Soil nematodes are abundant in the rhizosphere, showing a specific affinity with different crops. The nematodes are microscopic roundworms with an unsegmented body plan and a diverse mode of life in various ecological conditions. The genera Aphelenchus, Pratylenchus, Hoplolaimus and Tylenchorhynchus are ample in most of the soils and in terms of trophic mode, the free-living nematodes, Aphelenchus, Basiria and other beneficial nematodes play an essential role for the sustaining soil health and as natural predators of soil inhabiting insect pests.
       
Among the 10 most important plant parasitic nematodes, root knot nematodes (Meloidogyne spp.) are more abundant worldwide, affecting the major crops (Jones et al., 2013). Root-knot nematodes (RKN) are the most economically significant plant pathogens, causing substantial yield loss along with other plant pathogens (Topalović and Geisen, 2023). The plant growth promoting microbes (PGPM) secrete antibiosis, induced systemic resistance (ISR) in plants that may inhibit the nematode infection (Poveda et al., 2020). They produce secondary metabolites with nematicidal effects, like serine protease PR1 and antimicrobial peptides (Forghani and Hajihassani, 2020). Several Trichoderma species have recognized for their nematicidal activity, notably T. longibrachiatum, T. viridae, T. harzianum, T. hamatum, T. atroviridae, T. koningi (Yao et al., 2023; Eman et al., 2023). Their fermentation products have shown considerable inhibition of egg hatching and juvenile mortality in various nematode pests of economic importance. T. harzianum FB10, during the egg parasitism, the chitinase genes chi 18-5 and chi 18-12 are significantly upregulated, leading to increased chitinase content, which is crucial for the breakdown of nematode eggshells and facilitating egg destruction (Baazeem et al., 2021).
       
Plant-parasitic nematodes have developed resistance to conventional market-available nematicides, rendering them ineffective under actual field conditions, particularly across large areas. These nematicides do not have any label claim also. Therefore, this study assessed the impact of seed treatment with a new strain of T. asperellum (ITCC 12187.25) (@ 1×10⁸ CFUs/g) on rhizosphere soil nematode populations in green gram, black gram (kharif) and chickpea and field pea (rabi season). The evaluation was conducted in IPM and farmers’ practice (FP) fields in the Bundelkhand region of Uttar Pradesh-a region lacking prior studies of this nature-representing technology-driven impact research.

Study schedule
 
The study was carried out under IPM programme by ICAR-National Research Institute of Integrated Pest Management, New Delhi With the collaboration of the Division of Nematology, IARI, New Delhi and KVK Jalaun and IIPR Kanpur in pulse crops in two districts, Jhansi and Jalaun (Bundelkhand, U.P.), adopting two blocks in each district, during 2024-25 and the results of IPM were compared with FP (Farmers Practices, which were the traditional farming tactics). The eleven villages comprising six villages of Bangra and Bamaur blocks, (La. 25.6 -25.3°E and Lo. 79.0-79.1°N, dist. Jhansi) and five villages of the block, Jalaun and Dakore (La. 26.07-26.09°E, Lo. 79.4-79.5°N, dist. Jalaun) were selected for the study. Two IPM modules were followed, as IPM-1: applying T. asperellum (ITCC 12187.25) seed treatment (Trichogel, 1×10¹² CFUs/g, NRIIPM strain in form of gel formulation @ 1-2/kg seed). Another IPM module as IPM-2 was implemented in KVKs farm that was without any seed treatment, while other tactics were similar to the IPM-1. The findings of IPM-1 were compared with IPM-2 and FP fields. The IPM strategies included, field preparation, variety selection and other eco-friendly tactics for insect pest management in the pulse crops i.e. chickpea (cv JG-14), green gram (cv Shikha, IPM 410-03), black gram (cv PU-35) and field pea (cv IPFD-10-12). The studies were conducted during Rabi season (sampling during Dec-Jan 2023-24) in chickpea and field pea crop fields and similarly, during Kharif season (sampling during Aug., 2023) on green gram and black gram field at flowering periods.
 
