In-vitro evaluation of seed-dressing fungicides for the management of Sclerotium rolfsii Sacc. and enhancement of seed quality in groundnut

Introduction

Groundnut (Arachis hypogaea L.) considered as a major tropical and subtropical oilseed crop known for food, feed and edible oil because of its kernels are rich in protein, oil, vitamins, minerals and other bio-active compounds (Krishnan et al., 2025) [12]. It remains an economically important crop worldwide, with global production of about 54.2 million tonnes and India contributing nearly 10.1 million tonnes, establishing its importance in Indian agriculture (Jalla et al., 2024 and Krishnan et al., 2025) [9,12] . However, the productivity of groundnut is being affected by numerous abiotic and biotic stresses, among which fungal diseases are especially considered more damaging (Deepika et al., 2025) [4]. Among the various soil-borne diseases, groundnut stem rot or white mold caused by Sclerotium rolfsii is found to be one of the most destructive diseases of groundnut in India and other hot and humid regions (Deepika et al., 2025; Lavanya et al., 2024) [4,14] . The pathogen survives in soil for long periods through resilient sclerotia upon a broad host range and becomes difficult to manage under warm, humid conditions (Deepika et al., 2025; Meena et al., 2024) [4,18] . Yield losses commonly range from 10-25%, but severe infections can cause losses of 40-55% or even up to 80% (Jalla et al., 2024; Hotkar et al., 2025) [7,9] . Recent in vitro studies show strong activity of seed-treatment fungicides viz., carboxin + thiram and tebuconazole + trifloxystrobin against S. rolfsii (Meena et al., 2024; Surwade et al., 2024) [18,26] . Hence, the study was taken up to evaluate new-generation seed dressing fungicides against S. rolfsii and their effects on seed quality parameters.

Material and Methods

The study was carried out during kharif 2024 in the Seed Pathology Laboratory, Department of Seed Science and Technology, OUAT, Bhubaneswar.

The causal organism of stem rot of groundnut, Sclerotium rolfsii, was isolated from infected seeds of the high yielding variety Dharani collected from farmers' fields in Khordha district, Odisha. Seed-borne fungi were detected by using the standard blotter paper method. The pathogen was purified by the hyphal tip culture method (Kedar et al., 2025; Naikwade et al., 2025; Yadav et al., 2026) [10,19,29] .

Evaluation of seed dressing fungicides under laboratory condition

The S. rolfsii pure culture was maintained on potato dextrose agar (PDA) media. The fungicides were evaluated following the poisoned food technique (Table 1). Each fungicide with the required dose was mixed with sterilized PDA before pouring the medium into Petri plates (Aravind et al., 2024; Harshitha and Palakshappa, 2023; Satapathy and Sahoo, 2023) [2,6,23] . [2,15,23,25] . Measurement of colony diameter was taken in two directions at right angles and the mean value was obtained. Inhibition (%) of mycelial growth over the control was calculated using the following formula:

Where, C is the colony growth (mm) in the untreated control and T is the colony growth (mm) in the fungicide treatment (Aravind et al., 2024; Harshitha and Palakshappa, 2023) [2,6]

Seed quality assessment

The assessment of the effect of fungicidal seed dressing on seed quality of groundnut was observed at 7 and 12 days after treatment. Seed health parameters were examined by using the standard blotter paper method, while germination and seedling growth were evaluated by the paper towel method (Gupta et [5,19,20] . The experiment was conducted by laying out in a completely randomized design with four replications. Observations on different growth parameters viz., seed infection, germination and seedling were recorded regularly.

Growth and vigour measurements

Seeds treated with fungicides were evaluated for germination and seedling growth on the 12th day after sowing following the paper towel method. Each treatment was replicated four times taking 100 seeds each. Germination percentage was calculated by selecting only the number of normal seedlings as follows:

= Number of normal seedlings

Total number of seeds sown X 100

Ten normal seedlings from each treatment and the untreated control were selected randomly, and their root and shoot lengths were measured. By adding the mean root length and mean shoot length, the mean seedling length was calculated and expressed in centimetres (Khan et [11,13,20,21] .

To determine the seedling dry weight the same 10 seedlings were used. After removing the remnants of seed coats, the seedlings were dried in a hot-air oven at 80 °C for 24 h, then removed and cooled to room temperature and weighed.

Seedling dry weight was recorded and expressed in milligrams (mg) per seedling. The seedling lengths were averaged and mean seedling length observations were subsequently used and calculated the seed vigour indices.

Seed Vigour Indices (SVI) and statistical analysis

Seed vigour indices were calculated and assessed the effect of fungicidal seed treatments on seed germination and seedling growth. Seed Vigour Index-I (SVI-I) and Seed Vigour Index-II (SVI-II) were calculated following the method of Abdul-Baki and Anderson (1973) [1] . SVI-I = Germination (%) × Mean seedling length (cm) and SVI-II = Germination (%) × Mean seedling dry weight (mg)

The vigour indices obtained were used to compare the overall physiological performance of seedlings under different fungicidal seed treatments with the normal seedling length under untreated control. The recorded data were analyzed by computing the mean for each character in a Completely Randomized Design (CRD) and were subjected to analysis of variance (ANOVA) following the methods described by Rani et al. (2024) [22] . [26,27,28] . In contrast, thiophanate methyl 45% + pyraclostrobin 5% (T2) was the least effective treatment, with 64.16% inhibition. This lower efficacy is consistent with reports that some thiophanate-methyl-based treatments provide substantially lower suppression than the best-performing combinations and that thiophanate methyl alone can show only partial inhibition depending on concentration and formulation (Butt et [3,26,28] .

