A dataset on microbiome alterations in Drosophila melanogaster infected by entomopathogenic nematodes.

The widespread use of synthetic pesticides has raised concerns about their impacts on soil biodiversity and ecosystem functioning, increasing interest in plant-derived compounds as environmentally compatible alternatives. Cannabis sativa L. produces a diverse array of secondary metabolites, including cannabinoids and terpenes, which may act as chemical cues in belowground ecological interactions. This study examined the chemotactic responses of five nematode species - Heterorhabditis bacteriophora (Poinar), Steinernema carpocapsae (Weiser), Steinernema feltiae (Filipjev), Oscheius myriophilus (Poinar), and Phasmarhabditis papillosa (Schneider)- to various concentrations of crude flower extracts of C. sativa. Chemotaxis assays were conducted using an agar-based system, and nematode responses were quantified as overall motility and Chemotaxis Index (CI). Cannabinoid and terpene profiles were determined by HPLC-UV/DAD and GC-MS. Nematode species identity and extract concentration were the primary factors influencing motility and chemotactic behaviour, whereas chemotype extract identity had a minor effect. H. bacteriophora, S. feltiae, and P. papillosa showed positive chemotactic responses (higher CI values) in a species- and concentration-dependent manner, whereas S. carpocapsae and O. myriophilus exhibited mostly neutral responses. Chemotactic behaviour was strongly species-specific and concentration-dependent, with extract identity playing a minor role.

ABSTRACT The agave weevil, Scyphophorus acupunctatus , is a key pest of Agave spp. crops, causing severe damage and plant mortality. We evaluated the pathogenicity against adult and larval S. acupunctatus of isolates of the entomopathogenic fungi Beauveria bassiana , B. pseudobassiana , Metarhizium anisopliae and M. pingshaense ; and the entomopathogenic nematodes Steinernema feltiae , S. carpocapsae and Heterorhabditis bacteriophora . While fungal isolates caused low mortality in adults (< 20%), they induced moderate larval mortality (50%–80%) with no significant differences among isolates. Nematode‐induced mortality was also low in adults (< 10%) but significantly higher in larvae, especially with S. carpocapsae (~60%), compared with the other species (< 10%). In dual‐inoculation experiments, using isolates C18/11 ( B. bassiana ) and L21‐1 ( M. anisopliae ) with S. carpocapsae , additive effects were observed in adults, with higher mortality (up to 50%) than in single treatments. Against larvae, S. carpocapsae alone caused high mortality (~98%), which was similar to dual treatments, suggesting limited synergism. Overall, our findings suggest that biological control efforts should focus on the larval stage, where S. carpocapsae shows strong potential as a biological control agent. Future work should evaluate field application strategies to enhance nematode delivery and persistence in agave‐growing environments.

ABSTRACT Laboratory experiments were conducted to determine the efficacy of entomopathogenic nematodes (EPN), Steinernema kushidai and Heterorhabditis bacteriophora, against the adults of banana leaf and fruit scarring beetle, Basilepta subcostatum (Jacoby). Both EPN species were pathogenic against B. subcostatum, exhibiting time and dose-dependent mortality. The infective juveniles (IJs) of S. kushidai achieved 95% mortality, while those of H. bacteriophora caused 90% mortality at 300 IJs/adult after 120 h compared to 30% mortality in the control. S. kushidai was found to be more pathogenic than H. bacteriophora based on LD50 and LT50 values. The lowest LD50 and LT50 values obtained for S. kushidai were 97.3 IJs/adult and 53.9 h, respectively, while those of H. bacteriophora were 153.2 IJs/adult and 62.2 h, respectively. Considering the pathogenic potential at 72 h, the lethal dose, 265.0 IJs/adult for S. kushidai and 284.9 IJs/adult for H. bacteriophora were further evaluated in formulations with chitosan or Aloe vera gel and compared with imidacloprid 17.8 SL 0.3%, castor oil 3% and water (control). S. kushidai with chitosan 5 g/L H20 achieved 100% mortality after 168 h, which was significantly at par with imidacloprid 17.8 SL (100%) and castor oil (100%). H. bacteriophora with chitosan 5 g/L H20 resulted in 93.3% mortality after 168 h.

Efficacy of biological control agents against Popillia japonica: A strategy to reduce the risk of pest spread via potted plant substrates.

