Occurrence of Steinernema species in cabbage fields and the effect of inoculated S. feltiae on Delia radicum and its parasitoids

Abstract 1 Seven organically grown cabbage fields were surveyed for entomopathogenic nematodes in the autumn by baiting. Nematodes were obtained from three fields with bait larvae infection ranging from 1.3–4.0%. 2 Inoculation of Steinernema feltiae (Rhabditida: Steinernematidae) in the spring increased bait larvae infection to 16.0–32.0%. 3 Four different species were found (Steinernema affine, Steinernema bicornutum, S. feltiae and Steinernema C1). 4 The number of Delia radicum (L.) (Diptera: Anthomyiidae) puparia was significantly reduced at plants where S. feltiae had been inoculated. 5 Delia radicum parasitoids were also affected by S. feltiae. The results indicated that Aleochara species (Coleoptera: Staphylinidae) were more negatively affected than Trypliographa rapae (Westw.) (Hymenoptera: Cynipidae).     

Recycling of entomopathogenic nematodes in Delia radicum and in other insects from cruciferous crops

Larvae of Deliaradicum Linnaeus (Diptera: Anthomyiidae) wereexposed to Steinernema feltiae Filipjev(Rhabditida) to study the ability of thenematodes to fulfill their life cycle (recycle)in this host. In addition, the recycling ofSteinernema and Heterorhabditis(Rhabditida) species was studied in larvae ofthe following insects: Meligethes spp.Stephens (Coleoptera: Nitidulidae),Dasyneura brassicae Winnertz (Diptera:Cecidomyidae), Ceutorrhynchus assimilisPaykull, C. pallidactylus Marsham (Coleoptera: Curculionidae) and Mamestrabrassicae Linnaeus (Lepidoptera: Noctuidae).All larval instars of D. radicum werestudied (larvae live and pupate in soil)whereas the final instars of the remaininginsects were studied (larvae only pupate in soil).On average, the following numbers of infectivejuveniles were produced in the different hosts:1,200–1,400 in C. assimilis; 700–1,300 inMeligethes spp. and 47,000 in M.brassicae. In D. radicum, around 400infective juveniles were produced on average inthe smallest larvae and around 3,500 in thelargest larvae. The highest number of nematodesrecorded in a D. radicum larva (thirdinstar) was 9,500 infective juveniles. Thenumber of nematodes produced in D.radicum cadavers was positively related to thesize of the insect host, but large variationwas observed.     

Pathogenicity of three entomopathogenic nematodes against the onion thrips,Thrips tabaci Lind. (Thys.; Thripidae)

Pathogenicity of a native isolate of Steinernema feltiae (H1) and two exotic strains, Heterorhabditis bacteriophora and Steinernema carpocapsae was assessed under laboratory conditions using different concentrations i.e. 4000, 6000, 8000 and 10,000 infective juveniles/ml against second instar larvae, prepupa and pupa of Thrips tabaci Lindeman. The mortality data were recorded 24 and 48?h post-inoculation. The highest mortality rate was recorded for prepupa (62%) than second instar (12.5%) by H. bacteriophora and S. carpocapsae, respectively, 24?h after treatment. No significant differences were found in mortality between prepupa and pupa with increasing the nematodes concentrations (from 4000 to 10,000 nematode/ml) but increasing nematode concentrations increased the mortality of second instar. At the end of the experiment (48?h.), S. feltiae H1 caused the highest mortality on second instar larvae (74%), whereas all other species caused 80–83% mortalities on pupa. This study suggests that native isolate of S. feltiae (H1) had high potential to infect soil-dwelling stages of T. tabaci.     

Heterorhabditid Behavior in the Presence of the Cabbage Maggot,Delia radicum, and its Host Plants

The behavior of Heterorhabditis zealandica Poinar strain T327 was investigated in the presence of the cabbage maggot, Delia radicum L., and plants that are susceptible to D. radicum infestation. Newly formed puparia and freeze-killed third instar larvae were attractive to infective nematodes. Newly harvested infective nematodes did not respond to the puparia, whereas 1-month-old and 2-month-old nematodes reached the insect targets within 15 minutes. There were no significant differences in the ability of similar-sized, third instar larval D. radicum and Galleria mellonella L., the greater wax moth, to attract nematodes. There was a tendency for a greater number of insects to attract more nematodes. The roots of ball cabbage and radish were equally attractive to nematodes, but rutabaga roots neither attracted nor repelled the nematodes. Germinated seeds of radish attracted nematodes, and there was a tendency for more numerous germinated seeds to attract more nematodes.     

