The Mediterranean fruit fly Ceratitis capitata (Diptera; Tephrititade) is an important pest of citrus and other deciduous fruit trees. There is a need for sustainable pest management tools and the use of entomopathogenic nematodes have been explored for controlling the stages of medfly that occur in the soil. We have investigated further this approach by assessing the efficacy of commercially available entomopathogenic nematodes applied early on-season or off-season when the medfly populations are passing through their annual bottleneck period, aiming at reducing their population before the growing season. In laboratory experiments, the efficacy of commercial strains of Steinernema carpοcapsae, Steinernema feltiae and Heterοrhabditis bacteriοphοra and H. downesi at doses of (1.5 x 106 IJs/m2) and 2.5 x 106 IJs/m2, at 15C° and 25C° was assessed and Steinernema feltiae was found to result in up to 70% reduction of adult medfly emergence at a dose of 2.5 x 106 IJs/m2 and lower temperatures, confirming its superiority over other commercially available species. Further trials in citrus groves in Corinthos, Greece in Spring 2021 (early season) and Autumn 2021 (off-season) confirmed that a single application of S. feltiae at moderate dose regimes can provide about 62–65% suppression of adult medlfies. Therefore, a single, moderate dose application of entomopathogenic nematodes early or off-season, which is more economical feasible can provide significant suppression of overwintering medflies and can be safely integrated with other tools for medfly management.

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 carp omicron capsae, S. feltiae and Heter omicron rhabditis bacteri omicron ph omicron ra 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 degrees 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.

Entomopathogenic nematodes (EPNs) are effective against the immature stages of the large pine weevil Hylobius abietis. In three field trials we compared the efficacy of the application method of EPN for weevil suppression below the suggested threshold of 20 weevils per stump: applying the EPN suspension in the top edges of the stumps (‘top’) vs. drenching the soil around stumps (‘standard’). For Steinernema carpocapsae, weevil suppression was below the targeted threshold only when suspension was applied in the standard way (two of the three sites). On the other hand, weevil suppression was provided in all three cases of ‘top’ application of Heterorhabditis downesi suspension, whereas suppression in ‘standard’ application was observed in one site. Percentage parasitism of developing weevils in relation to depth and distance help explain EPN movement post-application. Weevil suppression relative to suggested thresholds can be improved by altering the method of EPN application depending on the nematode species.

The large pine weevil Hylobius abietis (L.), LPW, is a major pest of trees in replanted coniferous forests in northern Europe. The use of entomopathogenic nematodes (EPNs) applied against developing stages for population suppression is increasingly recognized as an effective alternative to plant protection using chemical pesticides. Here, we report results from a series of trials we conducted over 2 years using two species of EPN, Steinernema carpocapsae (Weiser) and Heterorhabitis downesi (Stock, Griffin, and Burnell) with different foraging strategies. Trials were conducted at lodgepole pine sites in Ireland on both mineral and peat soil type. EPN suspension was applied to the stumps of felled pine trees, and EPN efficacy was determined afterwards by directly assessing parasitism rates after debarking one quarter of the stumps and by collecting emerging adult weevils from traps erected over other treated and control stumps. Our results suggest that both species of EPN are equally effective in suppressing LPW populations to below the current, informal thresholds of economic damage. EPN were equally efficient in controlling LPW in peat and in mineral (lithosols/regosols and acid brown earth/brown podzolics) soils. Weevil density and distribution within pine stumps in peat versus mineral sites can explain patterns of LPW parasitism and suppression. Our results also suggest that infestation level (number of weevils per stump) can be an important factor in forecasting EPN application success as there is evidence of negative density-dependent parasitism when weevil densities were high.

The enduring stages of entomopathogenic nematodes of the genera Steinernema and Heterorhabditis are infective juveniles, which require a high humidity and sufficient ventilation for survival. Formulations must account for these requirements. Nematodes may be formulated inside the insects in which they reproduced or they need to be cleaned and mixed with a suitable binder to maintain humidity but allowing for gas exchange. Another method for formulation is the encapsulation in beads of Ca-alginate. Generic procedures for these formulation techniques are described.

