Biocontrol potential of entomopathogenic nematodes against the grey maize weevil Tanymecus dilaticollis (Coleoptera: Curculionidae) adults.

The grey maize weevil, Tanymecus dilaticollis, is a polyphagous species, which is among the most important pests of maize in Southeastern Europe. The efficacy of commercial products with two species of entomopathogenic nematodes (EPNs), Steinernema carpocapsae and Heterorhabditis bacteriophora, was investigated against adults of the grey maize weevil under laboratory conditions. Nemastar®, containing S. carpocapsae was more effective on T. dilaticollis adults than Nematop® containing H. bacteriophora, when applied uniformly to the surface of the soil, on Petri dishes containing T. dilaticollis adults. Results showed that S. carpocapsae rates of 83-333 infective juveniles/adult caused > 94% mortality in T. dilaticollis adults, whereas H. bacteriophora caused 27-61%, adult mortality, after exposure of insects to the commercial products of EPNs for 15 days. The infection rates of EPNs increased with concentration applied and ranged from 70-83% and 19-64% for Nemastar® and Nematop®, respectively. Subsequent field and semi-field tests were conducted with Nemastar® (application rate of 50 million S. carpocapsae per 100 m2) in maize crops with biological (mycoinsecticide Naturalis®, biofungicides and fertilizers) and chemical seed treatment (Gaucho® FS 600; active ingredient: imidacloprid) in Knezha, Bulgaria. Nematodes were found only in the dead specimens, in open plots and cages sprayed with the commercial nematode product. Nematode sprayings contributed for higher maize yields in the open maize plots in the fields with different seed treatments. We suggest that the use of powder formulation of S. carpocapsae in combination with biologically treated maize seeds can contribute to minimize the use of chemical insecticides against the grey maize weevil. The results obtained can be used as a base to further tests to ascertain the efficacy of EPNs products before they can be recommended for use in the integrated approach to T. dilaticollis management.

Behavioral and molecular response of the insect parasitic nematode Steinernema carpocapsae to plant volatiles.

Entomopathogenic nematodes (EPNs) use the chemical cues emitted by insects and insect-damaged plants to locate their hosts. Steinernema carpocapsae, a species of EPN, is an established biocontrol agent used against insect pests. Despite its promising potential, the molecular mechanisms underlying its ability to detect plant volatiles remain poorly understood. In this study, we investigated the response of S. carpocapsae infective juveniles (IJs) to 8 different plant volatiles. Among these, carvone was found to be the most attractive volatile compound. To understand the molecular basis of the response of IJs to carvone, we used RNA-Seq technology to identify gene expression changes in response to carvone treatment. Transcriptome analysis revealed 721 differentially expressed genes (DEGs) between carvone-treated and control groups, with 403 genes being significantly upregulated and 318 genes downregulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the responsive DEGs to carvone attraction were mainly involved in locomotion, localization, behavior, response to stimulus, and olfactory transduction. We also identified four upregulated genes of chemoreceptor and response to stimulus that were involved in the response of IJs to carvone attraction. Our results provide insights into the potential transcriptional mechanisms underlying the response of S. carpocapsae to carvone, which can be utilized to develop environmentally friendly strategies for attracting EPNs.

Influence of Asafoetida Extract on the Virulence of the Entomopathogenic Nematode Steinernema carpocapsae and Its Symbiotic Bacterium Xenorhabdus nematophila in the Host Pyrrhocoris apterus.

