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Current IssueSpecial IssueEditorial BoardOnline First ArticlesArchive

Full Research Article

Integrated Management of Fusarium Wilt in Red Gram (Cajanus cajan L.) Through Chemical, Biological and Organic Approaches under Controlled Conditions

A
Anandita Sahoo1
P
Priyanka Choudhary2
N
Nirakar Ranasingh3
U
Uttam Kumar Behera4
J
Jay Prakash Singh5
C
Chinmayee Mohapatra6
P
Priti Upadhyay7
S
Srujani Behera3,*
0000-0001-7764-9631
1Department of Plant Pathology, Odisha University of Agriculture and Technology, Bhubaneswar-751 003, Odisha, India.
2Department of Mycology and Plant Pathology, Institute of Agricultural Sciences, B.H. University, Varanasi- 221 005, Uttar Pradesh, India.
3Department of Plant Pathology, College of Agriculture, Odisha University of Agriculture and Technology, Bhawanipatna-766 001, Odisha, India.
4Seed Technology Research, AICRP on Seeds (Crops), Odisha University of Agriculture and Technology, Bhubaneswar-751 003, Odisha, India.
5Department of Plant Pathology, Faculty of Agriculture, SMMTD College, Ballia-277 001, Uttar Pradesh, India.
6Faculty of Agriculture, Sri Sri University, Arilo, Bidhyadharpur, Cuttack-754 006, Odisha, India.
7Division of Vegetable Science, ICAR- Indian Agricultural Research Institute, New Delhi-110 001, India.

  • Submitted26-11-2025|

  • Accepted28-04-2026|

  • First Online 12-05-2026|

  • doi 10.18805/LR-5610

ABSTRACT

Background: Red gram or pigeon pea (Cajanus cajan L.) is a major pulse crop cultivated in tropical and subtropical regions, whose productivity is hampered by the wilt from a soil-borne fungus, Fusarium oxysporum.

Methods: In vitro studies as well as controlled pot experiments were conducted during 2024-2025 that evaluated chemical fungicides viz., Tebuconazole50%+Trifloxystrobin 25% WG (Nativo®), Metalaxyl 8%+Mancozeb 64%WP (Ridomil G old MZ) and Carboxin 37.5% +Thiram 37.5% (Vitavax); biological antagonists viz., Trichoderma asperellum, T. viride, T. harzianum and Pseudomonas fluorescens and organic amendments viz., neem cake, groundnut cake and mustard cake, along with enriched farmyard manure, individually and in integration for effective management of Fusarium wilt on the susceptible red-gram variety PRG-176.

Result: In vitro screening revealed that around 100% inhibition of Fusarium mycelial growth was achieved by the fungicide combination tebuconazole 50% + trifloxystrobin 25% WG, while others showed moderate inhibition (60-78%). Among bioagents, Trichoderma asperellum (OKT-G) recorded the highest antagonistic activity (89.78% inhibition) with least mean radial growth (9.2 mm). Among organic amendments, Neem cake at 10% concentration gave the greatest inhibition (68.85%). Under pot trials the integrated module T3 (seed treatment @10 g/kg + soil application @ 2.5 g/kg of T. asperellum (OKT-G) with enriched FYM @ 100 kg/ha + neem cake @ 250 kg/ha) achieved the lowest disease incidence (16.7%), highest disease reduction (61.5%) and maximum yield (12.24 q/ha), outperforming other treatments. Present study demonstrates that combining soil health-based amendments offers an effective and sustainable strategy for managing fusarium wilt in red gram. The findings with field validation through multi location trails may provide a basis for adoption in puls­e based cropping systems.

INTRODUCTION

Leguminous crops are the principal element in food legume systems. They not only contribute plant-based protein, enhance soil fertility through the promotion of biological nitrogen fixation, but also help to maintain sustainable agriculture in marginal environments. Among the legumes, red gram (Cajanus cajan L.) is widely cultivated in the tropical and subtropical regions of the world and is one of the most important legumes in India, accounting for a major portion of total production. Its adaptability, nutritional value and yield are highly hampered due to a variety of diseases; among these, wilt caused by Fusarium spp. is one of the most serious (Sharma et al., 2016; Behera et al., 2020).
       
The pathogen Fusarium spp. infects red gram through the root system, colonizes its vascular tissue and blocks the transport of water and nutrients, leading to a sudden wilting, chlorosis and death of the plant (Upadhyay and Rai, 1983). Infected plants may lose up to 50% or more yield, particularly when infection occurs before pod formation. The pathogen persists for long periods in crop residues and soil as chlamydospores, making its control even more challenging. The epidemiology of fusarium wilt in crops is closely linked with the soil environment like soil temperature, moisture, texture as well as soil physico-chemical parameters such as pH, organic carbon, nutrient status and electrical conductivity influencing the pathogen’s saprophytic survival and host root colonization (Sharma et al., 2010; Yan et al., 2023). Increased soil temperature and poor drainage in Vertisols of semi-arid tropics often exacerbate Fusarium disease expression, highlighting the need for soil-based integrated strategies (Gaur et al., 2010).
       
Recent insights on plant disease management emphasizes ecological management aligning with sustainable intensification goals. In this context, the role of beneficial soil microbiota in disease suppression, where Trichoderma, Bacillus and actinobacteria improve soil enzymatic activity, nutrient cycling and systemic resistance in legumes (Sathya et al., 2016; Chandar et al., 2016; Choudhary et al., 2023; Attia et al., 2025) is in current research. Control of fusarium wilt in red gram has traditionally been banked on fungicide seed-treatment or soil-drenching, but repeated fungicide application is not only costly but may lead to the development of resistance influencing the ecological context of the soil borne pathogen (You et al., 2020). Various biological control agents have shown promising results in controlled conditions, including Trichoderma spp. (for example, T. viride, T. harzianum and T. asperellum), achieving high levels of inhibition of Fusarium spp. in vitro and under pot conditions (Mukhopadhyay and Kumar, 2020). Trichoderma asperellum suppressed maize stalk rot by Fusarium graminarium by antibiosis mechanisms (Li et al., 2016). Organic amendments, such as neem cake, have also provided effective control through the release of bioactive compounds and the enhancement of soil microbial communities, which may reduce fungal populations and disease incidence in crops infected by Fusarium oxysporum (Khanna et al., 2024; Haque et al., 2025).
       