ABSTRACT Biological pest control agents are increasingly used in agriculture, and their combined application with pesticides can enhance pest management efficacy. This study assessed, under laboratory conditions, the selectivity of herbicide, insecticide, and fungicide active ingredients on entomopathogenic nematodes, including Steinernema rarum (Doucet, 1986), Steinernema brazilense (Nguyen, Shapiro-Ilan, Mbata & Adams, 2007), Steinernema diaprepesi (Nguyen & Duncan, 2002), Steinernema glaseri (Steiner, 1929), and Heterorhabditis bacteriophora (Poinar, 1976), used in soybean pest control. We hypothesised differential impacts of agrochemicals on nematode viability. Sulfentrazone and carfentrazone caused high mortality in S. rarum, reaching up to 89.75% and 70.75% after 48 h of exposure, respectively, whereas glyphosate showed lower toxicity, with mortality not exceeding 54.50%. Among insecticides, spinetoram was the most toxic to S. brazilense, causing mortality up to 52.52% after four hours, while methomyl resulted in mortality below 2% and proved compatible. Fungicide exposure led to generally low mortality, with mancozeb and procymidone causing the highest effects, reaching up to 6.05% and 4.05%, respectively. These findings highlight the importance of selecting agrochemicals compatible with biological control agents to support integrated pest management and promote more sustainable agricultural practices.

Multi-component trail pheromones of the entomopathogenic nematodes Steinernema diaprepesi and Heterorhabditis bacteriophora.

Thrips parvispinus (Karny) (Thysanoptera: Thripidae) is an invasive pest increasingly affecting ornamental production in Florida, with a rapid expansion in North America, Europe, and Africa. Current management relies heavily on chemical control, highlighting the need for more sustainable alternatives, such as entomopathogenic nematodes (EPNs). We evaluated six EPN species ( Heterorhabditis bacteriophora , Heterorhabditis indica , Steinernema carpocapsae , Steinernema riobrave , Steinernema feltiae , and Steinernema kraussei ) under laboratory conditions, using a rate of 200 IJ/cm 2 . Subsequently, the four best-performing species ( H. bacteriophora , H. indica , S. carpocapsae , and S. riobrave ) were further tested under greenhouse conditions using mandevilla plants and soil applications at a rate of 100 IJ/cm 2 . Trials were conducted at 27 °C, reflecting the average temperatures of the ornamental growing and shipping seasons in southern Florida, while applications targeted the prepupal and pupal soil-dwelling stages of the pest. Observations of the surviving adults were recorded. In laboratory trials, the application of H. bacteriophora , S. riobrave , H. indica , and S. carpocapsae reduced the recovery adult thrips by 20 - 36% compared to controls. Greenhouse trials demonstrated reductions in all treatments where EPNs were applied, with reductions of up to 60% in adult recovery in S. riobrave and S. carpocapsae . Environmental factors, including warm temperatures and low substrate moisture, helped explain the EPN performance, favoring warm-adapted and desiccation-tolerant species. This is the first report demonstrating EPN efficacy against T. parvispinus , suggesting its potential for integration as an alternative tool within IPM programs.

Stomoxys calcitrans is a hematophagous dipteran known as a "stable fly", capable of parasitizing several animal species. The use of biological agents is an alternative for controlling S. calcitrans, with entomopathogenic nematodes (EPNs) being both resistant and virulent. The present study aimed to evaluate the effect of infection by EPNs subjected to spraying pressure on S. calcitrans larvae in sugarcane straw and vinasse under environmental conditions. Groups of 20 larvae of the stable fly were placed in plastic containers containing one kilogram of autoclaved soil and 200 g of sugarcane straw each. Subsequently, 200 JIs/larvae were sprayed onto the trays with larvae, and emergence traps were placed on the containers. In the control group, there were no EPNs; only vinasse was used. The experiment had three replicates and was monitored daily for 15 days. It was observed that the EPN Heterorhabditis bacteriophora HP88 showed a mortality rate of 33.3%, which was not statistically different from that of the control group (38.3%). The EPNs H. indica LPP30 and H. baujardi LPP7 caused mortality rates of 81.7% and 73.3%, respectively; both being higher than H. bacteriophora HP88 and the control group, but equal to each other under these conditions. It is concluded that H. indica LPP30 and H. baujardi LPP7 were effective against stable fly larvae under environmental conditions, whereas H. bacteriophora HP88 does not demonstrate similar efficacy.