A trial of entomopathogenic nematode(Heterorhabditis indica)in the dosages of 2000, 3000, 4000, 5000 and 10000IJs/pit (peanut plant) was conducted in this study for selecting an effective dosage to control chafer grubs in a peanut fields. The results showed that: 1) Damaged legumes (%) were significantly less in all treatments compared to the control treated with tap water only. The dosages of 4000, 5000 and 10000IJs/pit resulted in 4.1%, 5.7% and 7.5% damage, respectively, which was significantly lower in the positive control treated with 40% isofenphos-methyl (10.2%); 2) A 95.7% decrease of grub numbers was achieved dosages of 4000 and 5000IJs/pit, which was significantly higher than in the positive control (82.7%); 3) Peanut yield in the dosages of 4000IJs/pit was 327.6 kg/667m 2 , significantly higher than in the positive control (292.6kg/667m 2 ). The results demonstrated that H.indica could successfully control chafer grub and subsequently increase peanut yield and is superior to the chemical control strategy. The dosage of 4000IJs/pit was the best one.

While nematodes are most commonly known for their negative impact on plants, animals, and humans, there are a number of species which are commercially explored. This review highlights some of the most important success stories for the application of nematodes. They are used as bioindicators in ecological and toxicity studies, as model organisms for elucidating fundamental biological questions and for high throughput screening of drugs. Besides these indirect uses, direct applications include the use of Beddingia siricidicola against a major forest pest and the commercialization of Steinernema, Heterorhabditis, and Phasmarhabditis as biological pest control products. New directions for the commercialization of nematodes are the use as living food, specifically loaded with essential nutrients for various fish and shrimp larvae. Even human parasites or closely related species have been successfully used for curing autoimmune disorders and are currently in the process of being developed as drugs. With the striving development of life sciences, we are likely to see more applications for nematodes in the future. A prerequisite is that we continue to explore the vast number of yet undiscovered nematode species.

Field studies on the efficacy of entomopathogenic nematodes as biological pest control agents in field crops and over multiple seasons are rare. In the present study, we investigated the efficacy of Heterorhabditis bacteriophora (Rhabditida: Heterorhabditidae) at reducing maize root damage caused by the larvae of the western corn rootworm Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae). The study consisted of 21 field‐scale experiments in Hungary conducted between 2004 and 2007. Key factors behind successful control, such as application techniques, nematode dosage, soil environment and rainfall, were analyzed. Five of the six tested techniques for applying the nematodes using readily available farming machinery resulted in a reduction of root damage, as well as subsequent plant lodging. According to damage rating with the Iowa 1–6 scale, damage was reduced by 3–18%. According to the 0.00–3.00 node injury scale, damage was reduced by 14–54%. Approximately 7–62 % of plant lodging, and thus direct yield losses as a result of in‐harvestability, was prevented. An increase in nematode dosage and/or an increased rainfall during the period of applications increased the efficacy of H. bacteriophora at reducing root damage. Except for the obvious effect of moisture, the soil environment had little measurable influence on nematode efficacy. The present study demonstrates the potential for field‐scale nematode application to significantly reduce damage caused by D. v. virgifera larvae. It is recommended that the method of agent application should be selected on a case‐by‐case basis considering the efficacy data presented here, the available application machinery and its associated costs.

The three larval instars of western corn rootworm (Diabrotica virgifera virgifera LeConte, Coleoptera: Chrysomelidae) feed on the roots of maize, Zea mays (L.). The effects of six application techniques on the plant-scale efficacy of the entomopathogenic nematode species, Heterorhabditis bacteriophora Poinar (Rhabditida: Heterorhabditidae), in controlling D. v. virgifera populations were assessed in seven field plot experiments in southern Hungary between 2004 and 2007. Approximately 230 000 nematodes were applied per row metre using four different stream spray techniques; or, alternatively 400 000 nematodes per square metre using two different flat spray techniques. Nematode efficacy was assessed by comparing the number of emerging adult D. v. virgifera, and root damage between treatments and untreated controls. All tested nematode application techniques reduced D. v. virgifera density by at least 50% (on average across fields and years). The highest reduction in D. v. virgifera density was 68% and occurred when nematodes were applied into the soil together with maize sowing using a fluid solid stream. Rainfall, the day before application likely increased the control efficacy of H. bacteriophora. Using the 0.00-3.00 node injury damage rating scale, we estimated that potential root damage was prevented by 25-79% when H. bacteriophora was applied. Although, H. bacteriophora can effectively be applied with all of the techniques tested, for optimum performance and minimum costs, it is suggested that the nematodes be applied as follows: (i) as a stream requiring 8-10 times less volume of water than flat sprays, or as a granule requiring no water, and (ii) into the soil when sowing maize, requiring less water than soil surface sprays and avoiding the destruction of nematodes by UV radiation and additional machinery use.

