Drought Stress Enhances Mycorrhizal Colonization in Rice Landraces Across Agroecological Zones of Far-West Nepal

Abstract

Mycorrhizal symbiosis in rice enhances drought adaptation but there are limited studies regarding the frequency and amplitude of mycorrhizae colonization in traditional landraces. This study investigates mycorrhizal colonization frequency (FMS) and intensity (IRS) in 12 rice landraces across three agroecological zones (Tarai, Inner-Tarai, Mid-hill) of Far-West Nepal under drought stress. Field experiments exposed landraces to control, intermittent, and complete drought treatments, with soil properties and root colonization analyzed. Results revealed FMS and IRS variations driven by soil composition and genotype. Mid-hill soils (acidic, high organic matter) showed lower FMS but elevated IRS under drought, while neutral pH in Tarai and silt/clay-rich soils supported higher FMS. Sandy soil in Inner-Tarai also promoted FMS. Drought significantly increased IRS, particularly in Anjana and Sauthiyari (Tarai), Chiudi and Shanti (Inner-Tarai), and Chamade and Jhumke (Mid-hill), which exhibited IRS surges of 171–388%. These landraces demonstrated symbiotic resilience, linking mycorrhizal networks to enhanced nutrient/water uptake. Soil organic matter and nutrient levels amplified IRS responses, underscoring fertility’s role in adaptation. FMS ranged from 50 to 100%, and IRS 1.20–19.74%, with intensity being a stronger drought-tolerance indicator than frequency. The study highlights the conservation urgency for these landraces, as traditional varieties decline due to hybrid adoption. Their drought-inducible mycorrhizal symbiosis offers a sustainable strategy for climate-resilient rice production, emphasizing soil–genotype interactions in agroecological adaptation.

1. Introduction

Rice is a staple food to more than half of the world’s population, and, as nearly 21% of the caloric intake for the world, rice contributes significantly to quotidian caloric consumption, agricultural gross domestic product (GDP), and land cultivated to grow rice in Nepal [1,2,3]. However, climate change is making rice farming more challenging in Nepal, characterized by increasing temperatures, increasingly erratic weather, and increased extreme weather events. From the period of 1991–2020, the annual maximum temperature of Nepal increased by 0.063 °C, and the annual minimum shifted by 0.072 °C, thus contributing to more evapotranspiration, and less moisture in the soil, both of which are necessary to grow rice [4].

These impacts of climate change, such as prolonged drought events and lack of reliable rainfall patterns, are putting further stress on water resources, especially in rice farming communities around the world. Drought is increasingly recognized as a growing global crisis affecting ecosystems, economies, and food security. South Asian regions including Nepal are greatly impacted by unreliable monsoon rains, delayed rain, and extended drought periods that have decreased crop yields and are threatening food security and agricultural sustainability [4,5]. Solutions to these challenges will require adaptation to climate change with sustainable rice production development such as drought-tolerant varieties, improved water management, and climate-smart agricultural systems.

Mycorrhizal fungi and particularly arbuscular mycorrhizal fungi (AMF) are present in nearly all rice-producing areas and enter into symbiotic relationships with rice roots [6,7]. Mycorrhizal fungi form extensive hyphal networks that provide increased water and nutrient uptake capabilities, especially phosphorus, while also providing the plant with protection from pathogens, improving growth and resilience [8,9,10,11]. AMF not only improve the uptake of nutrients, but also help rice plants withstand (i.e., adapt to) biotic and abiotic stress (e.g., drought, salinity, nutrient deficiency) [7,12]. The success of AMF colonization is reliant on soil texture, fertility, and irrigation regime as well as the rice genotype [13,14]. For instance, soil pH, organic matter, and moisture content, as well as allometrically compatible varieties, can all exert influence on the level of colonization [15]. Thus, complex interactions must be understood to maximize AMF and benefit rice yield, especially in drought-prone environments.

Geographical diversity and climatic conditions in Nepal have allowed for many rice landraces to develop. The three agroecoregions of Nepal, Tarai, Mid-hill, and High Hill, contribute 68%, 28%, and 4% of rice production in Nepal, respectively [16]. Of an estimated number of around 2000 accessions of rice, 157 landraces are cultivated today, each with a group of adaptive traits [17,18,19]. Nevertheless, very little has been reported regarding their mycorrhizal association and stress tolerance. While mycorrhizal symbiosis was found to enhance rice drought adaptation [20,21], there have been fewer investigations on the frequency and amplitude of AMF colonization under drought stress in traditional landraces [22]. The clarification of these interactions is important for the identification of adaptive traits that can be used to inform sustainable agriculture.