Vine weevil (Otiorhynchus sulcatus) is a globally important pest of soft fruit and ornamental crops with larvae causing significant root damage. Entomopathogenic nematodes (EPNs) have emerged as a key biological control option for larvae following restrictions on synthetic chemical insecticides, but their reported efficacy varies considerably across studies. This variability has created uncertainty about EPN effectiveness and optimal deployment strategies, limiting evidence-based recommendations for growers. Across 162 comparisons from 23 studies, EPN applications significantly reduced vine weevil larval survival compared to untreated controls (Hedges' g = -1.60, 95% CI -1.85 to -1.36), equivalent to ≈63% fewer live larvae. All five EPN species tested (Steinernema carpocapsae, S. feltiae, S. kraussei, Heterorhabditis bacteriophora, H. megidis) were effective, with no significant differences among species. However, between-study heterogeneity was high (I2 ≈ 97%), indicating variability in outcomes despite strong average benefits. Univariate analyses identified soil temperature as the strongest moderator, with warmer temperatures (18-30 °C) associated with greater EPN efficacy. Protected cropping environments (glasshouses) also enhanced performance compared to outdoor applications. Application method (drench versus drip irrigation) and growing medium type showed no significant effects. However, when accounting for clustering of effects within studies using multilevel models with cluster-robust inference, these moderator effects were reduced and no longer statistically robust. Sensitivity analyses confirmed the overall efficacy estimate was robust to study quality concerns and potential publication bias. EPNs provide reliable biological control of vine weevil larvae under field and semi-field conditions, with effectiveness enhanced by warmer soil temperatures and protected growing environments. Although average effects are large and consistent, practitioners should expect considerable variability in outcomes and prioritise applications during warm conditions in protected environments where feasible. A lack of robust differences among species suggests that selection can be based on practical considerations such as cost and availability. © 2025 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The current investigation examined the pathogenicity of three indigenous entomopathogenic nematodes (EPNs) - Heterorhabditis indica (ICAR‑NBAIR strain), Heterorhabditis bacteriophora and Steinernema carpocapsae supplied as a powder formulation against the root grub of cardamom. In addition to assessing mortality, the study monitored the attachment and penetration rates of the infective juveniles (IJs) into the target insect. H. indica exhibited the highest penetration efficiency, outperforming H. bacteriophora, while S. carpocapsae showed the lowest entry rate; penetration ability was found to depend on the specific infection strategy of each EPN species. Application of the EPNs involved placing the novel powder at the base of the plant, a method that is rapid, simple and reduces labour requirements. The wettable‑powder (WP) formulation proved highly effective in controlling root grub, lowering production costs and decreasing reliance on chemical insecticides such as fipronil, chlorpyriphos and phorate in small‑scale cardamom systems. The results indicate that cardamom root grub is susceptible to all three tested EPNs, though their insect‑killing capacities differ. H. indica (ICAR‑NBAIR) and H. bacteriophora (ICAR‑NBAIR) emerged as the most promising candidates for grub management. These nematodes represent ecologically safe, efficient, sustainable and on‑farm recyclable “green” technologies for cardamom cultivation, offering a cost‑effective, value‑added approach to support sustainable agriculture in small‑holder cardamom plantations.

Effect of Heterorhabditis bacteriophora (HB EN01) and Steinernema rarum (PAM 25) nematodes on the control of cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae).

Biological characteristics of high-searching individuals of the Heterorhabditis bacteriophora HBH hybrid strain for use in biocontrol

Summary The transmission and retention of the bacterial symbiont Photorhabdus laumondii during endotokia matricida in Heterorhabditis bacteriophora were examined by transmission electron microscopy of first-stage juveniles (J1), pre-dauer juveniles (J2d), dauer juveniles (DJ), and hermaphrodites. A newly developed method was applied specifically for the fixation and cutting of the J1 stage. Bacteria were present in all developmental stages, except J1. In the J2d and young DJ stages symbiont cells were localised within the cardia, while in fully-developed infective DJ they were observed to divide in the intestinal lumen. In hermaphrodites, bacteria were found in vacuole-like structures in the terminal cells of the intestine near the rectum as well as throughout the intestine. The density of the bacterial cells varied considerably between stages, being lower in J2d than in young DJ. Prolonged storage of DJ resulted in degradation of bacterial cell morphology. These findings on bacterial retention across life stages sheds light on bacterial transmission and retention mechanisms during endotokia matricida in H. bacteriophora .