Susceptibility of Meligethes spp. and Dasyneura brassicae to entomopathogenic nematodes during pupation in soil

The susceptibility of pupating larvae of pollen beetles, Meligethes spp. Stephens (Coleoptera: Nitidulidae) and brassica pod midges, Dasyneura brassicae Winnertz (Diptera: Cecidomyidae) to entomopathogenic nematodes (Nematoda: Rhabditida) was studied in the laboratory. The results showed that brassica pod midge larvae were almost unaffected by the tested nematodes (Steinernema bicornutum, S. feltiae and Heterorhabditis bacteriophora) whereas successful pupation of pollen beetle larvae was reduced with increasing number of nematodes (S. bicornutum, S. carpocapsae, S. feltiae and H. bacteriophora). The exposed larvae had been collected in the field and some of the pollen beetle larvae were parasitised by parasitoid wasps. It appeared that parasitised larvae were less affected by nematodes than non-parasitised larvae.     

Predation of entomopathogenic nematodes by <Emphasis Type="Italic">Sancassania</Emphasis> sp. (Acari: Acaridae)

Predation of the entomopathogenic nematode, Steinernema feltiae (Rhabditida: Steinernematidae), by Sancassania sp. (Acari: Acaridae) isolated from field-collected scarab larvae was examined under laboratory conditions. Adult female mites consumed more than 80% of the infective juvenile (IJ) stage of S. feltiae within 24 h. When S. feltiae IJs were exposed to the mites for 24 h and then exposed to Galleria mellonella (Lepidoptera: Pyralidae) larvae, the number of nematodes penetrating into the larvae was significantly lower compared to S. feltiae IJs that were not exposed to mites (control). Soil type significantly affected the predation rate of IJs by the mites. Mites preyed more on nematodes in sandy soil than in loamy soil. We also observed that the mites consumed more S. feltiae IJs than Heterorhabditis bacteriophora (Rhabditida: Heterorhabditidae). No phoretic relationship was observed between mites and nematodes and the nematodes did not infect the mites.     

Stage-specific mortality of Rhagoletis indifferens (Diptera: Tephritidae) exposed to three species of Steinernema nematodes

Mortality of larval, pupal, and adult western cherry fruit fly, Rhagoletis indifferens (Tephritidae) exposed to the steinernematid nematodes Steinernema carpocapsae, Steinernema feltiae, and Steinernema intermedium, was determined in the laboratory and field. Larvae were the most susceptible stage, with mortality in the three nematode treatments ranging from 62 to 100%. S. carpocapsae and S. feltiae were equally effective against larvae at both 50 and 100 infective juveniles (IJs)/cm². S. intermedium was slightly less effective against larvae than the other two species. Mortalities of R. indifferens larvae at 0, 2, 4, and 6 days following their introduction into soil previously treated with S. carpocapsae and S. feltiae at 50 IJs/cm² were 78.6, 92.5, 95.0, and 77.5% and 87.5, 52.5, 92.5, and 70.0%, respectively, and at 100 IJs/cm² were 90.0, 92.0, 100.0, and 84.0% and 90.0, 50.0, 42.0, and 40.0%, respectively. There was no decline in mortality caused by S. carpocapsae as time progressed, whereas there was in one test with S. feltiae. Larval mortalities caused by the two species were the same in a 1:1:1 vermiculite:peat moss:sand soil mix and a more compact silt loam soil. In the field, S. carpocapsae and S. feltiae were equally effective against larvae. Pupae were not infected, but adult flies were infected by all three nematode species in the laboratory. S. carpocapsae was the most effective species at a concentration of 100 IJs/cm² and infected 11–53% of adults that emerged. The high pathogenicity of S. carpocapsae and S. feltiae against R. indifferens larvae and their persistence in soil as well as efficacy in different soil types indicate both nematodes hold promise as effective biological control agents of flies in isolated and abandoned lots or in yards of homeowners.     