Therefore, the current research aims to investigate the extent, variability, and functional relevance of mycorrhizal association in Nepal’s widely cultivated rice landraces under water stress. An important objective of this study is to identify specific rice landraces with strong mycorrhizal colonization that may contribute to sustainable and drought-resilient productivity.

2. Materials and Methods

2.1. Study Sites and Rice Landraces

The study was conducted in three distinct agroecological zones of the Far-West region (Sudur-paschim Province) of Nepal: Tarai, Inner-Tarai, and Mid-hill (Figure 1;

Table 1. Characteristics of the experimental sites.

S.N.	Agroecological Zone	District	Municipality/Site	Location	Elevation (masl.)
1.	Tarai	Kanchanpur	Belauri/Shantipur	28.696° N, 80.376° E	180
2.	Inner-Tarai	Dadeldhura	Parashuram/Parigaun	29.160° N, 80.286° E	353
3.	Mid-hill	Baitadi	Patan/Kagsyali	29.478° N, 80.617° E	1666

). These zones were selected to capture the region’s climatic and edaphic variability relevant to rice cultivation. The Tarai site (Kanchanpur) experiences a mean annual temperature of 27.1 °C and 1593 mm rainfall; Inner-Tarai (Dadeldhura) averages 20.8 °C and 1523 mm rainfall; and Mid-hill (Baitadi) has a mean temperature of 16.9 °C and 1156 mm rainfall. June is typically the hottest month, with July receiving the highest rainfall in all zones.

Twelve rice landraces (four per agroecological zone) were selected based on local farmer knowledge and prior field surveys, focusing on those reputed for drought tolerance and local adaptation (

Table 2. The rice varieties selected for the study.

S.N.	Location	Varieties Selected
1.	Tarai	Anjana, Ghiupuri, Lalchand, and Sauthyari
2.	Inner-Tarai	Batebudho, Chiudi, Jhini, and Shanti
3.	Mid-hill	Chamade, Jhumke, Ratomarso, and Temase

). This approach ensured representation of genetic and phenotypic diversity relevant to stress resilience.

2.2. Experimental Design

Field experiments were established in 2022 using a randomized complete block design with five replications per treatment. Each of the 12 landraces was grown in 1.5 m × 2 m plots within rainout shelters constructed to exclude natural rainfall while allowing ambient temperature and light. In total, 180 plots (12 landraces—4 at each location × 3 treatments × 5 replications) were used.

Three irrigation treatments were imposed: (i) Control: continuous irrigation, maintaining 90–100% soil moisture (water-flooded and fully saturated soils); (ii) Intermittent Drought Stress (IDS): irrigation withheld until soil moisture was 25–35%, then re-watered; (iii) Complete Drought Stress (DS): irrigated for 15 days post-transplantation, then withheld (soil moisture < 15%). Soil moisture was monitored using digital soil moisture sensors (model: WTPH01803, WANT Balance Instrument Co., Ltd., Changzhou, China) to ensure treatment consistency. A sensor was inserted up to a depth of 10–15 cm, and readings were taken from five random points of each plot. The readings were taken twice weekly. Landraces from the Mid-hill and Tarai matured at 120 days after seedling transplantation (DST), while Inner-Tarai landraces matured after 135 days.

2.3. Mycorrhizal Assessments

At harvest, three representative plants per plot were carefully uprooted to preserve root integrity. Fibrous roots were washed, cut into 1 cm segments, and fixed in 70% ethanol for transport to the Central Department of Botany, Tribhuvan University. Root processing followed McGonigle et al. [23] with minor modifications: roots were treated in 2% KOH (15 min, 90 °C), acidified in 2% HCl (3 min), stained with 0.05% trypan blue in lactoglycerol (3 min, 90 °C), and de-stained in 50% glycerol. For each sample, at least 30 root fragments were mounted on slides and examined under a stereomicroscope. Mycorrhizal colonization was scored (0–5 scale) following Trouvelot et al. [24]. Frequency (FMS) and intensity (IRS) of colonization were calculated as [23]:

