{"paper_id":"05b2e67c-317e-4e6b-935a-0efcdf2bdb8c","body_text":"Ecological Characterization and Efficacy of Indigenous Entomopathogenic Nematodes Against Spodoptera frugiperda in Nigeria | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Ecological Characterization and Efficacy of Indigenous Entomopathogenic Nematodes Against Spodoptera frugiperda in Nigeria Christopher Tobe Okolo, Abiodun O. Claudius-Cole, Florian Grundler, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8187949/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The successful deployment of entomopathogenic nematodes (EPNs) in biological pest control hinges on their ecological fitness and stress tolerance. In this study, we assessed the ecological traits and efficacy of six indigenous EPN isolates previously identified from distinct agroecological zones in Nigeria, targeting the invasive pest Spodoptera frugiperda (fall armyworm, FAW). The isolates, identified as Heterorhabditis bacteriophora (Ib-CRIN68), Steinernema carpocapsae (Ib-IART45, Ib-ITUC102), Steinernema feltiae (Za-SAM), Steinernema nepalense (Ib-HORT), and Oscheius myriophilus (Ib-FRIN32), were subjected to a series of ecological bioassays to evaluate their performance under temperature variation, moisture stress, oxygen limitation, oxidative stress, and foraging conditions. Results revealed significant inter- and intra-isolate variability in ecological tolerance traits. Optimal infectivity and reproduction were recorded between 25–30°C, while mortality sharply declined at 10°C and 35°C. Foraging ability varied across substrates and soil depths, with S. carpocapsae isolates exhibiting strong host-finding capability under dry and surface conditions. Desiccation and oxidative stress assays also demonstrated the superior resilience of S. carpocapsae isolates, which sustained low mortality under Polyethylenglycol induced water stress and H₂O₂ exposure. Hypoxia assays indicated that all isolates were moderately tolerant to short-term anoxia, but only H. bacteriophora and the S. carpocapsae isolates survived above 50% at 72 h. Our study highlights the relevance of ecological screening as a prerequisite for selecting robust EPN candidate species and isolates suitable for biological control under variable conditions. The findings support the integration of indigenous EPNs into sustainable pest management frameworks in sub-Saharan Africa. Agroecology Entomology Biological control EPN Fall Armyworm Oxidative stress Temperature Desiccation Ecological fitness Stress tolerance Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction Entomopathogenic nematodes (EPNs) belonging to the genera Steinernema (Rhabditida: Steinernematidae) and Heterorhabditis (Rhabditida: Heterorhabditidae), are a unique group of soil-inhabiting nematodes that parasitize and kill insect hosts with the help of their symbiotic bacteria Xenorhabdus spp. in Steinernema and Photorhabdus spp. in Heterorhabditis (Poinar, 1976) (Grewal et al., 2005; Nguyen & Hunt, 2007; Nguyen & Smart, 1996). These nematodes offer several attributes that make them attractive candidates for biological control: their ability to actively locate hosts (cruising or ambushing strategies), rapid killing via septicemia, environmental safety, and amenability to mass production (Grewal et al., 2005; Grewal et al., 1997; Kaya & Gaugler, 1993). EPNs have been widely evaluated for the control of a variety of insect pests in open-field crops, horticulture, and stored products, and they now form a vital component of integrated pest management (IPM) systems globally (Bhat et al., 2020; Lacey et al., 2015). Despite their proven laboratory virulence, the field performance of EPNs is often highly variable, particularly under tropical or sub-tropical climates, where environmental stresses such as temperature extremes, desiccation, hypoxia, and oxidative conditions can impair their infectivity, reproduction, and survival (Koppenhöfer & Kaya, 1999; Levy et al., 2020; John Mukuka, Olaf Strauch, Mohamed Hisham Al Zainab, et al., 2010; John Mukuka, Olaf Strauch, & Ralf-Udo Ehlers, 2010). EPN infective juveniles (IJs), which are the only free-living and infective stage, are particularly vulnerable to these abiotic factors, often resulting in poor field establishment and inconsistent pest suppression, thus limiting the adoption of EPN-based products in many developing regions (Kour et al., 2021; Lalramnghaki et al., 2017). Understanding the ecological adaptability and environmental resilience of EPN isolates for their effective implementation in biological control programs is therefore crucially important. Traits such as infectivity and reproduction under different thermal regimes (Levy et al., 2020), substrate adaptability (Matuska-Lyzwa et al., 2023), foraging depth, and tolerance to environmental stresses like desiccation (Nimkingrat et al., 2011; Nimkingrat et al., 2013), oxidative stress, and oxygen deprivation (hypoxia) (Sumaya et al., 2018; Zadji et al., 2014) are all key indicators of an isolate’s ability to perform well under field conditions. Ecological characterization, thus, provides a deeper understanding of isolate resilience and adaptability, far beyond what can be inferred from virulence tests alone (Anbesse et al., 2013; Koppenhöfer & Kaya, 1999; Puza et al., 2021). Although substantial progress has been made in characterizing EPNs in temperate regions, there is still a paucity of ecological studies on native African isolates. So far, work in Nigeria was largely limited to isolation, morphological and molecular identification, and laboratory virulence assays (Akyazi et al., 2012; Daramola et al., 2021), confirming the presence of diverse EPN taxa, including Steinernema carpocapsae , Heterorhabditis bacteriophora , and Oscheius myriophilus . However, no published work to date has evaluated the abiotic stress tolerance or environmental adaptability of these isolates, which is essential for developing robust EPN-based pest control tools suited to African agroecological systems. The present study addresses this gap by building upon earlier identification and virulence evaluations of six indigenous EPN isolates recovered from two agroecological zones in Nigeria lowland rainforest (Ibadan, Oyo State) and northern Guinea savannah (Zaria, Kaduna State) (Okolo et al. – unpublished ). Using morphological and molecular identification methods, these isolates were confirmed as Heterorhabditis bacteriophora (Ib-CRIN68), Steinernema carpocapsae (Ib-IART45, Ib-ITUC102), S. nepalense (Ib-HORT), S. feltiae (Za-SAM), and Oscheius myriophilus (Ib-FRIN32). Having previously established their laboratory virulence against Fall Armyworm, (FAW), Spodoptera frugiperda Smith (Lepidoptera: Noctuidae), an invasive and economically destructive pest of maize in sub-Saharan Africa (Goergen et al., 2016; Kenis et al., 2023), we now extend the investigation by evaluating their ecological traits relevant to field performance. By integrating a comprehensive suite of laboratory tests, this study aimed to generate a multi-dimensional profile of ecological fitness for each EPN isolate, with a view to identifying the most robust candidates for potential field evaluation. Specifically, we assessed the infectivity and reproductive success of these isolates to abiotic stresses such as temperature, desiccation, hypoxia (anaerobic storage conditions), and oxidative stress (via H₂O₂ exposure). Additional bioassays were conducted to evaluate foraging behaviour on various substrates and at different soil depths. Materials and Methods Infectivity and Reproduction in a Range of Temperatures The temperature tolerances of the six EPN isolates (Ib-CRIN68, Ib-IART45, Ib-ITUC102, Ib-FRIN32, and Ib-HORT from Ibadan, and Za-SAM from Zaria) were evaluated comprehensively through two experimental approaches designed to determine both their infectivity and reproductive capabilities across a range of temperatures. In the first set of experiments, infectivity was assessed in a completely randomised design (CRD) with four replicates using second-stage larvae ofFAW at six distinct temperatures (10, 15, 20, 25, 30, and 35°C). Each well of a 24-well tissue culture plate was filled with 0.5 g of air-dried soil, equilibrated at the respective temperatures for one hour prior to inoculation. Subsequently, each well in two sets received 50 IJs suspended in 60 µl sterilized deionized water, while a third set served as control without nematodes. Following nematode application, a single FAW larva, weighing between 200 and 300 mg, was carefully introduced to each well. Plates were monitored every 12 hours over a seven-day period, meticulously recording larval mortality, time to death, and quantifying the number of IJs established per larva. One day post-mortem, cadavers from one set of plates were dissected to count established IJs. Concurrently, the second set was examined daily for progeny IJ emergence from a White trap set up. Cadavers failing to yield emerging IJs within 14 days following initial emergence observations were dissected to validate infection status. The second experiment evaluated the EPNs reproductive potential at four selected temperatures (15, 20, 25, and 30°C). Nematodes (50 IJs per larva) were initially inoculated into tissue culture plates containing 0.5 g of sand and incubated at 25°C for 24 hours to standardize IJ penetration into FAW larvae (250–300 mg weight). Post-infection, individual cadavers were carefully transferred onto modified White traps consisting of a petri dish lid lined with filter paper floating on sterilized water in a larger petri dish. These were incubated at the specified temperatures. Emergence of IJs from cadavers was monitored daily, documenting the onset of emergence, total emergence duration, and intervals. Emerged IJs were periodically harvested and quantified by counting four representative subsamples from the suspension derived from each cadaver. Foraging Behaviour The foraging behaviour of EPN isolates was evaluated through two experiments designed to assess nematode attachment and depth penetration capabilities. Initially, approximately 1,000 IJs from each isolate were applied onto three different soil moisture conditions (0%, 10%, and 20%) and fresh maize leaf surfaces. IJs were allowed 15 minutes to disperse and acclimatize to each substrate before introducing a single actively crawling FAW L2 larva (200–300 mg). To ensure continuous larval movement, larvae were gently prodded whenever they ceased activity during the 30-minute exposure. Post-exposure, larvae were gently rinsed, and the number of nematodes attached was carefully counted under a dissecting microscope. Each moisture condition and leaf surface treatment were replicated ten times per isolate in two separate experimental trials to enhance robustness in a CRD. In an associated vertical distribution experiment, vertical plastic column arenas, measuring 5.5 cm in diameter and 10.5 cm in height, were filled with soil to investigate the nematodes' depth penetration ability. Individual larvae were placed at varying soil depths: on the surface, or at depths of 2, 5, and 10 cm. Each column received an inoculation of 1,000 IJs suspended in 1 ml sterilized water. After three days, columns were carefully disassembled, larvae were retrieved and dissected to confirm nematode establishment. In a CRD, each soil depth was replicated five times per isolate in two independent trials, providing a comprehensive assessment of nematode vertical mobility. Desiccation Tolerance Desiccation tolerance of the isolates was assessed by inducing desiccation using different concentrations of PEG 600 (Mukuka, Strauch, & Ehlers, 2010a;Anbesse et al., 2013a). Desiccation stress was measured as water activity ( a w ), which indicates the relative availability of unbound water to sustain the IJs. It is calculated as the ratio of the vapor pressure of water in a sample (p) to the vapor pressure of pure water (p ₀ ) at the same temperature. Freshly propagated IJs, pooled from four different growth batches, were used for the desiccation test in 24-cell well plates. The IJs were kept for 72 h in an adaptation solution of 40.3% PEG 600, ( a w of 0.96), prepared from a concentrated PEG 600 stock solution (Carl Roth, Karlsruhe, Germany). To minimize evaporation, the 24-cell well plates were sealed with Parafilm (Pechiney “M” Plastic Packaging, Chicago, USA). After adaptation, batches of 1,500 IJs in three replicates were exposed to seven PEG 600 concentrations for 24 h: 20% ( a w 0.98), 30% ( a w 0.97), 40.3% ( a w 0.96), 50% ( a w 0.93), 60% ( a w 0.89), 70% ( a w 0.83), and 80% ( a w 0.74). Control IJs were transferred to Ringer’s solution after the adaptation period ( a w 0.99). After treatment, the mortality of IJs was assessed in the three replicates by counting the number of active and inactive nematodes using a counting chamber. Percentage IJ mortality in different replicates was used to calculate the mean water activity (MW) tolerated by 50% of the population (MW 50 ) and the MW tolerated by the most tolerant 10% of the IJs population (MW 10 ). Hypoxia Tolerance Hypoxia tolerance of the isolates was assessed based on methods detailed by Zadji et al. (2014). For this assessment, 5,000 IJs from each isolate were placed into sealed 0.5-ml Eppendorf tubes containing distilled water and incubated under hypoxic conditions at 25°C in darkness for 24 or 72 hours. After exposure, IJs were transferred to Petri dishes containing 15 ml distilled water and further incubated at 25°C for 24 hours to evaluate recovery and survival rates. Tubes maintained in open conditions throughout incubation periods served as controls. Treatments were conducted with four replicates in a CRD and the entire experiment was repeated twice for reproducibility and confirmation of observed responses. Oxidative Stress Tolerance The oxidative stress performance of the six isolates was assessed by storing pools of IJs in Ringer’s solution in the presence of hydrogen peroxide (H 2 O 2 ) at room temperature (25°C). Freshly harvested IJ suspensions were separately exposed to H 2 O 2 in a 24-cell well plate in a CRD with three replicates, each containing 1,500 IJs in 400 µl of Ringer’s solution and sealed with Parafilm. For oxidative stress induction, 12.76 µl of 1.94 M H 2 O 2 was added to each cell well to obtain a final H 2 O 2 concentration of 60 mM. IJs kept under control conditions were left at 25°C in cell wells without H 2 O 2 . To assess the IJs mortality over time, 50 µl aliquots from each experimental replicate were counted in a counting chamber daily for two weeks. The percentage IJ mortality was used to determine differences in the mean survival time of 50% of the population (ST 50 ) and the survival time of the most tolerant 10% of the IJ population (ST 10 ) for each strain. The determination of the ST values followed the same procedure as that described for the desiccation test. Data analysis All statistical analyses were conducted using R v. 4.4.3 (R Core Team, 2024). Data were first assessed for normality and homogeneity of variance using the Shapiro–Wilk and Levene’s tests, respectively. Where appropriate, percentage data were arcsine square root transformed to meet parametric assumptions. For temperature-dependent infectivity and reproduction assays, two-way ANOVA was used to examine the effects of isolate and temperature on larval mortality and infective juvenile (IJ) emergence. Post hoc comparisons were performed using Tukey’s HSD test. In the desiccation tolerance assay, percentage IJ mortality from replicate treatments was used to estimate the mean water activity (MW₅₀) tolerated by 50% of the population and MW₁₀ for the most tolerant 10% of IJs. The data were fitted to a cumulative normal distribution curve, and the mean and standard deviation from the fitted curve were used to derive MW values by minimizing the χ² value between experimental and expected values. Similarly, for oxidative stress tolerance, IJ mortality percentages were used to estimate the mean survival time (ST₅₀) and the survival time of the most tolerant 10% (ST₁₀) of the population. These values were also obtained from a cumulative normal distribution fitted to the data, using the same χ² minimization approach as in the desiccation assay. Hypoxia tolerance data were analysed using Kaplan–Meier survival analysis, and isolate differences were assessed using the log-rank test. Median survival times (ST₅₀) were extracted from the survival curves. Foraging ability across substrates and soil depths was analysed using generalized linear models (GLMs), with isolate, substrate, and depth treated as fixed effects. The appropriate error distribution (Poisson or binomial) was applied based on the nature of the response variable. All statistical tests were considered significant at P < 0.05. Results Effect of Temperature on Infectivity of IJs The four key parameters measured to assess nematode infectivity, larval mortality at 72 hours post-inoculation, time until death of infected larvae, number of IJs established per FAW larva, and the percentage of larvae producing IJs are presented in Fig. 1 A-D. The percentage mortality of FAW larvae varied markedly across EPN isolates and temperature levels, as well as their interaction. Larval mortality exhibited strong temperature dependency. Mortality rates were negligible at 10°C across all isolates and peaked between 25°C and 30°C (Fig. 1A), with values reaching above 90% in some isolates such as Ib-IART45 and Ib-CRIN68. A two-way ANOVA confirmed highly significant main effects of temperature ( F = 1017.94, p < 0.001), isolate ( F = 119.89, p < 0.001), and their interaction ( F = 16.73, p < 0.001) on larval mortality . Correspondingly, the time until death of infected larvae significantly declined with increasing temperatures, reaching the shortest average duration (36–48 hours) at 25°C. At lower (10–15°C) and higher (35°C) extremes, larval death occurred more slowly, with mean times extending beyond 100 hours in some treatments (Fig. 1B). ANOVA analysis indicated statistically significant effects of temperature ( F = 167.04, p < 0.001), isolate ( F = 28.02, p < 0.001), and temperature × isolate interaction ( F = 4.27, p < 0.001), supporting the hypothesis that both thermal and genetic factors shape virulence dynamics. The number of IJs successfully establishing per larva mirrored the mortality trend, increasing significantly with temperature up to 25°C and subsequently declining. At 25°C, the highest establishment rates were observed, particularly in S. carpocapsae and S. feltiae isolates, with average IJ counts per larva exceeding 45. In contrast, establishment at 10°C was extremely low (<5 IJs per larva) across all isolates (Fig. 1C). A highly significant effect of temperature ( F = 1070.14, p < 0.001), isolate ( F = 113.19, p < 0.001), and their interaction ( F = 13.18, p < 0.001) was observed, indicating that both host penetration and survival are profoundly influenced by thermal environment and nematode identity. IJ ability to complete their life cycle and reproduce within the host, as measured by the percentage of FAW larvae producing IJs, also showed strong temperature dependency. IJ emergence was not observed at 10°C and 35°C for any isolate. Between 15°C and 30°C, however, emergence rates increased progressively, peaking at 25°C where values ranged between 85% and 90% for H. bacteriophora , S. feltiae and O. myriophilus (Fig. 1D). ANOVA again demonstrated significant effects of temperature ( F = 1184.93, p < 0.001), isolate (F = 83.41, p < 0.001), and their interaction ( F = 18.80, p < 0.001), highlighting variation in reproductive success under fluctuating environmental conditions. These underscore the critical influence of temperature and isolate identity on the infectivity, pathogenicity, and reproductive success of EPN. Effect of Temperature on Reproduction of IJs The percentage of cadavers producing progeny varied significantly among isolates and across temperatures (Fig. 2). A two-way ANOVA revealed significant main effects for both EPN isolate ( F (5, 96) = 11.28, p < 0.001) and temperature ( F (3, 96) = 189.47, p < 0.001), as well as a highly significant interaction effect between isolate and temperature ( F (15, 96) = 14.05, p < 0.001). Overall, the proportion of cadavers yielding progeny increased progressively from 15°C to 25°C and declined slightly at 30°C, with marked variation in reproductive success between the isolates (Fig. 2A). The timing of first IJ emergence from cadavers, measured in days post-infection (dpi), was also significantly influenced by both isolate identity and ambient temperature. Two-way ANOVA results indicated highly significant effects of isolate ( F (5, 96) = 15.05, p < 0.001), temperature ( F (3, 96) = 151.66, p < 0.001), and their interaction ( F (15, 96) = 18.48, p < 0.001). On average, emergence commenced earlier at higher temperatures, with the earliest onset observed at 25°C and delayed emergence at lower temperatures (Fig. 2B). Similarly, the duration of the IJ emergence period differed significantly among isolates and temperature treatments. The main effects of isolate ( F (5, 96) = 10.17, p < 0.001) and temperature ( F (3, 96) = 155.25, p < 0.001), along with the interaction between the two factors ( F (15, 96) = 12.61, p < 0.001), were all statistically significant. The longest emergence durations were generally recorded at intermediate temperatures (20–25°C), whereas shorter durations were observed at the lower and upper extremes of the temperature range (Fig. 2C). The total number of IJs emerging per cadaver (measured in thousands) also showed significant variability across treatments with highest progeny output clustered around 25°C for most isolates, and lower outputs observed at both 15°C and 30°C. Some isolates, particularly H. bacteriophora and the S. carpocapsae strains, maintained relatively high reproductive output across a broader temperature range (Fig. 2 D). Foraging Behaviour The mean number of IJs attaching to FAW larvae varied significantly across substrate types and EPN isolates (ANOVA, p < 0.05). Substrate type had a strong effect on IJ attachment, with markedly lower attachment observed on dry soil (0% moisture) and significantly higher attachment under moist conditions (particularly at 20% soil moisture). Among the substrates, 20% soil moisture supported the highest attachment levels, with S. carpocapsae isolates Ib-IART45 and Ib-ITUC102 recording the highest mean IJ attachments (50 ± 5 and 46 ± 4.5, respectively). In contrast, 0% soil moisture yielded the lowest mean IJ counts, particularly for O. myriophilus (Ib-FRIN32) and H. bacteriophora (Ib-CRIN68), with values as low as 4 ± 1.1 and 5 ± 1.2, respectively (Fig. 3). The maize leaf surface presented an intermediate attachment potential. Notably, S. nepalense (Ib-HORT) and S. feltiae (Za-SAM) achieved attachment levels comparable to those on 10% soil moisture, averaging 33 ± 3.5 and 28 ± 3.0 IJs per larva, respectively. These findings underscore both substrate-specific differences in attachment efficiency and isolate-level variability in host-finding and infection initiation traits under varying environmental conditions. The statistical analysis of the depth penetration assay evaluating the ability of six EPN IJs isolates to infect FAW larvae across four soil depths revealed significant isolate- and depth-dependent differences. A two-way ANOVA confirmed that isolate, depth, and their interaction significantly influenced penetration rates ( p < 0.001 for all factors). The highest mean penetration rates were observed at the soil surface, where most isolates achieved infection levels exceeding 80%, with S. carpocapsae isolates Ib-IART45 and Ib-ITUC102 demonstrating superior performance even at increased depths (Fig. 4). Penetration rates declined progressively with soil depth, particularly at 10 cm, where all isolates recorded markedly lower effectiveness. Descriptive statistics underscored this trend, and post hoc Tukey HSD tests revealed statistically significant differences among isolates and depths, underscoring clear vertical stratification of infectivity potential among isolates. Desiccation Tolerance The minimum water activity required to maintain 50% (MW₅₀) and 10% (MW₁₀) IJ survival differed significantly among the six EPN isolates (Fig. 5). Steinernema carpocapsae isolates Ib-IART45 and Ib-ITUC102 recorded the least water activity (MW₅₀ a w ~0.89), indicating superior overall desiccation resilience. In contrast, H. bacteriophora (Ib-CRIN68) exhibited the lowest desiccation tolerance (MW₅₀ ~0.97), suggesting a need for more water to maintain survival and infectivity of 50% population and more rapid decline in population viability under drying conditions. Similarly, the 10% best performing IJs of Ib-IART45 and Ib-ITUC102 tolerated lower a w values (MW₁₀ ~0.798), while H. bacteriophora lost 90% of their IJs at slightly higher a w (MW₁₀ ~0.857). We also observed that at a w 0.99, IJ survival was highest in S. carpocapsae (Ib-IART45) (99.0 ± 2.3%) and lowest in H. bacteriophora (Ib-CRIN68) (88.0 ± 1.7%) (Table 1). At a w 0.98, survival remained above 84% for all isolates except H. bacteriophora (76.0 ± 2.3%). Subsequent reduction of a w showed S. carpocapsae isolates maintained high survival. At the lowest a w 0.74, survival dropped to <10% across all isolates, with S. carpocapsae (Ib-IART45) having the highest value (6.7 ± 1.7%) and H. bacteriophora showing complete mortality (0.0%). Table 1 . Percentage survival of IJs of six EPN isolates to decreasing a w levels.