Diversity and distribution of entomopathogenic nematodes (Rhabditida: Steinernematidae and Heterorhabditidae) in Turkey

The diversity and distribution of entomopathogenic nematodes in thefamilies Steinernematidae and Heterorhabditidae were assessed throughout anextensive soil survey in Turkey during 1999 and 2000. Entomopathogenic nematodeswere recovered from six out of seven regions sampled, with 22 positive sites(2%) out of 1080 sites sampled. A single nematode isolate was recovered at eachof the positive sites, of which 15 were steinernematid isolates and seven wereheterorhabditid isolates representing a total of four species. Based onmorphometric and molecular data, the nematode species were identified asHeterorhabditis bacteriophora, Steinernemafeltiae, S. affine, andSteinernema n. sp. The most common species was S.feltiae, which was isolated from 10 sites in six regions, followed byH. bacteriophora from seven sites in five regions,S. affine from four sites in two regions, andSteinernema n. sp. from one site. Heterorhabditisbacteriophora and S. feltiae have been found inmany parts of the world, whereas S. affine, so far, hasonly been recovered in Europe until our survey. Steinernemaaffine was isolated from the European (Marmara) as well as theAsiatic region (Middle Anatolia) of Turkey. A new undescribedSteinernema sp. was isolated from the most eastern region(East Anatolia) of Turkey. Soils of the positive sites were classified as sandy,sandy loam, or loam (68.2%) and sandy–clay–loam or clay loam (31.8%) and the pHranged from 5.6 to 7.9. The habitats from which the entomopathogenic nematodeswere isolated were broadly classified as disturbed (59.1%), which includedagricultural fields and poplar planted for lumber and wind breaks, andundisturbed (40.9%), which included pine forest, grassland, marsh and reed sites.Steinernema feltiae, S. affine, andH. bacteriophora were recovered from both disturbed andundisturbed habitats. The new Steinernema sp. was recoveredfrom grassland. Our survey showed that these nematodes occur widely throughoutTurkey, but at a frequency below that reported for other parts of the world.     

Susceptibility of two Coconut Pests,Oryctes rhinoceros [Col.: Scarabaeidae] andTirathaba rufivena [Lep.: Pyralidae], to the entomoparasitic nematodeSteinernema feltiae [= Neoaplectana carpocapsae]

Bioassay indicated that 1 st instar larvae ofOryctes rhinoceros are resistant to infections by the nematodeSteinernema feltiae (estimated LD₅₀: 90 nematodes) but thatTirathaba rufivena larvae larger than 15 mm are highly susceptible (estimated LD₅₀: 3 nematodes). The “All” strain and the “mexican” strain ofS. feltiae gave very similar responses. In a field trial 300,000 nematodes were sprayed on each of 25 palms to controlT. rufivena. Although some mortality from nematodes was observed, the treatment failed to reduce the pest population significantly.     

Susceptibility of the Colorado Potato Beetle and the Sugarbeet Wireworm to Steinernema feltiae and S. glaseri

In laboratory tests, larvae of the Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say), and the sugarbeet wireworm (SBW), Limonius californicus (Mannerheim), were exposed to the nematodes Steinernema feltiae Filipjev (Mexican strain) (= Neoaplectana carpocapsae) and S. glaseri Steiner in soil. S. feltiae caused significantly higher mortality in SBW larvae than did S. glaseri, but both nematode species were equally effective against CPB larvae. The minimum concentration of S. feltiae for 100% mortality of CPB larvae after 13 days was 157 nematodes/cm² of soil, and the LC₅₀ based on 6-day mortality was 47.5 nematodes/cm²; in contrast, 100% mortality of SBW larvae was not achieved with even the highest concentration tested, 393 nematodes/cm². CPB adults emerging from nematode-contaminated soil were not infected. In field cage tests, S. feltiae applied to the soil surface at the rates of 155 and 310 nematodes/cm² soil caused 59% and 71% mortality, respectively, of late-fourth-instar spring-generation CPB, and 28% and 29% mortality, respectively, of SBW. No infection was obtained when larvae of summer generation CPB and SBW were placed in the same cages approximately 6 weeks after nematodes were applied to the soil. Inundative soil applications of S. feltiae, though cost prohibitive at present, were effective in reducing caged CPB and SBW field populations.     