$$\mathrm{Frequency}\mathrm{of}\mathrm{mycorrhiza}\mathrm{in}\mathrm{the}\mathrm{root}\mathrm{system}\left(\mathrm{F}\mathrm{M}\mathrm{S}\right)={\displaystyle \frac{\mathrm{Number}\mathrm{of}\mathrm{fragments}\mathrm{containing}\mathrm{mycorrhiza}}{\mathrm{Number}\mathrm{of}\mathrm{total}\mathrm{fragments}\mathrm{observed}}}\times 100$$

$$\mathrm{I}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{s}\mathrm{i}\mathrm{t}\mathrm{y} \mathrm{o}\mathrm{f} \mathrm{t}\mathrm{h}\mathrm{e} \mathrm{m}\mathrm{y}\mathrm{c}\mathrm{o}\mathrm{r}\mathrm{r}\mathrm{h}\mathrm{i}\mathrm{z}\mathrm{a}\mathrm{l} \mathrm{c}\mathrm{o}\mathrm{l}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{z}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n} \mathrm{i}\mathrm{n} \mathrm{t}\mathrm{h}\mathrm{e} \mathrm{r}\mathrm{o}\mathrm{o}\mathrm{t} \mathrm{s}\mathrm{y}\mathrm{s}\mathrm{t}\mathrm{e}\mathrm{m}\left(\mathrm{I}\mathrm{R}\mathrm{S}\right)={\displaystyle \frac{95\mathrm{n}5+70\mathrm{n}4+30\mathrm{n}3+5\mathrm{n}2+\mathrm{n}1}{\mathrm{N}\mathrm{u}\mathrm{m}\mathrm{b}\mathrm{e}\mathrm{r} \mathrm{o}\mathrm{f} \mathrm{t}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l} \mathrm{f}\mathrm{r}\mathrm{a}\mathrm{g}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{s} \mathrm{o}\mathrm{b}\mathrm{s}\mathrm{e}\mathrm{r}\mathrm{v}\mathrm{e}\mathrm{d}}}$$

where n5 = number of fragments rated 5; n4 = number of fragments rated 4; n3 = number of fragments rated 3; n2 = number of fragments rated 2; and n1 = number of fragments rated 1.

Relative frequency and intensity were calculated by dividing the frequency and intensity under stress treatment by control treatment and multiplying by 100. Also, the percentage changes in FMS and IRS were observed under intermittent drought and drought stress by using the formula:

$$\mathrm{P}\mathrm{e}\mathrm{r}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{g}\mathrm{e} \mathrm{c}\mathrm{h}\mathrm{a}\mathrm{n}\mathrm{g}\mathrm{e}={\displaystyle \frac{\mathrm{S}\mathrm{t}\mathrm{r}\mathrm{e}\mathrm{s}\mathrm{s} \mathrm{v}\mathrm{a}\mathrm{l}\mathrm{u}\mathrm{e}-\mathrm{C}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l} \mathrm{v}\mathrm{a}\mathrm{l}\mathrm{u}\mathrm{e}}{\mathrm{C}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l} \mathrm{v}\mathrm{a}\mathrm{l}\mathrm{u}\mathrm{e}}}\times 100\mathrm{\%}$$

where stress value = the value under stresses (i.e., intermittent drought stress and drought stress) and control value = the value under control (regular watering). A positive value represents an increase and a negative value represents a decrease in the percentage of mycorrhizae.

2.4. Soil Sampling and Analysis

Prior to planting, soil samples were collected from each experimental site. Five subsamples were taken from the corners and center of each 10 m × 10 m plot (depth: 0–15 cm), composited, and stored in airtight bags at 4 °C until analysis. Physical properties measured included bulk density (BD), pH, and texture (hydrometer method). Chemical analyses comprised soil organic carbon (SOC) [25], organic matter (SOM = SOC × 1.724), total nitrogen (TN) by the Kjeldahl method [26], available phosphorus (AP) [27], and available potassium (AK) [28]. All analyses were conducted at the Central Department of Botany, Tribhuvan University, following standard QA/QC procedures, including blanks, duplicates, and certified reference materials.

