An autodissemination station for the transfer of an insect growth regulator to mosquito oviposition sites.

A prototype autodissemination station to topically contaminate oviposition-seeking container-dwelling mosquitoes with the insect growth regulator, pyriproxyfen, was developed and tested in the laboratory. Our test subject was the Asian tiger mosquito, Aedes albopictus (Skuse) (Diptera: Culicidae), an urban species that colonizes small-volume cryptic larval habitats and is a skip ovipositor that visits multiple containers. The station consists of a water reservoir to attract gravid females, which is joined to a transfer chamber designed to contaminate visiting mosquitoes. The unit is easily constructed by moulding wet shredded cardboard using corn starch as a binder. The essential criteria that must be met to prove the efficacy of an autodissemination station require it to demonstrate effectiveness in attracting the target insect, in transferring the toxicant to the insect that will disperse the agent, and in facilitating the subsequent transfer of the toxicant from the insect to target habitats at a lethal concentration. Cage experiments demonstrated that the unit was readily accepted by gravid females as an oviposition site. A powder formulation of pyriproxyfen-impregnated silica particles adhered to visiting Ae. albopictus females (mean: 66 particles/female), although particles were lost over time. In cage (2.2 m(3) ) trials, pyriproxyfen-charged stations resulted in 100% inhibition of adult emergence, whereas in small-room (31.1 m(3) ) trials, 81% emergence inhibition was recorded. The venereal transfer of pyriproxyfen from contaminated males to virgin females was also observed, and pyriproxyfen was subsequently transferred to water-holding containers at concentrations that inhibited emergence. Key autodissemination station features include lack of maintenance requirements, biodegradable construction, low cost and low risk.

Characterization of immunosuppressive surface coat proteins from Steinernema glaseri that selectively kill blood cells in susceptible hosts.

Surface coat proteins (SCPs) of entomopathogenic nematodes are implicated in the suppression/evasion of host immune responses, which is required for successful host colonization. Steinernema glaseri NC strain SCPs suppressed immune responses in oriental beetle larvae (Exomala orientalis), a susceptible host for S. glaseri, in a dosage-dependent manner, thus protecting Heterorhabditis bacteriophora from being killed in the same host. Melanization of H. bacteriophora decreased from 92+/-5% in the untreated check to 1+/-3% when protected by injection of 230ng of S. glaseri SCPs. As the SCPs dosage increased, freely moving H. bacteriophora increased from 3+/-4% in the untreated group to 57+/-15% with an SCPs dose of 940ng. At 2h and in the absence of SCPs, 8% and 11% of hemocytes of E. orientalis were stained by propidium iodide and Hoechst, respectively. When exposed to 300ng/microl SCPs, 70% and 96% were stained, respectively. At 6h, propidium iodide stained 37% and 92% of the hemocytes without and with SCPs, respectively. In contrast, more than 90% of the cells were stained by Hoechst with or without SCPs. As native proteins, two isolated S. glaseri SCPs had an immunosuppressive effect; they were each composed of 38kDa (PI=4.6) and 56kDa (PI=3.6) subunits. SCP peptides were sequenced using LC-MS/MS and the mass fingerprints obtained with MALDI-TOF-MS; there were no significant matches found in peptide databases, which suggests that the SCPs studied are novel proteins. Twelve cDNA sequences were derived based on short peptides and 7 of them had no significant match against the Caenorhabditis elegans protein database. One of the cDNA matched an unknown C. elegans protein and the remaining 4 cDNAs matched proteins of C. elegans and Brugia malayi.

Relationship between the successful infection by entomopathogenic nematodes and the host immune response.