An integrated disease management approach combining biocontrol agents (Trichoderma spp. and Pseudomonas fluorescens), botanicals, fungicides and organic amendments effectively manage complex pathosystems in legumes. By addressing the complex host-pathogen-soil interactions, this strategy not only reduces disease incidence but also enhances beneficial microbial populations leading to improved yield, offering a sustainable and eco-friendly solution (Chandar et al., 2016; Behera et al., 2024; Khanna et al., 2024). Molecular characterization has revealed significant pathogenic variability among Fusarium spp. isolates across India (Naik et al., 2022), necessitating region-specific integrated management protocols rather than uniform recommendations. The integration of resistant cultivars with biological seed treatments and organic soil amendments provides synergistic and durable disease control while simultaneously improving soil fertility and crop profitability aligning with India’s National Mission on Sustainable Agriculture (NMSA) goals for eco-friendly plant protection (Singh and Gupta, 2024).
       
Regardless of the extensive research on Fusarium wilt management there remains inadequate evidence on the combined efficacy of Trichoderma asperellum, neem cake and enriched FYM under controlled soil conditions.

The present study, therefore, aims to evaluate integrated management strategies for optimizing wilt suppression and yield in red gram. The present study aimed at two objectives: (i) to evaluate the in vitro efficacy of some fungicides, bio-agents and organic amendments against Fusarium spp. and (ii) to examine an integrated management module comprising seed treatment, soil application of bio-agent with enriched FYM and neem-cake amendment on fusarium wilt incidence and yield of red gram under pot conditions. The work is expected to lead to the development of sustainable, soil health-based disease management strategies for red gram production systems.

MATERIALS AND METHODS

Experimental site and pathogen isolation
 
The present investigation was carried out at department of Plant Pathology, College of Agriculture, Odisha University of Agriculture and Technology (OUAT), Bhawanipatna-766001 during the year 2024-2025. The controlled pot experiments were carried out during the Rabi season on the susceptible red-gram variety PRG-176 to evaluate the efficacy of different management strategies against Fusarium oxysporum causing wilt in red gram (Cajanus cajan L.). The pathogen was isolated from infected red gram plants showing typical wilt symptoms and purified using the single spore technique on (concentration 2 × 106 / ml) Potato Dextrose Agar (PDA) medium. Pure cultures were maintained on PDA slants at 4oC for further studies. Molecular characterization of the pathogen revealed it to be Fusarium oxysporum, Accession Number PX606568.
 
In vitro evaluation of chemical fungicides
 
The efficacy of different fungicides was evaluated against Fusarium oxysporum using the poisoned food technique (Nene and Thapliyal, 1979). PDA medium was poisoned with each fungicide at concentrations of 0.1%, 0.15% and 0.2% and poured into sterilized Petri plates (20 ml plate-1). Mycelial discs (5 mm diameter) from 6-7-day-old cultures were inoculated at the centre of each plate. Control plates contained PDA without fungicides. Plates were incubated at 25±2oC and radial mycelial growth was recorded at 24 h intervals until the control plates reached the edge (Dhingra and Sinclair, 1995; Nene and Thapliyal, 2001). The percent inhibition (PI) of mycelial growth was calculated using the formula (Vincent, 1947):

 
Where
A = Colony diameter in control (mm).
B = Colony diameter in treatment plates (mm).
Each treatment was replicated thrice.
 
In vitro evaluation of bioagents
 
The antagonistic efficacy of selected fungal and bacterial bioagents was assessed using the dual culture method (Dennis and Webster, 1971). Trichoderma spp. were grown on PDA and bacterial isolates were cultured on nutrient agar. A 5 mm disc of Fusarium culture was placed on one side of the Petri plate and the antagonist was placed or streaked on the opposite side. Control plates contained only the pathogen. All treatments were replicated four times and incubated at 28±1oC for 8 days. The per cent inhibition over control was computed using Vincent’s (1947) formula.

 
Where,
PI = Per cent inhibition over control.
C = Mycelial growth of pathogen in control (mm).
T = Mycelial growth of pathogen in dual culture plate (mm).
 
In vitro evaluation of organic amendments
 
Three organic amendments viz., groundnut cake, neem cake and mustard cake were evaluated at 5% and 10% concentrations using the poisoned food technique. Each cake (100 g) was finely powdered, air-dried and extracted in 100 ml methanol for 24 h with intermittent shaking. Extracts were filtered and incorporated into molten sterilized PDA (5 ml extract + 95 ml PDA for 5% concentration).
       
After solidification, 5 mm mycelial discs of Fusarium oxysporum were inoculated at the plate centre and incubated at 25±2oC for 7-10 days. The radial growth and percent inhibition were calculated as per Vincent (1947). Control plates contained PDA without any amendment. Each treatment was replicated thrice.
 
Pot experiment
 
A pot experiment was conducted during Rabi 2024-25 to evaluate the combined efficacy of chemical fungicides, bioagents and organic amendments on wilt incidence and yield under artificially inoculated conditions.
 
Soil preparation and inoculation
 
A sterilized potting mixture of soil: Sand: FYM (2:1:1) was prepared and autoclaved for 1 hour on two consecutive days. Pots (5 kg capacity) were filled with the sterilized mixture.
       
The susceptible red gram variety ‘PRG-176’ was sown during the first week of October 2024. Each pot received an inoculum of Fusarium oxysporum (9 g pot-1) near the root zone at 21 days after sowing, followed by irrigation.
 
Treatment details
 
Seven treatments were tested in a completely randomized design (CRD) with three replications, including bioagents (Trichoderma asperellum, Pseudomonas fluorescens), organic amendments (neem cake) and fungicides (Carboxin + Thiram, Metalaxyl + Mancozeb, Tebuconazole + Trifloxystrobin).  Application modes included seed treatment (ST), soil application (SA) and soil drenching (SD).
 