2.5. Statistical Analysis

Descriptive analysis of data was carried out using the ‘dplyr’ package in R Studio. Soil parameters from the three experimental sites were analyzed using One-way Analysis of Variance (ANOVA). FMS and IRS were analyzed using Two-way ANOVA to show the effect of treatment, landrace, and their interaction. The ‘ggplot2’ package was deployed for data visualization. All statistical analyses were carried out using R Studio (R Core Team 2024) (version 4.3.3) [29]. For all analyses, p < 0.05 was considered significant (* p < 0.05; ** p < 0.01; *** p < 0.001).

3. Results

3.1. Soil Parameters

Bulk density (BD) was significantly higher in the soils of the Tarai and Inner-Tarai regions compared to the Mid-hill (p < 0.001,

Table 3. Soil parameters of experimental sites across three agroecological regions.

	BD (g/cm3)	Clay (%)	Silt (%)	Sand (%)	pH	SOM (%)	N (%)	P (P2O5 mg/kg)	K (kg/ha)
Mid-hill	0.88 ± 0.13 b	12.70 ± 1.4 ab	26.74 ± 2.9 b	60.56 ± 3.8 b	5.15 ± 0.12 c	5.81 ± 0.49 a	0.27 ± 0.05 a	0.62 ± 0.11 a	112.8 ± 37.6 a
Inner-Tarai	1.29 ± 0.13 a	9.22 ± 2.2 b	20.88 ± 3.3 c	69.90 ± 4.6 a	6.41 ± 0.14 b	4.58 ± 0.51 b	0.13 ± 0.05 b	0.29 ± 0.11 b	107.8 ± 24.3 a
Tarai	1.25 ± 0.09 a	12.76 ± 2.4 a	40.40 ± 1.7 a	46.84 ± 2.2 c	7.27 ± 0.31 a	1.87 ± 0.26 c	0.08 ± 0.01 b	0.18 ± 0.03 b	72.0 ± 20.7 a

BD: Bulk density, SOM: Soil organic matter, N: Nitrogen, P: Phosphorus, K: Potassium, P₂O₅: Phosphorus Pentaoxide. Letters in superscript ‘a, b, c’ denote significant difference in between agro-ecological zones.

). Soil texture also varied notably among agroecological zones: the Inner-Tarai had the highest sand content, followed by the Mid-hill and Tarai, whereas silt and clay were most abundant in the Tarai, with the lowest proportions found in the Inner-Tarai (p < 0.05 for clay, p < 0.001 for silt and sand; Table 3, Supplementary Table S1). Soil pH differed markedly between zones, with the Mid-hill exhibiting acidic conditions (pH 5.15), while the Tarai and Inner-Tarai were less acidic (pH 6.41 and 7.27, respectively; p < 0.001). The highest soil organic matter (SOM) content was observed in the Mid-hill (6%), followed by the Inner-Tarai and Tarai. Similarly, nitrogen and phosphorus concentrations were greater in the Mid-hill and Inner-Tarai than in the Tarai soils (p < 0.001). In contrast, potassium levels did not differ significantly between the three agroecological zones (p = 0.085).

3.2. Frequency of Mycorrhizal Colonization in Root System (FMS)

Analysis of variance revealed significant variation in FMS among the rice landraces in the Tarai (p < 0.001), but neither drought treatment nor the interaction between landrace and treatment had a significant effect (p = 0.118 and p = 0.177, respectively; Figure 2A). Among the Tarai landraces, Sauthiyari exhibited the highest FMS, followed by Anjana and Lalchand, while Ghiupuri showed the lowest values. In the Inner-Tarai, both landrace and treatment significantly influenced FMS (p < 0.001), though their interaction was not significant (p = 0.211; Figure 2B). Here, Jhini had the highest FMS, followed by Chiudi and Batebudho, with Shanti recording the lowest. In the Mid-hill region, significant differences in FMS were observed for the treatment and landrace, as well as their interaction (all p < 0.001; Supplementary Table S1). Jhumke displayed the highest FMS (95.5 ± 1.84%), with Jhumke, Ratomarso, and Temase not differing significantly from each other, but all three were significantly higher than Chamade (Figure 2C, Supplementary Table S2).