Nematode-microbe symbiosis plays a key role in determining pathogenesis against pests. The modulation of symbiotic bacteria may affect the virulence of entomopathogenic nematodes (EPNs) and the biological management of pests. We tested the influence of asafoetida (ASF) extract on the virulence of Steinernema carpocapsae and its symbiotic bacterium, Xenorhabdus nematophila, in Pyrrhocoris apterus. A total of 100 mg of ASF killed 30% of EPNs in 48 h, while P. apterus remained unaffected. The EPNs pre-treated with 100 mg of ASF influenced P. apterus's mortality by 24-91.4% during a period of 24 to 72 h. The topical application of ASF acted as a deterrent to S. carpocapsae, lowering host invasion to 70% and delaying infectivity with 30% mortality for 168 h. Interestingly, Steinernema's symbiotic bacterium, Xenorhabdus, remained unaffected by ASF. An in vitro turbidity test containing 100 mg of ASF in a medium increased the growth rate of Xenorhabdus compared to a control. A disc diffusion assay confirmed the non-susceptibility of Xenorhabdus to ASF compared to a positive control, streptomycin. Pro-phenol oxidase (PPO) and phenol oxidase (PO) upregulation showed that ASF influences immunity, while EPN/ASF showed a combined immunomodulatory effect in P. apterus. We report that ASF modulated the virulence of S. carpocapsae but not that of its symbiotic bacterium, X. nematophila, against P. apterus.

Parasitic nematode secreted phospholipase A2 suppresses cellular and humoral immunity by targeting hemocytes in Drosophila melanogaster.

A key aspect of parasitic nematode infection is the nematodes' ability to evade and/or suppress host immunity. This immunomodulatory ability is likely driven by the release of hundreds of excretory/secretory proteins (ESPs) during infection. While ESPs have been shown to display immunosuppressive effects on various hosts, our understanding of the molecular interactions between individual proteins released and host immunity requires further study. We have recently identified a secreted phospholipase A2 (sPLA2) released from the entomopathogenic nematode (EPN) Steinernema carpocapsae we have named Sc-sPLA2. We report that Sc-sPLA2 increased mortality of Drosophila melanogaster infected with Streptococcus pneumoniae and promoted increased bacterial growth. Furthermore, our data showed that Sc-sPLA2 was able to downregulate both Toll and Imd pathway-associated antimicrobial peptides (AMPs) including drosomycin and defensin, in addition to suppressing phagocytosis in the hemolymph. Sc-sPLA2 was also found to be toxic to D. melanogaster with the severity being both dose- and time-dependent. Collectively, our data highlighted that Sc-sPLA2 possessed both toxic and immunosuppressive capabilities.

Mass Spectrometry-Driven Discovery of Neuropeptides Mediating Nictation Behavior of Nematodes.

Neuropeptides regulate animal physiology and behavior, making them widely studied targets of functional genetics research. While the field often relies on differential -omics approaches to build hypotheses, no such method exists for neuropeptidomics. It would nonetheless be valuable for studying behaviors suspected to be regulated by neuropeptides, especially when little information is otherwise available. This includes nictation, a phoretic strategy of Caenorhabditis elegans dauers that parallels host-finding strategies of infective juveniles of many pathogenic nematodes. We here developed a targeted peptidomics method for the model organism C. elegans and show that 161 quantified neuropeptides are more abundant in its dauer stage compared with L3 juveniles. Many of these have orthologs in the commercially relevant pathogenic nematode Steinernema carpocapsae, in whose infective juveniles, we identified 126 neuropeptides in total. Through further behavioral genetics experiments, we identify flp-7 and flp-11 as novel regulators of nictation. Our work advances knowledge on the genetics of nictation behavior and adds comparative neuropeptidomics as a tool to functional genetics workflows.

Low-temperature exposure has immediate and lasting effects on the stress tolerance, chemotaxis and proteome of entomopathogenic nematodes.

Temperature is one of the most important factors affecting soil organisms, including the infective stages of parasites and entomopathogenic nematodes, which are important biological control agents. We investigated the response of 2 species of entomopathogenic nematodes to different storage regimes: cold (9°C), culture temperature (20°C) and temperature swapped from 9 to 20°C. For Steinernema carpocapsae, cold storage had profound effects on chemotaxis, stress tolerance and protein expression that were retained in temperature-swapped individuals. These effects included reversal of chemotactic response for 3 (prenol, methyl salicylate and hexanol) of the 4 chemicals tested, and enhanced tolerance to freezing (−10°C) and desiccation (75% RH). Label-free quantitative proteomics showed that cold storage induced widespread changes in S. carpocapsae, including an increase in heat-shock proteins and late embryogenesis abundant proteins. For Heterorhabditis megidis, cold storage had a less dramatic effect on chemotaxis (as previously shown for proteomic expression) and changes were not maintained on return to 20°C. Thus, cold temperature exposure has significant effects on entomopathogenic nematodes, but the nature of the change depends on the species. Steinernema carpocapsae, in particular, displays significant plasticity, and its behaviour and stress tolerance may be manipulated by brief exposure to low temperatures, with implications for its use as a biological control agent.