Soil sampling
 
The soil samples were collected from three places of the three selected fields of IPM-1, IPM-2 and three of FP fields from each village by a zigzag walking design. Thus, a total 11 samples from IPM-1 and 11 from FP fields were collected for the particular crop. Three composite samples of IPM-2 were collected separately from both the KVK farms. The samples were carried to the laboratory for analysis.
 
Extraction of soil nematodes
 
The nematodes were extracted by Cobb’s decanting and sieving technique described by Southey (1986). Modified Baermanns funnel method was followed for the extraction of nematodes. The population density was recorded under a stereoscope microscope. 
 
Soil physicochemical properties
 
Standard procedures were applied for soil physicochemical analysis such as, the maximum water-holding capacity (WHC), more commonly used method, organic matter (titration method), soil available phosphorus (Olsen and Sommers, 1982) and soil available nitrogen (Keeney and Nelson 1982). Flame photometer was used for the determination of potassium content and other cations. The soil pH and electrical conductivity were estimated by a pH meter. The weekly maximum and minimum temperatures, relative humidity and rainfall were recorded throughout the experimental periods at Jhansi and Jalaun, locations.
 
Statistical analysis
 
Data were analysed by the paired T-test for each field of IPM and FPs to determine the significant effect of the treatment on soil-inhabiting (rhizosphere) nematodes in different pulse crops. Significance was defined at (5%), p≤0.05 using Microsoft Excel (2016) to find out the P-values for all the samples. Per cent reduction in the number of nematodes in IPM-1 as compared to IPM-2 and FP fields were calculated using the following formula: 

Per cent reduction in IPM-1 in comparison of IPM-2 =

Per cent reduction in IPM-1 in comparison of FP =

 
Finally the average of above both reductions of nematode population in IPM-1 were compiled and designated in figures (Fig 1-8).

Soil physicochemical properties
 
Soil physical structure viz., percentage of sand, silt and clay indicated a slightly silt-loam soil with black colour at the KVK Jhansi. Whereas at the KVK Jalaun soil is a little bit sandy loam having 43.7±2.3% sandy share. Relatively, the pH value was more at KVK Jalaun than in area of KVK Jhansi. Contrarily, the greater values of cation exchange capacity (CEC) and water holding capacity (WHC) and organic C (%) were recorded at the Jhansi location. Moreover the soil available nutrients, available nitrogen (N), available phosphorus (P) and potassium were also recorded at both locations and variations were noted to some extent (Table 1). Few studies suggest a negative correlation between soil pH and total nematode abundance, meaning as pH increases, nematode populations tend to decrease. A pH range of 5-7, is considered favourable for nematode growth. Soil compaction can negatively affect nematode populations by reducing pore space, aeration and root growth. Thus soil properties, comprising temperature, moisture and water holding capacity, pH and electrical conductivity, play an essential role for soil functioning and distribution of soil nematodes (Castro et al., 1990). The chemical composition of soil affects population density of nematode communities (Cardoso et al., 2012). Additionally, soil type affects the composition of nematode species. Soil salinity and salt accumulation are other significant factors, which govern weather changes, irrigation and plant fertilization. Organic matter (OM) is a fundamental soil component that serves as a source of carbon and energy for soil fauna and flora. Soil OM also affects root infestation by nematodes (Al-Ghamdi, 2021).