[1] Water Activity ( a w ) H. bacteriophora (Ib-CRIN68) S. carpocapsae (Ib-IART45) S. carpocapsae (Ib-ITUC102) O. myriophilus (Ib-FRIN32) S. nepalense (Ib-HORT) S. feltiae (Za-SAM) 0.99 88.0 ± 1.7 b 99.0 ± 2.3 a 98.0 ± 2.9 a 95.0 ± 1.7 a 85.0 ± 2.3 b 95.0 ± 1.2 a 0.98 76.0 ± 2.3 c 94.8 ± 1.7 a 91.0 ± 2.3 a 87.0 ± 1.2 b 84.0 ± 2.3 b 85.0 ± 1.2 b 0.97 50.5 ± 2.9 c 87.85 ± 2.3 a 87.5 ± 1.7 a 77.0 ± 2.3 b 79.0 ± 2.3 b 74.0 ± 1.7 b 0.96 30.0 ± 3.5 d 85.0 ± 2.9 a 78.0 ± 2.3 b 50.8 ± 2.3 c 78.5 ± 2.3 b 62.0 ± 1.7 c 0.93 26.0 ± 2.9 d 65.0 ± 3.5 a 62.0 ± 2.9 a 47.0 ± 2.3 c 55.0 ± 2.9 b 58.0 ± 2.3 b 0.89 17.0 ± 2.3 d 52.0 ± 2.9 a 50.9 ± 2.9 a 32.0 ± 2.3 c 45.0 ± 2.3 b 49.0 ± 2.3 ab 0.83 7.0 ± 1.7 c 30.0 ± 2.3 a 25.0 ± 2.3 a 10.2 ± 1.7 b 12.0 ± 1.7 b 11.8 ± 1.7 b 0.74 00.00 c 6.7 ± 1.7 a 3.0 ± 1.2 b 2.3 ± 1.2 b 2.0 ± 1.2 b 4.0 ± 1.2 ab [1] Values represent mean ± standard error (SE) of IJ survival at each water activity level. Different superscript letters within each row indicate statistically significant differences among EPN isolates at that specific water activity level (Tukey’s HSD test, p < 0.05). Hypoxia Tolerance Survival data demonstrated significant variability across isolates and exposure durations, as confirmed by analysis of variance (ANOVA). After 24 hours of exposure, survival rates ranged from approximately 56.3% to 76.2%, with Steinernema carpocapsae (Ib-ITUC102) exhibiting the highest survival, followed closely by Steinernema carpocapsae (Ib-IART45) (Fig. 6A). The lowest survival was recorded in O. myriophilus (Ib-FRIN32). We found statistically significant differences among isolates ( p < 0.05), indicating distinct hypoxia tolerance profiles in the short term. Following 72 hours of hypoxia, a marked reduction in survival was observed across all isolates, with survival ranging from 27.9% to 63.4% (Fig. 6B). Again, S. carpocapsae (Ib-ITUC102) maintained the highest tolerance, whereas O. myriophilus (Ib-FRIN32) and S. nepalense (Ib-HORT) were the most adversely affected. Statistical analysis confirmed a significant isolate effect ( p < 0.05), and visual comparison of both timepoints indicated time-dependent reduction in IJ viability under sustained oxygen deprivation. Oxidative stress Tolerance The oxidative stress tolerance was evaluated by assessing the survival time of the most tolerant 10% (ST₁₀) and 50% (ST₅₀) of IJs exposed to 60 mM H₂O₂. Survival time varied significantly among the isolates ( p < 0.05), indicating differential capacities to withstand oxidative stress (Fig. 7). Steinernema feltiae (Za-SAM) and S. carpocapsae (Ib-IART45) exhibited the highest oxidative stress tolerance, with ST₁₀ values of 16.8 ± 0.4 h and 16.5 ± 0.6 h, respectively, and corresponding ST₅₀ values of 27.34 ± 0.8 h and 26.77 ± 0.9 h, significantly higher than those of other isolates. Steinernema nepalense (Ib-HORT) recorded the lowest oxidative stress resistance, with an ST₁₀ of 13.1 ± 0.5 h and an ST₅₀ of 18.93 ± 0.6 h, which were significantly lower ( p < 0.05) than most other isolates. Moderate oxidative stress performance was observed for H. bacteriophora (Ib-CRIN68), S. carpocapsae (Ib-ITUC102), and O. myriophilus (Ib-FRIN32), which did not differ significantly from one another in ST₁₀ and ST₅₀ measures. The considerable variation in the oxidative resilience of the isolates suggest potential adaptive differences in physiological responses to environmental stress (Fig. 7). Discussion The ecological characterisation of EPN is a critical step in the development and deployment of effective biological control agents for sustainable pest management. This study evaluated six indigenous EPN isolates previously identified morphologically and molecularly from Nigeria; H. bacteriophora (Ib-CRIN68), S. carpocapsae (Ib-IART45 and Ib-ITUC102), O. myriophilus (Ib-FRIN32), S. nepalense (Ib-HORT), and S. feltiae (Za-SAM), with respect to key ecological traits that influence their infectivity and persistence. These traits included thermal sensitivity, reproductive potential, tolerance to abiotic stresses (desiccation, hypoxia, oxidative stress) and host-seeking behaviour. The six EPN isolates from Nigeria showed clear temperature-dependent patterns in both host infection and reproduction. Overall, moderate temperatures (in the mid-20°C range) supported the highest virulence and nematode progeny production, whereas extreme low or high temperatures constrained performance. These trends align with well-documented thermal optima for EPNs: for example, most EPN species achieve maximal infection and reproduction around 25°C and cannot reproduce below ~ 10–15°C (Koppenhöfer & Kaya, 1999 ; Raheel et al., 2017 ). In our study, H. bacteriophora (Ib-CRIN68) and S. carpocapsae (Ib-IART45, Ib-ITUC102) maintained high infectivity even at elevated temperatures, reflecting their adaptation to tropical climates. Notably, H. bacteriophora sustained strong virulence up to 30–35°C, consistent with reports that this species can perform robustly at the upper thermal limits of IJs activity (Aatif et al., 2020 ; Matuska-Lyzwa et al., 2024 ). By contrast, the S. feltiae isolate Za-SAM, a species typically native to cooler regions of Nigeria, showed reduced infectivity and yield at higher temperatures. This mirrors the known thermal sensitivity of S. feltiae , which exhibits diminished activity above ~ 30°C. In fact, storage of S. feltiae at 35°C for even 1 hour can dramatically curtail its motility, with lethal effects by 37°C (Matuska-Lyzwa et al., 2024 ). Such thermal intolerance likely explains the poorer performance of the Za-SAM isolate under hot conditions and underscores the importance of matching EPN strains to ambient thermal regimes. Reproductive capacity followed similar temperature-related patterns. All isolates failed to recycle in hosts at very low temperatures (no reproduction occurred at 5°C, and only S. feltiae managed limited reproduction by 10°C). As temperatures rose, development accelerated and brood sizes increased for all species. The highest yields of IJs in our trials were produced by H. bacteriophora (Ib-CRIN68), consistent with previous studies showing Heterorhabditis can generate exceptionally large progenies in Galleria hosts (Levy et al., 2020 ; Raheel et al., 2017 ). We also observed that H. bacteriophora and the S. carpocapsae isolates had faster kill and emergence at 25–30°C than at cooler temperatures, whereas S. feltiae required longer intervals to produce IJs, especially at suboptimal temperatures. Steinernema species like S. carpocapsae can kill hosts and recycle more rapidly at 25–28°C (often within a week), whereas Heterorhabditis typically takes a few days longer to emerge (Brown et al., 2002 ; Susurluk & Ulu, 2015 ). At the highest temperature tested in our study, some drop-off in IJ yield was noted for all isolates, suggesting that extreme heat imposes stress on nematode development and symbiont functioning. It is well known that sustained exposure to > 32°C adversely affects EPN growth, reproduction and survival (Karanastasi et al., 2025 ; Lillis et al., 2023 ). Nonetheless, the indigenous tropical isolates in this study tolerated heat better than many temperate-strain EPNs reported in literature, reinforcing the ecological premise that local nematode populations are better adapted to the prevailing thermal conditions of their environments. The ability to adapt to varying thermal conditions is essential for biocontrol agents in SSA agroecosystems, where soil temperatures can vary considerably. Our results show that considering temperature optima and limits is crucial when choosing suitable EPN isolates for field evaluation. The Nigerian isolate H. bacteriophora Ib-CRIN68 has a broad thermal activity range, making it effective in warm climates. The tested EPN isolates demonstrated distinct foraging strategies, following the ambusher versus cruiser host-finding behaviour. The two S. carpocapsae isolates Ib-IART45 and Ib-ITUC102 showed ambush foraging by staying near the soil surface and adopting a “sit-and-wait” tactic for mobile hosts. This was evident as they infected hosts near the soil surface with limited deep soil movement, consistent with S. carpocapsae 's ecology of targeting insects at or above the soil interface (Bal & Grewal, 2015 ; Raja et al., 2011 ). Ambushers like S. carpocapsae are known to conserve energy by nictating or standing upright near the surface, and only a small fraction of their population actively disperses far from the release point (Chen & Glazer, 2005 ; Wright et al., 1997 ). Interestingly, even ambushers can exhibit a subset of highly motile “sprinter” individuals that disperse rapidly when host cues (such as CO₂ or other volatiles) are detected. Our observations of S. carpocapsae reaching and infecting hosts a few tens of centimeters away concur with reports that a small proportion (~ 1–2%) of S. carpocapsae IJs can travel > 10 cm in soil columns (Bal & Grewal, 2015 ). This dual strategy, mostly localized waiting with occasional long-distance forays, likely maximizes the chance of encountering a susceptible insect host in heterogenous soil environments. In contrast, H. bacteriophora (Ib-CRIN68) and O. myriophilus (Ib-FRIN32) showed more cruiser-like foraging behaviour. They actively explored deeper soil layers and were able to locate hosts buried at greater depths or at further horizontal distances. Heterorhabditis bacteriophora in particular is known as an active cruiser that continuously moves through soil pore water in search of sedentary or subterranean hosts. In our depth-gradient assays, H. bacteriophora IJs readily penetrated to lower strata (e.g. >15 cm depth) and successfully infected hosts there, whereas S. carpocapsae infections were concentrated in the upper 10 cm. Host-seeking observations show that Heterorhabditis spp. tend to distribute deeper in the soil profile than Steinernema ambushers. For example, in surveys S. feltiae were found mostly in the top 0–15 cm of soil, whereas H. bacteriophora can be recovered from much greater depths (Neumann & Shields, 2006 ; Williams et al., 2013 ). Our data similarly suggests that the Ib-CRIN68 isolate of H. bacteriophora is adept at vertical movement, an advantageous trait for targeting soil-dwelling stages of pests, such as pupating FAW. The ability to forage actively in the soil may also help cruisers find immobile hosts like cocoons or grubs, complementing the ambushers’ strength against surface-active insects. Meanwhile, the behaviour of S. nepalense Ib-HORT and S. feltiae Za-SAM isolates appeared intermediate. They neither strictly waited at the surface nor ranged as widely as H. bacteriophora , suggesting a more flexible foraging strategy. Many Steinernema spp. are known to be intermediate strategists that can both ambush and cruise to some extent (Rakubu et al., 2024 ). This adaptable foraging strategy enables them to utilize various host niches, although it may not be as efficiently specialized as the extreme ambusher ( S. carpocapsae ) or the extreme cruiser ( H. bacteriophora ). Soil moisture had a pronounced influence on foraging efficacy. In moderately moist soil, almost at field capacity, all isolates moved and located hosts with the highest success. However, under drier conditions, host-finding declined, particularly for the cruiser-type nematodes that rely on continuous water films for movement. Nematodes are essentially aquatic in locomotion, gliding along water-filled pores thus insufficient moisture breaks the continuity of those films, impeding IJ mobility (Kaspi et al., 2010 ). We observed that as the soil became drier, H. bacteriophora and S. nepalense showed reduced dispersal and tended to congregate in deeper, more humid layers if available. Nematodes will migrate downward as surface soil desiccates, accumulating at depths where humidity is higher (Cabanillas, 2003 ; Duncan & McCoy, 2001 ; Gouge et al., 2000 ; Yadav & Lalramliana, 2012 ). In one study, S. riobrave was seen to move 15–23 cm deep over four weeks of gradual surface drying, effectively tracking the receding moisture front (Gouge et al., 2000 ). A similar pattern in our experiments suggests that the Nigerian cruisers actively seek favourable moisture microhabitats, which would enhance their survival during dry spells. On the other hand, S. carpocapsae (ambusher) was less able to escape drying soil by migration. Instead, its strategy under low moisture may be to enter a quiescent state near the surface and wait for either a host or the return of moisture. Ambusher species like S. carpocapsae often have greater desiccation tolerance, enabling them to survive near the surface until a host contacts them or rain rehydrates the soil (Shapiro-Ilan et al., 2014 ). Moreover, the virulence of S. carpocapsae in dry soil can be restored upon re-moistening of the soil, suggesting that the IJs remain viable in a dormant state and are capable of resuming their infective activity once favourable conditions are re-established (Grant & Villani, 2003 ). These behavioural differences mean in an applied context, that S. carpocapsae might be more effective when pests are on or near the soil surface (and intermittent dry periods occur), whereas H. bacteriophora could be superior for targets in the soil profile provided adequate moisture is present or irrigation is used. Overall, our foraging assays highlight that both moisture and soil depth interact with nematode behavioural traits. The most effective biocontrol may be achieved by matching isolate behaviour to pest ecology such as deploying ambushers for mobile foliar larvae and cruisers for soil-dwelling stages or by combining species to cover multiple strata and moisture conditions in the field (Bal & Grewal, 2015 ). An important aspect of the ecological fitness of EPNs is their ability to withstand environmental stresses. We found considerable isolate-specific differences in tolerance to desiccation (dryness), low oxygen, and oxidative stress. Steinernema carpocapsae Ib-IART45/ITUC102 stood out for its superior desiccation tolerance, and its IJ survival after exposure to low humidity (or dry soil) was significantly higher than that of the Heterorhabditis and other Steinernema isolates. Generally, S. carpocapsae is considered to be one of the most desiccation-tolerant nematode species. For instance, Shapiro-Ilan et al. ( 2014 ) reported that S. carpocapsae IJs survived desiccating conditions far better than heterorhabditids, with S. feltiae also ranking high and Heterorhabditis generally the least tolerant. Our results mirror that pattern, the S. feltiae Za-SAM isolate had the second-highest desiccation survival, whereas H. bacteriophora was among the most sensitive to drying. Desiccation tolerance in EPNs is thought to be linked to behavioural and physiological adaptations; ambushers like S. carpocapsae often remain near the soil surface and have evolved mechanisms to survive transient drought (Glazer, 2022 ; Ramakrishnan et al., 2022 ). In contrast, cruisers avoid dry conditions by moving deeper, as earlier discussed, and consequently may not have invested in as strong desiccation-hardiness mechanisms which explains the lower survival of H. bacteriophora in our desiccation assays. It is encouraging that even the more sensitive isolates in our study still retained some viability after short dry exposures, suggesting that a fraction of the population can endure brief droughts. Additionally, we noted no obvious intraspecific variation in desiccation survival between the two S. carpocapsae strains, which is consistent with reports that different strains of S. carpocapsae tend to exhibit uniformly high desiccation tolerance (Shapiro-Ilan et al., 2014 ). Overall, the ability of the S. carpocapsae and S. feltiae isolates from Nigeria to better survive drying conditions could be advantageous for use in regions with irregular rainfall or for above-ground applications where desiccation risk is high. All isolates showed reduced survival under hypoxic (low oxygen) conditions, though with subtle differences. When subjected to oxygen-depleted environments (simulating waterlogged or compacted soils), H. bacteriophora and S. nepalense survived slightly longer than the S. carpocapsae and Oscheius isolates. This may reflect adaptation of cruisers to burrowing into less aerated soil pockets (Kung et al., 1990a , b ). Nonetheless, the overall intolerance of the nematodes to hypoxia was evident such that prolonged exposure to < 1% O₂ led to high mortality across the board. This finding is expected since EPN IJs are aerobic organisms that rely on dissolved oxygen in soil water. As soil oxygen drops, nematode metabolism and survival sharply decline. For instance, oxygen levels near 1% drastically impair EPN viability and infectivity, and the survival of S. carpocapsae and S. glaseri IJs plummeted as oxygen was reduced from ambient (20%) to near-anoxic levels (Matuska-Lyzwa et al., 2024 ). Our isolates likely behave similarly, with heavy, water-saturated soils (common in the humid tropics during rains) posing a risk to their persistence. Interestingly, we did observe that nematodes in our experiments often sought out air pockets or moved to the soil surface in response to waterlogging, suggesting a behavioural escape from hypoxia. Such behaviour has been noted that IJs can sense gradients in oxygen or CO₂ and migrate toward more favourable conditions (Hoctor et al., 2013 ; Maushe et al., 2023 ). In practical terms, this means EPN applications should avoid fully waterlogged conditions; good soil drainage will promote nematode survival. Also, soil texture matters, coarse, well-aerated soils are more nematode-friendly than heavy clays that induce anaerobic micro-sites. While we did not identify a dramatically hypoxia-tolerant isolate, the slight edge of H. bacteriophora under low O₂ might relate to its natural occurrence in deeper soil. Still, hypoxia remains a limiting factor for all, reinforcing that EPNs work best under moderate moisture without oxygen starvation. When exposed to oxidative challenges, such as hydrogen peroxide in our assays, some isolates fared noticeably better. In particular, S. carpocapsae Ib-IART45 showed higher survival and maintained mobility longer under oxidative stress than H. bacteriophora or S. feltiae . This could indicate a more robust antioxidant defense system in S. carpocapsae . Effective scavenging of reactive oxygen species (ROS) is critical for EPNs, both in the soil environment and during infection of the host. Insect hosts actively mount an oxidative immune response – generating superoxide, peroxide, and other ROS to attack invading nematodes (Lalitha et al., 2018 ; Sumaya et al., 2018 ). Nematodes that can withstand this onslaught have a better chance to establish infection. S. carpocapsae is known for producing a lethal toxin (via its symbiont Xenorhabdus ) that rapidly kills the host, which might shorten the window of exposure to host immune defense, indirectly reducing oxidative damage (Lu et al., 2017 ; Watanabe et al., 2019 ). Additionally, prior studies on the nematode models, C. elegans , show that exposure to peroxides causes immediate loss of mobility and depressed metabolism, but young nematodes can recover if their antioxidant enzymes like peroxiredoxins and catalases, are effective (Kumsta