Individual Coating of Entomopathogenic Nematodes with Titania (TiO2) Nanoparticles Based on Oil-in-Water Pickering Emulsion: A New Formulation for Biopesticides.

This study presents a new eco-friendly formulation of entomopathogenic nematodes (EPNs) based on individual coating of EPNs with titanium dioxide (TiO2) nanoparticles (NPs) and mineral oil via oil-in-water Pickering emulsions. Mineral oil-in-water emulsions stabilized by amine-functionalized titanium dioxide (TiO2-NH2) particles were prepared. 40:60 and 50:50 oil-water volume ratios using 2 wt % TiO2-NH2 particles were found to be the most stable emulsions with a droplet size suitable for the formulation and were further studied for their toxicity against the incorporated EPNs. Carboxyfluorescein was covalently bonded to TiO2-NH2 NPs, and the resulting composite was observed via fluorescence confocal microscopy. The dry coating was evaluated using SEM and confocal microscopy, which showed significant nematode coverage by the particles and oil. The final formulation was biocompatible with the studied EPNs, where the viability of the EPNs in the formulation was equivalent to control aqueous suspension after 120 days. Finally, yields of nematodes from infected Galleria mellonella cadavers collected for 150 days showed no significant differences (P > 0.05) using the tested emulsions compared to the control containing nematodes in water.

Pathogenicity of an Entomopathogenic Nematode, Steinernema carpocapsae on Alphitobius diaperinus (Coleoptera: Tenebrionidae) Strains From Turkey.

Alphitobius diaperinus Panzer (Coleoptera: Tenebrionidae) is an insect pest in poultry production systems. This insect has developed resistance to many chemical insecticides. As an alternative to chemicals, entomopathogenic nematodes (EPNs) are one of the most commonly used agents against several pest arthropods. The pathogenicity of the EPN Steinernema carpocapsae Weiser on four field strains of lesser mealworm, A. diaperinus, from Turkey was studied. In the experiments, larvae of A. diaperinus were found to be more sensitive than adults to S. carpocapsae infection. In terms of LC50 values in both larval and adult insects, Balikesir strain was found to be the most susceptible, Manisa strain the most resistant. The calculated LC50 values for adults were 85.9, 205.6, 135.4, and 418.8 IJs/ml, and for larvae 31.2, 39.8, 34.8, and 70.9 IJs/ml for the Balikesir, Canakkale, Izmir, and Manisa strains, respectively. This is the first report about the pathogenicity of EPNs against larvae and adults of lesser mealworm A. diaperinus strains from Turkey. We conclude that S. carpocapsae can be used as a control agent for lesser mealworms.

REVIEW OF THE ROLE OF PARASITIC NEMATODE EXCRETORY/SECRETORY PROTEINS IN HOST IMMUNOMODULATION.

Parasitic nematodes infect a variety of organisms including insects and vertebrates. To survive, they evade host immune responses to cause morbidity and mortality. Despite the vast clinical knowledge regarding nematode infections and their biological makeup, molecular understanding of the interactions between host and parasite remains poorly understood. The utilization of model systems has thus been employed to help elucidate the molecular interactions of the host immune response during parasitic nematode infection. Using model systems, it has been well established that parasitic nematodes evade host immunity by releasing excretory/secretory proteins (ESPs), which are involved in immunomodulation. Model systems have enabled researchers to characterize further the underlying mechanisms ESPs use to facilitate evasion and modulation of the host immune response. This review assessed notable ESPs from parasitic nematodes that infect vertebrates or insects and have been studied in mechanistic detail. Being able to characterize how ESPs affect the immune systems of hosts on a molecular level increases our understanding of host-parasite interactions and could lead to the identification of novel therapeutic targets and important molecular pathways.