Reproduction of entomopathogenic nematodes requires that they escape recognition by a host's immune system or that they have mechanisms to escape encapsulation and melanization. We investigated the immune responses of larvae for the greater wax moth (Galleria mellonella), tobacco hornworm (Manduca sexta), Japanese beetle (Popillia japonica), northern masked chafer (Cyclocephala borealis), oriental beetle (Exomala orientalis) and adult house crickets (Acheta domesticus), challenged with infective juveniles from different species and strains of entomopathogenic nematodes. The in vivo immune responses of hosts were correlated with nematode specificity and survival found by infection assays. In P. japonica, 45% of injected infective juveniles from Steinernema glaseri NC strain survived; whereas the hemocytes from the beetle strongly encapsulated and melanized the Heterorhabditis bacteriophora HP88 strain, S. glaseri FL strain, Steinernema scarabaei and Steinernema feltiae. Overall, H. bacteriophora was intensively melanized in resistant insect species (E. orientalis, P. japonica and C. borealis) and had the least ability to escape the host immune response. Steinernema glaseri NC strain suppressed the immune responses in susceptible hosts (M. sexta, E. orientalis and P. japonica), whereas S. glaseri FL strain was less successful. Using an in vitro assay, we found that hemocytes from G. mellonella, P. japonica, M. sexta and A. domestica recognized both nematode species quickly. However, many S. glaseri in M. sexta and H. bacteriophora in G. mellonella escaped from hemocyte encapsulation by 24h. These data indicate that, while host recognition underlies some of the differences between resistant and susceptible host species, escape from encapsulation following recognition can also allow successful infection. Co-injected surface-coat proteins from S. glaseri did not protect H. bacteriophora in M. sexta but did protect H. bacteriophora in E. orientalis larva; therefore, surface coat proteins do not universally convey host susceptibility. Comparisons of surface coat proteins by native and SDS-PAGE demonstrated different protein compositions between H. bacteriophora and S. glaseri and between the two strains of S. glaseri.

Production technology for entomopathogenic nematodes and their bacterial symbionts.

Entomopathogenic nematodes (genera Steinernema and Heterorhabditis) kill insects with the aid of mutualistic bacteria. The nematode-bacteria complex is mass produced for use as biopesticides using in vivo or in vitro methods, i.e., solid or liquid fermentation. In vivo production (culture in live insect hosts) is low technology, has low startup costs, and resulting nematode quality is high, yet cost efficiency is low. In vitro solid culture, i.e., growing the nematodes and bacteria on crumbled polyurethane foam, offers an intermediate level of technology and costs. In vivo production and solid culture may be improved through innovations in mechanization and streamlining. In vitro liquid culture is the most cost-efficient production method but requires the largest startup capital and nematode quality may be reduced. Liquid culture may be improved through progress in media development, nematode recovery, and bioreactor design. A variety of formulations is available to facilitate nematode storage and application.

Enhancement of entomopathogenic nematode production in in-vitro liquid culture of Heterorhabditis bacteriophoraby fed-batch culture with glucose supplementation.

Nematode yield is a decisive factor for successful large-scale commercial production of entomopathogenic nematode. Various carbon sources were tested in in-vitro liquid culture to improve the yield of the entomopathogenic nematode Heterorhabditis bacteriophora. Canola oil was the optimal carbon source for nematode culture compared to carbohydrates when applied as a sole carbon source. However, when some of carbohydrates were applied together with canola oil, significant increases in nematode yield were observed. When 25 mg glucose ml(-1) was supplemented to 25 mg oil-based liquid culture medium ml(-1), the highest nematode yield, 3.62 x 10(5) infective juveniles, was achieved at 12 days, but nematode growth was suppressed at higher than 75 mg glucose ml(-1). A fed-batch culture process was introduced in nematode liquid culture consisting of two growth phases: bacteria and nematode. In the oil fed-batch culture, in which only glucose was initially added and oil was fed to the culture after the bacterial growth phase concurrent with nematode inoculation, nematode yield increased up to 4.25 x 10(5) infective juveniles ml(-1), while the batch culture resulted in 3.60 x 10(5) infective juveniles ml(-1). These results indicate that glucose is a superior carbon source for the bacteria, whereas canola oil is optimal for the nematode. The application of fed-batch culture provides significant enhancement of nematode yield in in-vitro liquid culture.