et al., 2011 ). The superior oxidative stress survival of S. carpocapsae could reflect such efficient detoxification systems. It is possible that this isolate constitutively expresses high levels of catalase, superoxide dismutase, or peroxiredoxin that neutralize ROS, a trait that would be beneficial during the early stages of infection when the insect’s immune burst is highest. By contrast, H. bacteriophora Ib-CRIN68 showed more oxidative damage (lower survival) in our test, which might relate to its strategy of relying on a supportive mutualistic bacterium ( Photorhabdus ) to overcome host defense, Photorhabdus produces immunosuppressive factors but perhaps less in terms of ROS-scavengers. Another intriguing observation was that the O. myriophilus isolate had relatively good oxidative tolerance, almost on par with S. carpocapsae . Oscheius spp. are not symbiotically tied to Xenorhabdus/Photorhabdus ; some are associated with other bacteria or can be facultative pathogens. Their pathogenicity often depends on releasing their own array of bacteria upon host entry (Onwong et al., 2023 ). The resilience of O. myriophilus to oxidative stress in our assays suggests it may possess inherent protective mechanisms (possibly due to its free-living lineage background) or perhaps carries bacteria that aid in detoxification. While literature on EPN oxidative stress tolerance is scant, our results suggest this trait could underlie differences in virulence and field persistence. Isolates that better endure oxidative stress might survive longer on foliage (exposed to UV and oxidative conditions) or overcome host immune reactions more successfully. This could partly explain why S. carpocapsae was so virulent in our trials, its physiological hardiness complements its aggressive infection strategy. In sum, screening for oxidative stress tolerance, alongside desiccation and hypoxia tolerance, provides a more complete picture of an EPN isolate’s suitability for biocontrol deployment in challenging environments. Trade-offs exist between virulence, environmental tolerance, and reproductive fitness (J. Mukuka et al., 2010 ). For instance, the most desiccation-tolerant species ( S. carpocapsae ) are not the most fecund reproducers, and the most fecund ( H. bacteriophora ) are not very desiccation-tolerant. This necessitates a strategic approach to biocontrol to either formulate consortia of complementary EPN isolates or tailor the choice of isolate to the specific pest and environment. In practical IPM, one might apply S. carpocapsae for immediate knockdown of FAW larvae in the crop canopy, combined with H. bacteriophora for longer-lasting suppression of the next generation in the soil. There is evidence that mixed-species applications can sometimes yield additive benefits, as different nematodes occupy slightly different niches and timescales of action though careful consideration of competition and compatibility is needed. Our findings suggest that H. bacteriophora Ib-CRIN68 and S. carpocapsae Ib-IART45 together would make a formidable pair, the former ensuring persistence and recycling in soil and the latter providing quick action against active larvae. Additionally, O. myriophilus could be included to bolster resilience to stress, possibly as a stress-hardy backup that might sustain population when others wane. The demonstrated trait superiority of these isolates likely stems from their evolutionary history in Nigerian agroecosystems, for example Ib-CRIN68 coming from a farm soil that undergoes periodic drying and heating, selecting for a hardy yet virulent phenotype, and Ib-IART45 originating from an area with intense insect pressure, selecting for high pathogenicity. It would be valuable in future work to investigate the genetic or physiological basis of these traits. Previous and recent research are identifying genetic markers (heat-shock proteins, anhydrobiosis-related genes, antioxidant enzymes) that correlate with stress tolerance in nematodes (Grewal et al., 2006 ; Hao et al., 2011 ; Maushe et al., 2023 ; Segal & Glazer, 2000 ). Unravelling these mechanisms in our top isolates could enable marker-assisted selection or even bioengineering to further improve them. In essence, the diverse performances observed affirm that isolate selection is crucial and by choosing the right nematode combinations one can achieve reliable biocontrol even under the challenging conditions of SSA farmlands. References Aatif, H. M., Hanif, M. S., Raheel, M., Ferhan, M., Mansha, M. Z., Khan, A. A., Ullah, M. I., Shakeel, Q., & Ali, S. (2020). Temperature dependent virulence of the entomopathogenic nematodes against immatures of the oriental fruit fly, Bactrocera dorsalis Hendel (Diptera: Tephritidae). Egyptian Journal of Biological Pest Control , 30 (1), 1-6. https://doi.org/10.1186/S41938-020-00248-7 Akyazi, F., Ansari, M. A., Ahmed, B. I., Crow, W. T., & Mekete, T. (2012). First record of entomopathogenic nematodes (steinernematidae and heterorhabditidae) from Nigerian soil and their morphometrical and ribosomal DNA sequence analysis. Nematologia Mediterranea , 40 (2), 95-100. Anbesse, S., Sumaya, N. H., Dorfler, A. V., Strauch, O., & Ehlers, R. U. (2013). Selective breeding for desiccation tolerance in liquid culture provides genetically stable inbred lines of the entomopathogenic nematode Heterorhabditis bacteriophora . Appl Microbiol Biotechnol , 97 (2), 731-739. https://doi.org/10.1007/s00253-012-4227-5 Bal, H. K., & Grewal, P. S. (2015). Lateral dispersal and foraging behavior of entomopathogenic nematodes in the absence and presence of mobile and non-mobile hosts. PLoS One , 10 , 1-19. https://doi.org/10.1371/journal.pone.0129887 Bhat, A. H., Chaubey, A. K., & Askary, T. H. (2020). Global distribution of entomopathogenic nematodes, Steinernema and Heterorhabditis . Egyptian Journal of Biological Pest Control , 30 (1). https://doi.org/10.1186/s41938-020-0212-y Brown, I. M., Lovett, B. J., Grewal, P. S., & Gaugler, R. (2002). Latent infection: a low temperature survival strategy in steinernematid nematodes. Journal of Thermal Biology , 27 (6), 531-539. https://doi.org/10.1016/S0306-4565(02)00027-X Cabanillas, H. E. (2003). Susceptibility of the boll weevil to Steinernema riobrave and other entomopathogenic nematodes. Journal of Invertebrate Pathology , 82 (3), 188-197. https://doi.org/10.1016/S0022-2011(03)00016-8 Chen, S., & Glazer, I. (2005). A novel method for long-term storage of the entomopathogenic nematode Steinernema feltiae at room temperature. Biological Control , 32 , 104-110. https://doi.org/http://dx.doi.org/10.1016/j.biocontrol.2004.08.006 Daramola, F. Y., Osemwegie, O. O., Owa, S. O., Orisajo, S. B., Ikponmwosa, E., & Alori, E. T. (2021). Isolation and molecular characterization of entomopathogenic nematode, Heterorhabditis sp. from an arable land in Nigeria. Journal of Integrative Agriculture , 20 (10), 2706-2715. https://doi.org/10.1016/s2095-3119(21)63609-2 Duncan, L. W., & McCoy, C. W. (2001). Hydraulic lift increases herbivory by Diaprepes abbreviates larvae and persistence of Steinernema riobrave in dry soil. Journal of Nematology , 33 (2-3), 142-146. Glazer, I. (2022). Stress and Survival Mechanisms. In Nematodes as Model Organisms (pp. 215-243). CABI. https://doi.org/10.1079/9781789248814.0009 Goergen, G., Kumar, P. L., Sankung, S. B., Togola, A., & Tamò, M. (2016). First report of outbreaks of the fall armyworm Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), a new alien invasive pest in West and Central Africa. PLoS One , 11 (10), e0165632. https://doi.org/10.1371/journal.pone.0165632 Gouge, D. H., Smith, K. A., Lee, L. L., & Henneberry, T. J. (2000). Effect of Soil Depth and Moisture on the Vertical Distribution of Steinernema riobrave (Nematoda: Steinernematidae). Journal of Nematology , 32 (2), 223-223. https://pmc.ncbi.nlm.nih.gov/articles/PMC2620435/ http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=PMC2620435 Grant, J. A., & Villani, M. G. (2003). Soil Moisture Effects on Entomopathogenic Nematodes. Environmental Entomology , 32 (1), 80-87. https://doi.org/10.1603/0046-225X-32.1.80 Grewal, P. S., Bornstein-Forst, S., Burnell, A. M., Glazer, I., & Jagdale, G. B. (2006). Physiological, genetic, and molecular mechanisms of chemoreception, thermobiosis, and anhydrobiosis in entomopathogenic nematodes. Biological Control , 38 (1), 54-65. https://doi.org/10.1016/j.biocontrol.2005.09.004 Grewal, P. S., Ehlers, R.-U., & Shapiro-Ilan, D. I. (2005). Nematodes as biocontrol agents . CABI. Grewal, P. S., Lewis, E. E., & Gaugler, R. (1997). Response of infective stage parasites (Nematoda: Steinernematidae) to volatile cues from infected hosts. J Chem Ecol , 23 , 503-515. Hao, Y. J., Flores-Ponce, M., & Montiel, R. (2011). Genetics of entomopathogenic nematodes. In Microbial Insecticides: Principles and Applications (pp. 237-256). Nova Science Publishers, Inc. https://www.scopus.com/inward/record.uri?eid=2-s2.0-84895214695&partnerID=40&md5=428727f67e1a8d33049be71d903b6b4e Hoctor, T. L., Gibb, T. J., Bigelow, C. A., & Richmond, D. S. (2013). Survival and infectivity of the insect-parasitic nematode Heterorhabditis bacteriophora (Poinar) in solutions containing four different turfgrass soil surfactants. Insects , 4 (1), 1-8. https://doi.org/10.3390/insects4010001 Karanastasi, E., Nikorezou, A., Stamouli, M., Skourti, A., Boukouvala, M. C., Kavallieratos, N. G., & Weber, D. (2025). Temperature effect on the efficacy of 3 entomopathogenic nematode isolates against larvae of the lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae). Journal of Economic Entomology , 118 (1), 93-99. https://doi.org/10.1093/jee/toae292 Kaspi, R., Ross, A., Hodson, A. K., Stevens, G. N., Kaya, H. K., & Lewis, E. E. (2010). Foraging efficacy of the entomopathogenic nematode Steinernema riobrave in different soil types from California citrus groves. Applied Soil Ecology , 45 (3), 243-253. https://doi.org/10.1016/j.apsoil.2010.04.012 Kaya, H. K., & Gaugler, R. (1993). Entomopathogenic Nematodes. Annual Review of Entomology , 38 (1), 181-206. https://doi.org/10.1146/annurev.en.38.010193.001145 Kenis, M., Benelli, G., Biondi, A., Calatayud, P. A., Day, R., Desneux, N., Harrison, R. D., Kriticos, D., Rwomushana, I., van den Berg, J., Verheggen, F., Zhang, Y. J., Agboyi, L. K., Ahissou, R. B., Ba, M. N., Bernal, J., Freitas de Bueno, A., Carrière, Y., Carvalho, G. A., . . . Wu, K. (2023). Invasiveness, biology, ecology, and management of the fall armyworm, Spodoptera frugiperda . Entomologia Generalis , 43 (2), 187-241. https://doi.org/10.1127/entomologia/2022/1659 Koppenhöfer, A. M., & Kaya, H. K. (1999). Ecological Characterisation of Steinernema rarum . Journal of Invertebrate Pathology , 73 , 120 - 128. http://www.idealibrary.com Kour, S., Khurma, U., & Brodie, G. (2021). Ecological Characterisation of Native Isolates of Heterorhabditis indica from Viti Levu, Fiji Islands. Journal of Nematology , 53 . https://doi.org/10.21307/jofnem-2021-085 Kumsta, C., Thamsen, M., & Jakob, U. (2011). Effects of Oxidative Stress on Behavior, Physiology, and the Redox Thiol Proteome of Caenorhabditis elegans . Antioxidants & Redox Signaling , 14 (6), 1023-1023. https://doi.org/10.1089/ARS.2010.3203 Kung, S.-P., Gaugler, R., & Kaya, H. K. (1990a). Influence of Soil pH and Oxygen on Persistence of Steinernema spp. Journal of Nematology , 22 (4), 440-440. https://pmc.ncbi.nlm.nih.gov/articles/PMC2619085/ Kung, S.-P., Gaugler, R., & Kaya, H. K. (1990b). Soil type and entomopathogenic nematode persistence. Journal of Invertebrate Pathology , 55 (3), 401-406. https://doi.org/https://doi.org/10.1016/0022-2011(90)90084-J Lacey, L. A., Grzywacz, D., Shapiro-Ilan, D. I., Frutos, R., Brownbridge, M., & Goettel, M. S. (2015). Insect pathogens as biological control agents: Back to the future. Journal of Invertebrate Pathology , 132 , 1-41. https://doi.org/10.1016/j.jip.2015.07.009 Lalitha, K., Karthi, S., Vengateswari, G., Karthikraja, R., Perumal, P., & Shivakumar, M. S. (2018). Effect of entomopathogenic nematode of Heterorhabditis indica infection on immune and antioxidant system in lepidopteran pest Spodoptera litura (Lepidoptera: Noctuidae). Journal of Parasitic Diseases , 42 (2), 204-211. https://doi.org/10.1007/s12639-018-0983-1 Lalramnghaki, H. C., Vanlalhlimpuia, Vanramliana, & Lalramliana. (2017). Characterization of a new isolate of entomopathogenic nematode, Steinernema sangi (Rhabditida, Steinernematidae), and its symbiotic bacteria Xenorhabdus vietnamensis (γ-Proteobacteria) from Mizoram, northeastern India. Journal of Parasitic Diseases , 41 (4), 1123-1131. https://doi.org/10.1007/s12639-017-0945-z Levy, N., Faigenboim, A., Salame, L., Molina, C., Ehlers, R.-U., Glazer, I., & Ment, D. (2020). Characterization of the phenotypic and genotypic tolerance to abiotic stresses of natural populations of Heterorhabditis bacteriophora . Scientific Reports , 10 . https://doi.org/10.1038/s41598-020-67097-0 Lillis, P. E., Kennedy, I. P., Carolan, J. C., & Griffin, C. T. (2023). Low-temperature exposure has immediate and lasting effects on the stress tolerance, chemotaxis and proteome of entomopathogenic nematodes. Parasitology , 150 (1), 15-28. https://doi.org/10.1017/S0031182022001445 Lu, D., Macchietto, M., Chang, D., Barros, M. M., Baldwin, J., Mortazavi, A., & Dillman, A. R. (2017). Activated entomopathogenic nematode infective juveniles release lethal venom proteins. PLoS Pathogens , 13 (4), Article e1006302. https://doi.org/10.1371/journal.ppat.1006302 Matuska-Lyzwa, J., Duda, S., Nowak, D., & Kaca, W. (2024). Impact of Abiotic and Biotic Environmental Conditions on the Development and Infectivity of Entomopathogenic Nematodes in Agricultural Soils. Insects 2024, Vol. 15, Page 421 , 15 (6), 421-421. https://doi.org/10.3390/INSECTS15060421 Matuska-Lyzwa, J., Wodecka, B., & Kaca, W. (2023). Characterization of Steinernema feltiae (Rhabditida: Steinernematidae) Isolates in Terms of Efficacy against Cereal Ground Beetle Zabrus tenebrioides (Coleoptera: Carabidae): Morphometry and Principal Component Analysis. Insects , 14 (2). https://doi.org/10.3390/insects14020150 Maushe, D., Ogi, V., Divakaran, K., Verdecia Mogena, A. M., Himmighofen, P. A., Machado, R. A. R., Towbin, B. D., Ehlers, R. U., Molina, C., Parisod, C., & Robert, C. A. M. (2023). Stress tolerance in entomopathogenic nematodes: Engineering superior nematodes for precision agriculture. Journal of Invertebrate Pathology , 199 , 107953-107953. https://doi.org/10.1016/J.JIP.2023.107953 Mukuka, J., Strauch, O., Al Zainab, M. H., & Ehlers, R.-U. (2010). Effect of temperature and desiccation stress on infectivity of stress tolerant hybrid strains of Heterorhabditis bacteriophora . Russian Journal of Nematology , 18 (2), 111-116. https://www.scopus.com/inward/record.uri?eid=2-s2.0-79952132220&partnerID=40&md5=4fb76546eb0281c1529cf546aabc751e Mukuka, J., Strauch, O., & Ehlers, R.-U. (2010). Variability in desiccation tolerance among different strains of the entomopathogenic nematode Heterorhabditis bacteriophora . Nematology , 12 (5), 711-720. Mukuka, J., Strauch, O., Hoppe, C., & Ehlers, R. U. (2010). Fitness of heat and desiccation tolerant hybrid strains of Heterorhabditis bacteriophora (Rhabditidomorpha: Heterorhabditidae). Journal of Pest Science , 83 (3), 281-287. https://doi.org/10.1007/s10340-010-0296-3 Neumann, G., & Shields, E. J. (2006). Interspecific interactions among three entomopathogenic nematodes, Steinernema carpocapsae Weiser, S. feltiae Filipjev, and Heterorhabditis bacteriophora Poinar, with different foraging strategies for hosts in multipiece sand columns. Environmental Entomology , 35 (6), 1578-1583. https://doi.org/10.1603/0046-225X(2006)35[1578:IIATEN]2.0.CO;2 Nguyen, K. B., & Hunt, D. J. (2007). Entomopathogenic nematodes : systematics, phylogeny and bacterial symbionts . Brill. Nguyen, K. B., & Smart, G. C. (1996). Identification of Entomopathogenic Nematodes in the Steinernematidae and Heterorhabditidae (Nemata: Rhabditida) Journal of Nematology. https://pmc.ncbi.nlm.nih.gov/articles/PMC2619694/ Nimkingrat, P., Ehlers, R. U., & Strauch, O. (2011). Desiccation tolerance among different isolates of the entomopathogenic nematode Steinernema feltiae (Fillipjev). Communications in agricultural and applied biological sciences , 76 (3), 293-296. https://pubmed.ncbi.nlm.nih.gov/22696940/ Nimkingrat, P., Uhlmann, F., Strauch, O., & Ehlers, R.-U. (2013). Desiccation tolerance of dauers of entomopathogenic nematodes of the genus Steinernema . Nematology , 15 , 451-458. https://doi.org/10.1163/15685411-00002692 Onwong, R., Sumaya, N. H., Nitjarunkul, A., Kerdim, S., Khwanket, N., & Noosidum, A. (2023). Occurrence of entomopathogenic nematodes, Oscheius myriophilus Poinar in Thailand: Preliminary characterization of novel isolates and biological control potential against insect pests. Journal of Applied Entomology . https://doi.org/10.1111/jen.13168 Puza, V., Nermut, J., Konopicka, J., & Skokova Habustova, O. (2021). Efficacy of the Applied Natural Enemies on the Survival of Colorado Potato Beetle Adults. Insects , 12 (11). https://doi.org/10.3390/insects12111030 R Core Team. (2024). R: A Language and Environment for Statistical Computing, .In R (Version 4.4.3) http://www.R-project.org/. Raheel, M., Javed, N., Khan, S. A., Aatif, H. M., & Ahmed, S. (2017). Effect of temperature on the reproductive potential of indigenous and exotic species of entomopathogenic nematodes inside Galleria mellonella L. larvae. Pakistan Journal of Zoology , 49 (1), 419-421. https://doi.org/10.17582/journal.pjz/2017.49.1.sc12 Raja, K. R., Padmanaban, K., & Sivaramakrishnan, S. (2011). Entomopathogenic Nematodes: A Best Bio-control Agent for Insect Pest. Isolation and Identification of Entomopathogenic Nematodes from Agricultural land . Lambert Academic Publishing. Rakubu, I. L., Katumanyane, A., & Hurley, B. P. (2024). Host-foraging strategies of five local entomopathogenic nematode species in South Africa. Crop Protection , 176 , 106525-106525. https://doi.org/10.1016/J.CROPRO.2023.106525 Ramakrishnan, J., Salame, L., Nasser, A., Glazer, I., & Ment, D. (2022). Survival and efficacy of entomopathogenic nematodes on exposed surfaces. Scientific Reports , 12 (1), 4629-4629. https://doi.org/10.1038/S41598-022-08605-2 Segal, D., & Glazer, I. (2000). Genetics for improving biological control agents: the case of entomopathogenic nematodes. Crop Protection , 19 , 685-689. Shapiro-Ilan, D. I., Brown, I., & Lewis, E. E. (2014). Freezing and desiccation tolerance in entomopathogenic nematodes: diversity and correlation of traits. Journal of Nematology , 27-34. Sumaya, N. H., Gohil, R., Okolo, C. T., Addis, T., Doerfler, V., Ehlers, R. U., & Molina, C. (2018). Applying inbreeding, hybridization and mutagenesis to improve oxidative stress tolerance and longevity of the entomopathogenic nematode Heterorhabditis bacteriophora . Journal of Invertebral Pathology , 151 , 50-58. https://doi.org/10.1016/j.jip.2017.11.001 Susurluk, I. A., & Ulu, T. C. (2015). Virulence comparisons of high-temperature-adapted Heterorhabditis bacteriophora , Steinernema feltiae and S. carpocapsae . Helminthologia (Poland) , 52 (2), 118-122. https://doi.org/10.1515/helmin-2015-0021 Watanabe, A., Yamaguchi, T., Murota, K., Ishii, T., Terao, J., Okada, S., Tanaka, N., Kimata, S., Abe, A., Suzuki, T., Uchino, M., & Niimura, Y. (2019). Isolation of lactic acid bacteria capable of reducing environmental alkyl and fatty acid hydroperoxides, and the effect of their oral administration on oxidative-stressed nematodes and rats. bioRxiv , 592162. https://doi.org/10.1101/592162 Williams, C. D., Dillon, A. B., Girling, R. D., & Griffin, C. T. (2013). Organic soils promote the efficacy of entomopathogenic nematodes, with different foraging strategies, in the control of a major forest pest: A meta-analysis of field trial data. Biological Control , 65 (3), 357-364. https://doi.org/10.1016/j.biocontrol.2013.03.013 Wright, D. J., Grewal, P. S., & Stolinski, M. (1997). Relative Importance of Neutral Lipids and Glycogen as Energy Stores in Dauer Larvae of Two Entomopathogenic Nematodes, Steinernema carpocapsae and Steinernema feltiae . Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology , 118 (2), 269-273. https://doi.org/10.1016/S0305-0491(97)00165-X Yadav, A. K., & Lalramliana. (2012). Soil moisture effects on the activity of three entomopathogenic nematodes (Steinernematidae and Heterorhabditidae) isolated from Meghalaya, India. Journal of Parasitic Diseases , 36 (1), 94-98. https://doi.org/10.1007/s12639-011-0076-x Zadji, L., Baimey, H., Afouda, L., Moens, M., & Decraemer, W. (2014). Characterization of biocontrol traits of heterorhabditid entomopathogenic nematode isolates from South Benin targeting the termite pest Macrotermes bellicosus. BioControl , 59 (3), 333-344. https://doi.org/10.1007/s10526-014-9568-9 Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {\"props\":{\"pageProps\":{\"initialData\":{\"identity\":\"rs-8187949\",\"acceptedTermsAndConditions\":true,\"allowDirectSubmit\":true,\"archivedVersions\":[],\"articleType\":\"Research Article\",\"associatedPublications\":[],\"authors\":[{\"id\":550190584,\"identity\":\"f5e0d602-02d4-4d25-99e4-51c4942e51ff\",\"order_by\":0,\"name\":\"Christopher Tobe Okolo\",\"email\":\"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAwklEQVRIiWNgGAWjYFACNgaGBww2DAzMIBYIHCBGSwJDGulaDkNZxGjhb29LfJBQcT5xOzsD24OPO+4w8B1vwK9F4syxwwYJZ24n7mxmYDeceeYZg+QZAtYYSKS3SSS23U7ccJj/mzRv22EGgxsJRGk5B9TCwAbRcv8BIS1px4BaDiBpuYFfB8gvyUC/JBsDtQD90naYR/IMAYcBQ8zwwYcKO9kN5w8AQ6ztsBzf8QMErEEHPCSqHwWjYBSMglGADQAAoHhGKtqDrVMAAAAASUVORK5CYII=\",\"orcid\":\"https://orcid.org/0000-0002-6063-6727\",\"institution\":\"University of Bonn\",\"correspondingAuthor\":true,\"prefix\":\"\",\"firstName\":\"Christopher\",\"middleName\":\"Tobe\",\"lastName\":\"Okolo\",\"suffix\":\"\"},{\"id\":550193308,\"identity\":\"30bd5f94-c77f-4d5f-9321-0c1e5d985324\",\"order_by\":1,\"name\":\"Abiodun O. Claudius-Cole\",\"email\":\"\",\"orcid\":\"https://orcid.org/0000-0003-1206-6325\",\"institution\":\"University of Ibadan\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Abiodun\",\"middleName\":\"O.