Seed treatment (ST)
 
Seeds were surface sterilized with 0.1% HgCl2, soaked in bioagent suspension (5 × 10v  conidia ml-1) for 2 h and air-dried before sowing. For fungicide treatments, Carboxin + Thiram 75% WP was used at 2 g kg-1 seed.
 
Soil application (SA)
 
Bioagents (T. asperellum or P. fluorescens) were mixed with FYM at 2.5 kg/ha and incubated at 40% moisture for 15 days before incorporation. Neem cake 250 kg/ha was also incorporated before sowing where applicable.
 
Soil drenching (SD)
 
Bioagent formulations were applied as drenches thrice at 10-day intervals after inoculation (T. asperellum 100 ml plant-1, P. fluorescens 20 ml L-1). For chemical fungicides, Metalaxyl + Mancozeb (2 g L-1) and Tebuconazole + Trifloxystrobin (1 ml L-1) were similarly applied.
 
Observations and data recording
 
Per cent disease Incidence (PDI) was recorded at 60 and 90 days after sowing using the formula:

 
Statistical analysis
 
Data were analyzed using completely randomized design (CRD) as per Fisher’s method. OPSTAT was used of analysis of variance (ANOVA). The significance of differences among treatments was tested using the F-test at 5% probability level.

RESULTS AND DISCUSSION

In vitro evaluation of different fungicides against Fusarium oxysporum
 
The in vitro analyses demonstrated a concentration dependent inhibitory effect of all three fungicidal combinations on Fusarium oxysporum growth (Table 1, Fig 1). The triazole-strobilurin combination (Tebuconazole 50% + Trifloxystrobin 25% WG) achieved complete suppression (100% inhibition) at all test concentrations, indicating superior efficacy. Whereas, Metalaxyl 8% + Mancozeb 64% WP and Carboxin 37.5% + Thiram 37.5% WS achieved moderate inhibition levels (69-78% and 60-66% respectively). Similar results were recorded during the field trials (2018-2019) in northern India by Mohiddin et al. (2021) showing that fungicide combinations azoxystrobin + difenoconazole (9.19%) and azoxystrobin + tebuconazole (10.40%) performed markedly better over mancozeb + carbendazim (27.61%) against rice blast management. The markedly greater performance of the triazole-strobilurin combination likely stems from its dual-mode of action, the triazole interferes with ergosterol biosynthesis (cell membrane disruption) and the strobilurin inhibits mitochondrial respiration (electron transport blockage). This synergy enhances pathogen suppression even at lower concentrations (Shcherbakova, 2019). The chemical control approach although potent their repeated application risks the emergence of resistant pathogen strains and may carry non-target and environmental consequences. Thus, it is noteworthy that reliance solely on chemical control is not a long-term sustainable strategy for wilt management in red gram.

Table 1: In vitro efficacy of fungicides against Fusarium oxysporum.



Fig 1: In vitro efficacy of fungicides against radial growth of Fusarium oxysporum seven days after inoculation for various treatment codes (T1-T10).


 
In vitro evaluation of bioagents against Fusarium oxysporum
 
The bioagent experiments (Table 2; Fig 2) demonstrated significant antagonism against Fusarium oxysporum with Trichoderma asperellum (OKT-G) showing the highest suppression (89.78% inhibition; radial growth 9.20 mm), followed by T. viride (86.00%), T. harzianum (82.97%) and Pseudomonas fluorescens (53.89%).

Table 2: In vitro efficacy of organic amendments against Fusarium oxysporum.



Fig 2: In vitro efficacy of bioagents against radial growth of Fusarium oxysporum seven days after inoculation where, T1: Trichoderma asperellum OKT-G; T2: Trichoderma viride; T3: Trichoderma harzianum; T4: Pseudomonas fluorescens; T5: Control.


       
The superior performance of Trichoderma spp. is consistent with well-documented mechanisms including rapid substrate colonization, mycoparasitism, production of hydrolytic enzymes (e.g., chitinases, β-1,3-glucanases), secretion of secondary antifungal metabolites and induction of host resistance. Greenhouse experiments integrating transcriptomic and metabolomic analyses conducted by Zhang et al., (2022) revealed that Trichoderma asperellum M45a effectively colonized watermelon roots and enhanced defense responses against Fusarium oxysporum f. sp. niveum (FON) through increased resistance-related enzyme activities and upregulated defense genes such as MYB and PAL. Their reports on KEGG analysis displayed enrichment of phenylpropanoid pathways networked to lignin and cinnamic acid synthesis, supporting plant immunity. Their work on Metabolomic profiling ascertained four enriched pathways with upregulated metabolites, particularly galactinol and urea, which correlated positively with Trichoderma. In planta assays on wheat variety Khiar by Saadaoui et al., (2023) showed that Trichoderma seed coating significantly enhanced biomass, chlorophyll and nitrogen content in wheat plants. All strains exhibited bioprotective effects against Fusarium culmorum, with Th01 being most effective. Transcriptome analysis revealed activation of SA- and JA-dependent defense genes in roots and leaves, indicating robust growth promotion and disease resistance potential in modern wheat.
       
Apart from the robust antagonistic potential of Trichoderma asperellum in suppressing vascular fungal pathogens its ecological compatibility and potential to improve plant health beyond mere pathogen suppression make it a beneficial component for sustainable wilt control.
 
In vitro evaluation of organic Amendments against Fusarium oxysporum
 
Present study (Table 3) revealed that neem cake at 10% concentration recorded the greatest reduction in radial growth (28.00 mm; 68.89% inhibition), followed by 5% neem cake (61.85%) and then groundnut and mustard cakes. The agar well diffusion assay by Muthukumar et al. (2023) also revealed significant variation in the antifungal efficacy of different oilcake filtrates against Fusarium oxysporum f. sp. cepae. They reported that Neem cake filtrate showed the highest inhibition zone (1.29 cm at 15% concentration), followed by groundnut cake (1.36 cm at 30%). The lowest inhibition (2.28 cm) was recorded with pungam cake at 15% concentration, indicating its comparatively weaker antifungal activity. These findings are consistent with prior work on organic-amendment mediated suppression of soil-borne pathogens in pulses. The doctoral research by Appa, (2017) also demonstrated that Neem cake has one of the strongest antifungal effect and highest improvement in soil microbial activity and nutrient content, leading to enhanced plant vigor and lower disease incidence. The inhibitory effect likely derives from release of bioactive compounds (e.g., azadirachtin, limonoids) and enhanced soil microbial antagonism, which together create an unfavourable environment for Fusarium. Field trials by Tiyagi et al. (2001) showed that oil-seed cakes of neem, castor, linseed, groundnut, mustard and duan significantly reduced populations of plant-parasitic nematodes and pathogenic fungi in lentil and mungbean, while enhancing beneficial fungi like Trichoderma viride. Neem cake was most effectual, improving plant growth, chlorophyll content, nodulation and fertility. Also, the residual benefits persisted in the subsequent mungbean crop.