Liquid media development for Heterorhabditis bacteriophora: lipid source and concentration.

Lipids are important entomopathogenic nematode nutritional components because they are energy reserves and serve as indicators of nematode quality. The composition and concentration of the media lipid component determine bacterial and nematode yields. Heterorhabditis bacteriophora and its symbiont, Photorhabdus luminescens, were cultured in media containing various lipid sources. As lipid concentration increased from 2.5% to 8.0% (w/v), the final yield and productivity [calculated from the number of infective juveniles (IJ)] increased significantly from 2.1 x 10(5) IJ ml(-1) to 2.8 x 10(5) IJ ml(-1) (P < 0.05) and from 8.9 x 10(5) IJ l(-1) day(-1) to 11.8 x 10(5) IJ l(-1) day(-1) (P < 0.05), respectively. The nematode yield coefficient (IJ per gram of media), however, decreased from 2.8 x 10(6) to 2.2 x 10(6) (P < 0.05), while recovery increased from 45.3% to 58.0% (P < 0.05). Bacterial cell mass remained constant at 4.6 mg ml(-1) with changing lipid content (P > 0.05). The largest nematode yield (2.8 x 10(5) IJ ml(-1)) was achieved within 8 days, using a medium containing an 8% (w/v) olive and canola oil (50:50 w/v) combination. Moreover, developmental synchrony was achieved in this medium with 96% infective juveniles. In short, lipid sources rich in mono-unsaturated fatty acids and poor in saturated fatty acids produced optimal nematode growth.

Biological control agents for white grubs (Coleoptera: Scarabaeidae) in anticipation of the establishment of the Japanese beetle in California.

We tested biological control agents for the control of 3rd-instar scarab turfgrass pests, both for the masked chafer Cyclocephala hirta LeConte and the Japanese beetle, Popillia japonica Newman. The former species is endemic in California whereas the latter, although not yet established, constitutes a permanent serious threat to agriculture and horticulture in California. We conducted experiments using C. hirta in California and P. japonica in New Jersey. A field trial conducted in 2 different California turfgrass sites compared the field persistence in the absence of hosts of Bacillus thuringiensis Berliner subspecies japonensis Buibui strain, the milky disease bacterium, Paenibacillus (=Bacillus) popilliae (Dutky), and the entomopathogenic nematodes Steinernema kushidai Mamiya and Heterorhabditis bacteriophora Poinar to that of the organophosphate diazinon. Soil samples taken 0-70 d after applications were bio-assayed with P. japonica. Only diazinon and the entomopathogenic nematode S. kushidai caused substantial mortality and S. kushidai activity persisted significantly longer than diazinon activity. In greenhouse experiments, combinations of entomopathogenic nematode species usually resulted in additive mortality of scarab larvae. Combinations of S. kushidai and diazinon also resulted in additive mortality. In field trials, the efficacy of H. bacteriophora and especially S. kushidai and S. glaseri, was comparable to that of diazinon over 14-18 d. However, it is likely that at least S. kushidai would have outperformed diazinon over an extended period because of its longer persistence and potential for recycling in the hosts. S. kushidai, should it become commercially available, deserves further examination as an alternative to chemical white grub control especially as a highly compatible component of sustainable turfgrass management.

Genetic diversity in insect-parasitic nematodes (Rhabditida: heterorhabditidae)

Little is known about the genetic structure of various species and populations of entomopathogenic nematodes. We determined genetic variability within and among isolates of seven Heterorhabditis species using random amplified polymorphic DNA (RAPD) markers. We used 10 random primers which were previously identified as useful to quantify genetic variability among these species. Mean percentage similarity among the individuals of conspecific species was 96.25%, whereas, the mean value among different isolates for three species was 83.8%. Mean percentage similarity among different species was 31. 3%. The banding patterns produced by RAPDs positively correlated with described morphological classification; however, H. hawaiiensis could not be separated from H. indicus, or H. marelatus from H. hepialius. RAPD profiles placed an unidentified isolate (IS5) with H. indicus. Copyright 1998 Academic Press.