Effectiveness of entomopathogenic nematodes Steinernema feltiae and Heterorhabditis bacteriophora on the Colorado potato beetle Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae) under laboratory and greenhouse conditions

In laboratory and greenhouse studies, the invading ability, virulence, and mortality caused by Stinernema feltiae and Heterorhabditis bacteriophora were compared. After one and two days of exposure to either nematode species, the mortality of Colordo potato beetle (CPB) Leptinotarsa decemlineata larvae at different instars, third and fourth, was recorded and the number of nematodes invading cadavers was more than the number of nematodes inside the larvae at the late last instar (one day before pre-pupa). Two concentrations, 250 and 500 IJs/dish, infective juvenile nematodes/0.5 ml were tested on different CPB larval instar. S. feltiae was more effective, with fourth instar rather than third and late last instar. On the other hand, H. bacteriophora showed a very weak effect with L. decemlineata. Also it was clear that S. feltiae was more effective and faster than H. bacteriophora: more than 70% of larvae were killed within 24 hours compared with H. bacteriophora which killed 40% of larvae within 48–72 hours. A significant difference in invading efficiency was observed with concentration 2500 IJs/pot in the greenhouse test. The number of adult females found in the cadavers of L. decemlineata larvae was always higher than the number of males. Foliage application of S. feltiae and H. bacteriophora resulted in a significant reduction of the number of damaged leaves and a lower index of damage compared with that in the control. We conclude that S. feltiae has significant potential and can help in the management of the Colorado potato beetle.     

Selection of insect parasitic nematodes for biological control of the garden chafer,Phyllopertha horticola

Larvae ofPhyllopertha horticola L. (Coleoptera: Scarabaeidae) cause increasing problems on sports fields and lawns in NW-Europe. A biological control programme using insect parasitic nematodes is being developed. This paper contains the results of bioassays with various species and isolates of the nematode generaHeterorhabditis andSteinernema. In bioassays in small pots with moist sand, most of the nematode isolates gave 30–60% mortality against each of the three larval stages. The susceptibility of the grubs for nematode infection generally increased with larval development.H. bacteriophora, H. heliothidis, H. megidis, a DutchHeterorhabditis isolate NLH-E87.3 andS. glaseri 326 showed the highest mortality rates, with nearly 100% mortality of third instar grubs. The DutchHeterorhabditis isolate NLH-E87.3 andS. glaseri 326 were selected as candidates for further studies on their potential as biological control agents forP. horticola grubs in the field.     

Biological control of tobacco cutworm,Spodoptera litura Fabricius with entomopathogenic nematodes

The efficacies of several entomopathogenic nematodes ofSteinernema andHeterorhabditis spp. were examined against tobacco cutworm,Spodoptera litura Fabricius.H. bacteriophora HY showed 100% mortality after 20 h against 2nd instar of tobacco cutworm. In the case of 3–4th instar,S. carpocapsae PC.,H. bacteriophora HY andS. monticola CR showed 100% mortality after 47 h. In the case of 5–6th instar,S. carpocapsae PC proved more effective than the others. Generally, the number of nematodes harvested increased as their size decreased. Also, the highest number of nematodes was obtained in the 5–6th instar ofS. litura byH. bacteriophora HY, showing about 1.3×10⁶ nematodes per larva.In vitro culturedS. carpocapsae PG showed 100% mortality after 73 h against 5–6th instar tobacco cutworm, indicating that nematodes producedin vitro can be potentially used for the biological control ofS. litura instead of nematodesin vivo.     

Evaluation of entomopathogenic nematodes against the olive fruit fly, Bactrocera oleae (Diptera: Tephritidae)

Infectivity of six entomopathogenic nematode (EPNs) species against Bactrocera oleae was compared. Similar infection levels were observed when third-instar larvae were exposed to infective juveniles (IJs) on a sand-potting soil substrate. When IJs were sprayed over naturally infested fallen olives, many larvae died within treated olives as well as in the soil; Steinernema feltiae caused the highest overall mortality of 67.9%. In addition, three laboratory experiments were conducted to optimize a time period for S. feltiae field application. (1) Abundance of fly larvae inside fallen olives was estimated over the 2006–2007 season with the highest number of susceptible larvae (3 mm and larger) per 100 olives being observed during December, 2006. (2) S. feltiae efficacy against fly larvae dropped to the soil post-IJ-application was determined. B. oleae added to the substrate before and after nematode application were infected at similar levels. (3) Effect of three temperature regimes (min–max: 10–27, 6–18, and 3–12 °C) corresponding to October through December in Davis, California on S. feltiae survival and infectivity was determined. After 8 weeks, the IJs at the 3–12 °C treatment showed the highest survival rate. However, the cold temperature significantly limited S. feltiae infectivity. Our results demonstrate that B. oleae mature larvae are susceptible to EPN infection both in the soil and within infested olives. Being the most effective species, S. feltiae may have the potential to suppress overwintering populations of B. oleae. We suggest that November is the optimal time for S. feltiae field application in Northern California.     