Field Efficacy of Steinernema sp. (Rhabditida: Steinernematidae) on the Colorado Potato Beetle Overwintering Generation.

Colorado potato beetle (CPB) is an economic pest of potato that has developed resistance to all classes of chemical insecticides, thus requiring alternative control measures. As a potential solution, entomopathogenic nematodes (EPNs) have proven effective in suppressing this pest, but their efficacy against overwintering generations of CPB in Croatia has not been sufficiently researched. The aim of this two-year (2018-2019) field study was to determine the efficacy of Steinernema feltiae and Steinernema carpocapsae applied to overwintering CPB adults. EPNs were applied at three doses (7.5 mil./10 m2, 5.0 mil./10 m2 (the recommended dose) and 2.5 mil./10 m2) by watering the soil where the adults were overwintering. The first-year results were satisfactory for both EPNs: the efficacy of S. feltiae ranged from 79.03% to 100.00%, while the efficacy of S. carpocapsae ranged from 77.32% to 96.22%. In the second year, the highest efficacy (69.57%) was obtained using the recommended dose of S. feltiae. Although the results are not consistent across the two years of our study and suggest further research, they indicate that EPNs have great potential in controlling overwintering CPB generations to reduce first generation abundance and damage, and also to prevent the spread of new generations to surrounding potato growing areas.

Entomopathogenic nematode management of small hive beetles (Aethina tumida) in three native Alabama soils under low moisture conditions.

The goal was to determine the efficacy of entomopathogenic nematodes (EPNs) on Aethina tumida small hive beetle (SHB) in Alabama soils. The objectives were to (i) determine the pupation success of SHB wandering larvae; (ii) determine the efficacy of EPNs on SHB wandering larvae in natural and autoclaved soil; and (iii) determine the efficacy of EPNs on SHB wandering larvae in three Alabama soil types at typical low moisture levels. The Alabama soils were Kalmia loamy sand (KLS), Benndale fine sandy loam (BFSL), and Decatur silt loam (DSL). Heterorhabditis bacteriophora, H. indica, Steinernema carpocapsae, S. feltiae, S. kraussei, and S. riobrave were tested at population densities of 5, 10, 20, 40, and 80 third-stage infective EPN juveniles (IJ3) per 130 cm3 soil. Pupation success in SHB population densities of 5, 10, and 20 wandering larvae per Petri dish were similar. Of the six EPN species, S. carpocapsae achieved the highest efficacy across all EPN population densities in both natural and autoclaved soil. Steinernema riobrave and H. indica achieved the next highest efficacies; however, they were significantly less effective than S. carpocapsae. Steinernema carpocapsae parasitized 87% SHB wandering larvae across all population densities tested. Steinernema carpocapsae achieved the best efficacy colonizing 94% of the SHB in the KLS soil, 80% in the BFSL soil, and 47% in the DSL soil. In conclusions, S. carpocapsae is be a promising biological control EPN to implement into a management system on SHB.

The effect of chemical insecticides on the scavenging performance of Steinernema carpocapsae: Direct effects and exposure to insects killed by chemical insecticides.

Entomopathogenic nematodes are used widely in biological insect control. Entomopathogenic nematodes can infect live insects as well as dead insects (i.e., they can act as scavengers). It is important to determine compatibility of entomopathogenic nematodes with other pest management tactics such as chemical insecticides. We hypothesized that chemical insecticides have negative impact on scavenging nematodes. According to our hypothesis, we first investigated the effects of direct exposure of Steinernema carpocapsae infectivity juveniles (IJs) to three chemical insecticides, cypermethrin, spinosad or diflubenzuron in terms of nematode survival and virulence. Subsequently, using the same chemicals, we tested the effects of insecticide-killed insects on scavenger nematode penetration efficiency, time of emergence and the number of nematode progeny. Prior to our study, the impact of pesticides on scavenger nematode fitness had not been studied. Fall webworm, Hyphantria cunea, and greater wax moth, Galleria mellonella, larvae were used as host insects. The survival rate of IJs after direct exposure was 83% for cypermethrin and 93-97% for the other insecticides and control. There were no significant differences in the survival and virulence of the nematodes after 24 h exposure to insecticides. The number of nematodes that invaded the insecticide-killed host was significantly higher in cypermethrin and spinosad treated groups and live H. cunea than in the diflubenzoron treated group and freeze-killed control. However, no significant differences were observed in time of emergence. Significantly more progeny IJs emerged from Spinosad-killed insects than the freeze-killed control. In conclusion, we discovered that the fitness of scavenging IJs is not diminished by insecticides in insect cadavers. In fact, in some cases the exposure to chemical insecticides may enhance virulence.