Result and Discussion

Effect of seed dressing fungicides on the growth of Sclerotium rolfsii under in vitro condition

The complete inhibition produced by trifloxystrobin + tebuconazole agrees with previous studies in groundnut, where this combination gave maximum inhibition in vitro and also performed well under field conditions (Meena and Meena, 2023; Surwade et al., 2024) [17,26] . Similar evidence from another stem rot pathosystem found that trifloxystrobin + tebuconazole had the strongest inhibitory effect, with very low EC50 values of 0.13-0.14 µg/mL, supporting the high intrinsic antifungal activity of this mixture (Zhao and Huang, 2023) [31] . The strong activity of carboxin + thiram also corroborates earlier reports in groundnut, where this mixture consistently inhibited the pathogen completely and was considered among the most effective fungicides (Surwade et al., 2024; Yadav et al., 2025) [26,28] . Likewise, the penflufen + trifloxystrobin combination was reported to inhibit 100% mycelial growth of S. rolfsii across different concentrations, supporting the present finding that this seed-treatment combination is highly potent in vitro (Vijaykumar et al., 2023) [27] . The superior performance of these combinations may be due to their multiple modes of action. The disruption of mitochondrial respiration by strobilurin components such as trifloxystrobin and pyraclostrobin occurs through the blockage of electron transfer at cytochrome b, thereby halting fungal energy production (Yadav et al., 2025) [28] . Pyraclostrobin has also been shown to reduce sclerotial production, oxalate secretion and protein synthesis in S. rolfsii, which helps explain its preventive activity (You et al., 2021) [30] .

Effect of seed dressing fungicides on seed germination

Fungicide seed dressing significantly enhanced groundnut germination over the untreated control (56.75%). Carboxin 37.5% WS + thiram 37.5% WS (T4, 81.75%) and penflufen 13.28% + trifloxystrobin 13.2% FS (T1, 80.50%) were the best-performing fungicides. The present findings were aligned with Asha et al. (2011) [24] , who reported significantly higher seedling emergence (80.53-93.68%) with carboxin + thiram-treated groundnut seed over the untreated control (75.97%), and corroborated the earlier results of Jadon et al. (2015) [8] , who found carboxin-and tebuconazole-based seed dressings effective in improving groundnut stand establishment. The germination gain likely reflects suppression of seed-borne mycoflora causing preemergence rot, similar to Manikchandra et al. (2026) [16] , who recorded a rise in germination from 64.00% in the control to 90.67% following fungicidal seed dressing in Niger.

Effect of seed dressing fungicides on seedling length and seedling dry weight

Fungicide-treated groundnut seed produced longer seedlings than untreated controls, with carboxin 37.5% WS + thiram 37.5% WS (24.90 cm) recording the maximum length, followed by penflufen + trifloxystrobin (22.28 cm), while trifloxystrobin + tebuconazole gave the shortest seedlings (20.06 cm). This trend agrees with Asha et al. (2011) [24] , who reported that carboxin + thiram (Vitavax Power) seed treatment in groundnut significantly increased root and shoot length and plant vigour over the check, attributing this to reduced pre-and post-emergence rot and improved plant establishment.

Effect of seed dressing fungicides on seedling vigour indices

Seedling vigour indices (SVI-I and SVI-II) differed significantly among fungicide treatments, with carboxin 37.5% WS + thiram 37.5% WS recording the highest SVI-I (1793.93) and SVI-II (296.56), while trifloxystrobin 25% + tebuconazole 50% WG gave the lowest values. Since vigour indices are derived from germination percentage combined with seedling length and dry weight, the superiority of carboxin + thiram reflects its combined advantage in both germination and seedling growth traits observed earlier. This finding is aligned with Asha et al. (2011) [24] , who reported that carboxin + thiram seed treatment in groundnut improved plant vigour and establishment over untreated seed, attributing it to better protection of the germinating embryo from soil-and seed-borne pathogens. Similarly, Manikchandra et al. (2026) [16] observed that effective fungicidal seed dressing markedly raised the seedling vigour index over the untreated control in the rolled paper towel method.

Effect of fungicides on seedling length and seedling dry weight

Systemic fungicide seed treatments enhanced seedling growth over untreated seed, though the magnitude varied with the molecule. Carboxin 37.5% WS + thiram 37.5% WS produced the longest seedlings (24.90 cm), followed by penflufen + trifloxystrobin (22.28 cm) and thiophanate methyl + pyraclostrobin (20.73 cm), while trifloxystrobin + tebuconazole gave the shortest seedlings (20.06 cm). The results are in line with Asha et al. (2011) [24] , who reported that carboxin + thiram seed dressing in groundnut significantly increased root and shoot length and overall seedling vigour compared with the check, owing to reduced pre-and post-emergence rot.

Conclusion

The study concluded that systemic fungicide molecules differed markedly in their ability to suppress stem rot fungus and improve groundnut seed quality. Penflufen + trifloxystrobin, carboxin + thiram and trifloxystrobin + tebuconazole completely inhibited fungal growth under in vitro conditions. Among the seed quality treatments, carboxin + thiram performed best, recording the highest germination, seedling length and vigour indices. Penflufen + trifloxystrobin and thiophanate methyl + pyraclostrobin also improved seed performance over other treatments. In contrast, trifloxystrobin + tebuconazole showed comparatively lower values for seed quality parameters despite strong antifungal activity. Overall, carboxin + thiram emerged as the most promising fungicidal seed treatment for managing stem rot and enhancing seed quality.

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