Sex ratio and the infection process in entomopathogenic nematodes: are males the colonizing Sex?

In laboratory assays, male infective juveniles of some species of entomopathogenic nematodes in the genus Steinernema differ from females in their tendency to disperse and in their responses to the volatile cues emitted by parasitized and nonparasitized insects. These differences suggest that male infective juveniles might locate and establish in insect hosts before females, and that infection by males might render hosts suitable for nematode development and more attractive to females: "the male colonization hypothesis." We tested this hypothesis in laboratory experiments in which larvae of the greater wax moth, Galleria mellonella (L.), were exposed to infective juveniles of S. glaseri (Steiner) (NC strain) in sand columns for various periods of time. The hosts were dissected to determine the sex ratio of the adult nematodes that became established. We found that infective juveniles entered hosts over periods of up to at least 14 h, and that sex ratios varied among experiments. However, there were no temporal differences in colonization by males and females during the infection process: the proportion of males in host cadavers was not related to exposure time or to the total number of nematodes in host cadavers. This result is inconsistent with the male colonization hypothesis. We conclude that differential colonization of hosts over time by males and females either does not occur in S. glaseri, or occurs only rarely or under conditions differing from ours, or occurs to such a small extent that it is difficult to detect. In S. glaseri, male infective juveniles tend to emerge from host cadavers before females. This might give males an advantage over females in locating new hosts following a natural emergence. Nonetheless, to date, males have not been shown to colonize hosts before females in any steinernematid species. Copyright 1998 Academic Press.

Host and penetration site location by entomopathogenic nematodes against japanese beetle larvae

Entomopathogenic nematodes are soil-inhabiting parasites of insects. Behavioral responses to host and host environmental cues are critical steps in the infection process for some nematode species, such as Steinernema glaseri and Heterorhabditis bacteriophora, of finding, recognizing, and penetrating insects. We investigated the impact of host and host environmental cues on the infectivity of these two nematodes by testing their response to whole and wounded grass roots and gut fluid and hemolymph of Popillia japanica larvae. We also compared the influence of root presence on infectivity. Infective juveniles of both nematode species migrated to whole and wounded grass roots, suggesting that these nematodes may use cues from grass roots and root wounds for host habitat finding. Root presence enhanced the infectivity of S. glaseri, presumably because feeding larvae ingest more nematodes than nonfeeding ones. However, the infectivity of H. bacteriophora was not enhanced by grub feeding. This may due to the weak gut penetration ability of this species. We conclude that the two species respond similarly to host and host environmental cues but achieve infection differently via penetration: S. glaseri possesses superior gut penetration ability, whereas intersegmental areas such as leg and maxilla joints serve as cuticle penetration sites for H. bacteriophora. Copyright 1998 Academic Press.

Thermal response of Heterorhabditis bacteriophora transformed with the Caenorhabditis elegans hsp70 encoding gene.

A heat-shock response is induced when cells are exposed to temperatures slightly higher than their optimal physiological temperature. This response is based on the synthesis of heat-shock proteins encoded by the heat-shock genes. A correlation between the increased thermotolerance and production of 70-kDa heat-shock protein (hsp70) has been observed in many organisms. We tested this hypothesis by transferring a Caenorhabditis elegans heat-inducible hsp70 A-encoding gene into the entomopathogenic nematodes Heterorhabditis bacteriophora Hp88. Successful transformation of the gene was confirmed by Southern blot hybridization and polymerase chain reaction. Our blot studies showed that the transgenic nematodes contained five to ten copies per genome of the introduced hsp70 A gene. hsp70 mRNA transcripts were detected in both wild-type and transgenic nematodes. Transcripts increased severalfold in transgenic nematodes upon heat shock. Infective juveniles of both transgenic and wild-type nematodes that exposed to a sublethal heat treatment (35 degrees C) for 2 h followed by a normally lethal heat treatment (40 degrees C) for 1 h. More than 90% of transgenic nematodes survived heat treatment, compared to 2% to 3% of the wild-type strain. Our observations establish that overexpression of hsp70 A gene resulted an enhanced thermotolerance in the transgenic nematodes. The transgenic nematodes displayed normal growth and development.

