Compatibility of indigenous isolates HR1, HR2 of entomopathogenic nematodes, with low-toxicity insecticides for control of fall armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) and tomato leafminer, Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae)

Invasive species are a major danger to agronomic and natural ecosystems, and due to environmental concerns about pesticide use, EPNs have the potential to replace larvicidal action in pest management. The goal was to see how well local isolates of Steinernema feltiae (HR1) and Heterorhabditis bacteriophora (HR2) controlled invasive species when combined with low-toxicity pesticides. HR1 + Spinosad, chlorantraniliprole produced over 90% mortality in larvae at 96 hours, while HR2 + Spinosad, chlorantraniliprole caused over 95% mortality at 96 hours. After treatment, the high dose was regarded the least hazardous technique for controlling fall armyworm. At the high dose, HR1 + Spinosad, chlorantraniliprole produced larvae death of over 100 percent at 96 hours, and HR2 + Spinosad, chlorantraniliprole caused mortality of over 97.50 percent at 96 hours, and should be considered as a least hazardous strategy for T. absoluta management. Controlling larvae mortality of above 100% at 96 hours in combination with low-toxicity insecticide dosages should be included as a least harmful technique to control T. absoluta. The results showed that these HR2 strains have high pathogenicity against T. absoluta and S. frugiperda and have potential for control in integrated approaches, causing 100 percent and 90.00 percent mortality of T. absoluta and S. frugiperda at 96 hours at the high dose as a least toxic strategy to control.


Introduction
The biological control system creates an environment that is more tolerant of chemical control methods. When used properly, it can do a lot of good, last a long time, and kill a lot of insects. According to the instructions, entomopathogenic nematodes (EPNs) suppress these species: Coleoptera, Diptera, Hemiptera, Hymenoptera, Orthoptera, Lepidoptera, Siphonaptera, Thysanoptera, and Isoptera. In weed control, these pests interact with natural pesticides ( The most essential approach of FAW control is the use of chemical insecticides (Belay et al. 2012). However, this pest has been found to develop resistance to a wide range of commonly used pesticides (Zhu et al. 2015) as well as the chemical proteins of Bacillus thuringiensis Berlin (Bacillaceae) (Murua et al. 2019); as a result, safe, clean, and easy-to-manage control strategies are required and being developed. Biological control is a promising technique against a variety of pests, and ecologically friendly biopesticides are more effective than pesticides. Entomopathogenic nematodes (EPNs), like other biological control agents, are potential and promising pest control agents (Lacey and Georgis 2012; Bhairavi et al., 2021).
The Steinernematidae and Heterorhabditidae families of EPNs are natural insecticides (Abbas et al., 2021). Many researchers have indicated that EPN natural species are better at adapting to current environmental conditions than traditional problems (Bedding, 1990). Other employees did a study on the availability of S. frugiperda and other insect pests in EPNs (Viteri et al. 2018;.
Tomatoes (Solanum lycopersicum L.) are a South and Central American annual vegetable of the Solanaceae family. It rose to popularity as a result of its high food prices and consumption in the food sector (Canpolat, 2016;Golukcu et al. 2016). In India, tomatoes are the most productive vegetable on the planet. It is critical to safeguard its crops from diseases and pests because of their economic value. In locations where the climate is conducive to insects, red spider larvae, thrips, white ies, aphids, and leaf ies cause plant harm (Butut and Gocmen, 2000; Guncan et al. 2006). Invasive species pose a serious threat to the environment and agriculture. Tuta absoluta (Meyrick, 1917) (Lepidoptera: Gelechiidae), the tomato leafminer (TLM), is an invasive bug that has spread to many regions of the world in recent years, including India. T. absoluta larvae can penetrate and feed on all aerial parts of the host plant, including stems, leaves, shoots, and fruits (Desneux et al., 2010). Chemical management has been a prominent approach for pest control from its origins as an economic pest (Lietti et al., 2005). T. absoluta was rst discovered in India in October 2014 in the mid-western state of Maharashtra (Sridhar et al., 2014). Tomato pinworm / small leaf, T. Absoluta, an invasive new invasive insect, was rst discovered in Pune in 2014 in a tomato plant that grew in a eld and a thousand houses, and in southwestern India (Karnataka state) (ICAR 2015). To our knowledge, there have been no reports of this pest in eastern, northern, and eastern India. The insect was then discovered on a farmer's eld in major tomato-growing districts across the country, including Himachal Pradesh (Sharma and Gavkare, 2017). In response to carbon dioxide, insect movement, and insect faces, entomopathogenic nematodes (EPNs) are typically discovered in the surrounding soil (Kaya and Gaugler, 1993 More than 100 EPN species have been identi ed worldwide (about 80% are steinernematid), with at least 13 of them being marketed (Shapiro-Ilan et al. 2014). Inherent pathogenicity against various pest species varies widely among EPN species. Furthermore, the eld e cacy of EPNs can be determined by differences in host-seeking strategy and tolerance to environmental circumstances such as temperature and desiccation across EPN species (Martens et al. 2004). EPNs have been widely used in the biological control of a wide range of economically relevant pests in a variety of settings (Grewal et al. 2005). However, the use of adjuvants to increase leaf coverage and persistence of the IJs, or the formulation of EPNs to retard desiccation, has increased the usage of EPNs against foliar pests (