 
Population densities of soil nematodes in selected pulse crop fields
 
Chickpea
 
At the location near KVK Jhansi, the number of Aphelenchus was found 15.7±2.6, 19.7±1.5 and 19.3±2.1/100 cc soil in IPM-1, IPM-2 and FP fields respectively. Similarly, the number of Basiria was observed 17.3±2.5 in IPM-1, 19.3±1.8/100 cc soil in IPM-2 and 18.6±2.1/100 cc soil in FP fields. There were no significant (p≤0.05) variations in population densities of Aphelenchus and Basiria between IPM-1, IPM-2 and FP fields (Fig 1). No incidence of Meloidogyne was found in all IPM-1, IPM-2 and FP fields. The significant suppression in the population of other genera such as, Pratylenchus, Rotylenchulus, Tylenchorynchus and Tylenchus was also recorded in IPM-1 fields as compared to IPM-2 and FP fields and the number of all four above genera ranged from 9.0±2.0 to 159.0±7.0/100 cc soil in IPM-1 fields. Whereas in IPM-2 and FP fields it ranged from 19.3±1.5 to 311.3±55.3 and 21.3±5.8 to 298.7±37.5/100 cc soil respectively. Herein, 46-114% reduction of the nematodes was observed in IPM-1 fields as compared to the IPM-2 and FPs (Fig 1).    
       
Relatively, the lesser number of Aphelenchus was found in all IPM-1, IPM-2 and FPs at KVK Jalaun. In IPM-1 fields, the number of Aphelenchus was observed as 13.3±4.2/100 cc soil, while it was 15.0±2.0 and 13.7±6.1/100 cc soil in IPM-2 and FP fields respectively, indicating no significant (p≤0.05) difference among all the fields. The population densities of the genera Boleodorus, Filenchus, Helicotylenchus, Hoplolaimus, Meloidogyne Pratylenchus, Rotylenchulus, Tylenchorynchus and Tylenchus ranged from 4.3±1.5 to 126.7±25.1/100 cc soil in IPM-1 fields, whereas in IPM-2 and FP fields it varied from 14.3±3.1 to 369.0±58.0/100 cc soil, indicating a significant suppression of the nematode population from 61.0% to >200% in IPM-1 fields at both the KVKs (Fig 2). Hence, it can be interpreted that IPM-1 can consistently reduce the nematode population across both locations and several reductions were statistically different (p≤0.05), highlighting the effectiveness of T. asperellum (ITCC 12187.25) seed treatment. Trichoderma increased the nodulation, decreased the root knot nematode gall index and gave higher yields in chickpea compared to the control when applied (Singh et al., 2020; Joshi and Sunkad, 2025).
 
Green gram
 
A total 12-genera of the nematodes were detected in green gram fields at the selected locations. No incidence of Meloidogyne was found at KVK Jhansi, while its incidence was observed at KVK Jalaun. The number of galls in green gram roots was 5.0±1.0, 15.0±4.2 and 20.0±4.7/100 cc soil IPM-1, IPM-2 and in FP fields, thus a significant (p≤0.05) reduction in Meloidogyne incidence (>100%) was found in IPM-1 fields. Contrarily, the population of other nematodes such as, Boleodorus, Helicotylenchus,  Hoplolaimus, Pratylenchus, Rotylenchulus, Scutellonema, Tylenchorhynchus  and  Tylenchus were found suppressed (18-98%) in IPM-1 than IPM-2 and FP fields. The number of these nematodes ranged from 14.0±3.0 to 139.3±11.1/100 cc soil in IPM-1 and 20.7±2.1 to 247.3±39.0/100 cc soil in IPM-2 and FP fields, indicating a significant reduction in nematode population in IPM-1 as compared to IPM-2 and FPs (Fig 3). But more interestingly, significant suppression in Meloidogyne was observed at the Jalaun location. The number of galls was 5.0±1.0, 15.0±4.2 and 20.0±4.7/plant in IPM-1, IPM-2 and FP fields respectively, which specified a significant suppression of Meloidogyne incidence in IPM-1 fields (Fig 4). Mukhtar et al. (2021) recorded a plant growth promoter impact of T. harzianum in increasing the shoot length and root length by 45.5% and 45.1% respectively. Notably, the application decreased the number of galls (46%) and eggs (53%) and reduced the nematode reproductive factor by 78.0% indicating the strong suppression of M. javanica populations in green gram.
 