Evaluation of entomopathogenic nematodes for suppression of carrot weevil

The potential of entomopathogenic nematodes as biologicalcontrol agents for carrot weevil (Listronotus oregonensis) was evaluated throughboth laboratory and field experiments. In thelaboratory, Steinernema carpocapsae, S. riobrave, S. feltiae, Heterorhabditis megidis, H. bacteriophora, and a control (water only) werecompared in sand and muck soil against adults,and in sand against larvae. All nematodespecies produced high levels of larvalmortality. S. carpocapsae producedsignificantly greater adult mortality in sandthan other species or the untreated control. H. bacteriophora caused low adultmortality in sand, but the greatest adultmortality among treatments in a similar testthat used muck soil; S. carpocapsae wasranked second on muck soil. Other speciesconsistently produced intermediate (H.megidis and S. riobrave) or low (S.feltiae) levels of mortality on bothsubstrates. In the field, we compared theeffect of early season vs. late seasonapplications of H. bacteriophora or S. carpocapsae on carrot weevil mortality andparsley survival and yield. Significantdifferences among treatments in plant survivaland yield were not found; however treatmentsinvolving H. bacteriophora had higherplant survival than other treatments. Earlierapplication of this species was associated withhigher plant survival. S. carpocapsaetreatments had similar plant survival to thecontrol. Mortality of larvae and combinedstages of carrot weevil was significantlygreater at 1 week following H.bacteriophora application than for othertreatments. H. bacteriophora also showedgreater persistence than S. carpocapsaein treated plots. We conclude that H.bacteriophora is a good candidate for furtherevaluation as a biological control agentagainst carrot weevil on muck soils in theGreat Lakes region.     

Dose- and time-response assessments ofHeterorhabditis heliothidis andSteinernema feltiae [Nem.: Rhabitida] againstAedes aegypti larvae

Steinernema feltiae (=Neoaplectana carpocapsae) andHeterorhabditis heliothidis were tested against 3^rd instarAedes aegypti larvae in the laboratory. Different dosages of the nematodes and varying durations of exposure were assessed.H. heliothidis was more effective thanS. feltiae. Larval mortality showed a positive linear correlation with both nematode dosage and the duration of exposure. The number of nematodes of both species that gained access to the haemocoele of larvae was always low, but increased with dosage and exposure time. The rate of melanization of the nematodes in the larvae was correlated with dosage, but was not affected by the duration of exposure.      

Greenhouse Evaluation of Dose– and Time–Mortality Relationships of Two Nucleopolyhedroviruses for the Control of Beet Armyworm, Spodoptera exigua, on Chrysanthemum

Dose– and time–mortality relationships of baculoviruses in pest insects are important for the determination of effective spraying regimes. A series of experiments with Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) and Spodoptera exigua MNPV (SeMNPV) against synchronized populations of S. exigua larvae in greenhouse chrysanthemum was conducted. Dose– and time–mortality relationships of different virus concentrations and S. exigua target stages were determined and the area foliage consumption was measured. Crop injury was greatly reduced when S. exigua were controlled as second or third instar larvae, whereas virus applications against fourth instar larvae could not prevent considerable crop injury, even at high concentrations. SeMNPV was approximately 10 times as infectious as AcMNPV when applied on greenhouse chrysanthemum. The relative virulence of AcMNPV and SeMNPV corresponded reasonably well with previously published laboratory bioassay data. SeMNPV killed second and fourth instar S. exigua larvae approximately 12 h faster than did AcMNPV in chrysanthemum, but no difference in speed of action was found for third instar larvae. The relative speed of action of AcMNPV and SeMNPV determined in chrysanthemum and in laboratory bioassays did not correspond for third instar S. exigua larvae; laboratory bioassay data can therefore not simply be extrapolated to the crop level.     

