Interaction of microbial populations in Steinernema (Steinernematidae,Nematoda) infected Galleria mellonella larvae

Infection of Galleria mellonella larvae with the entomopathogenic nematodes Steinernema feltiae (A21 and R strains) and Steinernema glaseri (Dongrae) resulted in several species of bacteria, including the respective bacterial symbiont, Xenorhabdus spp., growing in the infected insect cadavers. These other bacteria were Enterococcus in all three nematode infections studied and Acinetobacter in the S. feltiae infections. The respective populations of these bacteria changed with time. Following infection of G. mellonella larvae with any one of the Steinernema sp., only Enterococcus bacteria were detected initially in the dead larvae. Between 30 and 50h post-infection Xenorhabdus bacteria were detected and concurrent with this Enterococcus population declined to zero. This was probably due to secondary metabolites with antibacterial properties that were produced by Xenorhabdus. In the S. feltiae (both R and A21 strains) infections a third bacterium, Acinetobacter, appeared at about 130h (in S. feltiae A21 infections) or 100h (in S. feltiae R infections) and increased in population size to approximately that of Xenorhabdus. It was demonstrated that Enterococcus, orginating from the G. mellonella digestive tract, was sensitive to the organically soluble antimicrobials produced by Xenorhabdus but Acinetobacter, which was carried by the nematode, was not.     

Dynamics of carbon dioxide release from insects infected with entomopathogenic nematodes

The quality of an insect as a host to an entomopathogenic nematode infective juvenile depends in part on whether or not the insect is already infected and on the stage of that infection. Previous research has shown that nematode response to hosts can change after infection and that, for uninfected hosts, CO(2) can be an important cue used by infective stage juveniles during attraction. We hypothesized that CO(2) production from an insect changes after it is infected, and that these changes could influence nematode infection decisions. Changes in CO(2) released by two insect species (Galleria mellonella and Tenebrio molitor) after infection by one of four nematode species (Steinernema carpocapsae, Steinernema feltiae, Steinernema glaseri, or Steinernema riobrave) were measured. Measurements were taken every 2h from time of initial exposure to nematodes up to 224 h after infection. Dead (freeze-killed) and live uninfected insects were used as controls. Infected G. mellonella showed two distinct peaks of CO(2) production: one between 20 and 30 h and the other between 70 and 115 h after exposure to the nematodes. Peaks were up to two times higher than levels produced by uninfected insects. Infected T. molitor showed only one peak between 25 and 50h. We found differences in peak height and timing among nematode and insect species combinations. The influence of these changes in CO(2) production on IJ attraction and infection behavior remains to be determined.     

Acclimation of entomopathogenic nematodes to novel temperatures: trehalose accumulation and the acquisition of thermotolerance

The effect of thermal acclimation on trehalose accumulation and the acquisition of thermotolerance was studied in three species of entomopathogenic nematodes adapted to either cold or warm temperatures. All three Steinernema species accumulated trehalose when acclimated at either 5 or 35 degrees C, but the amount of trehalose accumulation differed by species and temperature. The trehalose content of the cold adapted Steinernema feltiae increased by 350 and 182%, of intermediate Steinernema carpocapsae by 146 and 122% and of warm adapted Steinernema riobrave by 30 and 87% over the initial level (18.25, 27.24 and 23.97 microg trehalose/mg dry weight, respectively) during acclimation at 5 and 35 degrees C, respectively. Warm and cold acclimation enhanced heat (40 degrees C for 8h) and freezing (-20 degrees C for 4h) tolerance of S. carpocapsae and the enhanced tolerance was positively correlated with the increased trehalose levels. Warm and cold acclimation also enhanced heat but not freezing tolerance of S. feltiae and the enhanced heat tolerance was positively correlated with the increased trehalose levels. In contrast, warm and cold acclimation enhanced the freezing but not heat tolerance of S. riobrave, and increased freezing tolerance of only warm acclimated S. riobrave was positively correlated with the increased trehalose levels. The effect of acclimation on maintenance of original virulence by either heat or freeze stressed nematodes against the wax moth Galleria mellonella larvae was temperature dependent and differed among species. During freezing stress, both cold and warm acclimated S. carpocapsae (84%) and during heat stress, only warm acclimated S. carpocapsae (95%) maintained significantly higher original virulence than the non-acclimated (36 and 47%, respectively) nematodes. Both cold and warm acclimated S. feltiae maintained significantly higher original virulence (69%) than the non-acclimated S. feltiae (0%) during heat but not freezing stress. In contrast, both warm and cold acclimated S. riobrave maintained significantly higher virulence (41%) than the non-acclimated (14%) nematodes during freezing, but not during heat stress. Our data indicate that trehalose accumulation is not only a cold associated phenomenon but is a general response of nematodes to thermal stress. However, the extent of enhanced thermal stress tolerance conferred by the accumulated trehalose differs with nematode species.     