Insects
Under greenhouse circumstances, a T. absoluta colony was maintained on tomato plants. The colony was started with larvae collected in September 2020 from the vegetable science departmental form tomato greenhouse at Dr. Yashwant Singh Parmar University of Horticulture and Forestry, Solan, HP, India, which used local strains of S. feltiae (HR1) and H. bacteriophora (HR2) indigenous strains (Poinar) for pest control.

Entomopathogenic nematodes
In this investigation, two isolates of S. feltiae and H. bacteriophora were employed. They were utilised without culturing in the tests. G. mellonella larvae were used as nematode traps to extract the native isolate from soil samples collected in Rajgarh, Himachal Pradesh, India. This strain was cultivated on G. mellonella last instar larvae at 21 1°C using the technique (Kaya and Gaugler 1993). Infectious juveniles (IJs) were collected from white traps maintained at 4°C in distilled water for up to 14 days once they emerged during the rst ten days. Before being employed in the tests, the nematodes were acclimatised at room temperature for around 30 minutes.

Insect source
Larvae were taken from corn elds in the Department of Vegetable Science, Dr. YSPUHF, Nauni, Himachal Pradesh, India, to start the rst S. frugiperda colony. The young corn plant feed was then placed in test tubes measuring 15 cm in height and 1.5 cm in diameter. They were sexed and transported to PVC tubes 10 cm in diameter and 20 cm in height after transforming into pupae, with the tube extremities closed with a voile-type fabric and sealed with elastic. The tubes were lined with lter paper on the inside so that the females could oviposit shortly after the adults emerged. The moths were fed 10% honey isolates placed in a tube with cotton on a daily basis. The newly hatched caterpillars were placed in test tubes with the same arti cial diet after being sterilised Petri dishes with an arti cial diet were obtained.
TLM larvae and pupae were obtained from a greenhouse that had been invaded. T. absoluta was grown in a greenhouse at a temperature of 26°C, a relative humidity of 60%, and a photoperiod of L:D 8:16. The insects were raised on tomato plants in hardwood framed cages with 80 mesh organdy linen (S. lycopersicum L.). In the oviposition cage, the adults were fed a 10% sugar solution.

Methodology:
In a 20 mL plastic cup with a tiny amount of maize feed, one caterpillar was inserted (natural feed). Two biological agents, three insecticides, and an organic agent with low-toxic pesticides have their low-and high-dose labels (translated into lab dosage) registered (Table 1). A total of fteen larvae were given a topically applied pesticide solution of 200 litres each dose. Only distilled water was used for the control. The caught cups were housed in the lab at 18 to 20 ° C with a recording duration of 12:12 h in a randomly controlled 4-block construction (total n = 40 larvae per dosage) (L: D). Between 24-and 96-hours following application, mortality was measured. In different assays, the insecticide diagnosis of ¼x, ½x, x, ½y, and y insecticide was applied to 60 larvae per treatment to calculate the lethal doses (LD50) of chlorantraniliprole and Spinosad at 96 hours (x = dose low and y = maximum value is equal to the size registered and described in (Table 1) Table 1 To suppress fall armyworm and tomato leafminer larvae, researchers tested the e ciency of two biological agents, three synthetic insecticides, and the least toxic insecticide combinations. Combinations Manufacturers: DuPont (Coragen), Bayer (Indoxacarb), Syngenta (Spinosad).
Ten larvae were placed in an 8 cm petri dish with the natural diet of tomato leaves. Two biological agents, three synthetic insecticides, and the biological agent + low-toxicity insecticides were employed at low and high registered label dosages (converted to lab dosages) ( Table 1). Each dosage consisted of ten larvae given topically with 200 l of pesticide solution.
Only distilled water was used as a control. In the lab, treated Petri dishes were kept in a CRD design with four replications (total n = 40 larvae per dosage) at 18 to 20°C and photoperiod 12:12 h in a CRD design with four replications (total n = 40 larvae per dosage) (L:D). The mortality of larvae was measured between 24-and 96-hours following application.

Statistical analysis
The data was corrected for control larval mortality in the bioassays using Abbott's formula (Fleming and Retnakarn 1985), and PROBIT analysis was performed. LSD (P 0.05) values were also calculated to distinguish means between treatments.