Influence of cell density and phase variants of bacterial symbionts (Xenorhabdus spp.) on dauer juvenile recovery and development of biocontrol nematodes Steinernema carpocapsae and S. feltiae (Nematoda: Rhabditida)

The rhabditid nematodes Steinernema carpocapsae and Steinernema feltiae are used in biological control of insect pests. Mass production is done in liquid culture media pre-incubated with their bacterial symbionts Xenorhabdus nematophila and Xenorhabdus bovienii, respectively, before nematode dauer juveniles (DJs) are inoculated. As a response to food signals produced by the bacterial symbionts, the DJs exit from the developmentally arrested dauer stage (they recover development) and grow to adults, which produce DJ offspring. Variable DJ recovery after inoculation often causes process failure due to non-synchronous population development and low numbers of adult nematodes. This contribution investigated the influence of the bacterial cell density on DJ recovery and development to adults. At higher density of 10¹⁰ bacterial cells ml⁻¹, a higher percentage of DJ recovery was induced, and adults occurred earlier in both Steinernema spp. than at lower density of 10⁹ and 10⁸ cells ml⁻¹. Xenorhabdus symbionts produce phase variants. Recovery in bacteria-free supernatants was lower than in supernatants containing bacterial cells for both primary and secondary phase Xenorhabdus spp. and lower in secondary than in primary phase supernatants or cell suspensions. In general, recovery was lower for Steinernema feltiae and the time at which 50% of the population had recovered after exposure to the food signal was longer (RT₅₀ = 17.1 h) than for Steinernema carpocapsae (RT₅₀ = 6.6 h). Whereas >90% S. carpocapsae DJs recovered in hemolymph serum of the lepidopteran insect Galleria mellonella, recovery of S. feltiae only reached 31%. Penetration into a host insect prior to exposure to the insect’s food signal did not enhance DJ recovery. Consequences for liquid culture mass production of the nematodes and differences between species of the genera Steinernema and Heterorhabditis are discussed.     

Entomopathogenic nematodes control Plum Sawflies (Hoplocampa minuta and H. flava)

Plum sawflies are among the most damaging pests of European plum. Current control strategy implies insecticide application. Three species of entomopathogenic nematodes (EPN), Steinernema feltiae Filipjev, S. carpocapsae Weiser and Heterorhabditis bacteriophora Poinar were tested under laboratory and field conditions to assess effectiveness against larval and adult stages. Laboratory tests resulted in up to 100% mortality of last instar larvae before construction of a cocoon. However, the nematodes were not able to penetrate the cocoon. Foliar application did not result in plum sawflies larvae infestation by EPNs. Under field conditions, the nematodes reduced the number of emerging adults by application against sawfly larvae in the previous year before migration into the soil for overwintering by 62%–92%. Application of the nematodes against adults just before their anticipated emergence resulted in reduction of fruit infestation up to 100%. Mean results of 5 trials using caged trees were 47.8% with S. feltiae, 56.3% with S. carpocapsae and 62.9% with H. bacteriophora. In open field trails, control of adults obtained with S. feltiae at 0.5 million nematodes/m² was 98.2 and 67.8% and at 0.25 million m⁻² 41.7 and 41.2%. Forecasting adult emergence and optimal soil moisture conditions are essential for success of the nematode application.     

Comparative susceptibility of larval instars and pupae of the western corn rootworm to infection by three entomopathogenic nematodes

As a first step towards the development of an ecologically rational control strategy against western corn rootworm (WCR; Diabrotica virgifera virgifera LeConte, Coleoptera: Chrysomelidae) in Europe, we compared the susceptibility of the soil living larvae and pupae of this maize pest to infection by three entomopathogenic nematode (EPN) species. In laboratory assays using sand-filled trays, Heterorhabditis bacteriophora Poinar and H. megidis Poinar, Jackson & Klein (both Rhabditida: Heterorhabditidae) caused comparable mortality among all three larval instars and pupae of D. v. virgifera. In soil-filled trays, H. bacteriophora was slightly more effective against third larval instars and pupae, and H. megidis against third larval instars, compared to other developmental stages. In both sand and soil, Steinernema feltiae (Filipjev) (Rh.: Steinernematidae) was least effective against second instars. In conclusion, all larval instars of D. v. virgifera show susceptibility to infection by all three nematodes tested. It is predicted that early application against young larval instars would be most effective at preventing root feeding damage by D. v. virgifera. Applications of nematodes just before or during the time period when third instars are predominant in the field are likely to increase control efficacy. According to our laboratory assays, H. bacteriophora and H. megidis appear to be the most promising candidates for testing in the field. I. Hiltpold similarly contributed to this paper as the first author.