Combined Effect of Entomopathogens against Thrips tabaci Lindeman (Thysanoptera: Thripidae): Laboratory, Greenhouse and Field Trials.

Onion thrips, Thrips tabaci Lindeman (Thysanoptera: Thripidae) is one of the most damaging insect pests of onions, Allium cepa L., which is an economically important agricultural crop cultivated worldwide. In this study, the combined application of entomopathogenic nematodes with entomopathogenic fungi against different soil dwelling stages of T. tabaci was evaluated. The nematodes included Heterorhabditis bacteriophora (VS strain) and Steinernema feltiae (SN strain), and fungi included Beauveria bassiana (WG-11) and Metarhizium anisopliae (WG-02); all four paired combinations (nematode + fungus) were included. In a small cup bioassay, only the combined application of H. bacteriophora and B. bassiana (WG-11) caused a synergistic interaction against pre-pupae, while all other combinations were compatible in an additive manner against pupae and late second instars. In a larger arena, a potted soil bioassay, again, combined applications of both pathogens produced greater mortality compared to single applications of each pathogen; all the combinations exhibited additive interactions, with the highest mortality observed in pre-pupae, followed by pupae and late second instar larvae using H. bacteriophora and B. bassiana (WG-11). Additionally, in the potted plant bioassay, lower adult emergence was observed from treated groups compared to control groups. Under field conditions, lower numbers of adults and larvae were found in treated groups relative to controls. Overall, the pre-pupal stage was more susceptible to the pathogen treatments, followed by pupae and late second instar larvae, and also combined applications of both pathogens suppressed the adult population. Combined application of entomopathogenic nematodes and fungi could be used for integrated pest management (IPM) of T. tabaci in onion production systems.

Efficacy and residual activity of commercially available entomopathogenic nematode strains for Mediterranean fruit fly control and their ability to infect infested fruits.

BACKGROUND: Entomopathogenic nematodes (EPN) show potential in controlling larvae of the Mediterranean fruit fly (medfly) Ceratitis capitate, but previous studies mainly concern species and strains that are not commercially available. The use of EPN for control of Mediterranean fruit fly is further hampered by the cost of using nematodes. In this study, the efficacy and residual activity of commercial strains of three EPN species, Steinernema carpοcapsae, S. feltiae and Heterοrhabditis bacteriοphοra medfly) C. capitata, in the soil substrate and inside fruits were evaluated. RESULTS: Suspensions of these species were applied at a dose of 1.5 mi m-2 on a soil substrate wherein medfly larvae were added sequentially for a period of 4 weeks post application at 20 °C. S. feltiae provided the highest suppression up to 50% as assessed by adult medfly emergence because it had the highest immediate activity and long residual activity. Furthermore, S. feltiae, and to a lesser degree S. carpocapsae, were able to move and infect medfly larvae inside infested apples and oranges left in the surface of the substrate wherein EPN were applied, reducing significantly adult medfly emergence (60-78%). CONCLUSION: These results support the efficacy and feasibility of applying a single, relatively low dose of S. feltiae in autumn, off-season, targeting overwintering medfly larvae with the scope of reducing the number of adult medflies emerging later in the new season.

Behavioral and molecular response of the insect parasitic nematode Steinernema carpocapsae to cues emitted by a host, the red palm weevil, Rhynchophorus ferrugineus.