Influence of culture method on Steinernema glaseri lipids.

Entomopathogenic nematodes can be mass produced in artificial media for use as biological insecticides. Nematode in vitro media have been primarily developed on the basis of yield without fully considering nematode nutritional requirements. We investigated the quality and quantity of lipids in the entomopathogenic nematode Steinernema glaseri when grown in vivo in Popillia japonica (a natural host), Galleria mellonella (a factitious host), and in solid and liquid media. Nematode yield (infective juveniles per mg dry organic material) was 4 times higher in the in vivo compared with the in vitro cultures. Nematodes produced in vivo using P. japonica accumulated a significantly higher amount of lipids compared with nematodes grown using G. mellonella or in vitro solid and liquid methods, respectively. Fractionation of S. glaseri total lipids revealed that nematodes produced using P. japonica accumulated significantly higher phospholipids and sterols compared with other methods. C:18 fatty acids were the predominant class of lipids in S. glaseri irrespective of production method. In vivo-produced nematodes had oleic 18:1 acid as the major fatty acid, whereas in vitro-produced S. glaseri had a mixture of oleic 18:1 and linoleic 18:2 acids as the predominant fatty acids. We conclude that the lipid composition of entomopathogenic nematode is host or medium dependent. We suggest that adjusting the in vivo medium by addition of components similar to a natural host nutritional composition should improve nematode production.

Differential susceptibility of Dysmicoccus vaccinii (Homoptera: Pseudococcidae) to entomopathogenic nematodes (Rhabditida: Heterorhabditidae and Steinernematidae).

The susceptibility of the mealy bug, Dysmicoccus vaccinii Miller & Polavarapu, to infection by various species and strains of entomopathogenic nematodes was investigated in laboratory sand-dish and sand-column assays. Steinernemo carpocapsae (Weiser) (All strain), S. feltiae (Filipjev) (AB [Australia] strain), and S. glaseri (Steiner) (NC strain) were ineffective against individual mealybugs in sand-dish assays conducted in small petri dishes (1 cm high by 3.5 cm diameter) at 25 degrees C with doses of infective juvenile nematodes ranging up to 500 or 1,000 infective juveniles and exposure periods up to 5 d. However, Heterorhabditis bacteriophora Poinar (HP88 strain and 2 New Jersey isolates). H. hawaiiensis Gardner, Stock & Kaya (MG-13 strain), and H. indicus Poinar, Karunakar & David (EMS-13 strain) induced significant mortality (65.0-90.0%) at doses as low as 100 infective juveniles and an exposure of 5 d. H. bacteriophora (HBNJ strain) was effective at doses of 500 and 1,000 infective juveniles but, together with H. zealandica Poinar (V16 strain) and 4 other H. bacteriophora isolates from New Jersey, was ineffective at doses of 100 infective juveniles. Removal of the waxy coating from the mealybugs did not influence susceptibility to H. bacteriophora (HP88 strain). In the sand-column assay (5.5 cm high by 5 cm diameter, 5-d exposure, 25 degrees C), which more closely resembles host-finding in the field, H. bacteriophora (HP88 strain) induced no significant mortality against individual mealybugs at doses of 100 infective juveniles but produced 93.8% mortality at 500 infective juveniles, whereas H. indicus (EMS-13 strain) induced 56.3 and 100% mortality at 100 and 500 infective juveniles, respectively. H. bacteriophora (HP88 strain and some New Jersey isolates), H. hawaiiensis (MG-13 strain), and H. indicus (EMS-13 strain) successfully reproduced in and emerged from mealybug cadavers. This study demonstrates strong variability in the susceptibility of D. vaccinii to different species and strains of entomopathogenic nematodes, and implicates certain heterorhabditids as promising candidates for the biological control of this insect.