Distribution of the entomopathogenic nematodes from La Rioja (Northern Spain)

Entomopathogenic nematodes (EPNs) distribution in natural areas and crop field edges in La Rioja (Northern Spain) has been studied taking into account environmental and physical-chemical soil factors. Five hundred soil samples from 100 sites of the most representative habitats were assayed for the presence of EPNs. The occurrence of EPNs statistically fitted to a negative binomial distribution, which pointed out that the natural distribution of these nematodes in La Rioja was in aggregates. There were no statistical differences (p < or = 0.05) in the abundance of EPNs to environmental and physical-chemical variables, although, there were statistical differences in the altitude, annual mean air temperature and rainfall, potential vegetation series and moisture percentage recovery frequency. Twenty-seven samples from 14 sites were positive for EPNs. From these samples, twenty isolates were identified to a species level and fifteen strains were selected: 11 Steinernema feltiae, two S. carpocapsae and two S. kraussei strains. S. kraussei was isolated from humid soils of cool and high altitude habitats and S. carpocapsae was found to occur in heavy soils of dry and temperate habitats. S. feltiae was the most common species with a wide range of altitude, temperature, rainfall, pH and soil moisture, although this species preferred sandy soils. The virulence of nematode strains were assessed using G. mellonella as insect host, recording the larval mortality percentage and the time to insect die, as well as the number of infective juveniles produced to evaluate the reproductive potential and the time tooks to leave the insect cadaver to determinate the infection cycle length. The ecological trends and biological results are discussed in relationship with their future use as biological control.     

Distribution patterns of entomopathogenic nematodes applied through drip irrigation systems

The distribution of entomopathogenic nematodes applied by drip irrigation was evaluated by injecting small volumes of Steinernema carpocapsae (Weiser) All strain, Steinernema feltiae (Filipjev) SN strain, Steinernema glaseri Steiner, and Heterorhabditis bacteriophora HP 88 strain Poinar suspensions into drip irrigation lines. Additionally, Steinernema riobrave Cabanillas, Poinar, & Raulston, and S. carpocapsae were injected in a 10-liter volume of water with an injection pump. Overall, the nematodes were evenly distributed along the drip lines. The total number of nematodes recovered from drip emitters was variable ranging from 42 to 92%. However, drip irrigation lines have potential to deliver entomopathogenic nematodes efficiently into pest habitats.     

Influence of nematode age and culture conditions on morphological and physiological parameters in the bacterial vesicle of Steinernema carpocapsae (Nematoda: Steinernematidae)

Steinernema spp. third-stage infective juveniles (IJs) play a key role in the symbiotic partnership between these entomopathogenic nematodes and Xenorhabdus bacteria. Recent studies suggest that Steinernema carpocapsae IJs contribute to the nutrition and growth of their symbionts in the colonization site (vesicle) [Martens, E.C. and Goodrich-Blair, H., 2005. The S. carpocapsae intestinal vesicle contains a sub-cellular structure with which Xenorhabdus nematophila associates during colonization initiation. Cellular Microbiol. 7, 1723-1735.]. However, the morphological and physiological interactions between Xenorhabdus symbionts and Steinernema IJs are not understood in depth. This study was undertaken to assess the influence of culture conditions and IJ age on the structure, nutrition, and symbiont load (colonization level) of S. carpocapsae vesicles. Our observations indicate the vesicles of axenic IJs are shorter and wider than those of colonized IJs. Moreover, as colonized IJs age the vesicle becomes shorter and narrower and bacterial load declines. The colonization proficiency of several bacterial metabolic mutants was compared between two cultivation conditions: in vitro on lipid agar and in vivo in Galleria mellonella insects. Colonization defects were generally less severe in IJs cultivated in vivo versus those cultivated in vitro. However, IJs from both cultivation conditions exhibited similar declining bacterial load over time. These results suggest that although the vesicle forms in the absence of bacteria, the presence of symbionts within the vesicle may influence its fine structure. Moreover, these studies provide further evidence in support of the concept that the conditions under which steinernematid nematodes are cultivated and stored affect the nutritive content of the vesicle and the bacterial load, and therefore have an impact on the quality of the nematodes for their application as biological control agents.     