3.3. Intensity of Mycorrhizal Colonization in Root System (IRS)

For IRS, two-way ANOVA indicated significant effects of landrace, treatment, and their interaction in the Tarai (all p < 0.001; Supplementary Table S2). Under drought conditions, Anjana exhibited the highest IRS, while Ghiupuri, Lalchand, and Sauthiyari showed comparable levels (Figure 3A). In the Inner-Tarai, IRS was also significantly influenced by both landrace and treatment, as well as their interaction (all p < 0.001; Supplementary Table S2). Chiudi had the highest IRS among landraces. Notably, in Jhini and Shanti, IRS increased significantly under drought stress, whereas Batebudho did not show differences between treatments (Figure 3B). In the Mid-hill, IRS was significantly affected by landrace, treatment, and their interaction (all p < 0.001; Supplementary Table S2, Figure 3C). Temase had the highest IRS in control plants, followed by Ratomarso, Jhumke, and Chamade, with Chamade recording the lowest. Under drought, Jhumke showed a marked increase in IRS, while Ratomarso had lower values relative to the other landraces. Both the IDS and the DS treatments led to elevated IRS in Chamade and Jhumke (Figure 3C).

3.4. Percentage Change in FMS and IRS

The percentage change in FMS under both intermittent drought (IDS) and drought stress (DS) was assessed across all landraces in the Tarai (Figure 4A). Under IDS, Anjana exhibited an 11.97% increase in FMS compared to the control, followed by Sauthiyari, whereas Ghiupuri had lower FMS. Under DS, Anjana again showed the greatest increase (12.68%), while Lalchand experienced a 2.11% decrease. Sauthiyari and Ghiupuri also displayed modest increases (6.21% and 0.85%, respectively). For IRS, Anjana showed a dramatic increase under IDS (171.15%) and DS (387.18%) compared to control, followed by Sauthiyari and Ghiupuri, while Lalchand exhibited a decrease (41.30%) under IDS. In the Inner-Tarai, Batebudho had the largest percentage increase in FMS under IDS, followed by Chiudi, while Shanti and Jhini showed slight decreases. Under DS, Chiudi led with a 21.88% increase, followed by Batebudho and Shanti, with Jhini showing only a 5.38% rise (Figure 4B). For IRS, Shanti had a substantial increase under IDS (241.67%) and DS (387.5%), followed by Chiudi and Jhini, while Batebudho decreased by 13.35% (IDS) and 33.01% (DS). In the Mid-hill, Ratomarso showed the greatest increase in FMS under IDS (18.63%) and DS (24.22%), while Chamade and Jhumke had decreases. Temase and Jhumke had moderate increases under DS. IRS in Jhumke rose sharply under both IDS (206.72%) and DS (313.86%), with Chamade and Temase also showing increases under DS (227.22% and 88.01%, respectively; Figure 4C).

In the Inner-Tarai, a more substantial percentage change in the FMS of mycorrhizae under IDS was observed in Batebudho than the control, followed by Chiudi. In contrast, decreases of 0.98% and 2.15% in FMS were recorded in Shanti and Jhini, respectively. Under DS, Chiudi displayed a 21.88% higher FMS compared to control followed by Batebudho and Shanti, while Jhini recorded only a 5.38% increase (Figure 4B). The IRS under IDS was 241.67% higher in Shanti compared to the control, followed by Chiudi, while Jhini showed a slight decrease of 0.36%, and Batebudho had the smallest change, with a reduction of 13.35%. Under DS, IRS was 387.5% greater in Shanti compared to the control, followed by Chiudi and Jhini, which recorded percentage increases of 282.16% and 166.42%, respectively. In contrast, Batebudho exhibited a decrease of 33.01% in IRS (Figure 4B).

The Ratomarso landrace of the Mid-hill region showed an 18.63% increase in FMS under IDS than control, followed by Temase. In contrast, FMS decreased by 2.62% in Jhumke, followed by Chamade. Under DS, Ratomarso exhibited a 24.22% higher FMS, while Chamade showed a 1.06% decrease. Temase and Jhumke recorded increments of 11.24% and 3.66% higher FMS than the control under DS, respectively (Figure 4C). IRS levels in Jhumke increased by 206.72% and 313.86% under IDS and DS treatments, respectively, compared to control plants. Chamade exhibited a 154.45% increase in IRS, while Temase showed a 14.99% decrease under IDS. DS treatment resulted in a 12.28% increase in IRS for Ratomarso, while Chamade and Temase displayed increases of 227.22% and 88.01%, respectively (Figure 4C).