Diabroticina is a speciose subtribe of New World Chrysomelidae (Subfamily Galerucinae: Tribe Luperini) that includes pests such as corn rootworms, cucumber beetles and bean leaf beetles (e.g. Diabrotica, Acalymma, Cerotoma species). The evolution and spread of pesticide resistance, the European invasion of Diabrotica v. virgifera LeConte, and possible development of resistance due to the large-scale deployment of Diabrotica-active Bt maize in North America have generated a sense of urgency in developing biological control options against Diabroticina pests. In the present study, we review available knowledge on biological control options, including 290 publications on natural enemy-Diabroticina associations in the New World. Several natural enemy species or groups appear to be promising candidates for control strategies with different ecological rationales. We propose that future research should pursue: (1) development of inundative biological control products, particularly mass-produced entomopathogenic nematodes and fungi, (2) understanding of specific natural enemies of Diabroticina larvae throughout the Americas and of adults particularly in higher altitudes of Central America or northern South America including potential classical biological control agents against D. v. virgifera; (3) enhancement of natural enemies through cultural practices, i.e., reduced tillage, reduced weed control, cover crops, diversified crop rotations or soil amendments. Research and action must be coordinated to accelerate the exploration of biological control options.

Abstract Diabroticina is a speciose subtribe of New World Chrysomelidae (Subfamily Galerucinae: Tribe Luperini) that includes pests such as corn rootworms, cucumber beetles and bean leaf beetles (e.g. Diabrotica, Acalymma, Cerotoma species). The evolution and spread of pesticide resistance, the European invasion of Diabrotica v. virgifera LeConte, and possible development of resistance due to the large-scale deployment of Diabrotica-active Bt maize in North America have generated a sense of urgency in developing biological control options against Diabroticina pests. In the present study, we review available knowledge on biological control options, including 290 publications on natural enemy–Diabroticina associations in the New World. Several natural enemy species or groups appear to be promising candidates for control strategies with different ecological rationales. We propose that future research should pursue: (1) development of inundative biological control products, particularly mass-produced entomopathogenic nematodes and fungi, (2) understanding of specific natural enemies of Diabroticina larvae throughout the Americas and of adults particularly in higher altitudes of Central America or northern South America including potential classical biological control agents against D. v. virgifera; (3) enhancement of natural enemies through cultural practices, i.e., reduced tillage, reduced weed control, cover crops, diversified crop rotations or soil amendments. Research and action must be coordinated to accelerate the exploration of biological control options.

Entomopathogenic nematodes (EPNs) in the families Heterorhabditidae and Steinernematidae have considerable potential as biological control agents of soil-inhabiting insect pests. In the present study, the control potential of the EPNs Steinernema carpocapsae (TUR-S4). S. feltiae (Nemaplus), S. carpocapsae (Nemastar). S, feltiae (TUR-S3) and Heterorhabditis bacteriophora (Nematop) against a new longicorn pest, Dorcadion pseudopreissi Breuning, 1962 (Coleoptera: Cerambycidae), on turf was examined in laboratory studies. Pathogenicity test were performed at the following doses: 50, 100 and 150 Dauer Juveniles (DJs)/larva at 25 degrees C. Highest mortalities (75-92%) of the larvae were detected at the dose of 150 DJs/larva for all nematodes used. Reproduction capabilities of the used EPNs were examined at doses of 50, 75, 100 and 150 DJs/larva at 25 degrees C. S. carpocapsae (TUR-S4) had the most invasions (32 DJs/larva and reproduction (28042 DJs/larva) at the dose of 100 DJs, and the highest reproduction (per invaded DJ into a larva) was observed in H. bacteriophora (Nematop) (2402.85 DJs) at a dose of 50 DJs. The foraging behaviour of the nematodes in the presence of D. pseudopreissi and Galleria mellonella L. (Lepidoptera: Galleriidae) larvae was studied using a Petri dish filled with sand at 20 degrees C. All of the used nematodes accumulated near the larvae section of both insect species (32-53% of recovered DJs) with a higher percentage of S. carpocapsae (TUR-S4) (53%) and H. bacteriophora (48%) (Nematop) moving towards larvae of D. pseudopreissi, than the S. feltiae strains.

