Revolutionizing Garden Pea Yield: A Synergistic Strategy with Plant Extracts, Bio-control Agents and Fungicides for Root Rot Disease Management and Optimal Productivity

Pea, a prominent vegetable crop mainly grow in cooler regions globally, holds a substantial place in Indian economiy, In India, the menace of root rot disease poses a grave threat to pea cultivation, especially in the IIIrd A zone of Rajasthan, causing detrimental effects on the initial plant growth. This perilous disease is primarily caused by Fusarium solani (Mart.) Appel and Wolleweber f. sp. pisi (E. R. John) Snyder and Hansen and it remains a pervasive issue for pea (Pisum sativum L.) cultivation on a global scale. Fusarium root rot tends to be most devastating when the soil temperatures range between 26° to 27°C. Additionally, a scarcity of water exacerbates the severity of root rot Riley et al., (2016). Notzably, the disease manifests in field in patches Sinha et al., (2018).
       
The impact of this disease on pea crops is severe, affecting all stages of growth and resulting in staggering yield losses of up to 97 per cent due to this devastating disease El-Saadony et al., (2021). Moreover, the disease has made its presence felt in the Kashmir Valley, with incidence rates ranging from 14.8 to 64.7 per cent Nazir et al., (2022). Fusarium species, particularly F. solani and F. oxysporum, are the primary causal organism behind this damaging disease Chatterton et al., (2019).
       
Among fungal infections, Fusarium solani f. sp. pisi induced root rot poses a significant challenge, particularly due to its soil-borne nature. Therefore, this study has been initiated, aiming to design a more feasible and enduring strategy to combat Fusarium solani f. sp. pisi induced root rot, finally extenuating the losses endured by pea cultivators. This study aspires to offer practical insights into disease management, striving to enhance pea crop resilience and minimize the detrimental effects of root rot. By aligning with the study's fundamental goal, the research activities to contribute substantially to agricultural practices, ensuring a sustainable and prosperous future for pea cultivation while combatting the persistent challenge of Fusarium solani f. sp. pisi induced root rot.

Source of isolates
 
The isolates were taken from Rajasthan Agricultural Research Institute, Durgapura, Jaipur. A field experiment conducted at Rajasthan Agricultural Research Institute, Jaipur, Rajasthan, in the Rabi seasons of 2021-22 and 2022-23.
 
Efficacy of plant extract against F. solani f. sp. pisi
 
In this experiment, 9×12-inch sterilized clay pots were filled with autoclaved soil and each pot was inoculated with the pathogen. To do this, we mixed the top 5 cm of soil in each pot with the pathogen’s inoculum, which was previously multiplied on cereal grains (sorghum) at 25 g per pot.
       
Healthy plant parts (leaves, stems, cloves, rhizomes) were collected, washed and crushed with distilled water (100 g in 100 ml, w/v). The resulting extract was filtered through muslin cloth, yielding 100% concentration. This was further diluted 10% using distill water. Seeds soaked individually in a 10% plant extract solution (neem, giloy, turmeric, tulsi, clove and eucalyptus) for 30 minutes and sown at 10 seeds per pot. The experiments was conducted in CRD with three repetitions.
 
Efficacy of bio-agents against F. solani f. sp. pisi under greenhouse conditions
 
In this test, seeds were individually treated with bio-agents (namely Trichoderma harzianum, Trichoderma viride, Pseudomonas fluorescens and Bacillus subtilis) at a rate of 10 g/kg of seeds and then sown separately in pots. The experiment followed a Completely Randomized Design. A Control was also maintained which contained surface sterilized seeds without bio-agents. Pots were watered as needed, kept under steady conditions and root rot observations were recorded up to 60 days after sowing (DAS).
 
Efficacy of fungicide against F. solani f. sp. pisi
 
In this experiment, seeds received individual treatment with a fungicide (tebuconazole, azoxystrobin, hexaconazole, carboxin+thiram, tebuconazole+trifloxystrobin and carbendazim+mancozeb) at a rate of 2 gm/kg and 0.2 %/kg of seeds and were then sown separately in pots at a rate of 10 seeds per pot. The study utilized a completely randomized design (CRD) with three repetitions. Root rot incidence was carefully noted for 60 days after sowing (DAS).
 
Details of field experiment
 
A field experiment conducted at Rajasthan Agricultural Research Institute, Jaipur, Rajasthan, in the Rabi seasons of 2021-22 and 2022-23 aimed to develop a sustainable strategy to gather valuable insights and observations throughout the growing seasons to combat root rot in pea crops, specifically using the susceptible variety Arkel. Field experiments used a Randomized Block Design with three replicate plots of 3m×2m measurements and plants spaced 30cm×10cm apart. Organic soil amendments, were mixed into the soil two weeks prior to seed sowing. Following this, the soil was deliberately inoculated with a 25g inoculum /square meter at a depth of 5-8 cm to increase disease pressure. For seed treatment, bio-control agents (T. harzianum and T. viride) were individually cultured on Potato Dextrose Agar (PDA) followed by harvesting the sporulating colonies by suspending them in 20 ml of water in each Petri plate. This suspension was then mixed with 10 g of sterilized talc powder to form a slurry. This slurry was applied to the seeds at a rate of 10 g/kg. The treated seeds were placed in a moist chamber overnight to facilitate the establishment of the antagonists on them. For fungicide (tebuconazole, tebuconazole+trifloxystrobin) and plant extract (clove and neem) seed treatments, 0.2 % and 10% concentration employed respectively. The seeds were soaked separately in these solutions for 30 minutes, air-dried in the shade and then sown. Observations were meticulously recorded for root rot incidence and green pod yield and detailed data is available in Table 1.
 