Localization of a putative inositol 1,4,5-triphosphate receptor in the Limulus granulocyte.

The horseshoe crab (Limulus polyphemus) granulocyte (GR) degranulates upon contact with bacteria and release factors that mediate an immune response. Stimulated cells produce IP3, which binds to receptors (IP3R, M.W.240-300 kD) that function to release stored Ca2+ into the cytoplasm that mediates degranulation. This mechanism is believed to mediate exocytosis in the Limulus GR but IP3R in the GR has not been shown. The present study utilized monoclonal antibody 4C11 and a commercially available anti-IP3R antibody, both of which label amino acids of the N-terminal of all known isoforms. Electron microscopy, immunohistochemistry, SDS-PAGE, and Western blot analysis, which employed the use of the two antibodies, demonstrates that a putative IP3R exists in the: plasma membrane, smooth surfaced vesicles, nucleus and nuclear membrane. We hypothesize that this putative IP3R is involved in mediating the immune response of the Limulus GR.

Growth-mediated variations in fatty acids of Xenorhabdus sp.

The bacterium Xenorhabdus sp. is symbiotically associated with the entomopathogenic nematode Steinernema riobravis. This nematode is produced in monoxenic culture with Xenorhabdus sp. and is sold as a biological insecticide. Acceptable yields in fermentors can only be achieved in the presence of vigorous growth of the bacterium. We investigated the fatty acid composition of Xenorhabdus species when grown at 15, 20, 25 or 30 degrees C on media containing one of two primary carbon sources: glucose or lipids from the insect host, Galleria mellonella. Both temperature and primary carbon source significantly affected lipid quantity and quality in Xenorhabdus sp. Bacteria grown with insect lipids as a primary carbon source accumulated more lipids with greater proportion of longer chain fatty acids than bacteria grown with glucose as a primary carbon source. Cells grown with insect lipids at 15 degrees C had a lower lipid content than cells grown on the same media at 20, 25 or 30 degrees C. Increasing growth temperature increased saturated fatty acids and decreased unsaturated fatty acids, irrespective of carbon source. We recommend addition of complex fatty acid sources that resemble natural host lipids to growth medium for mass producing entomopathogenic nematodes. This could provide nematode quality similar to in vivo-produced nematodes.

Ecology in the service of biological control: the case of entomopathogenic nematodes.

Biological control manipulations of natural enemies to reduce pest populations represent large-scale ecological experiments that have both benefited from and contributed to various areas of modern ecology. Unfortunately, economic expediency and the need for rapid implementation often require that biological control programs be based more on trial and error than on sound ecological theory and testing. This approach has led to some remarkable successes but it has also produced dismal failures. This point is particularly well illustrated in the historical development and use of entomopathogenic nematodes for the biological control of insect pests. Intense effort has focused on developing these natural enemies as alternatives to chemical insecticides, in part because laboratory assays indicated that these nematodes possess a broad host range. This illusory attribute launched hundreds of field releases, many of which failed due to ecological barriers to infection that are not apparent from laboratory exposures, where conditions are optimal and host-parasite contact assured. For example, the entomopathogenic nematode Steinernema carpocapsae is a poor choice to control scarab larvae because this nematode uses an ambusher foraging strategy near the soil surface whereas the equally sedentary scarab remains within the soil profile, shows a weak host recognition response to scarabs, has difficulty overcoming the scarab immune response, and has low reproduction in this host. Conversely, two other nematodes, Heterorhabditis bacteriophora and S. glaseri, are highly adapted to parasitize scarabs: they use a cruising foraging strategy, respond strongly to scarabs, easily overcome the immune response, and reproduce well in these hosts. Increased understanding of the ecology of entomopathogenic nematodes has enabled better matches between parasites and hosts, and more accurate predictions of field performance. These results underline the importance of a strong partnership between basic and applied ecology in the area of biological control.