Host cadavers protect entomopathogenic nematodes during freezing

The entomopathogenic nematodes Heterorhabditis bacteriophora, Steinernema carpocapsae, Steinernema glaseri, and Steinernema feltiae were exposed to freezing while inside their hosts. Survival was assessed by observing live and dead nematodes inside cadavers and by counting the infective juveniles (IJs) that emerged after freezing. We (1) measured the effects of 24h of freezing at different times throughout the course of an infection, (2) determined the duration of freezing entomopathogenic nematodes could survive, (3) determined species differences in freezing survival. Highest stage-specific survival was IJs for S. carpocapsae, and adults for H. bacteriophora. When cadavers were frozen two or three days after infection, few IJs emerged from them. Freezing between five and seven days after infection had no negative effect on IJ production. No decrease in IJ production was measured for H. bacteriophora after freezing. H. bacteriophora also showed improved survival inside versus outside their host when exposed to freezing.     

Development of the one-on-one quality assessment assay for entomopathogenic nematodes.

The one-on-one bioassay was developed using Steinernema carpocapsae (All) nematodes against the wax moth larva, Galleria mellonella. The assay was used to develop and compare virulence profiles of both in vitro- and in vivo-produced nematodes and to provide a quality assessment 'standard' for in vitro-produced nematodes. The bioassay was subsequently used to develop virulence profiles for Steinernema carpocapsae (UK), S. feltiae (UK), S. feltiae (R1.5), S. feltiae (SN), S. glaseri (NJ-43), and S. riobrave (RGV). These profiles are unique for each species and isolate and are used as a standard of virulence in routine quality assessment of nematodes produced in liquid fermentation.     

Vibrations as a novel signal for host location by parasitic nematodes

Nematodes are known to parasitise all major invertebrate groups in soils, and it has been assumed that their host finding relies on attraction to chemical cues. We studied movement of three species of insect-parasitic nematodes, Steinernema carpocapsae, Steinernema feltiae and Heterorhabditis megidis in response to chemical and vibrational cues. All species showed strong, significant taxes towards the vibrations. We also show that in soils, the utility of chemical cues as attractants is substantially reduced by the presence of organic matter.     

Characterization of New Entomopathogenic Nematodes from Thailand: Foraging Behavior and Virulence to the Greater Wax Moth, Galleria mellonella L. (Lepidoptera: Pyralidae)

Entomopathogenic nematodes (EPNs) in the genera Steinernema and Heterorhabditis and their associated bacteria (Xenorhabdus spp. and Photorhabdus spp., respectively) are lethal parasites of soil dwelling insects. We collected 168 soil samples from five provinces, all located in southern Thailand. Eight strains of EPNs were isolated and identified to species using restriction profiles and sequence analysis. Five of the isolates were identified as Heterorhabditis indica, and one as Heterorhabditis baujardi. Two undescribed Steinernema spp. were also discovered which matched no published sequences and grouped separately from the other DNA restriction profiles. Behavioral tests showed that all Heterorhabditis spp. were cruise foragers, based on their attraction to volatile cues and lack of body-waving and standing behaviors, while the Steinernema isolates were more intermediate in foraging behavior. The infectivity of Thai EPN strains against Galleria mellonella larvae was investigated using sand column bioassays and the LC(50) was calculated based on exposures to nematodes in 24-well plates. The LC(50) results ranged from 1.99-6.95 IJs/insect. Nine centimeter columns of either sandy loam or sandy clay loam were used to determine the nematodes' ability to locate and infect subterranean insects in different soil types. The undescribed Steinernema sp. had the greatest infection rate in both soil types compared to the other Thai isolates and three commercial EPNs (Heterorhabditis bacteriophora, Steinernema glaseri and Steinernema riobrave).     