Bioassays with S. feltiae + S. frugiperda + Insecticides
High values (F = 61.77; df = 12; P0.05) were associated with the highest percentage mortality rate (Table 2). S. feltiae and Spinosad, on the other hand, produced 80 to 97 percent worm death in 96 hours at low and high rates. At the greatest rate, chlorantraniliprole and Spinosad consumption resulted in a mortality rate of up to 97.50 percent in 96 hours. These results differ from those reported by Belay et al. (2012), who found that Spinosad and chlorantraniliprole cause more than 80% of worm fatalities at the same time. In this investigation, the LD50 for chlorantraniliprole was 1600 ppm and for Spinosad was 400 ppm for 96 hours. The differences could be due to (1) long-term use of these active chemicals, resulting in a speci c level of resistance, (2) the study's stage, third or fth stage, or (3) variances between island larvae (Viteri et al., 2018). Indoxacarb, on the other hand, killed 75 and 90 percent of worms at low and high rates, respectively, after 96 hours. When S. feltiae was coupled with Spinosad and chlorantraniliprole, the percentage of mortality was higher by 24 hours than when these pesticides were applied alone, regardless of the dose employed (Table 2). Furthermore, at 72 hours, a very signi cant proportion of worm mortality (over 90%) was identi ed in high doses. Similarly, when larvae were exposed to

Bioassays with H. bacteriophora + S. frugiperda + Insecticides
High rates were associated with the highest percentage mortality rate (F = 67.09; df = 12; P0.05) ( Table 3). At low and high 96-hour rates, however, H. bacteriophora and Spinosad killed 72.50 to 95 percent of the larvae. At the greatest rate, chlorantraniliprole and Spinosad consumption resulted in a mortality rate of up to 92.50 percent in 96 hours. These effects differ from those previously described, in which Spinosad and chlorantraniliprole together killed more than 95 percent of foxes. In this investigation, the LD50 for Spinosad was 400 ppm and for chlorantraniliprole was 1600 ppm for 96 hours.
The discrepancy could be due to (1) long-term usage of these active components, resulting in a certain level of resistance,  In comparison to the use of these insecticides alone, H. bacteriophora in combination with Spinosad and chlorantraniliprole resulted in a higher percentage of mortality by 24 hours, regardless of the dose utilised (Table 3)  In comparison to the use of these insecticides alone, S. feltiae in combination with Spinosad and chlorantraniliprole resulted in higher percentages of mortality at 24 hours, regardless of the dosage utilised (Table 4). Furthermore, high doses at 72 hours resulted in the highest percentages of larval mortality (above 95%). Similarly, when compared to the lowest mortality produced by Spinosad (20.50 percent) or S. feltiae (45.00 percent) applied alone, the combination of Spinosad + S. feltiae was successful (70.00 percent of dead larvae) at the maximum dose at 96 hours (Viteri et al. 2018). Septicemia (S. feltiae) + lysed midgut epithelial cells (Spinosad), disturbed muscle regulation (chlorantraniliprole), or Indoxacarb neural transmission (spinetoram) exposed larvae to two separate modes of action at the same time (Viteri et al. 2018). Armyworm numbers from Florida peak in the fall (Yu 1991). However, more research is needed to con rm that this synergistic impact exists.

Bioassays with H. bacteriophora + T. absoluta + Insecticides
In general, higher doses resulted in a higher mean percent mortality (F = 72.98; df = 12; P 0.05). (  In comparison to the use of these insecticides alone, H. bacteriophora in combination with Spinosad and chlorantraniliprole resulted in higher percentages of mortality at 24 hours, regardless of the dosage utilised (Table 5). Furthermore, high doses at 72 hours resulted in the highest percentages of larval mortality (above 95.00 percent). Similarly, when compared to the lowest mortality produced by Spinosad (40.50 percent) or Hb (65.00 percent) applied alone, the combination of Hb+ Spinosad was effective (100 percent of dead larvae) with the high dose at 96 hours (Viteri et al. 2018). Septicemia (H. bacteriophora) + lysed midgut epithelial cells (Hb), poor muscle control (Spinosad), or aberrant neural transmission (spinetoram) in larvae exposed to two separate modes of action at the same time, (Viteri et al. 2018) caused their greater mortality. Armyworm populations peak in the fall (Yu 1991). However, more research is needed to con rm that this synergistic impact exists.

Discussion
The study involved evaluation on the susceptibility of S. frugiperda and T. absoluta to 2 species of EPNs isolated from Himachal Pradesh, North-Western India. Extra exposure duration is linked to higher mortality because it allows for more The amount of the fall armyworm larva differed between different species of nematodes, which is not unusual. (Andalo et al., 2010;Viteri et al., 2018) found that when Heterorhabditis sp. and S. arenarium were used to treat

Conclusions
Our ndings suggest that local indigenous strains HR1 and HR2 were acceptable hosts for lepidopteran, Spodoptera frugiperda, and Tuta absoluta. The nematode completed its life cycle and produced a large number of IJs, indicating that it has signi cant biological control potential in IPM. To control larvae in bioassays, in combination with low-toxicity pesticides at low and high dosages. HR1 + Spinosad, chlorantraniliprole, and HR2 + Spinosad or chlorantraniliprole caused larvae mortality of over 90% and 100% at 96 hours at the high dose, respectively, and should be considered as a least toxic control method.