Table 3: In vitro efficacy of bioagents against Fusarium oxysporum.


 
Efficacy of fungicides, biocontrol agents and organic amendments under pot conditions
 
The pot experiment further validated the integrated management strategy (Table 4). Among all treatments, T3, where, seed treatment @10 g/kg along with soil application @ 2.5 g/kg of T. asperellum (OKT-G) with enriched FYM @100 kg/ha and neem cake @ 250 kg/ha  recorded the lowest per cent disease incidence (16.67%), highest disease reduction (61.54%) and highest mean yield (12.24 q ha-1). This treatment performed very close to the completely chemical treatment T6 (seed treatment with Carboxin + Thiram + soil drench with Tebuconazole + Trifloxystrobin: PDI 20.00%, disease reduction 53.84%, yield 1120 kg/ha. Also, the comparable performance of the integrated biologically-oriented treatment (T3) and the chemical treatment (T6) suggests that IDM can deliver comparable disease suppression while posing benefits in terms of sustainability, soil health and reduced chemical dependency.

Table 4: Efficacy of fungicides, biocontrol agents and organic amendments against wilt disease under pot conditions.


       
The study by Akhter et al. (2015) identified Rhizoctonia solani isolate RS10 as most virulent and Trichoderma harzianum isolate T-3 as the most effective antagonist, inhibiting 77.22% mycelial growth. Bavistin 50 WP and Provax-200 entirely inhibited fungal growth, with Provax-200 displaying high compatibility with T. harzianum. Mustard oilcake delivered maximum inhibition (60.28%). Integrated use of T. harzianum T-3, mustard oilcake and Provax-200 considerably lessened seedling mortality and enhanced pea yield over single or dual treatments.
       
Chandar et al. (2016) evaluated the antagonistic efficacy of Trichoderma viride, T. harzianum and Pseudomonas fluorescens against Fusarium oxysporum f. sp. ciceri in vivo conditions. P. fluorescens showed the highest inhibition of mycelial growth, followed by T. harzianum and T. viride. Seed treatment with P. fluorescens most effectively reduced chickpea wilt incidence. Application of organic amendments farmyard manure, vermicompost and mustard cake enhanced disease suppression and improved antagonist populations in soil. Among these, mustard cake proved most effective in streng-thening biocontrol activity and disease control efficiency.
       
Study by Khanna et al. (2024) evaluated plant extracts, fungicides and bio-agents under laboratory and field conditions against Fusarium oxysporum f. sp. ciceris severely affecting chickpea by causing wilt. Neem leaf extract showed the highest inhibition of pathogen growth in vitro, followed by datura and garlic. Field seed treatment with neem and datura extracts (10%) reduced wilt incidence by up to 39% and improved seed yield by about 7%. Among fungicides, carbendazim 50 WP was most effective, followed by azoxystrobin 23 SC, reducing disease incidence by over 85% with significant yield gains. Trichoderma viride and T. harzianum were the most effective bio-agents, supporting integrated management of chickpea wilt.
       
Field studies (2022-2024) by Haque et al. (2025) revealed that soil application of Trichoderma harzianum enriched neem cake was most effective in managing Fusarium oxysporum f. sp. ciceris (FOC), reducing wilt severity by 69% and FOC population by 60%. This treatment also enhanced plant growth by 22-24% and yield by 17-23%. Treatments combining T. harzianum with carbendazim (½ dose) or neem cake reduced wilt by 52-55% and increased yield by 12-15%. The study highlights neem cake as a potential substrate for mass-multiplying Trichoderma spp., offering a sustainable, eco-friendly approach for chickpea wilt management.

CONCLUSION

The integrated application of chemical, biological and organic approaches for the management of Fusarium wilt in red gram (Cajanus cajan L.) was effectively evidenced through the present study. In vitro screening confirmed the strong antagonistic potential of Trichoderma asperellum (OKT-G) and the suppressive effects of organic amendment, neem cake offering an eco-sustainable strategy for the management of Fusarium wilt in red gram (Cajanus cajan L.).
       
Future studies should also be directed toward optimization of dosage, timing and formulation of Trichoderma-enriched organic substrates for scale-up application, testing their compatibility with eco-friendly fungicides and long-term impacts on soil health and microbial dynamics. This integrated approach holds promise and could be used to develop low-cost, farmer-adoptable and environmentally responsible disease management modules for pulse-based cropping systems.

ACKNOWLEDGEMENT

Work is M.Sc. (Ag.) thesis of Ms. Anandita Sahoo, under supervision of Dr. Srujani Behera, Assistant Professor (Plant Pathology), conducted at department of Plant Pathology, College of Agriculture, Bhawanipatna-766001, Odisha University of Agriculture and Technology, Bhubaneswar-751003. The authors immensely thank Dean, College of Agriculture, OUAT, Bhawanipatna for rendering necessary facilities and for their moral support during the time of investigation.
 
Disclaimers
 
The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
 
Ethics approval
 
(N/A).
 
Availability of data and material
 
The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

CONFLICT OF INTEREST

The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

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  24. Singh, A. and Gupta, A. (2024). Food Security Through Sustainable Agriculture: A Prospective Study in the Indian Context. In: Sustainability and Health Informatics: A Systems Approach to Address the Climate Action Induced Global Challenge Singapore: Springer Nature Singapore. (pp. 155-182). 