Comparing the relative frequency and intensity of mycorrhizal colonization in the root system of rice landraces, both were high in most of the landraces, with few exceptions (Figure 5). In all the agroecological sites, there was an increase in the relative values of the frequency of around almost a hundred percent, while the relative values of intensity varied with the landraces. In Tarai, both the values were high in Anjana under the stresses, followed by Sauthyari (Figure 5A). Although the relative frequency was high in Batebudo under both the stresses, inversely, the relative intensity was low (Figure 5B). The Shanti landrace had the highest relative intensity (487%) compared to other landraces of that ecological site under both the stresses. Under both the stresses, the relative intensity was the lowest in the Temase landrace and the highest in the Jhumke landrace, followed by Chamade (Figure 5C).

4. Discussion

4.1. The Effect of Ecological Zones and Rice Landraces on the FMS

Our findings demonstrate that the frequency of mycorrhizal colonization (FMS) in rice roots varied significantly between agroecological zones, with the highest values observed in Mid-hill landraces, and less pronounced differences between the Inner-Tarai and Tarai. This suggests that the Mid-hill environment supports more robust mycorrhizal associations, likely due to favorable soil fertility and higher organic matter content [30]. However, FMS is not solely determined by the overall richness of mycorrhizal fungi in the soil, as individual fungal species can form dense colonies within specific host roots [31]. Water availability is also critical, as mycorrhizal colonization generally declines in arid environments [32], while adequate moisture facilitates rapid root–fungal association and nutrient exchange [33]. The observed FMS rankings within each region—Sauthiyari > Anjana > Lalchand > Ghiupuri (Tarai), Jhini > Chiudi > Batebudho > Shanti (Inner-Tarai), and Jhumke > Chamade > Temase > Ratomarso (Mid-hill)—underscore the influence of host genotype. It is clear that the Mid-hill site has higher mycorrhizal frequency, suggesting that the fungi love acidic soils, but this could be the result of specific landrace genotype–fungus mutualistic association and adaptation because different sites have different landraces. Therefore, it cannot be reliably concluded that these differences are due to site (soil conditions) or landrace adaptability. Upland and highland landraces, adapted to periodic drought, may possess deeper root systems and other traits that promote stable yields and strong mycorrhizal associations [34]. Natural AMF populations in rain-fed, upland agroecosystems likely further enhance colonization, suggesting that Sauthiyari, Anjana, Jhini, Chiudi, Jhumke, and Chamade harbor traits indicative of drought tolerance.

4.2. FMS Response to Drought and Landrace-Specific Patterns

While we anticipated that drought stress would universally increase FMS due to the mutual benefits of enhanced water and nutrient uptake, our results revealed that this response was landrace-specific and varied by region. In the Inner-Tarai, all landraces exhibited increased FMS under drought, whereas in the Mid-hill, only Jhumke, Ratomarso, and Temase showed marked increases. These patterns suggest that drought can act as a trigger for mycorrhizal symbiosis, but the magnitude of response depends on both the rice genotype and the local fungal community. Enhanced mycorrhizal colonization under drought is associated with improved plant–water relations and photosynthetic efficiency [35], implying that landraces with higher FMS—such as Batebudho, Chiudi, Shanti, Ratomarso, and Temase—may be particularly well-suited for cultivation in drought-prone environments.

4.3. FMS and Drought: Percentage Change and Conservation Implications

Analysis of percentage changes in FMS under drought revealed that in the Tarai, Anjana and Sauthiyari exhibited positive responses, while Ghiupuri and Lalchand declined, reinforcing the notion that not all landraces benefit equally from mycorrhizal associations during stress [36]. In the Inner-Tarai, although Jhini had the highest baseline FMS, Batebudho and Chiudi showed greater increases under drought, suggesting that the capacity for adaptive mycorrhizal recruitment may be more critical than absolute colonization levels. Similarly, in the Mid-hill, Ratomarso maintained strong colonization under drought, while Chamade and Jhumke declined. These findings indicate that high FMS under optimal conditions does not guarantee resilience under stress. Importantly, many of the landraces showing promising mycorrhizal responses are at risk of being lost due to the spread of hybrid varieties. Our results underscore the need to conserve these traditional landraces, as they may harbor unique adaptations to drought-prone environments.