Black gram
 
There was no incidence of Meloidogyne at KVK Jhansi, but it was recorded at KVK Jalaun. The number of galls in black gram roots was 6.6±1.8/plant in IPM-1, 17.0±3.3/plant in IPM-2 and 10.3±3.5/plant in FP fields, showing a significantly (p≤0.05) higher Meloidogyne incidence in IPM-2 and FP fields at KVK Jalaun. In the Jhansi area, the number of Aphelenchus was recorded as 21.0±2.6, 24.7±3.2 and 25.0±1.0/100 cc soil in IPM-1, IPM-2 and FP fields. As well, the population of Basiria was also not affected in the treated fields as compared to IPM-2 and FP fields (Fig 5). The number of galls of root knot nematodes in the roots of black gram was found to be 6.6±1.8 galls/plant, 17.0±3.3 galls/plant and 10.3±3.5 galls/plant in IPM-1, IPM-2 and FP fields respectively. Thus, a significant suppression in Meloidogyne incidence was observed in IPM-1 fields (157.6%). Findings also exhibited a successful suppression in population densities of other concerned ecto-and semi-endoparasitic nematodes; and their population densities varied from 11.3±3.1 to 123.3±6.4/100 cc soil in IPM-1 fields, which was significantly lesser than their IPM-2 and FP-fields viz., which ranged from 24.0±2.6 to 339.3±57.5/100 cc soil (Fig 6). 
 
Field pea
 
Studies were continued in field pea crop, harvested at the mature stage during Feb.-March. Vigorous plant growth was observed in the treated fields across all selected locations. In the Jhansi area, the Aphelenchus population was found 23.0±2.6 and 26.7±2.5/100 cc soil in IPM-1 and IPM-2 fields respectively, while in FP fields it was 26.3±4.5/100 cc soil. Similarly, the number of Basiria was recorded 15.0±0.5/100 cc soil in IPM-1 and 14.0±2.3/100 cc soil in IPM-2 fields (Fig 7). The incidence of Meloidogyne was observed at both the KVK locations in IPM and FP trials. The number of galls in IPM-1 fields was 6.0±1.7/plant compared to 12.3±1.5 galls/plant in IPM-2 and 11.7±4.0 galls/plant in FP fields, representing a 105% reduction in Meloidogyne incidence at KVK Jhansi. Comparatively, greater incidence of Meloidogyne was observed at KVK Jalaun such as, the number galls in IPM fields were 9.7±2.1 galls/plant and 15.3±1.5 galls/plant in FP fields that signified 57.7% reduction the incidence in IPM-1 fields. However, almost similar findings for Aphelenchus and Basiria were recorded in all the IPM-1, IPM-2 and FPs fields with no significant differences at the location of KVK Jalaun (Fig 8). 
 
Yield and economics of the selected crops
 
Approximately, similar yields of the selected crops could be achieved in IPM fields, as the standard levels of the corresponding varieties reported upon. Relatively in the scenario of yield increase (%) in IPM-1 and IPM-2 over FP fields in chickpea field was achieved up to 20.2% and 13.2% respectively. Likewise in green gram, 23.1% and 10.5% yield increase was achieved in IPM-1 and IPM-2 fields respectively over the FP fields (Table 2). Almost similar yield increase (%) was recorded in black gram and field pea crops with lesser input costs than the FPs (Table 3).  Comparatively, the total gross return was also found greater in IPM fields than the FPs, that relates to the more net return with greater B:C ratio in IPM fields. The B:C ratio in IPM-1 fields was found as 4.2, 3.9, 3.9 and 4.1 in chickpea, green gram, black gram and field pea respectively (Table 2 and 3). However, IPM-2 indicated slightly lesser B:C ratios than the IPM-1. Whose FP fields exhibited the B:C ratios as 2.9, 2.7, 2.8 and 2.9 in the consistent crops.