  25. Singh, S.K., Singh, R.H. and Dutta, S. (2002). Integrated management of pigeonpea wilt by biotic agents and biopesticides. Ann. Plant Prot. Sci. 10: 323-326. 

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    Integrated Management of Fusarium Wilt in Red Gram (Cajanus cajan L.) Through Chemical, Biological and Organic Approaches under Controlled Conditions

    A
    Anandita Sahoo1
    P
    Priyanka Choudhary2
    N
    Nirakar Ranasingh3
    U
    Uttam Kumar Behera4
    J
    Jay Prakash Singh5
    C
    Chinmayee Mohapatra6
    P
    Priti Upadhyay7
    S
    Srujani Behera3,*
    0000-0001-7764-9631
    1Department of Plant Pathology, Odisha University of Agriculture and Technology, Bhubaneswar-751 003, Odisha, India.
    2Department of Mycology and Plant Pathology, Institute of Agricultural Sciences, B.H. University, Varanasi- 221 005, Uttar Pradesh, India.
    3Department of Plant Pathology, College of Agriculture, Odisha University of Agriculture and Technology, Bhawanipatna-766 001, Odisha, India.
    4Seed Technology Research, AICRP on Seeds (Crops), Odisha University of Agriculture and Technology, Bhubaneswar-751 003, Odisha, India.
    5Department of Plant Pathology, Faculty of Agriculture, SMMTD College, Ballia-277 001, Uttar Pradesh, India.
    6Faculty of Agriculture, Sri Sri University, Arilo, Bidhyadharpur, Cuttack-754 006, Odisha, India.
    7Division of Vegetable Science, ICAR- Indian Agricultural Research Institute, New Delhi-110 001, India.

    • Submitted26-11-2025|

    • Accepted28-04-2026|

    • First Online 12-05-2026|

    • doi 10.18805/LR-5610

    ABSTRACT

    Background: Red gram or pigeon pea (Cajanus cajan L.) is a major pulse crop cultivated in tropical and subtropical regions, whose productivity is hampered by the wilt from a soil-borne fungus, Fusarium oxysporum.

    Methods: In vitro studies as well as controlled pot experiments were conducted during 2024-2025 that evaluated chemical fungicides viz., Tebuconazole50%+Trifloxystrobin 25% WG (Nativo®), Metalaxyl 8%+Mancozeb 64%WP (Ridomil G old MZ) and Carboxin 37.5% +Thiram 37.5% (Vitavax); biological antagonists viz., Trichoderma asperellum, T. viride, T. harzianum and Pseudomonas fluorescens and organic amendments viz., neem cake, groundnut cake and mustard cake, along with enriched farmyard manure, individually and in integration for effective management of Fusarium wilt on the susceptible red-gram variety PRG-176.

    Result: In vitro screening revealed that around 100% inhibition of Fusarium mycelial growth was achieved by the fungicide combination tebuconazole 50% + trifloxystrobin 25% WG, while others showed moderate inhibition (60-78%). Among bioagents, Trichoderma asperellum (OKT-G) recorded the highest antagonistic activity (89.78% inhibition) with least mean radial growth (9.2 mm). Among organic amendments, Neem cake at 10% concentration gave the greatest inhibition (68.85%). Under pot trials the integrated module T3 (seed treatment @10 g/kg + soil application @ 2.5 g/kg of T. asperellum (OKT-G) with enriched FYM @ 100 kg/ha + neem cake @ 250 kg/ha) achieved the lowest disease incidence (16.7%), highest disease reduction (61.5%) and maximum yield (12.24 q/ha), outperforming other treatments. Present study demonstrates that combining soil health-based amendments offers an effective and sustainable strategy for managing fusarium wilt in red gram. The findings with field validation through multi location trails may provide a basis for adoption in puls­e based cropping systems.

    INTRODUCTION

    Leguminous crops are the principal element in food legume systems. They not only contribute plant-based protein, enhance soil fertility through the promotion of biological nitrogen fixation, but also help to maintain sustainable agriculture in marginal environments. Among the legumes, red gram (Cajanus cajan L.) is widely cultivated in the tropical and subtropical regions of the world and is one of the most important legumes in India, accounting for a major portion of total production. Its adaptability, nutritional value and yield are highly hampered due to a variety of diseases; among these, wilt caused by Fusarium spp. is one of the most serious (Sharma et al., 2016; Behera et al., 2020).
           
    The pathogen Fusarium spp. infects red gram through the root system, colonizes its vascular tissue and blocks the transport of water and nutrients, leading to a sudden wilting, chlorosis and death of the plant (Upadhyay and Rai, 1983). Infected plants may lose up to 50% or more yield, particularly when infection occurs before pod formation. The pathogen persists for long periods in crop residues and soil as chlamydospores, making its control even more challenging. The epidemiology of fusarium wilt in crops is closely linked with the soil environment like soil temperature, moisture, texture as well as soil physico-chemical parameters such as pH, organic carbon, nutrient status and electrical conductivity influencing the pathogen’s saprophytic survival and host root colonization (Sharma et al., 2010; Yan et al., 2023). Increased soil temperature and poor drainage in Vertisols of semi-arid tropics often exacerbate Fusarium disease expression, highlighting the need for soil-based integrated strategies (Gaur et al., 2010).
           
    Recent insights on plant disease management emphasizes ecological management aligning with sustainable intensification goals. In this context, the role of beneficial soil microbiota in disease suppression, where Trichoderma, Bacillus and actinobacteria improve soil enzymatic activity, nutrient cycling and systemic resistance in legumes (Sathya et al., 2016; Chandar et al., 2016; Choudhary et al., 2023; Attia et al., 2025) is in current research. Control of fusarium wilt in red gram has traditionally been banked on fungicide seed-treatment or soil-drenching, but repeated fungicide application is not only costly but may lead to the development of resistance influencing the ecological context of the soil borne pathogen (You et al., 2020). Various biological control agents have shown promising results in controlled conditions, including Trichoderma spp. (for example, T. viride, T. harzianum and T. asperellum), achieving high levels of inhibition of Fusarium spp. in vitro and under pot conditions (Mukhopadhyay and Kumar, 2020). Trichoderma asperellum suppressed maize stalk rot by Fusarium graminarium by antibiosis mechanisms (Li et al., 2016). Organic amendments, such as neem cake, have also provided effective control through the release of bioactive compounds and the enhancement of soil microbial communities, which may reduce fungal populations and disease incidence in crops infected by Fusarium oxysporum (Khanna et al., 2024; Haque et al., 2025).
           