 
The per cent disease incidence was calculated by using the following formula:
 

Effect of different plant extracts against root rot disease
 
In green house condition, all plant extracts, demonstrated significant superiority over the control. The data revealed that treating the seeds with clove extract at a concentration of 10% resulted in the lowest disease incidence (29.31%), followed by neem extract (30.84%) (Table 2).
 

       
This superior effect of clove aqueous extract can be due to the presence of mainly phenolic compounds and their derivatives reported by Shan et al., (2005). Eugenol is the main bioactive compound of clove, Neveu et al., (2010) known for its ability to inhibit fungal growth and spore production. The aqueous extract of Azadirachta indica inhibits the growth of soil borne pathogens, indicating the presence of antifungal substances in the plant material Mondall et al., (2009). A similar conclusion, highlighting the in vitro effectiveness of various plant extracts, including Azadirachta indica, in controlling brinjal wilt fungal pathogens Babu Joseph et al., (2008). seed treatment with extracts of neem at 10% concentration were most effective against F. oxysporum Khanna et al., (2021).
 
Effect of different bio-agents against root rot disease
 
The minimum disease incidence percentage (19.27%) was recorded in the case of T. harzianum, closely followed by T. viride (21.38%). Additional details can be found in Table 3. Trichoderma harzianum demonstrated significant interactions with the activities of the pathogen’s phenol oxidizing enzymes Chakraborty et al., (2007). Notably, a reduction in the activities of phenol oxidizing enzymes was observed after 21 days of incubation when plants were treated with both the pathogen and the antagonist. The potential of Trichoderma harzianum in managing diseases caused by R. solani has been demonstrated in mothbean cultivation Godara and Singh (2021).
 

 
Effect of different fungicides against root rot disease
 
The data highlighted that seed treatment with tebuconazole + trifloxystrobin resulted in the lowest percentage of disease incidence (7.57%). Our findings of green house and field experiment are consistent with the results of Attri et al., (2019) Fusarium wilt of bell pepper was also managed by using fungicides and mycelial growth of pathogen was significantly inhibited by all tested fungicide. Carbendazim and tebuconazole + trifloxystrobin effectively control root rot of pea reported by Anita and Ratnoo (2015) Table 4.
 

 
Field experiment under in vivo conditions
 
The application of plant extracts, bio-agents and fungicides combined with neem cake resulted in a significant reduction in root rot incidence across all years, as depicted in Table 5. The findings at 60 days after sowing indicated the lowest disease incidence, with 6.74% and 9.32% root rot incidence during 2022 and 2023, respectively, observed in seed treatment with tebuconazole  + trifloxystrobin, along with soil application of Neem cake. Leading to an impressive 80.71% reduction in root rot incidence compared to untreated control plots. Following closely, seed treatment with tebuconazole, coupled with soil application of neem cake, was identified as the next best option, reducing root rot incidence by 75.65% compared to the untreated control plots.
 

       
The results at 90 days after sowing clearly indicate that the lowest disease incidence, at 7.68% and 10.41% root rot incidence during 2022 and 2023, respectively, was observed in seed treatment involving tebuconazole + trifloxystrobin, along with soil application of Neem cake. Following closely, seed treatment with tebuconazole, combined with soil application of neem cake, proved to be the next best option, reducing root rot incidence by 78.51% compared to the untreated control plots.
       
It was evident that the maximum pod yield of 75.80 q/ha was observed in seed treatment involving tebuconazole + trifloxystrobin, combined with soil application of neem cake. Following closely, seed treatment with tebuconazole, coupled with soil application of neem cake, yielded 75.09 q/ha of pods.
       
Further analyzing the pod yield data, the maximum increase in green pod yield was observed with seed treatment involving tebuconazole 25%+ trifloxystrobin 50% WG at 0.1% concentration, combined with soil application of neem cake at 5q/ha, resulting in a 93.77% increase. Tebuconazole is categorized as triazole fungicides which fungicides operate as inhibitors of demethylase (DMI) Amoghavarsha et al., (2021). It disrupt the fungal cell wall formation process, ultimately impeding the fungus’s ability to reproduce and grow. The recently introduced trifloxystrobin, has been shown to inhibit broad spectrum fungi Margot et al., (1998). The evaluated fungicides showed high activity against Fusarium in vitro and demonstrated great efficacy for reducing the disease in the greenhouse trial Ezrari et al., (2022). Trifloxystrobin belongs to the strobilurin class of fungicides which possess improved safety and environmental characteristics Neem cake was highly effective in controlling crown rot of cucumber under in-vivo conditions, further validating our result Nehra et al., (2020). Neem cake proved to be the most effective treatment, achieving the highest disease reduction rates of 79.15% and 70.07% against root rot of clusterbean Shivran et al., (2023).

In recent years, the cultivation of peas has witnessed a decline primarily due to the significant constraint of root rot disease caused by Fusarium solani f. sp. pisi. Our study highlighted the high effectiveness of T. harzianum as bio-agents, as well as clove extracts and tebuconazole + trifloxystrobin in inhibiting disease incidence under greenhouse conditions. All the promising treatments incorporating an organic amendment demonstrated superiority in reducing disease incidence and promoting higher pod yield. Notably, neem cake applied at 5q/ha emerged as the most effective, showcasing minimal disease incidence and significantly enhancing pod yield compared to treatments involving mustard cake at 5q/ha.

On behalf of all authors of this manuscript, I, [Yogita Nain], declare that none of the authors have any conflicts of interest that could have influenced the work reported in this manuscript. All authors have agreed to this submission and have approved the final manuscript. There are no financial, personal, or professional relationships that could be perceived as potential conflicts of interest.