4.4. IRS Varied with Soil Moisture and Landraces

Mycorrhizal colonization intensity (IRS) was strongly influenced by both treatment and landrace—all rice landraces increased IRS under drought. However, the degree of response was variable, indicating the intricate interactions between plants and fungi under stress. For example, in the Tarai, Anjana, Ghiupuri, and Sauthiyari showed big increases in IRS under drought—only Lalchand went down. This indicates that landraces have different strategies to adapt to drought. In the Inner-Tarai, Shanti showed a significant increase in IRS under drought conditions—even though FMS went down. This shows that we need to consider intensity and not just frequency of mycorrhizae because some genotypes may be using intensity to develop their resilience to drought. In the Mid-hill, Jhumke produced the highest IRS score under drought, again indicating diverse adaptive capacity. We hypothesize that variation in adaptive phenotypic traits may stem from differences in root exudate chemistry and architecture, which are genetic traits [37] coupled with genotype by environment [38]. There may not be a direct relationship between IRS and improved growth response; studies have noted that greater IRS under drought connotes improved nutrient acquisition and stress tolerance [39], confirming that IRS is an essential trait related to adaptation.

4.5. Soil Properties, FMS, and IRS

Soil physico-chemical properties, particularly pH, texture, and nutrient status, played a significant role in shaping both FMS and IRS across regions [40]. In the Tarai, higher pH and silt/clay content were associated with elevated FMS in landraces like Sauthiyari, while the more acidic soils of the Mid-hill were linked to lower FMS, consistent with findings that acidification impacts AM fungal community composition [41]. Sandy soils in the Inner-Tarai and Mid-hill can promote mycorrhizal colonization due to increased porosity and root proliferation [42], though exceptions were observed among landraces with unique root traits. Despite higher SOM and NPK in the Mid-hill and Inner-Tarai, FMS remained above 75% in most landraces, suggesting that nutrient levels alone do not fully explain colonization patterns. In contrast, increases in IRS under drought—particularly in Anjana (Tarai), Chiudi (Inner-Tarai), and Jhumke (Mid-hill)—were more closely aligned with high SOM and nutrient availability, supporting the view that these soil properties facilitate intensive mycorrhizal symbiosis [43,44].

4.6. Drought-Tolerant Landraces and Conservation

Our data highlight Anjana and Sauthiyari (Tarai), Chiudi and Shanti (Inner-Tarai), and Chamade and Jhumke (Mid-hill) as landraces with notably high IRS under drought, suggesting a strong reliance on mycorrhizal symbiosis for nutrient and water uptake [45]. Although high frequency of mycorrhizal colonization (FMS) was found in several but not all landraces, intensity of colonization appears to be tied to drought tolerance most closely. Comparative analysis of the mycorrhizal responses in non-native and improved rice cultivars should be carried out to evaluate their symbiotic compatibility. The complex interaction of factors such as soil pH, nutrient status, and organic matter makes prediction of mycorrhizal associations very difficult and emphasizes the need for more information regarding the genotype by environment interaction. The discovery and conservation of these high-IRS landraces for current breeding and future sustainable agriculture, especially in arid regions, will be an important challenge for agriculture in the future. This study has evaluated a limited set of parameters for mycorrhizal association. Future studies could include a greater number of rice genotypes, which would enhance the scope of the findings. Additionally, use of statistical techniques such as correlation or multivariate analyses or cluster analysis would likely have yielded more comprehensive and insightful results. This study has significance as a preliminary exploration to provide local or region-specific insights and forms the basis for more extensive studies involving broader genetic and environmental conditions.

5. Conclusions

In summary, our observations indicate that both the FMS and IRS of certain rice landraces are controlled by an interaction of genes, agroecological environments, and soil properties. Comparatively lower soil pH and silt content is associated with greater FMS, indicating that the colonization of crops is high under acidic conditions, with FMS being consistently greatest under drought stress, especially for landraces which gave indications of high soil organic matter (SOM) and nutrient status. Despite encountering an overall trend in soil pH and SOM, the landraces that showed high IRS under drought conditions were Anjana, Sauthiyari, Chiudi, Shanti, Chamade, and Jhumke, which showed potential for a source of drought tolerance. Their conservation and further characterization are a must, given their decline in cultivation and their potential use for climate-resilient rice cultivation.