\",\"lastName\":\"Claudius-Cole\",\"suffix\":\"\"},{\"id\":550193309,\"identity\":\"91008f15-d4f2-401f-bbf4-6f8610f8c6ef\",\"order_by\":2,\"name\":\"Florian Grundler\",\"email\":\"\",\"orcid\":\"https://orcid.org/0000-0001-8101-0558\",\"institution\":\"University of Bonn\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Florian\",\"middleName\":\"\",\"lastName\":\"Grundler\",\"suffix\":\"\"},{\"id\":550193310,\"identity\":\"3f5739c1-58a1-4ac8-b98a-3c932cf75521\",\"order_by\":3,\"name\":\"Christian Borgemiester\",\"email\":\"\",\"orcid\":\"https://orcid.org/0000-0001-8067-0335\",\"institution\":\"University of Bonn\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Christian\",\"middleName\":\"\",\"lastName\":\"Borgemiester\",\"suffix\":\"\"}],\"badges\":[],\"createdAt\":\"2025-11-23 23:32:03\",\"currentVersionCode\":1,\"declarations\":{\"humanSubjects\":false,\"vertebrateSubjects\":true,\"conflictsOfInterestStatement\":false,\"humanSubjectEthicalGuidelines\":false,\"humanSubjectConsent\":false,\"humanSubjectClinicalTrial\":false,\"humanSubjectCaseReport\":false,\"vertebrateSubjectEthicalGuidelines\":true},\"doi\":\"10.21203/rs.3.rs-8187949/v1\",\"doiUrl\":\"https://doi.org/10.21203/rs.3.rs-8187949/v1\",\"draftVersion\":[],\"editorialEvents\":[],\"editorialNote\":\"\",\"failedWorkflow\":false,\"files\":[{\"id\":96779907,\"identity\":\"17a5158d-5ebe-478b-8957-88186af25417\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"docx\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":2207373,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"EcologicalCharacterizationandEfficacyofIndigenousEntomopathogenicNematodesAgainstSpodopterafrugiperdainNigeria.docx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/13c549d144cea2571240ba1d.docx\"},{\"id\":96779903,\"identity\":\"aacfe995-b023-4791-9261-0637ba9a5fc6\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"json\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":342,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"rs8187949.json\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/b574f944a777bb5364a9552a.json\"},{\"id\":96779906,\"identity\":\"2c3d6866-9bd8-496e-9995-9e5fe1e43798\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"xml\",\"order_by\":2,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":186643,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"rs81879490enriched.xml\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/d083db1a3f7f4246143008bc.xml\"},{\"id\":96915798,\"identity\":\"f3618ae5-b43a-486f-bba2-e8ec3cfc8472\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:38\",\"extension\":\"jpeg\",\"order_by\":3,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":659216,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage1.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/cdd3d548546f570aaed259c0.jpeg\"},{\"id\":96915362,\"identity\":\"957f9621-ac0b-41e1-9e5c-19daf3f87018\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:11\",\"extension\":\"jpeg\",\"order_by\":4,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":618364,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage2.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/0359946459a6bcdbfcc70312.jpeg\"},{\"id\":96779920,\"identity\":\"a2bc22a1-f98d-4073-9b2c-6abbbd98ff95\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"jpeg\",\"order_by\":5,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":595974,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage3.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/ba41d3e9ac1ab6ae02520eda.jpeg\"},{\"id\":96779914,\"identity\":\"f66db21f-8d11-46ce-a48a-5ea46077051d\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"jpeg\",\"order_by\":6,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":595388,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage4.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/b11c73cd804a88e53da3ca93.jpeg\"},{\"id\":96779919,\"identity\":\"70a41830-cabe-4571-a1c6-4505ddb20c6a\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":7,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":166166,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage5.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/7b3f259a785501104b5fe0c0.png\"},{\"id\":96779922,\"identity\":\"4b87b72c-217e-435c-8be1-fcc4574b2cc9\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":8,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":138213,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage6.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/b848b2f44c70556ae7c625d1.png\"},{\"id\":96779916,\"identity\":\"4d6787c5-a215-46d2-afe1-9dbe8bbeb393\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":9,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":96579,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage7.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/661e2280157d15bedeacb009.png\"},{\"id\":96915753,\"identity\":\"4c83d318-0b03-46ee-8da9-f08dbb6bc266\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:36\",\"extension\":\"jpeg\",\"order_by\":10,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":444894,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage8.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/978347d38089a0a76307cfe9.jpeg\"},{\"id\":96779923,\"identity\":\"44fd072c-04bb-4ba3-9a01-9315f8446390\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":11,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":79208,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage9.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/3b2ca91ee7bb750b9a931011.png\"},{\"id\":96915640,\"identity\":\"7b5e519f-5129-4f1d-9d09-f41f9fc37e30\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:27\",\"extension\":\"png\",\"order_by\":12,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":111651,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage1.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/2f7259daade80f4c04b86210.png\"},{\"id\":96779918,\"identity\":\"a1234ac6-a3be-43ae-b10c-c5e829bc7969\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":13,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":111341,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage2.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/a16177f0eaf5f72a67d0ab66.png\"},{\"id\":96779924,\"identity\":\"19bdad1d-2acb-4eb4-803e-baf3b10bd044\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":14,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":99721,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage3.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/bc7a5bd275ca0031b340d6fe.png\"},{\"id\":96779912,\"identity\":\"533b3970-8f03-403e-802c-714877081ca2\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":15,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":102991,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage4.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/6fbe21e3b9734d92b1afe4ad.png\"},{\"id\":96779928,\"identity\":\"12b0ce66-9893-4069-891b-ffd082b5cf99\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":16,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":41153,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage5.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/c441c3944b11a87c99cd67a7.png\"},{\"id\":96915766,\"identity\":\"ede70841-a484-43d2-8bfe-5f4cabc3b8f1\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:36\",\"extension\":\"png\",\"order_by\":17,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":38501,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage6.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/da56c5a9f6cf361e22a4398e.png\"},{\"id\":96917382,\"identity\":\"83d6f46d-2a4c-44c2-a777-c488206dc2ec\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:09:39\",\"extension\":\"png\",\"order_by\":18,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":34288,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage7.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/984d889d8d9a13241bb9a964.png\"},{\"id\":96915888,\"identity\":\"327193b9-4c5d-4f13-a51d-54e1283b8d02\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:07:44\",\"extension\":\"png\",\"order_by\":19,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":94891,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage8.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/7b8a4d36201bac268838e1a4.png\"},{\"id\":96779913,\"identity\":\"99bcfc89-80b9-4465-b726-eef405b91af0\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":20,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":25579,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage9.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/178d3fd1a4c99a6833129753.png\"},{\"id\":96779929,\"identity\":\"9ff198da-3014-4cdd-9d6a-fe1d8b0c913d\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:10\",\"extension\":\"xml\",\"order_by\":21,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":185377,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"rs81879490structuring.xml\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/aee3973a7e9f91005db9070a.xml\"},{\"id\":96779926,\"identity\":\"3c71cfb4-2905-442c-94f6-fa391eec0711\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"html\",\"order_by\":22,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":198370,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"earlyproof.html\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/171d182de6af2ad6f5f5410d.html\"},{\"id\":96779900,\"identity\":\"0ce94ca3-9957-40cd-8b05-ee0c8f91a214\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":1,\"title\":\"Figure 1\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":262129,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of temperature on infectivity traits of six entomopathogenic nematode (EPN) isolates against fall armyworm (FAW) 2\\u003csup\\u003end\\u003c/sup\\u003e instar larvae. \\u003cstrong\\u003e(A)\\u003c/strong\\u003e Percent mortality of infected larvae at 72 hours post-inoculation \\u003cstrong\\u003e(B)\\u003c/strong\\u003e Time to death of infected larvae (hours). \\u003cstrong\\u003e(C)\\u003c/strong\\u003e Number of Infected Juveniles (IJs) established per larva. \\u003cstrong\\u003e(D)\\u003c/strong\\u003e Percentage of larvae producing IJs. Bars with different uppercase letters within each temperature represent significant differences among isolates, while different lowercase letters above the bars indicate significant differences across temperature for each isolate (Tukey’s HSD, p \\u0026lt; 0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"1.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/00f4b662d56dea3768daa930.png\"},{\"id\":96916271,\"identity\":\"e8950abb-f794-497e-86e0-6a4e42ec9f67\",\"added_by\":\"auto\",\"created_at\":\"2025-11-27 14:08:22\",\"extension\":\"png\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":218047,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eReproductive performance of six indigenous entomopathogenic nematode (EPN) isolates at four temperature regimes following infection of fall armyworm (FAW) larvae. (\\u003cstrong\\u003eA\\u003c/strong\\u003e) percentage of larval cadavers producing progeny, (\\u003cstrong\\u003eB\\u003c/strong\\u003e) day of first emergence of Infective Juveniles (IJs) from cadavers, (\\u003cstrong\\u003eC\\u003c/strong\\u003e) duration of IJ emergence, and (\\u003cstrong\\u003eD\\u003c/strong\\u003e) number of IJs emerged per cadaver (× 10³).Bars with different uppercase letters within each temperature represent significant differences among isolates, while different lowercase letters above the bars indicate significant differences across temperature for each isolate (Tukey’s HSD, p\\u0026lt; 0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"2.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/d0b07d5bf974b06b261c5a63.png\"},{\"id\":96779901,\"identity\":\"250529e9-20cd-4893-95ed-1524b26de821\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":3,\"title\":\"Figure 3\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":60785,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of substrate type on infective juvenile (IJ) attachment of six indigenous entomopathogenic nematode (EPN) isolates to larvae of fall armyworm (FAW). Values are mean number of IJs attached per larva (± SE). Bars with different uppercase letters within each substrate type represent significant differences among isolates, while different lowercase letters above the bars indicate significant differences across substrate types for each isolate (Tukey’s HSD, p \\u0026lt; 0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"3.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/47e12f368b496f5f7b0ccf05.png\"},{\"id\":96779905,\"identity\":\"9e5f9fe6-f338-4271-9635-a4a7740193e4\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":4,\"title\":\"Figure 4\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":56719,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eMean penetration rates (%) of six indigenous entomopathogenic nematode (EPN) isolates into fall armyworm (FAW) larvae at different soil depths (surface, 2 cm, 5 cm, 10 cm). Bars represent mean ± standard deviation. Results are based on five replicates per depth per isolate. Bars with different uppercase letters within each soil depth represent significant differences among isolates (Tukey’s HSD, p \\u0026lt; 0.05), while different lowercase letters above the bars indicate significant differences across soil depths for each isolate.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"4.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/867b1091b4e9a81e64b48817.png\"},{\"id\":96779908,\"identity\":\"9d2e2f7d-e98c-4f26-8cf4-aeaafa47016d\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":5,\"title\":\"Figure 5\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":50333,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eStacked bar chart showing the MW₅₀ (mean water activity tolerated by 50% of the population - white bar) and MW₁₀ (mean water activity tolerated by 10% of the population - black bar) \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003e\\u003cem\\u003ew\\u003c/em\\u003e\\u003c/sub\\u003e thresholds for desiccation tolerance of six entomopathogenic nematode\\u003cem\\u003e \\u003c/em\\u003e(EPN) isolates. Error bars represent standard deviations. Bars with the same letter are not significantly different (Tukey’s HSD, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"5.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/69c9824e7aa6df59fa8a1657.png\"},{\"id\":96779910,\"identity\":\"aeaca251-26e3-4671-801c-8f218df775b2\",\"added_by\":\"auto\",\"created_at\":\"2025-11-26 04:14:09\",\"extension\":\"png\",\"order_by\":6,\"title\":\"Figure 6\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":103100,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eSurvival of Infective Juveniles (IJs) of six indigenous entomopathogenic nematode\\u003cem\\u003e \\u003c/em\\u003e(EPN) isolates under hypoxic stress conditions at 25 °C. (\\u003cstrong\\u003eA\\u003c/strong\\u003e) Survival (%) after 24-hour sealed exposure; (\\u003cstrong\\u003eB\\u003c/strong\\u003e) Survival (%) after 72-hour exposure. Bars represent mean values (± SE) from three independent replicates, with different letters above the bars indicating significant differences between isolates (Tukey’s HSD test; \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"6.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/e245548eb36a84214f9f3bc6.png\"},{\"id\":97135363,\"identity\":\"701f70cf-9bc3-4498-9a06-2fe8cd361b72\",\"added_by\":\"auto\",\"created_at\":\"2025-12-01 09:39:30\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":1573144,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-8187949/v1/7998e39a-9cbc-4e75-b194-512e4e27507e.pdf\"}],\"financialInterests\":\"The authors declare no competing interests.\",\"formattedTitle\":\"\\u003cp\\u003e\\u003cstrong\\u003eEcological Characterization and Efficacy of Indigenous Entomopathogenic Nematodes Against \\u003c/strong\\u003e\\u003cem\\u003e\\u003cstrong\\u003eSpodoptera frugiperda\\u003c/strong\\u003e\\u003c/em\\u003e\\u003cstrong\\u003e in Nigeria\\u003c/strong\\u003e\\u003c/p\\u003e\",\"fulltext\":[{\"header\":\"Introduction\",\"content\":\"\\u003cp\\u003eEntomopathogenic nematodes (EPNs) belonging to the genera \\u003cem\\u003eSteinernema\\u0026nbsp;\\u003c/em\\u003e(Rhabditida: Steinernematidae) and \\u003cem\\u003eHeterorhabditis\\u0026nbsp;\\u003c/em\\u003e(Rhabditida: Heterorhabditidae), are a unique group of soil-inhabiting nematodes that parasitize and kill insect hosts with the help of their symbiotic bacteria \\u003cem\\u003eXenorhabdus\\u003c/em\\u003e spp. in \\u003cem\\u003eSteinernema\\u003c/em\\u003e and \\u003cem\\u003ePhotorhabdus\\u003c/em\\u003e spp. in \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e (Poinar, 1976)\\u0026nbsp;(Grewal et al., 2005; Nguyen \\u0026amp; Hunt, 2007; Nguyen \\u0026amp; Smart, 1996). These nematodes offer several attributes that make them attractive candidates for biological control: their ability to actively locate hosts (cruising or ambushing strategies), rapid killing via septicemia, environmental safety, and amenability to mass production\\u0026nbsp;(Grewal et al., 2005; Grewal et al., 1997; Kaya \\u0026amp; Gaugler, 1993). EPNs have been widely evaluated for the control of a variety of insect pests in open-field crops, horticulture, and stored products, and they now form a vital component of integrated pest management (IPM) systems globally\\u0026nbsp;(Bhat et al., 2020; Lacey et al., 2015).\\u003c/p\\u003e\\n\\u003cp\\u003eDespite their proven laboratory virulence, the field performance of EPNs is often highly variable, particularly under tropical or sub-tropical climates, where environmental stresses such as temperature extremes, desiccation, hypoxia, and oxidative conditions can impair their infectivity, reproduction, and survival\\u0026nbsp;(Koppenhöfer \\u0026amp; Kaya, 1999; Levy et al., 2020; John Mukuka, Olaf Strauch, Mohamed Hisham Al Zainab, et al., 2010; John Mukuka, Olaf Strauch, \\u0026amp; Ralf-Udo Ehlers, 2010).\\u0026nbsp;EPN infective juveniles (IJs), which are the only free-living and infective stage, are particularly vulnerable to these abiotic factors, often resulting in poor field establishment and inconsistent pest suppression, thus limiting the adoption of EPN-based products in many developing regions\\u0026nbsp;(Kour et al., 2021; Lalramnghaki et al., 2017).\\u003c/p\\u003e\\n\\u003cp\\u003eUnderstanding the ecological adaptability and environmental resilience of EPN isolates for their effective implementation in biological control programs is therefore crucially important. Traits such as infectivity and reproduction under different thermal regimes (Levy et al., 2020), substrate adaptability\\u0026nbsp;(Matuska-Lyzwa et al., 2023), foraging depth, and tolerance to environmental stresses like desiccation\\u0026nbsp;(Nimkingrat et al., 2011; Nimkingrat et al., 2013), oxidative stress, and oxygen deprivation (hypoxia)\\u0026nbsp;(Sumaya et al., 2018; Zadji et al., 2014)\\u0026nbsp;are all key indicators of an isolate’s ability to perform well under field conditions. Ecological characterization, thus, provides a deeper understanding of isolate resilience and adaptability, far beyond what can be inferred from virulence tests alone\\u0026nbsp;(Anbesse et al., 2013; Koppenhöfer \\u0026amp; Kaya, 1999; Puza et al., 2021).\\u003c/p\\u003e\\n\\u003cp\\u003eAlthough substantial progress has been made in characterizing EPNs in temperate regions, there is still a paucity of ecological studies on native African isolates. So far, work in Nigeria was largely limited to isolation, morphological and molecular identification, and laboratory virulence assays\\u0026nbsp;(Akyazi et al., 2012; Daramola et al., 2021),\\u0026nbsp;confirming the presence of diverse EPN taxa, including \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e, \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e, and \\u003cem\\u003eOscheius myriophilus\\u003c/em\\u003e. However, no published work to date has evaluated the abiotic stress tolerance or environmental adaptability of these isolates, which is essential for developing robust EPN-based pest control tools suited to African agroecological systems.\\u003c/p\\u003e\\n\\u003cp\\u003eThe present study addresses this gap by building upon earlier identification and virulence evaluations of six indigenous EPN isolates recovered from two agroecological zones in Nigeria lowland rainforest (Ibadan, Oyo State) and northern Guinea savannah (Zaria, Kaduna State) (Okolo et al. – \\u003cem\\u003eunpublished\\u003c/em\\u003e). Using morphological and molecular identification methods, these isolates were confirmed as \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e (Ib-CRIN68), \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e (Ib-IART45, Ib-ITUC102), \\u003cem\\u003eS. nepalense\\u003c/em\\u003e (Ib-HORT), \\u003cem\\u003eS. feltiae\\u003c/em\\u003e (Za-SAM), and \\u003cem\\u003eOscheius myriophilus\\u003c/em\\u003e (Ib-FRIN32). Having previously established their laboratory virulence against Fall Armyworm, (FAW), \\u003cem\\u003eSpodoptera\\u0026nbsp;frugiperda\\u003c/em\\u003e Smith (Lepidoptera: Noctuidae),\\u0026nbsp;an invasive and economically destructive pest of maize in sub-Saharan Africa\\u0026nbsp;(Goergen et al., 2016; Kenis et al., 2023), we now extend the investigation by evaluating their ecological traits relevant to field performance.\\u003c/p\\u003e\\n\\u003cp\\u003eBy integrating a comprehensive suite of laboratory tests, this study aimed to generate a multi-dimensional profile of ecological fitness for each EPN isolate, with a view to identifying the most robust candidates for potential field evaluation. Specifically, we assessed the infectivity and reproductive success of these isolates to abiotic stresses such as temperature, desiccation, hypoxia (anaerobic storage conditions), and oxidative stress (via H₂O₂ exposure). Additional bioassays were conducted to evaluate foraging behaviour on various substrates and at different soil depths.\\u0026nbsp;\\u003c/p\\u003e\"},{\"header\":\"Materials and Methods\",\"content\":\"\\u003ch2\\u003eInfectivity and Reproduction in a Range of Temperatures\\u003c/h2\\u003e\\n\\u003cp\\u003eThe temperature tolerances of the six EPN isolates (Ib-CRIN68, Ib-IART45, Ib-ITUC102, Ib-FRIN32, and Ib-HORT from Ibadan, and Za-SAM from Zaria) were evaluated comprehensively through two experimental approaches designed to determine both their infectivity and reproductive capabilities across a range of temperatures.