    An integrated disease management approach combining biocontrol agents (Trichoderma spp. and Pseudomonas fluorescens), botanicals, fungicides and organic amendments effectively manage complex pathosystems in legumes. By addressing the complex host-pathogen-soil interactions, this strategy not only reduces disease incidence but also enhances beneficial microbial populations leading to improved yield, offering a sustainable and eco-friendly solution (Chandar et al., 2016; Behera et al., 2024; Khanna et al., 2024). Molecular characterization has revealed significant pathogenic variability among Fusarium spp. isolates across India (Naik et al., 2022), necessitating region-specific integrated management protocols rather than uniform recommendations. The integration of resistant cultivars with biological seed treatments and organic soil amendments provides synergistic and durable disease control while simultaneously improving soil fertility and crop profitability aligning with India’s National Mission on Sustainable Agriculture (NMSA) goals for eco-friendly plant protection (Singh and Gupta, 2024).
           
    Regardless of the extensive research on Fusarium wilt management there remains inadequate evidence on the combined efficacy of Trichoderma asperellum, neem cake and enriched FYM under controlled soil conditions.

    The present study, therefore, aims to evaluate integrated management strategies for optimizing wilt suppression and yield in red gram. The present study aimed at two objectives: (i) to evaluate the in vitro efficacy of some fungicides, bio-agents and organic amendments against Fusarium spp. and (ii) to examine an integrated management module comprising seed treatment, soil application of bio-agent with enriched FYM and neem-cake amendment on fusarium wilt incidence and yield of red gram under pot conditions. The work is expected to lead to the development of sustainable, soil health-based disease management strategies for red gram production systems.

    MATERIALS AND METHODS

    Experimental site and pathogen isolation
     
    The present investigation was carried out at department of Plant Pathology, College of Agriculture, Odisha University of Agriculture and Technology (OUAT), Bhawanipatna-766001 during the year 2024-2025. The controlled pot experiments were carried out during the Rabi season on the susceptible red-gram variety PRG-176 to evaluate the efficacy of different management strategies against Fusarium oxysporum causing wilt in red gram (Cajanus cajan L.). The pathogen was isolated from infected red gram plants showing typical wilt symptoms and purified using the single spore technique on (concentration 2 × 106 / ml) Potato Dextrose Agar (PDA) medium. Pure cultures were maintained on PDA slants at 4oC for further studies. Molecular characterization of the pathogen revealed it to be Fusarium oxysporum, Accession Number PX606568.
     
    In vitro evaluation of chemical fungicides
     
    The efficacy of different fungicides was evaluated against Fusarium oxysporum using the poisoned food technique (Nene and Thapliyal, 1979). PDA medium was poisoned with each fungicide at concentrations of 0.1%, 0.15% and 0.2% and poured into sterilized Petri plates (20 ml plate-1). Mycelial discs (5 mm diameter) from 6-7-day-old cultures were inoculated at the centre of each plate. Control plates contained PDA without fungicides. Plates were incubated at 25±2oC and radial mycelial growth was recorded at 24 h intervals until the control plates reached the edge (Dhingra and Sinclair, 1995; Nene and Thapliyal, 2001). The percent inhibition (PI) of mycelial growth was calculated using the formula (Vincent, 1947):

     
    Where
    A = Colony diameter in control (mm).
    B = Colony diameter in treatment plates (mm).
    Each treatment was replicated thrice.
     
    In vitro evaluation of bioagents
     
    The antagonistic efficacy of selected fungal and bacterial bioagents was assessed using the dual culture method (Dennis and Webster, 1971). Trichoderma spp. were grown on PDA and bacterial isolates were cultured on nutrient agar. A 5 mm disc of Fusarium culture was placed on one side of the Petri plate and the antagonist was placed or streaked on the opposite side. Control plates contained only the pathogen. All treatments were replicated four times and incubated at 28±1oC for 8 days. The per cent inhibition over control was computed using Vincent’s (1947) formula.

     
    Where,
    PI = Per cent inhibition over control.
    C = Mycelial growth of pathogen in control (mm).
    T = Mycelial growth of pathogen in dual culture plate (mm).
     
    In vitro evaluation of organic amendments
     
    Three organic amendments viz., groundnut cake, neem cake and mustard cake were evaluated at 5% and 10% concentrations using the poisoned food technique. Each cake (100 g) was finely powdered, air-dried and extracted in 100 ml methanol for 24 h with intermittent shaking. Extracts were filtered and incorporated into molten sterilized PDA (5 ml extract + 95 ml PDA for 5% concentration).
           
    After solidification, 5 mm mycelial discs of Fusarium oxysporum were inoculated at the plate centre and incubated at 25±2oC for 7-10 days. The radial growth and percent inhibition were calculated as per Vincent (1947). Control plates contained PDA without any amendment. Each treatment was replicated thrice.
     
    Pot experiment
     
    A pot experiment was conducted during Rabi 2024-25 to evaluate the combined efficacy of chemical fungicides, bioagents and organic amendments on wilt incidence and yield under artificially inoculated conditions.
     
    Soil preparation and inoculation
     
    A sterilized potting mixture of soil: Sand: FYM (2:1:1) was prepared and autoclaved for 1 hour on two consecutive days. Pots (5 kg capacity) were filled with the sterilized mixture.
           
    The susceptible red gram variety ‘PRG-176’ was sown during the first week of October 2024. Each pot received an inoculum of Fusarium oxysporum (9 g pot-1) near the root zone at 21 days after sowing, followed by irrigation.
     
    Treatment details
     
    Seven treatments were tested in a completely randomized design (CRD) with three replications, including bioagents (Trichoderma asperellum, Pseudomonas fluorescens), organic amendments (neem cake) and fungicides (Carboxin + Thiram, Metalaxyl + Mancozeb, Tebuconazole + Trifloxystrobin).  Application modes included seed treatment (ST), soil application (SA) and soil drenching (SD).
     