As the larvae of the date palm pest, the red palm weevil, Rhynchophorus ferrugineus, feeds on the host tissue, they emit a distinctive sound which can be recorded outside of the infected tree. We evaluated the response of infective juveniles (IJs) of the entomopathogenic nematodes Steinernema carpocapsae to the R. ferrugineus larvae and it's sound source, separately. In the presence of the insect larvae, 50.2 % of total IJs moved toward those larvae. Recorded insect larvae sound emitted by the speaker resulted in 7% of total IJs near the sound source. RNA-Seq data indicated that more genes were downregulated in S. carpocapsae IJs exposed to insect and speaker compared to non-stimulated IJs. IJs exposed to insect exhibited more up-regulated genes than IJs exposed to speaker. Enriched pathways and biological processes in IJs were similar for both stimuli. The inhibition of locomotion, regulation of neurotransmitter secretion, response to biotic stimulus, and cellular response to chemical stimuli were enriched with unique GO terms for speaker treatment. The regulation of localization, sodium ion transmembrane transport, regulation of response to stress and response to organic substances were the GO categories enriched unique to insect. The host-parasitic interaction was regulated by the differential expression of Ras/MAP kinase, TGF-beta signaling, insulin signaling, AMPK signaling, PPAR signaling pathways and many developmental pathways. More prominent R. ferrugineus host localization by S. carpocapsae was primarily due to the differential transcriptional regulation of olfactory signal transduction, FOXO-family proteins, calcium signaling, WNT and mTOR signaling pathway. The neural basis for the nematode attraction to insect host is based on the chemosensation and the mechanosensation. Many neuropeptides and neuromodulators are involved in regulating the foraging behavior of S. carpocapsae. The results of this study provide new insights into the molecular mechanisms that allow these nematodes to seek insect hosts. Our finding, especially the molecular ones suggest that chemical cues emitted by the active insect host are stimulants of nematodes attraction. Whereas the sound emitted by the insect has minor effects on the nematode behavior.

Susceptibility of Various Developmental Stages of the Fall Armyworm, Spodoptera frugiperda, to Entomopathogenic Nematodes.

The fall armyworm, Spodoptera frugiperda, which is native to Central and South America, has recently invaded Africa and Asia, causing serious damage to various crops. Although management to date has been largely unsuccessful, entomopathogenic nematodes (EPNs) are a potential biological control agent that could be used to control the late larval and pupal stages of S. frugiperda that dwell under the ground. Here, we compared the virulence of seven EPNs against larval and pupal stages of S. frugiperda. In a Petri dish assay, both Heterorhabditis indica and Steinernema carpocapsae were highly virulent against younger larvae, whereas S. arenarium and S. longicaudum were highly virulent against older larvae. In contrast, H. bacteriophora, Heterorhabditis sp., and S. kushidai showed low virulence against all larval stages. In soil column and pot assays, H. indica, S. carpocapsae, and S. longicaudum were highly virulent against late larval and pupal stages compared with the other EPN species. Thus, H. indica, S. carpocapsae, and S. longicaudum are recommended for the biological control of S. frugiperda. Our study provides important information of EPNs for the practical application of biological control of fall armyworm.

Transcriptome Profile Changes Associated With Heat Shock Reaction in the Entomopathogenic Nematode, Steinernema carpocapsae.

The entomopathogenic nematode Steinernema carpocapsae has been used for control of soil insects. However, S. carpocapse is sensitive to environmental factors, particularly temperature. We studied an S. carpocapse group that was shocked with high temperature. We also studied the transcriptome-level responses associated with temperature stress using a BGIseq sequencing platform. We de novo assembled the reads from the treatment and control groups into one transcriptome consisting of 43.9 and 42.9 million clean reads, respectively. Based on the genome database, we aligned the clean reads to the Nr, Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases and analyzed the differentially expressed genes (DEGs). Compared with the control, the heat-shocked group had significant differential expression of the heat shock protein (HSP) family, antioxidase [glutathione S-transferases (GSTs) and superoxide dismutase (SOD)], monooxygenase (P450), and transcription factor genes (DAF-16 and DAF-2). These DEGs were demonstrated to be part of the Longevity pathway and insulin/insulin-like signaling pathway. The results revealed the potential mechanisms, at the transcriptional level, of S. carpocapsae under thermal stress.