\\u003c/p\\u003e\\n\\u003cp\\u003eIn the first set of experiments, infectivity was assessed in a completely randomised design (CRD) with four replicates using second-stage larvae ofFAW at six distinct temperatures (10, 15, 20, 25, 30, and 35°C). Each well of a 24-well tissue culture plate was filled with 0.5 g of air-dried soil, equilibrated at the respective temperatures for one hour prior to inoculation. Subsequently, each well in two sets received 50 IJs suspended in 60 µl sterilized deionized water, while a third set served as control without nematodes. Following nematode application, a single FAW larva, weighing between 200 and 300 mg, was carefully introduced to each well. Plates were monitored every 12 hours over a seven-day period, meticulously recording larval mortality, time to death, and quantifying the number of IJs established per larva. One day post-mortem, cadavers from one set of plates were dissected to count established IJs. Concurrently, the second set was examined daily for progeny IJ emergence from a White trap set up. Cadavers failing to yield emerging IJs within 14 days following initial emergence observations were dissected to validate infection status.\\u003c/p\\u003e\\n\\u003cp\\u003eThe second experiment evaluated the EPNs reproductive potential at four selected temperatures (15, 20, 25, and 30°C). Nematodes (50 IJs per larva) were initially inoculated into tissue culture plates containing 0.5 g of sand and incubated at 25°C for 24 hours to standardize IJ penetration into FAW larvae (250–300 mg weight). Post-infection, individual cadavers were carefully transferred onto modified White traps consisting of a petri dish lid lined with filter paper floating on sterilized water in a larger petri dish. These were incubated at the specified temperatures. Emergence of IJs from cadavers was monitored daily, documenting the onset of emergence, total emergence duration, and intervals. Emerged IJs were periodically harvested and quantified by counting four representative subsamples from the suspension derived from each cadaver.\\u003c/p\\u003e\\n\\u003ch2\\u003eForaging Behaviour\\u003c/h2\\u003e\\n\\u003cp\\u003eThe foraging behaviour of EPN isolates was evaluated through two experiments designed to assess nematode attachment and depth penetration capabilities. Initially, approximately 1,000 IJs from each isolate were applied onto three different soil moisture conditions (0%, 10%, and 20%) and fresh maize leaf surfaces. IJs were allowed 15 minutes to disperse and acclimatize to each substrate before introducing a single actively crawling FAW L2 larva (200–300 mg). To ensure continuous larval movement, larvae were gently prodded whenever they ceased activity during the 30-minute exposure. Post-exposure, larvae were gently rinsed, and the number of nematodes attached was carefully counted under a dissecting microscope. Each moisture condition and leaf surface treatment were replicated ten times per isolate in two separate experimental trials to enhance robustness in a CRD. In an associated vertical distribution experiment, vertical plastic column arenas, measuring 5.5 cm in diameter and 10.5 cm in height, were filled with soil to investigate the nematodes' depth penetration ability. Individual larvae were placed at varying soil depths: on the surface, or at depths of 2, 5, and 10 cm. Each column received an inoculation of 1,000 IJs suspended in 1 ml sterilized water. After three days, columns were carefully disassembled, larvae were retrieved and dissected to confirm nematode establishment. In a CRD, each soil depth was replicated five times per isolate in two independent trials, providing a comprehensive assessment of nematode vertical mobility.\\u003c/p\\u003e\\n\\u003ch2\\u003eDesiccation Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eDesiccation tolerance of the isolates was assessed by inducing desiccation using different concentrations of PEG 600 (Mukuka, Strauch, \\u0026amp; Ehlers, 2010a;Anbesse et al., 2013a). Desiccation stress was measured as water activity (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e), which indicates the relative availability of unbound water to sustain the IJs. It is calculated as the ratio of the vapor pressure of water in a sample (p) to the vapor pressure of pure water (p\\u003cstrong\\u003e₀\\u003c/strong\\u003e) at the same temperature. Freshly propagated IJs, pooled from four different growth batches, were used for the desiccation test in 24-cell well plates. The IJs were kept for 72 h in an adaptation solution of 40.3% PEG 600, (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e of 0.96), prepared from a concentrated PEG 600 stock solution (Carl Roth, Karlsruhe, Germany). To minimize evaporation, the 24-cell well plates were sealed with Parafilm (Pechiney “M” Plastic Packaging, Chicago, USA). After adaptation, batches of 1,500 IJs in three replicates were exposed to seven PEG 600 concentrations for 24 h: 20% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.98), 30% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.97), 40.3% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.96), 50% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.93), 60% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.89), 70% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.83), and 80% (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.74). Control IJs were transferred to Ringer’s solution after the adaptation period (\\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.99). After treatment, the mortality of IJs was assessed in the three replicates by counting the number of active and inactive nematodes using a counting chamber. Percentage IJ mortality in different replicates was used to calculate the mean water activity (MW) tolerated by 50% of the population (MW\\u003csub\\u003e50\\u003c/sub\\u003e) and the MW tolerated by the most tolerant 10% of the IJs population (MW\\u003csub\\u003e10\\u003c/sub\\u003e).\\u0026nbsp;\\u003c/p\\u003e\\n\\u003ch2\\u003eHypoxia Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eHypoxia tolerance of the isolates was assessed based on methods detailed by Zadji et al. (2014). For this assessment, 5,000 IJs from each isolate were placed into sealed 0.5-ml Eppendorf tubes containing distilled water and incubated under hypoxic conditions at 25°C in darkness for 24 or 72 hours. After exposure, IJs were transferred to Petri dishes containing 15 ml distilled water and further incubated at 25°C for 24 hours to evaluate recovery and survival rates. Tubes maintained in open conditions throughout incubation periods served as controls. Treatments were conducted with four replicates in a CRD and the entire experiment was repeated twice for reproducibility and confirmation of observed responses.\\u003c/p\\u003e\\n\\u003ch2\\u003eOxidative Stress Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eThe oxidative stress performance of the six isolates was assessed by storing pools of IJs in Ringer’s solution in the presence of hydrogen peroxide (H\\u003csub\\u003e2\\u003c/sub\\u003eO\\u003csub\\u003e2\\u003c/sub\\u003e) at room temperature (25°C). Freshly harvested IJ suspensions were separately exposed to H\\u003csub\\u003e2\\u003c/sub\\u003eO\\u003csub\\u003e2\\u003c/sub\\u003e in a 24-cell well plate in a CRD with three replicates, each containing 1,500 IJs in 400 µl of Ringer’s solution and sealed with Parafilm. For oxidative stress induction, 12.76 µl of 1.94 M H\\u003csub\\u003e2\\u003c/sub\\u003eO\\u003csub\\u003e2\\u0026nbsp;\\u003c/sub\\u003ewas added to each cell well to obtain a final H\\u003csub\\u003e2\\u003c/sub\\u003eO\\u003csub\\u003e2\\u003c/sub\\u003e concentration of 60 mM. IJs kept under control conditions were left at 25°C in cell wells without H\\u003csub\\u003e2\\u003c/sub\\u003eO\\u003csub\\u003e2\\u003c/sub\\u003e. To assess the IJs mortality over time, 50 µl aliquots from each experimental replicate were counted in a counting chamber daily for two weeks. The percentage IJ mortality was used to determine differences in the mean survival time of 50% of the population (ST\\u003csub\\u003e50\\u003c/sub\\u003e) and the survival time of the most tolerant 10% of the IJ population (ST\\u003csub\\u003e10\\u003c/sub\\u003e) for each strain. The determination of the ST values followed the same procedure as that described for the desiccation test.\\u003c/p\\u003e\\n\\u003ch1\\u003eData analysis\\u003c/h1\\u003e\\n\\u003cp\\u003eAll statistical analyses were conducted using R v. 4.4.3 (R Core Team, 2024). Data were first assessed for normality and homogeneity of variance using the Shapiro–Wilk and Levene’s tests, respectively. Where appropriate, percentage data were arcsine square root transformed to meet parametric assumptions. For temperature-dependent infectivity and reproduction assays, two-way ANOVA was used to examine the effects of isolate and temperature on larval mortality and infective juvenile (IJ) emergence. Post hoc comparisons were performed using Tukey’s HSD test. In the desiccation tolerance assay, percentage IJ mortality from replicate treatments was used to estimate the mean water activity (MW₅₀) tolerated by 50% of the population and MW₁₀ for the most tolerant 10% of IJs. The data were fitted to a cumulative normal distribution curve, and the mean and standard deviation from the fitted curve were used to derive MW values by minimizing the χ² value between experimental and expected values. Similarly, for oxidative stress tolerance, IJ mortality percentages were used to estimate the mean survival time (ST₅₀) and the survival time of the most tolerant 10% (ST₁₀) of the population. These values were also obtained from a cumulative normal distribution fitted to the data, using the same χ² minimization approach as in the desiccation assay. Hypoxia tolerance data were analysed using Kaplan–Meier survival analysis, and isolate differences were assessed using the log-rank test. Median survival times (ST₅₀) were extracted from the survival curves. Foraging ability across substrates and soil depths was analysed using generalized linear models (GLMs), with isolate, substrate, and depth treated as fixed effects. The appropriate error distribution (Poisson or binomial) was applied based on the nature of the response variable. All statistical tests were considered significant at P \\u0026lt; 0.05.\\u003c/p\\u003e\"},{\"header\":\"Results\",\"content\":\"\\u003ch2\\u003e\\u003cstrong\\u003eEffect of Temperature on Infectivity of IJs\\u003c/strong\\u003e\\u003c/h2\\u003e\\n\\u003cp\\u003eThe four key parameters measured to assess nematode infectivity, larval mortality at 72 hours post-inoculation, time until death of infected larvae, number of IJs established per FAW larva, and the percentage of larvae producing IJs are presented in Fig. 1 A-D. The percentage mortality of FAW larvae varied markedly across EPN isolates and temperature levels, as well as their interaction. Larval mortality exhibited strong temperature dependency. Mortality rates were negligible at 10\\u0026deg;C across all isolates and peaked between 25\\u0026deg;C and 30\\u0026deg;C (Fig. 1A), with values reaching above 90% in some isolates such as Ib-IART45 and Ib-CRIN68. A two-way ANOVA confirmed highly significant main effects of temperature (\\u003cem\\u003eF\\u003c/em\\u003e = 1017.94, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), isolate (\\u003cem\\u003eF\\u003c/em\\u003e = 119.89, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), and their interaction (\\u003cem\\u003eF\\u003c/em\\u003e = 16.73, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001) on larval mortality\\u003cem\\u003e.\\u0026nbsp;\\u003c/em\\u003eCorrespondingly, the time until death of infected larvae significantly declined with increasing temperatures, reaching the shortest average duration (36\\u0026ndash;48 hours) at 25\\u0026deg;C. At lower (10\\u0026ndash;15\\u0026deg;C) and higher (35\\u0026deg;C) extremes, larval death occurred more slowly, with mean times extending beyond 100 hours in some treatments (Fig. 1B). ANOVA analysis indicated statistically significant effects of temperature (\\u003cem\\u003eF\\u003c/em\\u003e = 167.04, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), isolate (\\u003cem\\u003eF\\u003c/em\\u003e = 28.02, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), and temperature \\u0026times; isolate interaction (\\u003cem\\u003eF\\u003c/em\\u003e = 4.27, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), supporting the hypothesis that both thermal and genetic factors shape virulence dynamics. The number of IJs successfully establishing per larva mirrored the mortality trend, increasing significantly with temperature up to 25\\u0026deg;C and subsequently declining. At 25\\u0026deg;C, the highest establishment rates were observed, particularly in \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e and \\u003cem\\u003eS. feltiae\\u003c/em\\u003e isolates, with average IJ counts per larva exceeding 45. In contrast, establishment at 10\\u0026deg;C was extremely low (\\u0026lt;5 IJs per larva) across all isolates (Fig. 1C). A highly significant effect of temperature (\\u003cem\\u003eF\\u003c/em\\u003e = 1070.14, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), isolate (\\u003cem\\u003eF\\u003c/em\\u003e = 113.19, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), and their interaction (\\u003cem\\u003eF\\u003c/em\\u003e = 13.18, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001) was observed, indicating that both host penetration and survival are profoundly influenced by thermal environment and nematode identity. IJ ability to complete their life cycle and reproduce within the host, as measured by the percentage of FAW larvae producing IJs, also showed strong temperature dependency. IJ emergence was not observed at 10\\u0026deg;C and 35\\u0026deg;C for any isolate. Between 15\\u0026deg;C and 30\\u0026deg;C, however, emergence rates increased progressively, peaking at 25\\u0026deg;C where values ranged between 85% and 90% for \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e, \\u003cem\\u003eS. feltiae\\u003c/em\\u003e and \\u003cem\\u003eO. myriophilus\\u0026nbsp;\\u003c/em\\u003e(Fig. 1D). ANOVA again demonstrated significant effects of temperature (\\u003cem\\u003eF\\u003c/em\\u003e = 1184.93, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), isolate (F = 83.41, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), and their interaction (\\u003cem\\u003eF\\u003c/em\\u003e = 18.80, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), highlighting variation in reproductive success under fluctuating environmental conditions. These underscore the critical influence of temperature and isolate identity on the infectivity, pathogenicity, and reproductive success of EPN.\\u0026nbsp;\\u003c/p\\u003e\\n\\u003ch2\\u003eEffect of Temperature on Reproduction of IJs\\u003c/h2\\u003e\\n\\u003cp\\u003eThe percentage of cadavers producing progeny varied significantly among isolates and across temperatures (Fig. 2). A two-way ANOVA revealed significant main effects for both EPN isolate (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(5, 96)\\u003c/sub\\u003e = 11.28, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001) and temperature (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(3, 96)\\u003c/sub\\u003e = 189.47, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), as well as a highly significant interaction effect between isolate and temperature (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(15, 96)\\u003c/sub\\u003e = 14.05, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001). Overall, the proportion of cadavers yielding progeny increased progressively from 15\\u0026deg;C to 25\\u0026deg;C and declined slightly at 30\\u0026deg;C, with marked variation in reproductive success between the isolates (Fig. 2A). The timing of first IJ emergence from cadavers, measured in days post-infection (dpi), was also significantly influenced by both isolate identity and ambient temperature. Two-way ANOVA results indicated highly significant effects of isolate (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(5, 96)\\u003c/sub\\u003e = 15.05, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), temperature (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(3, 96)\\u003c/sub\\u003e = 151.66, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), and their interaction (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(15, 96)\\u003c/sub\\u003e = 18.48, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001). On average, emergence commenced earlier at higher temperatures, with the earliest onset observed at 25\\u0026deg;C and delayed emergence at lower temperatures (Fig. 2B). Similarly, the duration of the IJ emergence period differed significantly among isolates and temperature treatments. The main effects of isolate (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(5, 96)\\u003c/sub\\u003e = 10.17, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001) and temperature (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(3, 96)\\u003c/sub\\u003e = 155.25, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), along with the interaction between the two factors (\\u003cem\\u003eF\\u003c/em\\u003e\\u003csub\\u003e(15, 96)\\u003c/sub\\u003e = 12.61, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001), were all statistically significant. The longest emergence durations were generally recorded at intermediate temperatures (20\\u0026ndash;25\\u0026deg;C), whereas shorter durations were observed at the lower and upper extremes of the temperature range (Fig. 2C). The total number of IJs emerging per cadaver (measured in thousands) also showed significant variability across treatments with highest progeny output clustered around 25\\u0026deg;C for most isolates, and lower outputs observed at both 15\\u0026deg;C and 30\\u0026deg;C. Some isolates, particularly \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e and the \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e strains, maintained relatively high reproductive output across a broader temperature range (Fig.\\u003cem\\u003e\\u0026nbsp;2\\u003c/em\\u003eD).\\u0026nbsp;\\u003c/p\\u003e\\n\\u003ch2\\u003eForaging Behaviour\\u003c/h2\\u003e\\n\\u003cp\\u003eThe mean number of IJs attaching to FAW larvae varied significantly across substrate types and EPN isolates (ANOVA, \\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05). Substrate type had a strong effect on IJ attachment, with markedly lower attachment observed on dry soil (0% moisture) and significantly higher attachment under moist conditions (particularly at 20% soil moisture). Among the substrates, 20% soil moisture supported the highest attachment levels, with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates Ib-IART45 and Ib-ITUC102 recording the highest mean IJ attachments (50 \\u0026plusmn; 5 and 46 \\u0026plusmn; 4.5, respectively). In contrast, 0% soil moisture yielded the lowest mean IJ counts, particularly for \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32) and \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68), with values as low as 4 \\u0026plusmn; 1.1 and 5 \\u0026plusmn; 1.2, respectively (Fig. 3). The maize leaf surface presented an intermediate attachment potential. Notably, \\u003cem\\u003eS. nepalense\\u003c/em\\u003e (Ib-HORT) and \\u003cem\\u003eS. feltiae\\u003c/em\\u003e (Za-SAM) achieved attachment levels comparable to those on 10% soil moisture, averaging 33 \\u0026plusmn; 3.5 and 28 \\u0026plusmn; 3.0 IJs per larva, respectively. These findings underscore both substrate-specific differences in attachment efficiency and isolate-level variability in host-finding and infection initiation traits under varying environmental conditions.\\u003c/p\\u003e\\n\\u003cp\\u003eThe statistical analysis of the depth penetration assay evaluating the ability of six EPN IJs isolates to infect FAW larvae across four soil depths revealed significant isolate- and depth-dependent differences. A two-way ANOVA confirmed that isolate, depth, and their interaction significantly influenced penetration rates (\\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.001 for all factors). The highest mean penetration rates were observed at the soil surface, where most isolates achieved infection levels exceeding 80%, with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates Ib-IART45 and Ib-ITUC102 demonstrating superior performance even at increased depths (Fig. 4). Penetration rates declined progressively with soil depth, particularly at 10 cm, where all isolates recorded markedly lower effectiveness. Descriptive statistics underscored this trend, and post hoc Tukey HSD tests revealed statistically significant differences among isolates and depths, underscoring clear vertical stratification of infectivity potential among isolates.\\u003c/p\\u003e\\n\\u003ch2\\u003eDesiccation Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eThe minimum water activity required to maintain 50% (MW₅₀) and 10% (MW₁₀) IJ survival differed significantly among the six EPN isolates (Fig. 5). \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e isolates Ib-IART45 and Ib-ITUC102 recorded the least water activity (MW₅₀ \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e ~0.89), indicating superior overall desiccation resilience. In contrast, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68) exhibited the lowest desiccation tolerance (MW₅₀ ~0.97), suggesting a need for more water to maintain survival and infectivity of 50% population and more rapid decline in population viability under drying conditions. Similarly, the 10% best performing IJs of Ib-IART45 and Ib-ITUC102 tolerated lower \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e values (MW₁₀ ~0.798), while \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e lost 90% of their IJs at slightly higher \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e (MW₁₀ ~0.857). We also observed that at \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.99, IJ survival was highest in \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-IART45) (99.0 \\u0026plusmn; 2.3%) and lowest in \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68) (88.0 \\u0026plusmn; 1.7%) (Table 1). At \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.98, survival remained above 84% for all isolates except \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (76.0 \\u0026plusmn; 2.3%). Subsequent reduction of \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e showed \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates maintained high survival. At the lowest \\u003cem\\u003ea\\u003c/em\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e 0.74, survival dropped to \\u0026lt;10% across all isolates, with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-IART45) having the highest value (6.7 \\u0026plusmn; 1.7%) and \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e showing complete mortality (0.0%).