adaptive, and environmentally sound agricultural systems.