Photorhabdus bacteria live in mutualistic relationships with Heterorhabditis nematodes, and together, they act as effective insect pathogens. These bacteria produce a diverse array of lectins, sugar-binding proteins that are believed to play crucial roles in the complex tripartite interaction among Photorhabdus, nematodes, and their insect hosts. One such lectin, Photorhabdus laumondii tumor necrosis factor (TNF)-like lectin (PLTL), identified in Photorhabdus laumondii subsp. laumondii TTO1, exhibits notable sequence similarity to the N-terminal domain of the BC2L-C lectin (BC2L-CN), a TNF-like lectin recognized for its specificity toward fucosylated glycans associated with human embryonic stem cells and certain cancers. Through glycan array analysis and surface plasmon resonance, we identified PLTL's binding preference for branched histo-blood group oligosaccharides. The crystallographic structure of PLTL in complex with the BLeb pentasaccharide reveals a network of direct and water-mediated hydrogen bonds simultaneously stabilizing the Fucα1-2 and Galα1-3 moieties, which define its narrow glycan specificity. A combination of mass spectrometry, protein crystallography, and analytical ultracentrifugation showed a unique hexameric PLTL architecture stabilized by intermolecular disulfide bridges. Our data suggest that PLTL may contribute to the mutualistic relationship between Photorhabdus and its nematode symbiont, Heterorhabditis bacteriophora, rather than playing a role in the interaction with the insect host. This study provides a structural and functional characterization of PLTL, a newly identified member of the TNF-like lectin family. Comparative analysis with BC2L-CN highlights both conserved and distinct structural features, suggesting potential applications in glycan recognition-based diagnostics or biotechnological tools beyond its biological role. Our findings underscore its complex glycan specificity and offer insights into its potential role in Photorhabdus-nematode symbiosis.

As a staple food, potato (Solanum tuberosum L.) (Solanaceae) is one of the most produced food crops to ensure food security. The potato tuber moth (PTM), Phthorimaea operculella (Zeller, 1873) (Lepidoptera: Gelechiidae), is a major pest of potato, damaging both the growing and storage processes. In recent years, green pest control strategies have been gaining importance to reduce the adverse effects of chemicals and protect the environment. Entomopathogenic nematodes (EPNs) and their bacterial endosymbionts (Xenorhabdus and Photorhabdus spp.) have been one of the top topics studied in sustainable pest control approaches. In the present study, the two most common EPN species, Steinernema feltiae and Heterorhabditis bacteriophora, and their bacterial associates, Xenorhabdus bovienii and Photorhabdus luminescens subsp. kayaii were evaluated against PTM larvae separately and in combination with spinosad. The survival rates of infective juveniles (IJs) of EPNs were over 92% after 72 h of direct exposure to spinosad. Co-application of EPNs and bioactive compounds (BACs) of endosymbiotic bacteria with spinosad induced synergistic interactions and achieved the maximum mortality (100%) in PTM larvae 48 h post-treatment. Spinosad and BAC combinations were highly efficient in controlling the PTM larvae and provided LT50 values below 23.0 h. Gas chromatography mass spectrometry (GC-MS) analysis identified 29 compounds in total, 20 of which belonged to P. luminescens subsp. kayaii. The results indicate that the integration of EPNs and BACs of endosymbiotic bacteria with spinosad presents a synergistic interaction and enhances pest control efficacy.

Relative virulence, host finding ability, and reproductive capacity of entomopathogenic nematodes for control of the goat biting louse Bovicola caprae (Phthiraptera: Trichodectidae).