Immune Response of Drosophila suzukii Larvae to Infection with the Nematobacterial Complex Steinernema carpocapsae-Xenorhabdus nematophila.

Entomopathogenic nematodes have been proposed as biological agents for the control of Drosophila suzukii, an invasive pest of small-stone and soft-skinned fruits. Larvae of the fly are susceptible to Steinernema carpocapsae infection but the reaction of immune defenses of the host are unknown. To determine the immune response, larvae were infected with S. carpocapsae and Xenorhabdus nematophila to evaluate the effector mechanisms of both humoral and cellular processes. The symbiont bacteria presented an inhibitory effect on the phenoloxidase cascade with a low level of melanization. Besides, X. nematophila activated the synthesis of putative antimicrobial peptides on the hemolymph of infected larvae. However, those peptides presented a lower antimicrobial activity compared to hemolymph from larvae infected with non-symbiont bacteria. Xenorhabdus nematophila avoided also the phagocytosis response of hemocytes. During in vitro and in vivo assays, S. carpocapsae was not encapsulated by cells, unless the cuticle was damaged with a lipase-treatment. Hemocyte counts confirmed differentiation of lamellocytes in the early phase of infection despite the unrecognition of the nematodes. Both X. nematophila and S. carpocapsae avoided the cellular defenses of D. suzukii larvae and depressed the humoral response. These results confirmed the potential of entomopathogenic nematodes to control D. suzukii.

Partner-specific induction of Spodoptera frugiperda immune genes in response to the entomopathogenic nematobacterial complex Steinernema carpocapsae-Xenorhabdus nematophila.

The Steinernema carpocapsae-Xenorhabdus nematophila association is a nematobacterial complex used in biological control of insect crop pests. The infection success of this dual pathogen strongly depends on its interactions with the host's immune system. Here, we used the lepidopteran pest Spodoptera frugiperda to analyze the respective impact of each partner in the induction of its immune responses. First, we used previously obtained RNAseq data to construct the immunome of S. frugiperda and analyze its induction. We then selected representative genes to study by RT-qPCR their induction kinetics and specificity after independent injections of each partner. We showed that both X. nematophila and S. carpocapsae participate in the induction of stable immune responses to the complex. While X. nematophila mainly induces genes classically involved in antibacterial responses, S. carpocapsae induces lectins and genes involved in melanization and encapsulation. We discuss putative relationships between these differential inductions and the pathogen immunosuppressive strategies.

Assessing immunocompetence in red palm weevil adult and immature stages in response to bacterial challenge and entomopathogenic nematode infection.

Parasites and pathogens can follow different patterns of infection depending on the host developmental stage or sex. In fact, immune function is energetically costly for hosts and trade-offs exist between immune defenses and life history traits as growth, development and reproduction and organisms should thus optimize immune defense through their life cycle according to their developmental stage. Identifying the most susceptible target and the most virulent pathogen is particularly important in the case of insect pests, in order to develop effective control strategies targeting the most vulnerable individuals with the most effective control agent. Here, we carried out laboratory tests to identify the most susceptible target of infection by infecting different stages of the red palm weevil Rhynchophorus ferrugineus (larvae, pupae, male, and female adults) with both a generic pathogen, antibiotic-resistant Gram-negative bacteria Escherichia coli XL1-Blue, and two specific strains of entomopathogenic nematodes (EPNs), Steinernema carpocapsae ItS-CAO1 and Heterorhabditis bacteriophora ItH-LU1. By evaluating bacterial clearance, host mortality and parasite progeny release, we demonstrate that larvae are more resistant than adults to bacterial challenge and they release less EPNs progeny after infection despite a higher mortality compared to adults. Considering the two EPN strains, S. carpocapsae was more virulent than H. bacteriophora both in terms of host mortality and more abundant progeny released by hosts after death. The outcomes attained with unspecific and specific pathogens provide useful information for a more efficient and sustainable management of this invasive pest.