Effectiveness of different species of entomopathogenic nematodes for biocontrol of the Mediterranean flatheaded rootborer, Capnodis tenebrionis (Linné) (Coleoptera: Buprestidae) in potted peach tree

The susceptibility of larvae of the Mediterranean flatheaded rootborer (Capnodis tenebrionis) to 13 isolates of entomopathogenic nematodes was examined using GF-677 potted trees (peachxalmond hybrid) as the host plant. The nematode strains tested included nine Steinernema feltiae, one S. affine, one S. carpocapsae and two Heterorhabditis bacteriophora. Nematodes showed the ability to locate and kill larvae of C. tenebrionis just after they enter into the roots of the tree. S. feltiae strains provided an efficacy ranging from 79.68% to 88.24%. H. bacteriophora strains resulted in control of 71.66-76.47%. S. carpocapsae (B14) and S. affine (Gspe3) caused lower control of C. tenebrionis larvae (62.03% and 34.76%, respectively). The influence of foraging strategy and the use of autochthonous nematodes to control C. tenebrionis larvae inside the roots is discussed.     

Differences between the pathogenic processes induced by Steinernema and Heterorhabditis (Nemata: Rhabditida) in Pseudaletia unipuncta (Insecta: Lepidoptera)

Larvae of Pseudaletia unipuncta are moderately susceptible to infections caused by entomopathogenic nematodes, being a desirable host to study pathogenic processes caused by Heterorhabditis bacteriophora, Steinernema carpocapsae, and Steinernema glaseri and their associated bacteria. The ability of the infective stage of these nematodes to invade hosts is quite different. S. carpocapsae invades the highest number of insects and presents the highest penetration rate, followed by H. bacteriophora. Regression analysis between the number of insects parasitized and the number of IJs counted per insect, over time, showed a high correlation for S. carpocapsae whereas for H. bacteriophora it was low. Dose-response was most evident at a concentration below 100 IJs per insect on H. bacteriophora, whereas on S. carpocapsae it was found for doses ranging from 100 to 2,000 IJs. Student's t test analysis of dose-response showed parallel, yet unequal, slopes for both strains of H. bacteriophora, whereas distinct regressions were obtained for S. carpocapsae and S. glaseri, thus, evidencing each species develop a distinct pathogenic process. Insects injected with Photorhabdus luminescens died within 50 h after injection, whereas those treated with X. nematophila died much later. Moreover, the mortality in insects exposed to H. bacteriophora complex and injected with P. luminescens was close, but insects injected with bacteria died faster. Insect mortality in treatments with complexes S. carpocapsae and S. glaseri was significantly higher than that which was observed in insects injected with symbiotic bacteria.     

Directional movement of entomopathogenic nematodes in response to electrical field: effects of species, magnitude of voltage, and infective juvenile age

Entomopathogenic nematodes respond to a variety of stimuli when foraging. Previously, we reported a directional response to electrical fields for two entomopathogenic nematode species; specifically, when electrical fields were generated on agar plates Steinernema glaseri (a nematode that utilizes a cruiser-type foraging strategy) moved to a higher electric potential, whereas Steinernema carpocapsae, an ambush-type forager, moved to a lower potential. Thus, we hypothesized that entomopathogenic nematode directional response to electrical fields varies among species, and may be related to foraging strategy. In this study, we tested the hypothesis by comparing directional response among seven additional nematode species: Heterorhabditis bacteriophora, Heterorhabditis georgiana, Heterorhabditis indica, Heterorhabditis megidis, Steinernema feltiae, Steinernema riobrave, and Steinernema siamkayai. S. carpocapsae and S. glaseri were also included as positive controls. Heterorhabditids tend toward cruiser foraging approaches whereas S. siamkayai is an ambusher and S. feltiae and S. riobrave are intermediate. Additionally, we determined the lowest voltage that would elicit a directional response (tested in S. feltiae and S. carpocapsae), and we investigated the impact of nematode age on response to electrical field in S. carpocapsae. In the experiment measuring diversity of response among species, we did not detect any response to electrical fields among the heterorhabditids except for H. georgiana, which moved to a higher electrical potential; S. glaseri and S. riobrave also moved to a higher potential, whereas S. carpocapsae, S. feltiae, and S. siamkayai moved to a lower potential. Overall our hypothesis that foraging strategy can predict directional response was supported (in the nematodes that exhibited a response). The lowest electric potential that elicited a response was 0.1 V, which is comparable to electrical potential associated with some insects and plant roots. The level of response to electrical potential diminished with nematode age. These results expand our knowledge of electrical fields as cues that may be used by entomopathogenic nematodes for host-finding or other aspects of navigation in the soil.     