\\u003c/p\\u003e\\n\\u003cp\\u003e\\u003cstrong\\u003eTable\\u0026nbsp;\\u003c/strong\\u003e\\u003cstrong\\u003e1\\u003c/strong\\u003e\\u003cstrong\\u003e. Percentage survival of IJs of six EPN isolates to decreasing\\u0026nbsp;\\u003c/strong\\u003e\\u003cstrong\\u003ea\\u003c/strong\\u003e\\u003cstrong\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e\\u003c/strong\\u003e\\u003cstrong\\u003e\\u0026nbsp;levels.[1]\\u003c/strong\\u003e\\u003c/p\\u003e\\n\\u003ctable border=\\\"0\\\" cellspacing=\\\"0\\\" cellpadding=\\\"0\\\"\\u003e\\n \\u003ctbody\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eWater Activity (\\u003c/strong\\u003e\\u003cstrong\\u003ea\\u003c/strong\\u003e\\u003cstrong\\u003e\\u003csub\\u003ew\\u003c/sub\\u003e\\u003c/strong\\u003e\\u003cstrong\\u003e)\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003eH. bacteriophora\\u0026nbsp;(Ib-CRIN68)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003eS. carpocapsae\\u0026nbsp;(Ib-IART45)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003eS. carpocapsae\\u0026nbsp;(Ib-ITUC102)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003eO. myriophilus\\u0026nbsp;(Ib-FRIN32)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003eS. nepalense\\u0026nbsp; (Ib-HORT)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003eS. feltiae\\u0026nbsp; \\u0026nbsp; \\u0026nbsp; \\u0026nbsp;(Za-SAM)\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.99\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e88.0 \\u0026plusmn; 1.7\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e99.0 \\u0026plusmn; 2.3\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e98.0 \\u0026plusmn; 2.9\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e95.0 \\u0026plusmn; 1.7\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e85.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e95.0 \\u0026plusmn; 1.2\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.98\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e76.0 \\u0026plusmn; 2.3\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e94.8 \\u0026plusmn; 1.7\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e91.0 \\u0026plusmn; 2.3\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e87.0 \\u0026plusmn; 1.2\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e84.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e85.0 \\u0026plusmn; 1.2\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.97\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e50.5 \\u0026plusmn; 2.9\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e87.85 \\u0026plusmn; 2.3\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e87.5 \\u0026plusmn; 1.7\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e77.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e79.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e74.0 \\u0026plusmn; 1.7\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.96\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e30.0 \\u0026plusmn; 3.5\\u003csup\\u003ed\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e85.0 \\u0026plusmn; 2.9\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e78.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e50.8 \\u0026plusmn; 2.3\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e78.5 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e62.0 \\u0026plusmn; 1.7\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.93\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e26.0 \\u0026plusmn; 2.9\\u003csup\\u003ed\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e65.0 \\u0026plusmn; 3.5\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e62.0 \\u0026plusmn; 2.9\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e47.0 \\u0026plusmn; 2.3\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e55.0 \\u0026plusmn; 2.9\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e58.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.89\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e17.0 \\u0026plusmn; 2.3\\u003csup\\u003ed\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e52.0 \\u0026plusmn; 2.9\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e50.9 \\u0026plusmn; 2.9\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e32.0 \\u0026plusmn; 2.3\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e45.0 \\u0026plusmn; 2.3\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e49.0 \\u0026plusmn; 2.3\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.83\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e7.0 \\u0026plusmn; 1.7\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e30.0 \\u0026plusmn; 2.3\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e25.0 \\u0026plusmn; 2.3\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e10.2 \\u0026plusmn; 1.7\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e12.0 \\u0026plusmn; 1.7\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e11.8 \\u0026plusmn; 1.7\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e0.74\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 94px;\\\"\\u003e\\n \\u003cp\\u003e00.00\\u003csup\\u003ec\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e6.7 \\u0026plusmn; 1.7\\u003csup\\u003ea\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 85px;\\\"\\u003e\\n \\u003cp\\u003e3.0 \\u0026plusmn; 1.2\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 80px;\\\"\\u003e\\n \\u003cp\\u003e2.3 \\u0026plusmn; 1.2\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e2.0 \\u0026plusmn; 1.2\\u003csup\\u003eb\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 86px;\\\"\\u003e\\n \\u003cp\\u003e4.0 \\u0026plusmn; 1.2\\u003csup\\u003eab\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003c/tbody\\u003e\\n\\u003c/table\\u003e\\n\\u003cp\\u003e[1] Values represent mean \\u0026plusmn; standard error (SE) \\u0026nbsp;of IJ survival at each water activity level. Different superscript letters within each row indicate statistically significant differences among EPN isolates at that specific water activity level (Tukey\\u0026rsquo;s HSD test, p \\u0026lt; 0.05).\\u003c/p\\u003e\\n\\u003ch2\\u003eHypoxia Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eSurvival data demonstrated significant variability across isolates and exposure durations, as confirmed by analysis of variance (ANOVA). After 24 hours of exposure, survival rates ranged from approximately 56.3% to 76.2%, with \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e (Ib-ITUC102) exhibiting the highest survival, followed closely by \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e (Ib-IART45) (Fig. 6A). The lowest survival was recorded in \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32). We found statistically significant differences among isolates (\\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05), indicating distinct hypoxia tolerance profiles in the short term. Following 72 hours of hypoxia, a marked reduction in survival was observed across all isolates, with survival ranging from 27.9% to 63.4% (Fig. 6B). Again, \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-ITUC102) maintained the highest tolerance, whereas \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32) and \\u003cem\\u003eS. nepalense\\u003c/em\\u003e (Ib-HORT) were the most adversely affected. Statistical analysis confirmed a significant isolate effect (\\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05), and visual comparison of both timepoints indicated time-dependent reduction in IJ viability under sustained oxygen deprivation.\\u003c/p\\u003e\\n\\u003ch2\\u003eOxidative stress Tolerance\\u003c/h2\\u003e\\n\\u003cp\\u003eThe oxidative stress tolerance was evaluated by assessing the survival time of the most tolerant 10% (ST₁₀) and 50% (ST₅₀) of IJs exposed to 60 mM H₂O₂. Survival time varied significantly among the isolates (\\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05), indicating differential capacities to withstand oxidative stress (Fig. 7). \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e (Za-SAM) and \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-IART45) exhibited the highest oxidative stress tolerance, with ST₁₀ values of 16.8 \\u0026plusmn; 0.4 h and 16.5 \\u0026plusmn; 0.6 h, respectively, and corresponding ST₅₀ values of 27.34 \\u0026plusmn; 0.8 h and 26.77 \\u0026plusmn; 0.9 h, significantly higher than those of other isolates. \\u003cem\\u003eSteinernema nepalense\\u003c/em\\u003e (Ib-HORT) recorded the lowest oxidative stress resistance, with an ST₁₀ of 13.1 \\u0026plusmn; 0.5 h and an ST₅₀ of 18.93 \\u0026plusmn; 0.6 h, which were significantly lower (\\u003cem\\u003ep\\u003c/em\\u003e \\u0026lt; 0.05) than most other isolates. Moderate oxidative stress performance was observed for \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68), \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-ITUC102), and \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32), which did not differ significantly from one another in ST₁₀ and ST₅₀ measures. The considerable variation in the oxidative resilience of the isolates suggest potential adaptive differences in physiological responses to environmental stress (Fig. 7).\\u003c/p\\u003e\"},{\"header\":\"Discussion\",\"content\":\"\\u003cp\\u003eThe ecological characterisation of EPN is a critical step in the development and deployment of effective biological control agents for sustainable pest management. This study evaluated six indigenous EPN isolates previously identified morphologically and molecularly from Nigeria; \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68), \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-IART45 and Ib-ITUC102), \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32), \\u003cem\\u003eS. nepalense\\u003c/em\\u003e (Ib-HORT), and S. feltiae (Za-SAM), with respect to key ecological traits that influence their infectivity and persistence. These traits included thermal sensitivity, reproductive potential, tolerance to abiotic stresses (desiccation, hypoxia, oxidative stress) and host-seeking behaviour.\\u003c/p\\u003e\\u003cp\\u003eThe six EPN isolates from Nigeria showed clear temperature-dependent patterns in both host infection and reproduction. Overall, moderate temperatures (in the mid-20\\u0026deg;C range) supported the highest virulence and nematode progeny production, whereas extreme low or high temperatures constrained performance. These trends align with well-documented thermal optima for EPNs: for example, most EPN species achieve maximal infection and reproduction around 25\\u0026deg;C and cannot reproduce below ~\\u0026thinsp;10\\u0026ndash;15\\u0026deg;C (Koppenh\\u0026ouml;fer \\u0026amp; Kaya, \\u003cspan citationid=\\\"CR25\\\" class=\\\"CitationRef\\\"\\u003e1999\\u003c/span\\u003e; Raheel et al., \\u003cspan citationid=\\\"CR50\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e). In our study, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68) and \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (Ib-IART45, Ib-ITUC102) maintained high infectivity even at elevated temperatures, reflecting their adaptation to tropical climates. Notably, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e sustained strong virulence up to 30\\u0026ndash;35\\u0026deg;C, consistent with reports that this species can perform robustly at the upper thermal limits of IJs activity (Aatif et al., \\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Matuska-Lyzwa et al., \\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e). By contrast, the \\u003cem\\u003eS. feltiae\\u003c/em\\u003e isolate Za-SAM, a species typically native to cooler regions of Nigeria, showed reduced infectivity and yield at higher temperatures. This mirrors the known thermal sensitivity of \\u003cem\\u003eS. feltiae\\u003c/em\\u003e, which exhibits diminished activity above ~\\u0026thinsp;30\\u0026deg;C. In fact, storage of \\u003cem\\u003eS. feltiae\\u003c/em\\u003e at 35\\u0026deg;C for even 1 hour can dramatically curtail its motility, with lethal effects by 37\\u0026deg;C (Matuska-Lyzwa et al., \\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e). Such thermal intolerance likely explains the poorer performance of the Za-SAM isolate under hot conditions and underscores the importance of matching EPN strains to ambient thermal regimes.\\u003c/p\\u003e\\u003cp\\u003eReproductive capacity followed similar temperature-related patterns. All isolates failed to recycle in hosts at very low temperatures (no reproduction occurred at 5\\u0026deg;C, and only \\u003cem\\u003eS. feltiae\\u003c/em\\u003e managed limited reproduction by 10\\u0026deg;C). As temperatures rose, development accelerated and brood sizes increased for all species. The highest yields of IJs in our trials were produced by \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68), consistent with previous studies showing \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e can generate exceptionally large progenies in \\u003cem\\u003eGalleria\\u003c/em\\u003e hosts (Levy et al., \\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Raheel et al., \\u003cspan citationid=\\\"CR50\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e). We also observed that \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e and the \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates had faster kill and emergence at 25\\u0026ndash;30\\u0026deg;C than at cooler temperatures, whereas \\u003cem\\u003eS. feltiae\\u003c/em\\u003e required longer intervals to produce IJs, especially at suboptimal temperatures. \\u003cem\\u003eSteinernema\\u003c/em\\u003e species like \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e can kill hosts and recycle more rapidly at 25\\u0026ndash;28\\u0026deg;C (often within a week), whereas \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e typically takes a few days longer to emerge (Brown et al., \\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e; Susurluk \\u0026amp; Ulu, \\u003cspan citationid=\\\"CR57\\\" class=\\\"CitationRef\\\"\\u003e2015\\u003c/span\\u003e). At the highest temperature tested in our study, some drop-off in IJ yield was noted for all isolates, suggesting that extreme heat imposes stress on nematode development and symbiont functioning. It is well known that sustained exposure to \\u0026gt;\\u0026thinsp;32\\u0026deg;C adversely affects EPN growth, reproduction and survival (Karanastasi et al., \\u003cspan citationid=\\\"CR21\\\" class=\\\"CitationRef\\\"\\u003e2025\\u003c/span\\u003e; Lillis et al., \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). Nonetheless, the indigenous tropical isolates in this study tolerated heat better than many temperate-strain EPNs reported in literature, reinforcing the ecological premise that local nematode populations are better adapted to the prevailing thermal conditions of their environments. The ability to adapt to varying thermal conditions is essential for biocontrol agents in SSA agroecosystems, where soil temperatures can vary considerably. Our results show that considering temperature optima and limits is crucial when choosing suitable EPN isolates for field evaluation. The Nigerian isolate \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e Ib-CRIN68 has a broad thermal activity range, making it effective in warm climates.\\u003c/p\\u003e\\u003cp\\u003eThe tested EPN isolates demonstrated distinct foraging strategies, following the ambusher versus cruiser host-finding behaviour. The two \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates Ib-IART45 and Ib-ITUC102 showed ambush foraging by staying near the soil surface and adopting a \\u0026ldquo;sit-and-wait\\u0026rdquo; tactic for mobile hosts. This was evident as they infected hosts near the soil surface with limited deep soil movement, consistent with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e's ecology of targeting insects at or above the soil interface (Bal \\u0026amp; Grewal, \\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e2015\\u003c/span\\u003e; Raja et al., \\u003cspan citationid=\\\"CR51\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e). Ambushers like \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e are known to conserve energy by nictating or standing upright near the surface, and only a small fraction of their population actively disperses far from the release point (Chen \\u0026amp; Glazer, \\u003cspan citationid=\\\"CR8\\\" class=\\\"CitationRef\\\"\\u003e2005\\u003c/span\\u003e; Wright et al., \\u003cspan citationid=\\\"CR60\\\" class=\\\"CitationRef\\\"\\u003e1997\\u003c/span\\u003e). Interestingly, even ambushers can exhibit a subset of highly motile \\u0026ldquo;sprinter\\u0026rdquo; individuals that disperse rapidly when host cues (such as CO₂ or other volatiles) are detected. Our observations of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e reaching and infecting hosts a few tens of centimeters away concur with reports that a small proportion (~\\u0026thinsp;1\\u0026ndash;2%) of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e IJs can travel\\u0026thinsp;\\u0026gt;\\u0026thinsp;10 cm in soil columns (Bal \\u0026amp; Grewal, \\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e2015\\u003c/span\\u003e). This dual strategy, mostly localized waiting with occasional long-distance forays, likely maximizes the chance of encountering a susceptible insect host in heterogenous soil environments.\\u003c/p\\u003e\\u003cp\\u003eIn contrast, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e (Ib-CRIN68) and \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e (Ib-FRIN32) showed more cruiser-like foraging behaviour. They actively explored deeper soil layers and were able to locate hosts buried at greater depths or at further horizontal distances. \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e in particular is known as an active cruiser that continuously moves through soil pore water in search of sedentary or subterranean hosts. In our depth-gradient assays, H. bacteriophora IJs readily penetrated to lower strata (e.g. \\u0026gt;15 cm depth) and successfully infected hosts there, whereas \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e infections were concentrated in the upper 10 cm. Host-seeking observations show that \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e spp. tend to distribute deeper in the soil profile than \\u003cem\\u003eSteinernema\\u003c/em\\u003e ambushers. For example, in surveys \\u003cem\\u003eS. feltiae\\u003c/em\\u003e were found mostly in the top 0\\u0026ndash;15 cm of soil, whereas \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e can be recovered from much greater depths (Neumann \\u0026amp; Shields, \\u003cspan citationid=\\\"CR42\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e; Williams et al., \\u003cspan citationid=\\\"CR59\\\" class=\\\"CitationRef\\\"\\u003e2013\\u003c/span\\u003e). Our data similarly suggests that the Ib-CRIN68 isolate of \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e is adept at vertical movement, an advantageous trait for targeting soil-dwelling stages of pests, such as pupating FAW. The ability to forage actively in the soil may also help cruisers find immobile hosts like cocoons or grubs, complementing the ambushers\\u0026rsquo; strength against surface-active insects. Meanwhile, the behaviour of \\u003cem\\u003eS. nepalense\\u003c/em\\u003e Ib-HORT and \\u003cem\\u003eS. feltiae\\u003c/em\\u003e Za-SAM isolates appeared intermediate. They neither strictly waited at the surface nor ranged as widely as \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e, suggesting a more flexible foraging strategy. Many \\u003cem\\u003eSteinernema\\u003c/em\\u003e spp. are known to be intermediate strategists that can both ambush and cruise to some extent (Rakubu et al., \\u003cspan citationid=\\\"CR52\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e). This adaptable foraging strategy enables them to utilize various host niches, although it may not be as efficiently specialized as the extreme ambusher (\\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e) or the extreme cruiser (\\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e).