Three new peptides, xenortides A and B and xenematide, were isolated from a culture of the nematode-associated entomopathogenic bacterium Xenorhabdus nematophilus. Their structures were elucidated using NMR, MS, and chemical derivatization methods. Xenortides A and B are the N-phenethylamide and tryptamide derivatives, respectively, of the dipeptide (NMe-L-Leu-NMe-L-Phe). The cyclodepsipeptide xenematide has the sequence (Thr-Trp-Trp-Gly), with a 2-phenylacetamide substituent at the threonine residue and one d-tryptophan. The new peptides and the two known compounds xenocoumacin II and nematophin were tested for antibacterial, antifungal, insecticidal, and anti-Artemia salina activities. Xenematide and xenocoumacin II showed moderate antibacterial activities. Xenocoumacin II, nematophin, and the two xenortides were active in the Artemia salina assay, and xenematide acted weakly insecticidal.

Entomopathogenic nematodes are often found naturally infecting codling moth larvae. The effect of an autumn treatment with S. feltiae on the fruit damage in the following summer was evaluated by treating 4 different apple orchards in October 2004 and 2005 at application rates of 3.75; 2 and 1.5 billion nematodes in 4000 l / ha. In three of the treated orchards, one treated with 3.75x109 nematodes/ha the other two treated with 2e9 nematode/ha, reduction in fruit damage was around 50%. In the most heavily infested orchard, which was treated with 1.5x109 nematode/ha only 33% reduction in fruit damage was achieved. Compared to previous studies, this was the first assessing the effect on the fruit damage in the summer following the treatment rather than assessing the mortality of sentinel larvae fixed to the treated tree trunks.

The western corn rootworm (Diabrotica virgifera virgifera LeConte, Coleoptera: Chrysomelidae) is an invasive maize (Zea mays L.) pest in Europe. Crop yield is significantly impacted by the feeding of all three larval instars on maize roots, making them prime targets for control measures. Therefore, the control efficacy of three entomopathogenic nematodes (EPNs), Steinernema feltiae (Filipjev), Heterorhabditis bacteriophora Poinar, and H. megidis Poinar, Jackson and Klein (Nematoda: Rhabditida), was studied in four field plot experiments in southern Hungary in 2005 and 2006. All EPN species significantly reduced D. v. virgifera independently, whether applied as a row spray with a solid stream into the soil at sowing or onto the soil along maize rows in June. When applied at maize sowing, H. bacteriophora was more effective at reducing D. v. virgifera (81%) than H. megidis (49%) and S. feltiae (36%). When applied in June, H. bacteriophora and H. megidis were more effective at reducing D. v. virgifera (around 70%) than S. feltiae (32%). All tested EPN species significantly reduced damage on maize roots independently, whether they were applied at sowing or in June. Damage, however, was not totally prevented. The use of H. bacteriophora for the development of a biological control product for inundative releases against D. v. virgifera larvae is suggested.

The use of entomopathogenic nematodes (EPN) is a promising approach to control the cherry fruit fly, Rhagoletis cerasi L.. We already demonstrated the high potential of EPN to infect larvae after leaving the cherry for pupation in the soil in laboratory and field experiments. For practice, an application technique is needed, that is both, grower- and EPN friendly. We tested a tractor mounted spray boom for treatment under the canopy area. The achieved rate of EPN in the soil met the expectations. The activity of EPN in soil samples was high after application, but dropped to 60% of the initial activity within one week. Exact forecasting of larval drop from cherries is another major challenge. To obtain basic data, we recorded the phenology of infestation and larval emergence on trees which were not harvested. Sequential infestation on the same cherry variety was observed and larvae dropped from individual trees for several weeks.