Virulence of entomopathogenic nematodes to pecan weevil larvae, Curculio caryae (Coleoptera: Curculionidae), in the laboratory

The pecan weevil, Curculio caryae (Horn), is a key pest of pecans in the Southeast. Entomopathogenic nematodes have been shown to be pathogenic toward the larval stage of this pest. Before this research, only three species of nematodes had been tested against pecan weevil larvae. In this study, the virulence of the following nine species and 15 strains of nematodes toward fourth-instar pecan weevil was tested: Heterorhabditis bacteriophora Poinar (Baine, HP88, Oswego, NJ1, and Tf strains), H. indica Poinar, Karunakar & David (original and Homl strains), H. marelatus Liu & Berry (IN and Point Reyes strains), H. megidis Poinar, Jackson & Klein (UK211 strain), H. zealandica Poinar (NZH3 strain), Steinernema riobrave Cabanillas, Poinar & Raulston (355 strain), S. carpocapsae (Weiser) (All strain), S. feltiae (Filipjev) (SN strain), and S. glaseri (Steiner) (NJ43 strain). No significant difference in virulence was detected among nematode species or strains. Nematode-induced mortality was not significantly greater than control mortality (in any of the experiments conducted) for the following nematodes: H. bacteriophora (Baine), H. zealandica (NZH3), S. carpocapsae (All), S. feltiae (SN), S. glaseri (NJ43), and S. riobrave (355). All other nematodes caused greater mortality than the control in at least one experiment. Heterorhabditis megidis (UK211) but not H. indica (original) displayed a positive linear relationship between nematode concentration and larval mortality. Results suggested that, as pecan weevil larvae age, they may have become more resistant to infection with entomopathogenic nematodes.     

Steinernema feltiae (DD-136) and S. glaseri: Persistence in Soil and Bark Compost and Their Influence on Native Nematodes

Infective juveniles (J3) of the entomogenous nematodes Steinernema feltiae DD-136 (ca. 10,000 J3/100 ml) and S. glaseri (ca. 2,500 J3/100 ml) were incubated in steam-sterilized and nonsterilized sandy soil and bark compost for 8 weeks at 25 C. The nematodes were recovered by a two-step extraction procedure at 1-week intervals, and their infectivity to lepidopterous larvae (Spodoptera litura and Galleria mellonella) and their effect on the population and community of native nematodes in soil were determined. Survival of inoculated nematodes and mortality of insects were enhanced in sterilized media. Nonsterilized bark compost proved to be equally as suitable a medium as sterilized compost. In nonsterilized soil, the survival curve of S.feltiae declined more rapidly than that or S. glaseri which was less infective to insects despite its greater persistence even in nonsterilized soil. Soon after the addition of steinernematids to soil, the population of native nematodes showed a fluctuation with an increase in rhabditids and a decrease in other kinds of nematodes.     

Laboratory Evaluation of Seven Pakistani Strains of Entomopathogenic Nematodes Against a Stored Grain Insect Pest, Pulse beetle Callosobruchus chinensis (L.)

Seven Pakistani strains of entomopathogenic nematodes belonging to the genera Steinernema and Heterorhabditis were tested against last instar and adult stages of the pulse beetle, Callosobruchus chinensis (L.). These nematodes included Steinernema pakistanense Shahina, Anis, Reid and Maqbool (Ham 10 strain); S. asiaticum Anis, Shahina, Reid and Rowe (211 strain); S. abbasi Elawad, Ahmad and Reid (507 strain); S. siamkayai Stock, Somsook and Reid (157 strain); S. feltiae Filipjev (A05 strains); Heterorhabditis bacteriophora Poinar (1743 strain); and H. indica Poinar, Karunakar and David (HAM-64 strain). Activity of all strains was determined at four different nematode densities in Petri dishes and in concrete containers. A significant nematode density effect was detected for all nematode species tested. Overall, Heterorhabditis bacteriophora, S. siamkayai, and S. pakistanense were among those that showed the highest virulence to pulse beetle larvae and adults. For all nematode species, the last larval stage of the pulse beetle seems to be more susceptible than the adult. LC(50) values in Petri dish and concrete containers were 14-340 IJs/larvae and 41-441 IJs/larvae, respectively, and 59-1376 IJs/adult and 170-684/adult, respectively.     