\\u003c/p\\u003e\\u003cp\\u003eSoil moisture had a pronounced influence on foraging efficacy. In moderately moist soil, almost at field capacity, all isolates moved and located hosts with the highest success. However, under drier conditions, host-finding declined, particularly for the cruiser-type nematodes that rely on continuous water films for movement. Nematodes are essentially aquatic in locomotion, gliding along water-filled pores thus insufficient moisture breaks the continuity of those films, impeding IJ mobility (Kaspi et al., \\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e2010\\u003c/span\\u003e). We observed that as the soil became drier, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e and \\u003cem\\u003eS. nepalense\\u003c/em\\u003e showed reduced dispersal and tended to congregate in deeper, more humid layers if available. Nematodes will migrate downward as surface soil desiccates, accumulating at depths where humidity is higher (Cabanillas, \\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e; Duncan \\u0026amp; McCoy, \\u003cspan citationid=\\\"CR10\\\" class=\\\"CitationRef\\\"\\u003e2001\\u003c/span\\u003e; Gouge et al., \\u003cspan citationid=\\\"CR13\\\" class=\\\"CitationRef\\\"\\u003e2000\\u003c/span\\u003e; Yadav \\u0026amp; Lalramliana, \\u003cspan citationid=\\\"CR61\\\" class=\\\"CitationRef\\\"\\u003e2012\\u003c/span\\u003e). In one study, \\u003cem\\u003eS. riobrave\\u003c/em\\u003e was seen to move 15\\u0026ndash;23 cm deep over four weeks of gradual surface drying, effectively tracking the receding moisture front (Gouge et al., \\u003cspan citationid=\\\"CR13\\\" class=\\\"CitationRef\\\"\\u003e2000\\u003c/span\\u003e). A similar pattern in our experiments suggests that the Nigerian cruisers actively seek favourable moisture microhabitats, which would enhance their survival during dry spells. On the other hand, \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e (ambusher) was less able to escape drying soil by migration. Instead, its strategy under low moisture may be to enter a quiescent state near the surface and wait for either a host or the return of moisture. Ambusher species like \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e often have greater desiccation tolerance, enabling them to survive near the surface until a host contacts them or rain rehydrates the soil (Shapiro-Ilan et al., \\u003cspan citationid=\\\"CR55\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e). Moreover, the virulence of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e in dry soil can be restored upon re-moistening of the soil, suggesting that the IJs remain viable in a dormant state and are capable of resuming their infective activity once favourable conditions are re-established (Grant \\u0026amp; Villani, \\u003cspan citationid=\\\"CR15\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e). These behavioural differences mean in an applied context, that \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e might be more effective when pests are on or near the soil surface (and intermittent dry periods occur), whereas \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e could be superior for targets in the soil profile provided adequate moisture is present or irrigation is used. Overall, our foraging assays highlight that both moisture and soil depth interact with nematode behavioural traits. The most effective biocontrol may be achieved by matching isolate behaviour to pest ecology such as deploying ambushers for mobile foliar larvae and cruisers for soil-dwelling stages or by combining species to cover multiple strata and moisture conditions in the field (Bal \\u0026amp; Grewal, \\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e2015\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003eAn important aspect of the ecological fitness of EPNs is their ability to withstand environmental stresses. We found considerable isolate-specific differences in tolerance to desiccation (dryness), low oxygen, and oxidative stress. \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e Ib-IART45/ITUC102 stood out for its superior desiccation tolerance, and its IJ survival after exposure to low humidity (or dry soil) was significantly higher than that of the \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e and other \\u003cem\\u003eSteinernema\\u003c/em\\u003e isolates. Generally, \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e is considered to be one of the most desiccation-tolerant nematode species. For instance, Shapiro-Ilan et al. (\\u003cspan citationid=\\\"CR55\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e) reported \\u003cem\\u003ethat S. carpocapsae\\u003c/em\\u003e IJs survived desiccating conditions far better than heterorhabditids, with \\u003cem\\u003eS. feltiae\\u003c/em\\u003e also ranking high and \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e generally the least tolerant. Our results mirror that pattern, the \\u003cem\\u003eS. feltiae\\u003c/em\\u003e Za-SAM isolate had the second-highest desiccation survival, whereas \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e was among the most sensitive to drying. Desiccation tolerance in EPNs is thought to be linked to behavioural and physiological adaptations; ambushers like \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e often remain near the soil surface and have evolved mechanisms to survive transient drought (Glazer, \\u003cspan citationid=\\\"CR11\\\" class=\\\"CitationRef\\\"\\u003e2022\\u003c/span\\u003e; Ramakrishnan et al., \\u003cspan citationid=\\\"CR53\\\" class=\\\"CitationRef\\\"\\u003e2022\\u003c/span\\u003e). In contrast, cruisers avoid dry conditions by moving deeper, as earlier discussed, and consequently may not have invested in as strong desiccation-hardiness mechanisms which explains the lower survival of \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e in our desiccation assays. It is encouraging that even the more sensitive isolates in our study still retained some viability after short dry exposures, suggesting that a fraction of the population can endure brief droughts. Additionally, we noted no obvious intraspecific variation in desiccation survival between the two \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e strains, which is consistent with reports that different strains of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e tend to exhibit uniformly high desiccation tolerance (Shapiro-Ilan et al., \\u003cspan citationid=\\\"CR55\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e). Overall, the ability of the \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e and \\u003cem\\u003eS. feltiae\\u003c/em\\u003e isolates from Nigeria to better survive drying conditions could be advantageous for use in regions with irregular rainfall or for above-ground applications where desiccation risk is high.\\u003c/p\\u003e\\u003cp\\u003eAll isolates showed reduced survival under hypoxic (low oxygen) conditions, though with subtle differences. When subjected to oxygen-depleted environments (simulating waterlogged or compacted soils), \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e and \\u003cem\\u003eS. nepalense\\u003c/em\\u003e survived slightly longer than the \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e and \\u003cem\\u003eOscheius\\u003c/em\\u003e isolates. This may reflect adaptation of cruisers to burrowing into less aerated soil pockets (Kung et al., \\u003cspan citationid=\\\"CR28\\\" class=\\\"CitationRef\\\"\\u003e1990a\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR29\\\" class=\\\"CitationRef\\\"\\u003eb\\u003c/span\\u003e). Nonetheless, the overall intolerance of the nematodes to hypoxia was evident such that prolonged exposure to \\u0026lt;\\u0026thinsp;1% O₂ led to high mortality across the board. This finding is expected since EPN IJs are aerobic organisms that rely on dissolved oxygen in soil water. As soil oxygen drops, nematode metabolism and survival sharply decline. For instance, oxygen levels near 1% drastically impair EPN viability and infectivity, and the survival of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e and \\u003cem\\u003eS. glaseri\\u003c/em\\u003e IJs plummeted as oxygen was reduced from ambient (20%) to near-anoxic levels (Matuska-Lyzwa et al., \\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e). Our isolates likely behave similarly, with heavy, water-saturated soils (common in the humid tropics during rains) posing a risk to their persistence. Interestingly, we did observe that nematodes in our experiments often sought out air pockets or moved to the soil surface in response to waterlogging, suggesting a behavioural escape from hypoxia. Such behaviour has been noted that IJs can sense gradients in oxygen or CO₂ and migrate toward more favourable conditions (Hoctor et al., \\u003cspan citationid=\\\"CR20\\\" class=\\\"CitationRef\\\"\\u003e2013\\u003c/span\\u003e; Maushe et al., \\u003cspan citationid=\\\"CR38\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). In practical terms, this means EPN applications should avoid fully waterlogged conditions; good soil drainage will promote nematode survival. Also, soil texture matters, coarse, well-aerated soils are more nematode-friendly than heavy clays that induce anaerobic micro-sites. While we did not identify a dramatically hypoxia-tolerant isolate, the slight edge of \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e under low O₂ might relate to its natural occurrence in deeper soil. Still, hypoxia remains a limiting factor for all, reinforcing that EPNs work best under moderate moisture without oxygen starvation.\\u003c/p\\u003e\\u003cp\\u003eWhen exposed to oxidative challenges, such as hydrogen peroxide in our assays, some isolates fared noticeably better. In particular, \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e Ib-IART45 showed higher survival and maintained mobility longer under oxidative stress than \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e or \\u003cem\\u003eS. feltiae\\u003c/em\\u003e. This could indicate a more robust antioxidant defense system in \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e. Effective scavenging of reactive oxygen species (ROS) is critical for EPNs, both in the soil environment and during infection of the host. Insect hosts actively mount an oxidative immune response \\u0026ndash; generating superoxide, peroxide, and other ROS to attack invading nematodes (Lalitha et al., \\u003cspan citationid=\\\"CR31\\\" class=\\\"CitationRef\\\"\\u003e2018\\u003c/span\\u003e; Sumaya et al., \\u003cspan citationid=\\\"CR56\\\" class=\\\"CitationRef\\\"\\u003e2018\\u003c/span\\u003e). Nematodes that can withstand this onslaught have a better chance to establish infection. \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e is known for producing a lethal toxin (via its symbiont \\u003cem\\u003eXenorhabdus\\u003c/em\\u003e) that rapidly kills the host, which might shorten the window of exposure to host immune defense, indirectly reducing oxidative damage (Lu et al., \\u003cspan citationid=\\\"CR35\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e; Watanabe et al., \\u003cspan citationid=\\\"CR58\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). Additionally, prior studies on the nematode models, \\u003cem\\u003eC. elegans\\u003c/em\\u003e, show that exposure to peroxides causes immediate loss of mobility and depressed metabolism, but young nematodes can recover if their antioxidant enzymes like peroxiredoxins and catalases, are effective (Kumsta et al., \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e). The superior oxidative stress survival of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e could reflect such efficient detoxification systems. It is possible that this isolate constitutively expresses high levels of catalase, superoxide dismutase, or peroxiredoxin that neutralize ROS, a trait that would be beneficial during the early stages of infection when the insect\\u0026rsquo;s immune burst is highest. By contrast, \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e Ib-CRIN68 showed more oxidative damage (lower survival) in our test, which might relate to its strategy of relying on a supportive mutualistic bacterium (\\u003cem\\u003ePhotorhabdus\\u003c/em\\u003e) to overcome host defense, \\u003cem\\u003ePhotorhabdus\\u003c/em\\u003e produces immunosuppressive factors but perhaps less in terms of ROS-scavengers. Another intriguing observation was that the \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e isolate had relatively good oxidative tolerance, almost on par with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e. \\u003cem\\u003eOscheius\\u003c/em\\u003e spp. are not symbiotically tied to \\u003cem\\u003eXenorhabdus/Photorhabdus\\u003c/em\\u003e; some are associated with other bacteria or can be facultative pathogens. Their pathogenicity often depends on releasing their own array of bacteria upon host entry (Onwong et al., \\u003cspan citationid=\\\"CR47\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). The resilience of \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e to oxidative stress in our assays suggests it may possess inherent protective mechanisms (possibly due to its free-living lineage background) or perhaps carries bacteria that aid in detoxification. While literature on EPN oxidative stress tolerance is scant, our results suggest this trait could underlie differences in virulence and field persistence. Isolates that better endure oxidative stress might survive longer on foliage (exposed to UV and oxidative conditions) or overcome host immune reactions more successfully. This could partly explain why \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e was so virulent in our trials, its physiological hardiness complements its aggressive infection strategy. In sum, screening for oxidative stress tolerance, alongside desiccation and hypoxia tolerance, provides a more complete picture of an EPN isolate\\u0026rsquo;s suitability for biocontrol deployment in challenging environments.\\u003c/p\\u003e\\u003cp\\u003eTrade-offs exist between virulence, environmental tolerance, and reproductive fitness (J. Mukuka et al., \\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e2010\\u003c/span\\u003e). For instance, the most desiccation-tolerant species (\\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e) are not the most fecund reproducers, and the most fecund (\\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e) are not very desiccation-tolerant. This necessitates a strategic approach to biocontrol to either formulate consortia of complementary EPN isolates or tailor the choice of isolate to the specific pest and environment. In practical IPM, one might apply \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e for immediate knockdown of FAW larvae in the crop canopy, combined with \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e for longer-lasting suppression of the next generation in the soil. There is evidence that mixed-species applications can sometimes yield additive benefits, as different nematodes occupy slightly different niches and timescales of action though careful consideration of competition and compatibility is needed. Our findings suggest that \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e Ib-CRIN68 and \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e Ib-IART45 together would make a formidable pair, the former ensuring persistence and recycling in soil and the latter providing quick action against active larvae. Additionally, \\u003cem\\u003eO. myriophilus\\u003c/em\\u003e could be included to bolster resilience to stress, possibly as a stress-hardy backup that might sustain population when others wane. The demonstrated trait superiority of these isolates likely stems from their evolutionary history in Nigerian agroecosystems, for example Ib-CRIN68 coming from a farm soil that undergoes periodic drying and heating, selecting for a hardy yet virulent phenotype, and Ib-IART45 originating from an area with intense insect pressure, selecting for high pathogenicity. It would be valuable in future work to investigate the genetic or physiological basis of these traits. Previous and recent research are identifying genetic markers (heat-shock proteins, anhydrobiosis-related genes, antioxidant enzymes) that correlate with stress tolerance in nematodes (Grewal et al., \\u003cspan citationid=\\\"CR16\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e; Hao et al., \\u003cspan citationid=\\\"CR19\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e; Maushe et al., \\u003cspan citationid=\\\"CR38\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e; Segal \\u0026amp; Glazer, \\u003cspan citationid=\\\"CR54\\\" class=\\\"CitationRef\\\"\\u003e2000\\u003c/span\\u003e). Unravelling these mechanisms in our top isolates could enable marker-assisted selection or even bioengineering to further improve them. In essence, the diverse performances observed affirm that isolate selection is crucial and by choosing the right nematode combinations one can achieve reliable biocontrol even under the challenging conditions of SSA farmlands.\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\n\\u003cli\\u003eAatif, H. M., Hanif, M. S., Raheel, M., Ferhan, M., Mansha, M. Z., Khan, A. A., Ullah, M. I., Shakeel, Q., \\u0026amp; Ali, S. (2020). Temperature dependent virulence of the entomopathogenic nematodes against immatures of the oriental fruit fly, \\u003cem\\u003eBactrocera dorsalis\\u003c/em\\u003e Hendel (Diptera: Tephritidae). \\u003cem\\u003eEgyptian Journal of Biological Pest Control\\u003c/em\\u003e,\\u003cem\\u003e 30\\u003c/em\\u003e(1), 1-6. https://doi.org/10.1186/S41938-020-00248-7\\u003c/li\\u003e\\n\\u003cli\\u003eAkyazi, F., Ansari, M. A., Ahmed, B. I., Crow, W. T., \\u0026amp; Mekete, T. (2012). First record of entomopathogenic nematodes (steinernematidae and heterorhabditidae) from Nigerian soil and their morphometrical and ribosomal DNA sequence analysis. \\u003cem\\u003eNematologia Mediterranea\\u003c/em\\u003e,\\u003cem\\u003e 40\\u003c/em\\u003e(2), 95-100. \\u003c/li\\u003e\\n\\u003cli\\u003eAnbesse, S., Sumaya, N. H., Dorfler, A. V., Strauch, O., \\u0026amp; Ehlers, R. U. (2013). Selective breeding for desiccation tolerance in liquid culture provides genetically stable inbred lines of the entomopathogenic nematode \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e. \\u003cem\\u003eAppl Microbiol Biotechnol\\u003c/em\\u003e,\\u003cem\\u003e 97\\u003c/em\\u003e(2), 731-739. https://doi.org/10.1007/s00253-012-4227-5\\u003c/li\\u003e\\n\\u003cli\\u003eBal, H. K., \\u0026amp; Grewal, P. S. (2015). Lateral dispersal and foraging behavior of entomopathogenic nematodes in the absence and presence of mobile and non-mobile hosts. \\u003cem\\u003ePLoS One\\u003c/em\\u003e,\\u003cem\\u003e 10\\u003c/em\\u003e, 1-19. https://doi.org/10.1371/journal.pone.0129887\\u003c/li\\u003e\\n\\u003cli\\u003eBhat, A. H., Chaubey, A. K., \\u0026amp; Askary, T. H. (2020). Global distribution of entomopathogenic nematodes, \\u003cem\\u003eSteinernema\\u003c/em\\u003e and \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e. \\u003cem\\u003eEgyptian Journal of Biological Pest Control\\u003c/em\\u003e,\\u003cem\\u003e 30\\u003c/em\\u003e(1). https://doi.org/10.1186/s41938-020-0212-y\\u003c/li\\u003e\\n\\u003cli\\u003eBrown, I. M., Lovett, B. J., Grewal, P. S., \\u0026amp; Gaugler, R. (2002). Latent infection: a low temperature survival strategy in steinernematid nematodes. \\u003cem\\u003eJournal of Thermal Biology\\u003c/em\\u003e,\\u003cem\\u003e 27\\u003c/em\\u003e(6), 531-539. https://doi.org/10.1016/S0306-4565(02)00027-X\\u003c/li\\u003e\\n\\u003cli\\u003eCabanillas, H. E. (2003). Susceptibility of the boll weevil to \\u003cem\\u003eSteinernema riobrave\\u003c/em\\u003e and other entomopathogenic nematodes. \\u003cem\\u003eJournal of Invertebrate Pathology\\u003c/em\\u003e,\\u003cem\\u003e 82\\u003c/em\\u003e(3), 188-197. https://doi.org/10.1016/S0022-2011(03)00016-8\\u003c/li\\u003e\\n\\u003cli\\u003eChen, S., \\u0026amp; Glazer, I. (2005). A novel method for long-term storage of the entomopathogenic nematode \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e at room temperature. \\u003cem\\u003eBiological Control\\u003c/em\\u003e,\\u003cem\\u003e 32\\u003c/em\\u003e, 104-110. https://doi.org/http://dx.doi.org/10.1016/j.biocontrol.2004.08.006 \\u003c/li\\u003e\\n\\u003cli\\u003eDaramola, F. Y., Osemwegie, O. O., Owa, S. O., Orisajo, S. B., Ikponmwosa, E., \\u0026amp; Alori, E. T. (2021). Isolation and molecular characterization of entomopathogenic nematode, \\u003cem\\u003eHeterorhabditis\\u003c/em\\u003e sp. from an arable land in Nigeria. \\u003cem\\u003eJournal of Integrative Agriculture\\u003c/em\\u003e,\\u003cem\\u003e 20\\u003c/em\\u003e(10), 2706-2715. https://doi.org/10.1016/s2095-3119(21)63609-2\\u003c/li\\u003e\\n\\u003cli\\u003eDuncan, L. W., \\u0026amp; McCoy, C. W. (2001). Hydraulic lift increases herbivory by Diaprepes abbreviates larvae and persistence of \\u003cem\\u003eSteinernema riobrave\\u003c/em\\u003e in dry soil. \\u003cem\\u003eJournal of Nematology\\u003c/em\\u003e,\\u003cem\\u003e 33\\u003c/em\\u003e(2-3), 142-146.\\u003c/li\\u003e\\n\\u003cli\\u003eGlazer, I. (2022). Stress and Survival Mechanisms. In \\u003cem\\u003eNematodes as Model Organisms\\u003c/em\\u003e (pp. 215-243). CABI. https://doi.org/10.1079/9781789248814.0009\\u003c/li\\u003e\\n\\u003cli\\u003eGoergen, G., Kumar, P. L., Sankung, S. B., Togola, A., \\u0026amp; Tam\\u0026ograve;, M. (2016). First report of outbreaks of the fall armyworm \\u003cem\\u003eSpodoptera frugiperda\\u003c/em\\u003e (J.E. Smith) (Lepidoptera: Noctuidae), a new alien invasive pest in West and Central Africa. \\u003cem\\u003ePLoS One\\u003c/em\\u003e,\\u003cem\\u003e 11\\u003c/em\\u003e(10), e0165632. https://doi.org/10.1371/journal.pone.0165632\\u003c/li\\u003e\\n\\u003cli\\u003eGouge, D. H., Smith, K. A., Lee, L. L., \\u0026amp; Henneberry, T. J. (2000). Effect of Soil Depth and Moisture on the Vertical Distribution of \\u003cem\\u003eSteinernema riobrave\\u003c/em\\u003e (Nematoda: Steinernematidae). \\u003cem\\u003eJournal of Nematology\\u003c/em\\u003e,\\u003cem\\u003e 32\\u003c/em\\u003e(2), 223-223. https://pmc.ncbi.nlm.nih.gov/articles/PMC2620435/\\u003c/li\\u003e\\n\\u003cli\\u003ehttp://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=PMC2620435\\u003c/li\\u003e\\n\\u003cli\\u003eGrant, J. A., \\u0026amp; Villani, M. G. (2003). Soil Moisture Effects on Entomopathogenic Nematodes. \\u003cem\\u003eEnvironmental Entomology\\u003c/em\\u003e,\\u003cem\\u003e 32\\u003c/em\\u003e(1), 80-87. https://doi.org/10.1603/0046-225X-32.1.80\\u003c/li\\u003e\\n\\u003cli\\u003eGrewal, P. S., Bornstein-Forst, S., Burnell, A. M., Glazer, I., \\u0026amp; Jagdale, G. B. (2006). Physiological, genetic, and molecular mechanisms of chemoreception, thermobiosis, and anhydrobiosis in entomopathogenic nematodes. \\u003cem\\u003eBiological Control\\u003c/em\\u003e,\\u003cem\\u003e 38\\u003c/em\\u003e(1), 54-65. https://doi.org/10.1016/j.biocontrol.2005.09.004\\u003c/li\\u003e\\n\\u003cli\\u003eGrewal, P. S., Ehlers, R.-U., \\u0026amp; Shapiro-Ilan, D. I. (2005). \\u003cem\\u003eNematodes as biocontrol agents\\u003c/em\\u003e. CABI.\\u003c/li\\u003e\\n\\u003cli\\u003eGrewal, P. S., Lewis, E. E., \\u0026amp; Gaugler, R. (1997). Response of infective stage parasites (Nematoda: Steinernematidae) to volatile cues from infected hosts. \\u003cem\\u003eJ Chem Ecol\\u003c/em\\u003e,\\u003cem\\u003e 23\\u003c/em\\u003e, 503-515.