SummaryThe abundant resources provided by the host provide an evolutionary rationale for parasitism and drive the metabolic and developmental divergence of parasitic and free-living animals. Two evolutionally distant nematode genera, Steinernema and Heterorhabditis, independently evolved an entomopathogenic lifestyle, in which they invade insects and kill them with the assistance of specifically associated symbiotic pathogenic bacteria. It had been generally assumed that the worm, being a bacterivore, feeds on its symbiotic bacteria, which rapidly reproduce while consuming the insect host. The evolutionary adaptations of entomopathogenic nematodes to a parasitic lifestyle developmentally, and the symbiotic relationships of entomopathogenicity, remain largely unknown. We developed an axenic culture medium that allows for robust and sustained growth of Steinernema hermaphroditum, allowing finite control of nutrients available to the nematodes. We found that, uniquely among nematodes tested, the hatchlings of S. hermaphroditum cannot endure in a nutrient-poor environment; this ability is impaired but still present in Heterorhabditis bacteriophora. Similarly, the ability to forage for food is completely lost in H. bacteriophora hatchlings and severely compromised in S. hermaphroditum. We reasoned that these traits were lost because they are unnecessary to obligate parasites that always hatch in a resource-rich host. We further found that Steinernema and, to a limited extent, Heterorhabditis nematodes can successfully invade, develop, and reproduce inside a living insect host independent of their symbiotic bacteria, apparently feeding on the hemolymph, and emerge carrying bacteria found within, explaining the evolutionary origins of entomopathogenic nematodes.

Entomopathogenic nematodes (EPNs) are valuable biological control agents and research models in agriculture and ecology. Traditional extraction and rearing methods for EPNs, such as the Baermann funnel and White trap, work well but have limitations in efficiency and practicality. The NEMA Device, constructed from PVC components, was designed to address these limitations by combining extraction and rearing into a single tool with improved portability, scalability, and ease of use. The efficiency of the NEMA Device was evaluated by comparing it to the conventional Baermann funnel extraction method and the White trap method for nematode multiplication. Validation of the instrument was performed using two nematode species, Steinernema khuongi and Heterorhabditis bacteriophora, which vary in size and can be used for pest control. Our results demonstrated that the NEMA Device achieved higher recovery rates of both S. khuongi and H. bacteriophora compared to the Baermann method. Additionally, the production rate of nematodes using the NEMA Device was comparable to that of the White trap method, with no significant difference observed between the two methods. The NEMA Device offers a standardized, cost-effective methodology for the extraction and multiplication of EPNs, enhancing the accessibility and efficiency of studying these agriculturally important nematodes and potentially improving biological control outcomes.

The efficacy of three native isolates of entomopathogenic nematodes: Heterorhabditis bacteriophora (H), H. indica (Fn) and Steinernema affine (313) was assessed against 3rd and 4th instar larvae (both outside and inside leaf tunnels) and pupae of Tuta absoluta, a destructive pest in Syria, at 25°C. In the laboratory, the isolates were tested on larvae outside leaf tunnels, at different doses 1, 5, 10, 15, 25 and 50 infective juveniles IJs/larva. The results obtained showed the susceptibility of larval instars to nematode infection, with variation according to the isolate, the concentration, and the larval instar stage inoculated. The isolate H exhibited the highest mortality rates across all treatments, followed by the isolate Fn, while the isolate 313 had the least virulence. The 50% lethal doses (LD50) for the three isolates against the 3rd and 4th larval instars, were 9.62 and 7.57 IJs/larva for isolate H; 12.33 and 8.24 for isolate Fn; 30.36 and 24.20 for isolate 313, respectively. The two most efficient isolates from the previous experiment, H and Fn, demonstrated the capability to access and kill larvae within the leaf tunnels with no significant difference between them. The H isolate exhibited mortality rates of 37.12 and 45.66%, whereas the Fn isolate produced mortality rates of 34.33 and 41.12% for 3rd and 4th instar larvae, respectively. Nevertheless, their efficacy on pupae was comparatively lower, with the H isolate displaying the highest mortality rate of 12.33%. In pot experiments, two concentrations (500 and 1000 IJs/ml) of H and Fn isolates were applied, with no significant differences observed between the two isolates. Although their efficacy was limited on pupae, the virulence on larvae instars was evident and increased with heightened concentrations and instar progression, achieving mortality rates for the 4th instar of 94.27 and 83.87% outside and inside leaf tunnels of the H isolate, and 91.66 and 80.10% for the Fn isolate, respectively. These findings suggest that entomopathogenic nematodes adapted to moderately warm temperatures are considered promising effective biological control agents against Tuta absoluta. Further in-depth research and practical applications within greenhouses are needed. Keywords: Biological control, entomopathogenic nematodes, Heterorhabditis, Steinernema, Tuta absoluta, LD50.