    Seed treatment (ST)
     
    Seeds were surface sterilized with 0.1% HgCl2, soaked in bioagent suspension (5 × 10v  conidia ml-1) for 2 h and air-dried before sowing. For fungicide treatments, Carboxin + Thiram 75% WP was used at 2 g kg-1 seed.
     
    Soil application (SA)
     
    Bioagents (T. asperellum or P. fluorescens) were mixed with FYM at 2.5 kg/ha and incubated at 40% moisture for 15 days before incorporation. Neem cake 250 kg/ha was also incorporated before sowing where applicable.
     
    Soil drenching (SD)
     
    Bioagent formulations were applied as drenches thrice at 10-day intervals after inoculation (T. asperellum 100 ml plant-1, P. fluorescens 20 ml L-1). For chemical fungicides, Metalaxyl + Mancozeb (2 g L-1) and Tebuconazole + Trifloxystrobin (1 ml L-1) were similarly applied.
     
    Observations and data recording
     
    Per cent disease Incidence (PDI) was recorded at 60 and 90 days after sowing using the formula:

     
    Statistical analysis
     
    Data were analyzed using completely randomized design (CRD) as per Fisher’s method. OPSTAT was used of analysis of variance (ANOVA). The significance of differences among treatments was tested using the F-test at 5% probability level.

    RESULTS AND DISCUSSION

    In vitro evaluation of different fungicides against Fusarium oxysporum
     
    The in vitro analyses demonstrated a concentration dependent inhibitory effect of all three fungicidal combinations on Fusarium oxysporum growth (Table 1, Fig 1). The triazole-strobilurin combination (Tebuconazole 50% + Trifloxystrobin 25% WG) achieved complete suppression (100% inhibition) at all test concentrations, indicating superior efficacy. Whereas, Metalaxyl 8% + Mancozeb 64% WP and Carboxin 37.5% + Thiram 37.5% WS achieved moderate inhibition levels (69-78% and 60-66% respectively). Similar results were recorded during the field trials (2018-2019) in northern India by Mohiddin et al. (2021) showing that fungicide combinations azoxystrobin + difenoconazole (9.19%) and azoxystrobin + tebuconazole (10.40%) performed markedly better over mancozeb + carbendazim (27.61%) against rice blast management. The markedly greater performance of the triazole-strobilurin combination likely stems from its dual-mode of action, the triazole interferes with ergosterol biosynthesis (cell membrane disruption) and the strobilurin inhibits mitochondrial respiration (electron transport blockage). This synergy enhances pathogen suppression even at lower concentrations (Shcherbakova, 2019). The chemical control approach although potent their repeated application risks the emergence of resistant pathogen strains and may carry non-target and environmental consequences. Thus, it is noteworthy that reliance solely on chemical control is not a long-term sustainable strategy for wilt management in red gram.

    Table 1: In vitro efficacy of fungicides against Fusarium oxysporum.



    Fig 1: In vitro efficacy of fungicides against radial growth of Fusarium oxysporum seven days after inoculation for various treatment codes (T1-T10).


     
    In vitro evaluation of bioagents against Fusarium oxysporum
     
    The bioagent experiments (Table 2; Fig 2) demonstrated significant antagonism against Fusarium oxysporum with Trichoderma asperellum (OKT-G) showing the highest suppression (89.78% inhibition; radial growth 9.20 mm), followed by T. viride (86.00%), T. harzianum (82.97%) and Pseudomonas fluorescens (53.89%).

    Table 2: In vitro efficacy of organic amendments against Fusarium oxysporum.



    Fig 2: In vitro efficacy of bioagents against radial growth of Fusarium oxysporum seven days after inoculation where, T1: Trichoderma asperellum OKT-G; T2: Trichoderma viride; T3: Trichoderma harzianum; T4: Pseudomonas fluorescens; T5: Control.


           
    The superior performance of Trichoderma spp. is consistent with well-documented mechanisms including rapid substrate colonization, mycoparasitism, production of hydrolytic enzymes (e.g., chitinases, β-1,3-glucanases), secretion of secondary antifungal metabolites and induction of host resistance. Greenhouse experiments integrating transcriptomic and metabolomic analyses conducted by Zhang et al., (2022) revealed that Trichoderma asperellum M45a effectively colonized watermelon roots and enhanced defense responses against Fusarium oxysporum f. sp. niveum (FON) through increased resistance-related enzyme activities and upregulated defense genes such as MYB and PAL. Their reports on KEGG analysis displayed enrichment of phenylpropanoid pathways networked to lignin and cinnamic acid synthesis, supporting plant immunity. Their work on Metabolomic profiling ascertained four enriched pathways with upregulated metabolites, particularly galactinol and urea, which correlated positively with Trichoderma. In planta assays on wheat variety Khiar by Saadaoui et al., (2023) showed that Trichoderma seed coating significantly enhanced biomass, chlorophyll and nitrogen content in wheat plants. All strains exhibited bioprotective effects against Fusarium culmorum, with Th01 being most effective. Transcriptome analysis revealed activation of SA- and JA-dependent defense genes in roots and leaves, indicating robust growth promotion and disease resistance potential in modern wheat.
           
    Apart from the robust antagonistic potential of Trichoderma asperellum in suppressing vascular fungal pathogens its ecological compatibility and potential to improve plant health beyond mere pathogen suppression make it a beneficial component for sustainable wilt control.
     