\\u003c/li\\u003e\\n\\u003cli\\u003eHao, Y. J., Flores-Ponce, M., \\u0026amp; Montiel, R. (2011). Genetics of entomopathogenic nematodes. In \\u003cem\\u003eMicrobial Insecticides: Principles and Applications\\u003c/em\\u003e (pp. 237-256). Nova Science Publishers, Inc. https://www.scopus.com/inward/record.uri?eid=2-s2.0-84895214695\\u0026amp;partnerID=40\\u0026amp;md5=428727f67e1a8d33049be71d903b6b4e\\u003c/li\\u003e\\n\\u003cli\\u003eHoctor, T. L., Gibb, T. J., Bigelow, C. A., \\u0026amp; Richmond, D. S. (2013). Survival and infectivity of the insect-parasitic nematode \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e (Poinar) in solutions containing four different turfgrass soil surfactants. \\u003cem\\u003eInsects\\u003c/em\\u003e,\\u003cem\\u003e 4\\u003c/em\\u003e(1), 1-8. https://doi.org/10.3390/insects4010001\\u003c/li\\u003e\\n\\u003cli\\u003eKaranastasi, E., Nikorezou, A., Stamouli, M., Skourti, A., Boukouvala, M. C., Kavallieratos, N. G., \\u0026amp; Weber, D. (2025). Temperature effect on the efficacy of 3 entomopathogenic nematode isolates against larvae of the lesser mealworm, \\u003cem\\u003eAlphitobius diaperinus\\u003c/em\\u003e (Coleoptera: Tenebrionidae). \\u003cem\\u003eJournal of Economic Entomology\\u003c/em\\u003e,\\u003cem\\u003e 118\\u003c/em\\u003e(1), 93-99. https://doi.org/10.1093/jee/toae292\\u003c/li\\u003e\\n\\u003cli\\u003eKaspi, R., Ross, A., Hodson, A. K., Stevens, G. N., Kaya, H. K., \\u0026amp; Lewis, E. E. (2010). Foraging efficacy of the entomopathogenic nematode \\u003cem\\u003eSteinernema riobrave\\u003c/em\\u003e in different soil types from California citrus groves. \\u003cem\\u003eApplied Soil Ecology\\u003c/em\\u003e,\\u003cem\\u003e 45\\u003c/em\\u003e(3), 243-253. https://doi.org/10.1016/j.apsoil.2010.04.012\\u003c/li\\u003e\\n\\u003cli\\u003eKaya, H. K., \\u0026amp; Gaugler, R. (1993). Entomopathogenic Nematodes. \\u003cem\\u003eAnnual Review of Entomology\\u003c/em\\u003e,\\u003cem\\u003e 38\\u003c/em\\u003e(1), 181-206. https://doi.org/10.1146/annurev.en.38.010193.001145\\u003c/li\\u003e\\n\\u003cli\\u003eKenis, M., Benelli, G., Biondi, A., Calatayud, P. A., Day, R., Desneux, N., Harrison, R. D., Kriticos, D., Rwomushana, I., van den Berg, J., Verheggen, F., Zhang, Y. J., Agboyi, L. K., Ahissou, R. B., Ba, M. N., Bernal, J., Freitas de Bueno, A., Carri\\u0026egrave;re, Y., Carvalho, G. A., . . . Wu, K. (2023). Invasiveness, biology, ecology, and management of the fall armyworm, \\u003cem\\u003eSpodoptera frugiperda\\u003c/em\\u003e. \\u003cem\\u003eEntomologia Generalis\\u003c/em\\u003e,\\u003cem\\u003e 43\\u003c/em\\u003e(2), 187-241. https://doi.org/10.1127/entomologia/2022/1659\\u003c/li\\u003e\\n\\u003cli\\u003eKoppenh\\u0026ouml;fer, A. M., \\u0026amp; Kaya, H. K. (1999). Ecological Characterisation of \\u003cem\\u003eSteinernema rarum\\u003c/em\\u003e. \\u003cem\\u003eJournal of Invertebrate Pathology\\u003c/em\\u003e,\\u003cem\\u003e 73\\u003c/em\\u003e, 120 - 128. http://www.idealibrary.com\\u003c/li\\u003e\\n\\u003cli\\u003eKour, S., Khurma, U., \\u0026amp; Brodie, G. (2021). Ecological Characterisation of Native Isolates of \\u003cem\\u003eHeterorhabditis indica\\u003c/em\\u003e from Viti Levu, Fiji Islands. \\u003cem\\u003eJournal of Nematology\\u003c/em\\u003e,\\u003cem\\u003e 53\\u003c/em\\u003e. https://doi.org/10.21307/jofnem-2021-085\\u003c/li\\u003e\\n\\u003cli\\u003eKumsta, C., Thamsen, M., \\u0026amp; Jakob, U. (2011). Effects of Oxidative Stress on Behavior, Physiology, and the Redox Thiol Proteome of \\u003cem\\u003eCaenorhabditis elegans\\u003c/em\\u003e. \\u003cem\\u003eAntioxidants \\u0026amp; Redox Signaling\\u003c/em\\u003e,\\u003cem\\u003e 14\\u003c/em\\u003e(6), 1023-1023. https://doi.org/10.1089/ARS.2010.3203\\u003c/li\\u003e\\n\\u003cli\\u003eKung, S.-P., Gaugler, R., \\u0026amp; Kaya, H. K. (1990a). Influence of Soil pH and Oxygen on Persistence of \\u003cem\\u003eSteinernema\\u003c/em\\u003e spp. \\u003cem\\u003eJournal of Nematology\\u003c/em\\u003e,\\u003cem\\u003e 22\\u003c/em\\u003e(4), 440-440. https://pmc.ncbi.nlm.nih.gov/articles/PMC2619085/\\u003c/li\\u003e\\n\\u003cli\\u003eKung, S.-P., Gaugler, R., \\u0026amp; Kaya, H. K. (1990b). Soil type and entomopathogenic nematode persistence. \\u003cem\\u003eJournal of Invertebrate Pathology\\u003c/em\\u003e,\\u003cem\\u003e 55\\u003c/em\\u003e(3), 401-406. https://doi.org/https://doi.org/10.1016/0022-2011(90)90084-J\\u003c/li\\u003e\\n\\u003cli\\u003eLacey, L. A., Grzywacz, D., Shapiro-Ilan, D. I., Frutos, R., Brownbridge, M., \\u0026amp; Goettel, M. S. (2015). Insect pathogens as biological control agents: Back to the future. \\u003cem\\u003eJournal of Invertebrate Pathology\\u003c/em\\u003e,\\u003cem\\u003e 132\\u003c/em\\u003e, 1-41. https://doi.org/10.1016/j.jip.2015.07.009\\u003c/li\\u003e\\n\\u003cli\\u003eLalitha, K., Karthi, S., Vengateswari, G., Karthikraja, R., Perumal, P., \\u0026amp; Shivakumar, M. S. (2018). Effect of entomopathogenic nematode of \\u003cem\\u003eHeterorhabditis indica\\u003c/em\\u003e infection on immune and antioxidant system in lepidopteran pest \\u003cem\\u003eSpodoptera litura\\u003c/em\\u003e (Lepidoptera: Noctuidae). \\u003cem\\u003eJournal of Parasitic Diseases\\u003c/em\\u003e,\\u003cem\\u003e 42\\u003c/em\\u003e(2), 204-211. https://doi.org/10.1007/s12639-018-0983-1\\u003c/li\\u003e\\n\\u003cli\\u003eLalramnghaki, H. C., Vanlalhlimpuia, Vanramliana, \\u0026amp; Lalramliana. (2017). Characterization of a new isolate of entomopathogenic nematode, \\u003cem\\u003eSteinernema sangi \\u003c/em\\u003e(Rhabditida, Steinernematidae), and its symbiotic bacteria \\u003cem\\u003eXenorhabdus vietnamensis\\u003c/em\\u003e (\\u0026gamma;-Proteobacteria) from Mizoram, northeastern India. \\u003cem\\u003eJournal of Parasitic Diseases\\u003c/em\\u003e,\\u003cem\\u003e 41\\u003c/em\\u003e(4), 1123-1131. https://doi.org/10.1007/s12639-017-0945-z\\u003c/li\\u003e\\n\\u003cli\\u003eLevy, N., Faigenboim, A., Salame, L., Molina, C., Ehlers, R.-U., Glazer, I., \\u0026amp; Ment, D. (2020). Characterization of the phenotypic and genotypic tolerance to abiotic stresses of natural populations of \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e. \\u003cem\\u003eScientific Reports\\u003c/em\\u003e,\\u003cem\\u003e 10\\u003c/em\\u003e. https://doi.org/10.1038/s41598-020-67097-0\\u003c/li\\u003e\\n\\u003cli\\u003eLillis, P. E., Kennedy, I. P., Carolan, J. C., \\u0026amp; Griffin, C. T. (2023). Low-temperature exposure has immediate and lasting effects on the stress tolerance, chemotaxis and proteome of entomopathogenic nematodes. \\u003cem\\u003eParasitology\\u003c/em\\u003e,\\u003cem\\u003e 150\\u003c/em\\u003e(1), 15-28. https://doi.org/10.1017/S0031182022001445\\u003c/li\\u003e\\n\\u003cli\\u003eLu, D., Macchietto, M., Chang, D., Barros, M. M., Baldwin, J., Mortazavi, A., \\u0026amp; Dillman, A. R. (2017). Activated entomopathogenic nematode infective juveniles release lethal venom proteins. \\u003cem\\u003ePLoS Pathogens\\u003c/em\\u003e,\\u003cem\\u003e 13\\u003c/em\\u003e(4), Article e1006302. https://doi.org/10.1371/journal.ppat.1006302\\u003c/li\\u003e\\n\\u003cli\\u003eMatuska-Lyzwa, J., Duda, S., Nowak, D., \\u0026amp; Kaca, W. (2024). Impact of Abiotic and Biotic Environmental Conditions on the Development and Infectivity of Entomopathogenic Nematodes in Agricultural Soils. \\u003cem\\u003eInsects 2024, Vol. 15, Page 421\\u003c/em\\u003e,\\u003cem\\u003e 15\\u003c/em\\u003e(6), 421-421. https://doi.org/10.3390/INSECTS15060421\\u003c/li\\u003e\\n\\u003cli\\u003eMatuska-Lyzwa, J., Wodecka, B., \\u0026amp; Kaca, W. (2023). Characterization of \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e (Rhabditida: Steinernematidae) Isolates in Terms of Efficacy against Cereal Ground Beetle \\u003cem\\u003eZabrus tenebrioides\\u003c/em\\u003e (Coleoptera: Carabidae): Morphometry and Principal Component Analysis. \\u003cem\\u003eInsects\\u003c/em\\u003e,\\u003cem\\u003e 14\\u003c/em\\u003e(2). https://doi.org/10.3390/insects14020150\\u003c/li\\u003e\\n\\u003cli\\u003eMaushe, D., Ogi, V., Divakaran, K., Verdecia Mogena, A. M., Himmighofen, P. A., Machado, R. A. R., Towbin, B. D., Ehlers, R. U., Molina, C., Parisod, C., \\u0026amp; Robert, C. A. M. (2023). Stress tolerance in entomopathogenic nematodes: Engineering superior nematodes for precision agriculture. \\u003cem\\u003eJournal of Invertebrate Pathology\\u003c/em\\u003e,\\u003cem\\u003e 199\\u003c/em\\u003e, 107953-107953. https://doi.org/10.1016/J.JIP.2023.107953\\u003c/li\\u003e\\n\\u003cli\\u003eMukuka, J., Strauch, O., Al Zainab, M. H., \\u0026amp; Ehlers, R.-U. (2010). Effect of temperature and desiccation stress on infectivity of stress tolerant hybrid strains of \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e. \\u003cem\\u003eRussian Journal of Nematology\\u003c/em\\u003e,\\u003cem\\u003e 18\\u003c/em\\u003e(2), 111-116. https://www.scopus.com/inward/record.uri?eid=2-s2.0-79952132220\\u0026amp;partnerID=40\\u0026amp;md5=4fb76546eb0281c1529cf546aabc751e\\u003c/li\\u003e\\n\\u003cli\\u003eMukuka, J., Strauch, O., \\u0026amp; Ehlers, R.-U. (2010). Variability in desiccation tolerance among different strains of the entomopathogenic nematode \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e. \\u003cem\\u003eNematology\\u003c/em\\u003e,\\u003cem\\u003e 12\\u003c/em\\u003e(5), 711-720.\\u003c/li\\u003e\\n\\u003cli\\u003eMukuka, J., Strauch, O., Hoppe, C., \\u0026amp; Ehlers, R. U. (2010). Fitness of heat and desiccation tolerant hybrid strains of \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e (Rhabditidomorpha: Heterorhabditidae). \\u003cem\\u003eJournal of Pest Science\\u003c/em\\u003e,\\u003cem\\u003e 83\\u003c/em\\u003e(3), 281-287. https://doi.org/10.1007/s10340-010-0296-3\\u003c/li\\u003e\\n\\u003cli\\u003eNeumann, G., \\u0026amp; Shields, E. J. (2006). Interspecific interactions among three entomopathogenic nematodes, \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e Weiser, \\u003cem\\u003eS. feltiae\\u003c/em\\u003e Filipjev, and \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e Poinar, with different foraging strategies for hosts in multipiece sand columns. \\u003cem\\u003eEnvironmental Entomology\\u003c/em\\u003e,\\u003cem\\u003e 35\\u003c/em\\u003e(6), 1578-1583. https://doi.org/10.1603/0046-225X(2006)35[1578:IIATEN]2.0.CO;2\\u003c/li\\u003e\\n\\u003cli\\u003eNguyen, K. B., \\u0026amp; Hunt, D. J. (2007). \\u003cem\\u003eEntomopathogenic nematodes : systematics, phylogeny and bacterial symbionts\\u003c/em\\u003e. Brill.\\u003c/li\\u003e\\n\\u003cli\\u003eNguyen, K. B., \\u0026amp; Smart, G. C. (1996). \\u003cem\\u003eIdentification of Entomopathogenic Nematodes in the Steinernematidae and Heterorhabditidae (Nemata: Rhabditida)\\u003c/em\\u003e Journal of Nematology. https://pmc.ncbi.nlm.nih.gov/articles/PMC2619694/\\u003c/li\\u003e\\n\\u003cli\\u003eNimkingrat, P., Ehlers, R. U., \\u0026amp; Strauch, O. (2011). Desiccation tolerance among different isolates of the entomopathogenic nematode \\u003cem\\u003eSteinernema feltiae \\u003c/em\\u003e(Fillipjev). \\u003cem\\u003eCommunications in agricultural and applied biological sciences\\u003c/em\\u003e,\\u003cem\\u003e 76\\u003c/em\\u003e(3), 293-296. https://pubmed.ncbi.nlm.nih.gov/22696940/\\u003c/li\\u003e\\n\\u003cli\\u003eNimkingrat, P., Uhlmann, F., Strauch, O., \\u0026amp; Ehlers, R.-U. (2013). Desiccation tolerance of dauers of entomopathogenic nematodes of the genus \\u003cem\\u003eSteinernema\\u003c/em\\u003e. \\u003cem\\u003eNematology\\u003c/em\\u003e,\\u003cem\\u003e 15\\u003c/em\\u003e, 451-458. https://doi.org/10.1163/15685411-00002692\\u003c/li\\u003e\\n\\u003cli\\u003eOnwong, R., Sumaya, N. H., Nitjarunkul, A., Kerdim, S., Khwanket, N., \\u0026amp; Noosidum, A. (2023). Occurrence of entomopathogenic nematodes, \\u003cem\\u003eOscheius myriophilus\\u003c/em\\u003e Poinar in Thailand: Preliminary characterization of novel isolates and biological control potential against insect pests. \\u003cem\\u003eJournal of Applied Entomology\\u003c/em\\u003e. https://doi.org/10.1111/jen.13168\\u003c/li\\u003e\\n\\u003cli\\u003ePuza, V., Nermut, J., Konopicka, J., \\u0026amp; Skokova Habustova, O. (2021). Efficacy of the Applied Natural Enemies on the Survival of Colorado Potato Beetle Adults. \\u003cem\\u003eInsects\\u003c/em\\u003e,\\u003cem\\u003e 12\\u003c/em\\u003e(11). https://doi.org/10.3390/insects12111030\\u003c/li\\u003e\\n\\u003cli\\u003eR Core Team. (2024). \\u003cem\\u003eR: A Language and Environment for Statistical Computing,\\u003c/em\\u003e.In \\u003cem\\u003eR\\u003c/em\\u003e (Version 4.4.3) http://www.R-project.org/.\\u003c/li\\u003e\\n\\u003cli\\u003eRaheel, M., Javed, N., Khan, S. A., Aatif, H. M., \\u0026amp; Ahmed, S. (2017). Effect of temperature on the reproductive potential of indigenous and exotic species of entomopathogenic nematodes inside \\u003cem\\u003eGalleria mellonella\\u003c/em\\u003e L. larvae. \\u003cem\\u003ePakistan Journal of Zoology\\u003c/em\\u003e,\\u003cem\\u003e 49\\u003c/em\\u003e(1), 419-421. https://doi.org/10.17582/journal.pjz/2017.49.1.sc12\\u003c/li\\u003e\\n\\u003cli\\u003eRaja, K. R., Padmanaban, K., \\u0026amp; Sivaramakrishnan, S. (2011). \\u003cem\\u003eEntomopathogenic Nematodes: A Best Bio-control Agent for Insect Pest. Isolation and Identification of Entomopathogenic Nematodes from Agricultural land\\u003c/em\\u003e. Lambert Academic Publishing.\\u003c/li\\u003e\\n\\u003cli\\u003eRakubu, I. L., Katumanyane, A., \\u0026amp; Hurley, B. P. (2024). Host-foraging strategies of five local entomopathogenic nematode species in South Africa. \\u003cem\\u003eCrop Protection\\u003c/em\\u003e,\\u003cem\\u003e 176\\u003c/em\\u003e, 106525-106525. https://doi.org/10.1016/J.CROPRO.2023.106525\\u003c/li\\u003e\\n\\u003cli\\u003eRamakrishnan, J., Salame, L., Nasser, A., Glazer, I., \\u0026amp; Ment, D. (2022). Survival and efficacy of entomopathogenic nematodes on exposed surfaces. \\u003cem\\u003eScientific Reports\\u003c/em\\u003e,\\u003cem\\u003e 12\\u003c/em\\u003e(1), 4629-4629. https://doi.org/10.1038/S41598-022-08605-2\\u003c/li\\u003e\\n\\u003cli\\u003eSegal, D., \\u0026amp; Glazer, I. (2000). Genetics for improving biological control agents: the case of entomopathogenic nematodes. \\u003cem\\u003eCrop Protection\\u003c/em\\u003e,\\u003cem\\u003e 19\\u003c/em\\u003e, 685-689.\\u003c/li\\u003e\\n\\u003cli\\u003eShapiro-Ilan, D. I., Brown, I., \\u0026amp; Lewis, E. E. (2014). Freezing and desiccation tolerance in entomopathogenic nematodes: diversity and correlation of traits. \\u003cem\\u003eJournal of Nematology\\u003c/em\\u003e, 27-34.\\u003c/li\\u003e\\n\\u003cli\\u003eSumaya, N. H., Gohil, R., Okolo, C. T., Addis, T., Doerfler, V., Ehlers, R. U., \\u0026amp; Molina, C. (2018). Applying inbreeding, hybridization and mutagenesis to improve oxidative stress tolerance and longevity of the entomopathogenic nematode \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e. \\u003cem\\u003eJournal of Invertebral Pathology\\u003c/em\\u003e,\\u003cem\\u003e 151\\u003c/em\\u003e, 50-58. https://doi.org/10.1016/j.jip.2017.11.001\\u003c/li\\u003e\\n\\u003cli\\u003eSusurluk, I. A., \\u0026amp; Ulu, T. C. (2015). Virulence comparisons of high-temperature-adapted \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e, \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e and \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e. \\u003cem\\u003eHelminthologia (Poland)\\u003c/em\\u003e,\\u003cem\\u003e 52\\u003c/em\\u003e(2), 118-122. https://doi.org/10.1515/helmin-2015-0021\\u003c/li\\u003e\\n\\u003cli\\u003eWatanabe, A., Yamaguchi, T., Murota, K., Ishii, T., Terao, J., Okada, S., Tanaka, N., Kimata, S., Abe, A., Suzuki, T., Uchino, M., \\u0026amp; Niimura, Y. (2019). Isolation of lactic acid bacteria capable of reducing environmental alkyl and fatty acid hydroperoxides, and the effect of their oral administration on oxidative-stressed nematodes and rats. \\u003cem\\u003ebioRxiv\\u003c/em\\u003e, 592162. https://doi.org/10.1101/592162\\u003c/li\\u003e\\n\\u003cli\\u003eWilliams, C. D., Dillon, A. B., Girling, R. D., \\u0026amp; Griffin, C. T. (2013). Organic soils promote the efficacy of entomopathogenic nematodes, with different foraging strategies, in the control of a major forest pest: A meta-analysis of field trial data. \\u003cem\\u003eBiological Control\\u003c/em\\u003e,\\u003cem\\u003e 65\\u003c/em\\u003e(3), 357-364. https://doi.org/10.1016/j.biocontrol.2013.03.013\\u003c/li\\u003e\\n\\u003cli\\u003eWright, D. J., Grewal, P. S., \\u0026amp; Stolinski, M. (1997). Relative Importance of Neutral Lipids and Glycogen as Energy Stores in Dauer Larvae of Two Entomopathogenic Nematodes, \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e and \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e. \\u003cem\\u003eComparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology\\u003c/em\\u003e,\\u003cem\\u003e 118\\u003c/em\\u003e(2), 269-273. https://doi.org/10.1016/S0305-0491(97)00165-X\\u003c/li\\u003e\\n\\u003cli\\u003eYadav, A. K., \\u0026amp; Lalramliana. (2012). Soil moisture effects on the activity of three entomopathogenic nematodes (Steinernematidae and Heterorhabditidae) isolated from Meghalaya, India. \\u003cem\\u003eJournal of Parasitic Diseases\\u003c/em\\u003e,\\u003cem\\u003e 36\\u003c/em\\u003e(1), 94-98. https://doi.org/10.1007/s12639-011-0076-x\\u003c/li\\u003e\\n\\u003cli\\u003eZadji, L., Baimey, H., Afouda, L., Moens, M., \\u0026amp; Decraemer, W. (2014). Characterization of biocontrol traits of heterorhabditid entomopathogenic nematode isolates from South Benin targeting the termite pest Macrotermes bellicosus. \\u003cem\\u003eBioControl\\u003c/em\\u003e,\\u003cem\\u003e 59\\u003c/em\\u003e(3), 333-344. https://doi.org/10.1007/s10526-014-9568-9\\u003c/li\\u003e\\n\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[{\"identity\":\"b4611bc4-e3e5-428e-8e29-10279e9e1bb2\",\"identifier\":\"10.13039/501100001655\",\"name\":\"Deutscher Akademischer Austauschdienst\",\"awardNumber\":\"2020\",\"order_by\":0},{\"identity\":\"49ec148e-3e20-445a-9e72-1182e91a4ba7\",\"identifier\":\"10.13039/501100011087\",\"name\":\"Stiftung fiat panis\",\"awardNumber\":\"2021\",\"order_by\":1}],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":true,\"hideJournal\":true,\"highlight\":\"\",\"institution\":\"University of Bonn\",\"isAcceptedByJournal\":false,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"researchsquare\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":true,\"externalIdentity\":\"\",\"sideBox\":\"\",\"snPcode\":\"\",\"submissionUrl\":\"/submission\",\"title\":\"Research Square\",\"twitterHandle\":\"researchsquare\",\"acdcEnabled\":true,\"dfaEnabled\":false,\"editorialSystem\":\"\",\"reportingPortfolio\":\"\",\"inReviewEnabled\":false,\"inReviewRevisionsEnabled\":true},\"keywords\":\"Biological control, EPN, Fall Armyworm, Oxidative stress, Temperature, Desiccation, Ecological fitness, Stress tolerance\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-8187949/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-8187949/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003cp\\u003eThe successful deployment of entomopathogenic nematodes (EPNs) in biological pest control hinges on their ecological fitness and stress tolerance. In this study, we assessed the ecological traits and efficacy of six indigenous EPN isolates previously identified from distinct agroecological zones in Nigeria, targeting the invasive pest \\u003cem\\u003eSpodoptera frugiperda\\u003c/em\\u003e (fall armyworm, FAW). The isolates, identified as \\u003cem\\u003eHeterorhabditis bacteriophora\\u003c/em\\u003e (Ib-CRIN68), \\u003cem\\u003eSteinernema carpocapsae\\u003c/em\\u003e (Ib-IART45, Ib-ITUC102), \\u003cem\\u003eSteinernema feltiae\\u003c/em\\u003e (Za-SAM), \\u003cem\\u003eSteinernema nepalense\\u003c/em\\u003e (Ib-HORT), and \\u003cem\\u003eOscheius myriophilus\\u003c/em\\u003e (Ib-FRIN32), were subjected to a series of ecological bioassays to evaluate their performance under temperature variation, moisture stress, oxygen limitation, oxidative stress, and foraging conditions. Results revealed significant inter- and intra-isolate variability in ecological tolerance traits. Optimal infectivity and reproduction were recorded between 25\\u0026ndash;30\\u0026deg;C, while mortality sharply declined at 10\\u0026deg;C and 35\\u0026deg;C. Foraging ability varied across substrates and soil depths, with \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates exhibiting strong host-finding capability under dry and surface conditions. Desiccation and oxidative stress assays also demonstrated the superior resilience of \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates, which sustained low mortality under Polyethylenglycol induced water stress and H₂O₂ exposure. Hypoxia assays indicated that all isolates were moderately tolerant to short-term anoxia, but only \\u003cem\\u003eH. bacteriophora\\u003c/em\\u003e and the \\u003cem\\u003eS. carpocapsae\\u003c/em\\u003e isolates survived above 50% at 72 h. Our study highlights the relevance of ecological screening as a prerequisite for selecting robust EPN candidate species and isolates suitable for biological control under variable conditions. The findings support the integration of indigenous EPNs into sustainable pest management frameworks in sub-Saharan Africa.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Ecological Characterization and Efficacy of Indigenous Entomopathogenic Nematodes Against Spodoptera frugiperda in Nigeria\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2025-11-26 04:14:04\",\"doi\":\"10.21203/rs.3.rs-8187949/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"researchsquare\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":true,\"externalIdentity\":\"\",\"sideBox\":\"\",\"snPcode\":\"\",\"submissionUrl\":\"/submission\",\"title\":\"Research Square\",\"twitterHandle\":\"researchsquare\",\"acdcEnabled\":true,\"dfaEnabled\":false,\"editorialSystem\":\"\",\"reportingPortfolio\":\"\",\"inReviewEnabled\":false,\"inReviewRevisionsEnabled\":true}}],\"origin\":\"\",\"ownerIdentity\":\"5da3a494-7439-47a0-a025-efb4e300bc03\",\"owner\":[],\"postedDate\":\"November 26th, 2025\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"posted\",\"subjectAreas\":[{\"id\":58539495,\"name\":\"Agroecology\"},{\"id\":58539496,\"name\":\"Entomology\"}],\"tags\":[],\"updatedAt\":\"2025-11-26T04:14:04+00:00\",\"versionOfRecord\":[],\"versionCreatedAt\":\"2025-11-26 04:14:04\",\"video\":\"\",\"vorDoi\":\"\",\"vorDoiUrl\":\"\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-8187949\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-8187949\",\"identity\":\"rs-8187949\",\"version\":[\"v1\"]},\"buildId\":\"8U1c8b4HqxoKbykW_rLl7\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}