Soil mediates the interaction of coexisting entomopathogenic nematodes with an insect host

We tested for soil substrate effects on the movement and infectivity of naturally co-occurring entomopathogenic nematodes Steinernema feltiae and Heterorhabditis marelatus, alone and in combination. We manipulated the presence and bulk density of soil and added Galleria mellonella baits within capped and perforated 15mL centrifuge tubes. Sampling tubes were then deployed in situ into field and laboratory settings as experimental traps for infective juveniles. In comparisons with standard soil collections from Lupinus arboreus rhizospheres, sampling tubes were equally sensitive to the presence of H. marelatus and more sensitive to S. feltiae. In laboratory microcosms, both EPN species infected Galleria at high frequencies in tubes lacking soil and in the absence of heterospecifics. Infection frequency of S. feltiae was unaffected by the presence of H. marelatus, but it declined with higher soil bulk density inside tubes. In contrast, detectable infection frequency by H. marelatus was reduced only marginally by the presence of soil but severely by the presence of S. feltiae. Thus, the presence of soil in tubes reversed the identity of dominant species infecting Galleria in tubes, an effect magnified when soils were compacted. Moreover, S. feltiae rarely moved into tubes lacking Galleria baits, whereas H. marelatus colonized unbaited tubes 4- to 5-fold more frequently than S. feltiae. In situ, sampling tubes acted as filters to reduce interference and contamination by fungal pathogens common in field soils. The method allows precision sampling with minimal soil disturbance while protecting bait insects from scavengers. Manipulation of tube design may allow selective sampling of EPN species, depending on the abiotic characteristics of soils, and the biology, behavior, and interspecific interactions of coexisting species.     

Facultative scavenging as a survival strategy of entomopathogenic nematodes

Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria mellonella larvae, we demonstrated that these nematodes use scavenging as an alternative survival strategy. We consider scavenging as the ability of entomopathogenic nematodes to penetrate, develop and produce offspring in insects which have been killed by causes other than the nematode-bacteria complex. Six Steinernema and two Heterorhabditis species scavenged but there were differences among them in terms of frequency of colonisation and in the time after death of G. mellonella larvae that cadavers were penetrated. The extremes of this behaviour were represented by Steinernema glaseri which was able to colonise cadavers which had been freeze-killed 240 h earlier and Heterorhabditis indica which only colonised cadavers which had been killed up to 72 h earlier. Also, using an olfactometer, we demonstrated that entomopathogenic nematodes were attracted to G. mellonella cadavers.     

Parasitism of bark scorpion Centruroides exilicauda (Scorpiones: Buthidae) by entomopathogenic nematodes (Rhabditida: Steinernematidae; Heterorhabditidae)

In laboratory bioassays, Steinernema glaseri Steiner, Steinernema riobrave Cabanillas, Poinar & Raulston, Heterorhabditis bacteriophora Poinar, and Heterorhabditis marelatus Liu & Berry were capable of infecting and killing the bark scorpion, Centruroides exilicauda (Wood). Steinernema feltiae (Filipjev) and Steinernema carpocapsae (Weiser) failed to infect C. exilicauda at 22 degrees C. S. glaseri, H. marelatus, and H. bacteriophora caused significant mortality at 22 degrees C, indicating the potential role of these parasites as a biocontrol option. Efficacy of S. glaseri and H. bacteriophora was reduced in an assay conducted at 25 degrees C. Only S. glaseri was able to reproduce in the target host. Dissection of scorpions at the end of the experimental periods revealed inactive juvenile S. riobrave, H. marelatus, and H. bacteriophora nematodes. Both mermithid and oxyurid nematodes have been documented as nematode parasites of scorpions, but rhabditids have not been reported until now. Field studies are warranted to assess the usefulness of entomopathogenic nematodes as biocontrol agents of bark scorpions.     

Effect of entomopathogenic nematodes on Plectrodera scalator (Fabricius) (Coleoptera: Cerambycidae)

Entomopathogenic nematodes were screened for efficacy against the cottonwood borer, Plectrodera scalator (Fabricius). Steinernema feltiae SN and S. carpocapsae All killed 58 and 50% of larvae, respectively, in filter paper bioassays but less than 10% in diet cup bioassays. S. glaseri NJ, S. riobrave TX, and H. indica MG-13 killed less than 10% of larvae in both assays. H. marelata IN was ineffective in the diet cup bioassay and killed 12.9% of larvae in a filter paper bioassay. The nematode isolates we tested are not suitable for use as biological control agents against P. scalator.