    In vitro evaluation of organic Amendments against Fusarium oxysporum
     
    Present study (Table 3) revealed that neem cake at 10% concentration recorded the greatest reduction in radial growth (28.00 mm; 68.89% inhibition), followed by 5% neem cake (61.85%) and then groundnut and mustard cakes. The agar well diffusion assay by Muthukumar et al. (2023) also revealed significant variation in the antifungal efficacy of different oilcake filtrates against Fusarium oxysporum f. sp. cepae. They reported that Neem cake filtrate showed the highest inhibition zone (1.29 cm at 15% concentration), followed by groundnut cake (1.36 cm at 30%). The lowest inhibition (2.28 cm) was recorded with pungam cake at 15% concentration, indicating its comparatively weaker antifungal activity. These findings are consistent with prior work on organic-amendment mediated suppression of soil-borne pathogens in pulses. The doctoral research by Appa, (2017) also demonstrated that Neem cake has one of the strongest antifungal effect and highest improvement in soil microbial activity and nutrient content, leading to enhanced plant vigor and lower disease incidence. The inhibitory effect likely derives from release of bioactive compounds (e.g., azadirachtin, limonoids) and enhanced soil microbial antagonism, which together create an unfavourable environment for Fusarium. Field trials by Tiyagi et al. (2001) showed that oil-seed cakes of neem, castor, linseed, groundnut, mustard and duan significantly reduced populations of plant-parasitic nematodes and pathogenic fungi in lentil and mungbean, while enhancing beneficial fungi like Trichoderma viride. Neem cake was most effectual, improving plant growth, chlorophyll content, nodulation and fertility. Also, the residual benefits persisted in the subsequent mungbean crop.

    Table 3: In vitro efficacy of bioagents against Fusarium oxysporum.


     
    Efficacy of fungicides, biocontrol agents and organic amendments under pot conditions
     
    The pot experiment further validated the integrated management strategy (Table 4). Among all treatments, T3, where, seed treatment @10 g/kg along with soil application @ 2.5 g/kg of T. asperellum (OKT-G) with enriched FYM @100 kg/ha and neem cake @ 250 kg/ha  recorded the lowest per cent disease incidence (16.67%), highest disease reduction (61.54%) and highest mean yield (12.24 q ha-1). This treatment performed very close to the completely chemical treatment T6 (seed treatment with Carboxin + Thiram + soil drench with Tebuconazole + Trifloxystrobin: PDI 20.00%, disease reduction 53.84%, yield 1120 kg/ha. Also, the comparable performance of the integrated biologically-oriented treatment (T3) and the chemical treatment (T6) suggests that IDM can deliver comparable disease suppression while posing benefits in terms of sustainability, soil health and reduced chemical dependency.

    Table 4: Efficacy of fungicides, biocontrol agents and organic amendments against wilt disease under pot conditions.


           
    The study by Akhter et al. (2015) identified Rhizoctonia solani isolate RS10 as most virulent and Trichoderma harzianum isolate T-3 as the most effective antagonist, inhibiting 77.22% mycelial growth. Bavistin 50 WP and Provax-200 entirely inhibited fungal growth, with Provax-200 displaying high compatibility with T. harzianum. Mustard oilcake delivered maximum inhibition (60.28%). Integrated use of T. harzianum T-3, mustard oilcake and Provax-200 considerably lessened seedling mortality and enhanced pea yield over single or dual treatments.
           
    Chandar et al. (2016) evaluated the antagonistic efficacy of Trichoderma viride, T. harzianum and Pseudomonas fluorescens against Fusarium oxysporum f. sp. ciceri in vivo conditions. P. fluorescens showed the highest inhibition of mycelial growth, followed by T. harzianum and T. viride. Seed treatment with P. fluorescens most effectively reduced chickpea wilt incidence. Application of organic amendments farmyard manure, vermicompost and mustard cake enhanced disease suppression and improved antagonist populations in soil. Among these, mustard cake proved most effective in streng-thening biocontrol activity and disease control efficiency.
           
    Study by Khanna et al. (2024) evaluated plant extracts, fungicides and bio-agents under laboratory and field conditions against Fusarium oxysporum f. sp. ciceris severely affecting chickpea by causing wilt. Neem leaf extract showed the highest inhibition of pathogen growth in vitro, followed by datura and garlic. Field seed treatment with neem and datura extracts (10%) reduced wilt incidence by up to 39% and improved seed yield by about 7%. Among fungicides, carbendazim 50 WP was most effective, followed by azoxystrobin 23 SC, reducing disease incidence by over 85% with significant yield gains. Trichoderma viride and T. harzianum were the most effective bio-agents, supporting integrated management of chickpea wilt.
           
    Field studies (2022-2024) by Haque et al. (2025) revealed that soil application of Trichoderma harzianum enriched neem cake was most effective in managing Fusarium oxysporum f. sp. ciceris (FOC), reducing wilt severity by 69% and FOC population by 60%. This treatment also enhanced plant growth by 22-24% and yield by 17-23%. Treatments combining T. harzianum with carbendazim (½ dose) or neem cake reduced wilt by 52-55% and increased yield by 12-15%. The study highlights neem cake as a potential substrate for mass-multiplying Trichoderma spp., offering a sustainable, eco-friendly approach for chickpea wilt management.

    CONCLUSION

    The integrated application of chemical, biological and organic approaches for the management of Fusarium wilt in red gram (Cajanus cajan L.) was effectively evidenced through the present study. In vitro screening confirmed the strong antagonistic potential of Trichoderma asperellum (OKT-G) and the suppressive effects of organic amendment, neem cake offering an eco-sustainable strategy for the management of Fusarium wilt in red gram (Cajanus cajan L.).
           
    Future studies should also be directed toward optimization of dosage, timing and formulation of Trichoderma-enriched organic substrates for scale-up application, testing their compatibility with eco-friendly fungicides and long-term impacts on soil health and microbial dynamics. This integrated approach holds promise and could be used to develop low-cost, farmer-adoptable and environmentally responsible disease management modules for pulse-based cropping systems.

    ACKNOWLEDGEMENT

    Work is M.Sc. (Ag.) thesis of Ms. Anandita Sahoo, under supervision of Dr. Srujani Behera, Assistant Professor (Plant Pathology), conducted at department of Plant Pathology, College of Agriculture, Bhawanipatna-766001, Odisha University of Agriculture and Technology, Bhubaneswar-751003. The authors immensely thank Dean, College of Agriculture, OUAT, Bhawanipatna for rendering necessary facilities and for their moral support during the time of investigation.
     
    Disclaimers
     
    The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
     
    Ethics approval
     
    (N/A).
     
    Availability of data and material
     
    The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

    CONFLICT OF INTEREST

    The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

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