       
Trichoderma mycelium may attack on egg masses, juveniles (J2) and adults. The mycelium secrete enzymes that include proteases, cellulases, hemicellulases, chitinases and glucanases (Keswani et al., 2013). Castillo et al., (2008) reported Pratylenchus damage in patches showing stunting, chlorosis and necrosis in leaves and roots and reduced root growth and shoot weight in chickpea. Pratylenchus has also been reported earlier in chickpea grown in heavy soils from Bundelkhand (Singh, 2014) and Madhya Pradesh (Tiwari et al. 1992). Application of T. asperellum (ITCC 12187.25) indicated a significant suppression of the population of Pratylenchus in various selected pulse crops. De Oliveira et al. (2023) reported that the gene expression of T. harzianum exhibits a complex transcriptome response when interacting with the plant-parasitic nematode P. brachyurus and the T. harzianum genome were differentially expressed in the presence of P. brachyurus, in relation to the absence of the nematode. The population density of Hoplolaimus was found reduced in T. asperellum (ITCC 12187.25) seed treated fields might be due to reducing the number of juveniles, eggs and cysts in the soil. A significant reduction in the population of  Rotylenchulus was also recorded in the pulse crops. Likewise, Amin et al., (2002) pointed out the reduced population of  Rotylenchulus, semi-endoparasitic nematodes in sunflower fields after the T. asperellum (ITCC 12187.25) treatment.
       
The major concern of the study was to evaluate the incidence of root knot nematode in the pulse crops. In Bundelkhand, the pulses are grown mostly on black and clay soils, which leads to poor drainage and aeration that may be unfavourable for the survival of root-knot nematodes. No incidence of Meloidogyne was found in chickpea, green gram and black gram fields and little bit the incidence was recorded in field pea at the Jhansi location. However at the Jalaun location, the incidence of Meloidogyne was observed in all the selected crops, which could be managed significantly in IPM fields i.e. applying T. asperellum (ITCC 12187.25) as seed treatment. Likewise, Sharon et al. (2001) noticed the ability T. harzianum to attack and colonize eggs and egg masses of the root-knot nematode (M. javanica) under field conditions. The T. harzianum induces resistance towards RKN by increasing secondary metabolite synthesis and hastens the defence-related mechanism. The RKN infection increased the levels of reactive oxygen species (ROS; H₂O₂ and O₂>) and lipid peroxidation in tomato roots, which are the immune related defence mechanism of plants. The colonization of T. harzianum  significantly reduced the levels of ROS, malondialdehyde and electrolyte leakage, which was associated with increased accumulation of multiple secondary metabolites such as, flavonoids, phenols, lignin and cellulose 75/ days after inoculation with M. incognita (Yan et al., 2021). A similar report was pointed out towards the application T. harzianum along with soil organic amendment; reduced RKN infestation in chickpea and improved the plants growth parameters, nodulation and grain yield (Singh et al., 2020). Saharan et al., (2023) reported that application of T. asperellum FbMi6 enriched neem cake (1-ton ha^-1) increased the okra yield by 28.3% and decreased nematode population by 57.1% as compared with contro due to higher polyphenol content. Nevertheless, several reports mentioned that the Trichoderma spp. have lethal effects against  Meloidogyne sp. such as T. harzianum, T. koningii and T. asperellum  against M. incognita (Fan et al., 2020); T. harzianum, T. asperellum  and T. koningii against M. javanica (Elgorban et al., 2014), yet furthermore investigation is required in this direction. Also T. viride isolates that were derived from peptone + ammonium sulphate was found to be significantly effective on the nodule enumeration like number of nodule, nodule biomass, nutrient content in seed and straw and yield of chickpea (Chalie-U et al., 2025).

Seed treatment with T. asperellum (ITCC 12187.25) has indicated a significant control over all the endo-parasite, semi-endo-parasites, ecto-parasites and free-living nematodes. At the Jhansi location soil is black and clay loam, which is the least suitable for the growth and survival of soil nematodes. Whereas, at the location Jalaun relatively more population of the nematodes were recorded that may be due to the sandy loam soil at the Jalaun location. Moreover, investigation is needed in this line that would be an absolute direction to achieve the tedious objective for nematode management using PGP microbes. Herein, the asperellum (ITCC 12187.25) can be applied as an important tool in the form of gel formulation for nematodes along with other plant pathogen management in pulse crops.

The authors declare that they have no competing financial interests or personal relationships that could have influenced the work reported in this paper.