Compatibility of soil amendments with entomopathogenic nematodes.

The impact of inorganic and organic fertilizers on the infectivity, reproduction, and population dynamics of entomopathogenic nematodes was investigated. Prolonged (10- to 20-day) laboratory exposure to high inorganic fertilizer concentrations inhibited nematode infectivity and reproduction, whereas short (1-day) exposures increased infectivity. Heterorhabditis bacteriophora was more sensitive to adverse effects than were two species of Steinernema. In field studies, organic manure resulted in increased densities of a native population of Steinernema feltiae, whereas NPK fertilizer suppressed nematode densities regardless of manure applications. Inorganic fertilizers are likely to be compatible with nematodes in tank mixes and should not reduce the effectiveness of nematodes used for short-term control as biological insecticides, but may interfere with attempts to use nematodes as inoculative agents for long-term control. Organic manure used as fertilizer may encourage nematode establishment and recycling.

Host recognition behaviour predicts host suitability in the entomopathogenic nematode Steinernema carpocapsae (Rhabditida:Steinernematidae).

Steinernema carpocapsae (Rhabditida: Steinernematidae) host recognition behaviour was assessed and compared with 2 measures of host suitability. Previous research showed that S. carpocapsae infective juveniles respond to host cues in a hierarchical order, with attraction to Galleria mellonella volatiles being stimulated by contact with G. mellonella cuticle. We measured host recognition behaviour by calculating the percentage response of S. carpocapsae infective juveniles to volatiles produced by G. mellonella last instars after the nematodes were exposed to the cuticle of 11 candidate arthropod hosts and 2 control surfaces. Host suitability was measured by nematode-induced mortality to candidate hosts at 2 nematode doses and the level of reproduction supported by each host. The highest recognition response was scored for Agrotis ipsilon (Lepidoptera: Noctuidae). This insect also incurred nearly 100% mortality due to nematode infection and supported the highest level of reproduction. Non-insect arthropods tested (Chilopoda and Isopoda) stimulated no behavioural response and were not susceptible to nematode infection. Other insect species elicited intermediate levels of the recognition response. There were significant correlations between behavioural response and nematode-induced mortality at the lower dose. The level of reproduction supported by the candidate hosts was also correlated with S. carpocapsae behavioural response.

Entomopathogenic nematode (Heterorhabditidae and Steinernematidae) spatial distribution in turfgrass.

Understanding the temporal and spatial distribution of entomopathogenic nematodes is essential for determining the role of these insect parasites in soil communities and ultimately for their use in suppression of pest insect populations. We measured the vertical and horizontal distribution of endemic populations of entomopathogenic nematodes (Steinernema carpocapsae and Heterorhabditis bacteriophora) in turfgrass. Vertical distribution was determined by taking soil cores every 3 h from 05.00 to 23.00 h, over 4 days, and dividing the cores into 8, 1 cm deep sections. Steinernema carpocapsae was recovered primarily near the soil surface: 50% of positive sections were recovered in the thatch or first 1 cm of soil. S. carpocapsae recovery was lower during the middle of the day and none were recovered in the upper section. H. bacteriophora was recovered uniformly throughout the top 8 cm of soil and its vertical distribution did not change over the course of the day. Horizontal distribution was measured as the number of nematodes recovered from cores taken from 12 randomly selected 0.3 x 0.8 m sections from within four 15.3 x 15.3 m plots. Samples were collected biweekly over a 9-month period. H. bacteriophora had a patchier distribution than S. carpocapsae and both nematode species had more patchy distributions then their potential hosts. Our results support the hypothesis that these two species of nematode utilize different foraging strategies; S. carpocapsae primarily a surface adapted ambusher and H. bacteriophora as a cruise forager.