Impact of Entomopathogenic Nematodes on Non-target Hosts

Biological pest control has been thought to be ecologically safe for many years. More recently, it has been questioned whether entomopathogens and beneficial arthropods or nematodes truly have no impact on non-target species. Only a few studies deal with the action of entomopathogenic nematodes on non-target animals, although a broad spectrum of species has been tested in the laboratory. Entomopathogenic nematodes do not affect vertebrates under natural conditions. Mortality caused by the release of entomopathogenic nematodes among non-target arthropod populations can occur, but will only be temporary, will be spatially restricted and will affect only part of a population. In plots treated with entomopathogenic nematodes, the impact on the non-target fauna proved to be negligible. The possible impact of introduced exotic nematode species is discussed and regulatory measures for the release are proposed.     

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.     

Factors limiting short-term persistence of entomopathogenic nematodes

Abstract:  Entomopathogenic nematodes are commonly applied to soil and crops for biocontrol of insects. Typically, when nematodes are applied, recoverable numbers decline quickly. Because ultra-violet light is known to kill entomopathogenic nematodes in the laboratory, many researchers blame poor field persistence on the action of ultra-violet light. We conducted a field experiment to test this hypothesis, using Heterorhabditis bacteriophora applied to turf as a model system. We compared persistence of surface-applied nematodes with subsurface-applied nematodes and found no difference in persistence. Numbers of potential nematode antagonists (mites and collembola) were also monitored. Poor persistence was positively correlated with numbers of mites and collembola in plots where nematodes were surface-applied, but not in plots where they were subsurface-applied.     

Effects of Crop Residue on the Persistence of Steinernema carpocapsae

We determined the effects of crop residue on the persistence of an entomopathogenic nematode, Steinernema carpocapsae. During 2 consecutive years, nematodes were applied at rates of 2.5 × 10₄ and 1.0 × 10⁵ infective juveniles/m² to small field plots planted with corn. Nematode persistence was monitored by exposing Galleria mellonella larvae to soil samples from plots with and without crop residue (approximately 75% coverage of soybean stubble). Persistence of S. carpocapsae was significantly greater in crop residue plots than in plots without residue. In crop residue plots that received the higher rate of nematode application, larval mortality did not significantly decrease during the study period (3 to 5 days) and remained above 85%. In nematode-treated plots without crop residue, however, larval mortality fell from over 96% to below 11% and 35% in the first and second trials, respectively. The increased crop residue may have benefited nematode persistence through protection from desiccation or ultraviolet light. We conclude that increased ground cover in cropping systems (e.g., due to reduced tillage) may lead to increased insect pest suppression with entomopathogenic nematodes.     

Effect of neonicotinoid synergists on entomopathogenic nematode fitness

In previous greenhouse and field studies, the neonicotinoid insecticide imidacloprid interacted synergistically with five entomopathogenic nematode species against five scarab species. Two other neonicotinoids, thiamethoxam and acetamiprid, showed a weaker interaction with nematodes in scarab larvae. Entomopathogenic nematodes have the potential to recycle in hosts after inundative applications, thereby increasing the persistence of nematodes and insect control. Thus we investigated the effect of neonicotinoids on nematode fitness after tank mixing and after combined applications. Tank mixing only had a negative effect on nematode survival and infectivity in a few nematode–insecticide combinations and only if both insecticide concentration and exposure time were several times higher than typical for field applications. Combined application of nematodes with imidacloprid generally had no negative effect on the percentage of scarab cadavers producing progeny or the number of nematode progeny emerging per cadaver. In experiments with a synergistic increase in scarab mortality, the total number of progeny in combination treatments was up to four times higher than in nematodes only treatments. Similarly, nematode populations in soil from combination treatments were 13.2 times greater than for nematodes only treatments at 28 days after treatment. Combined imidacloprid–nematode applications did not affect the pathogenicity or infectivity of the nematode progeny. Combining thiamethoxam with nematodes had no negative effects on nematode reproduction in the majority of treatments. However, due to the weaker interaction of thiamethoxam and nematodes on scarab mortality, the total number of nematode progeny per treatment generally did not increase compared with nematodes only treatments. The demonstrated tank mix compatibility of imidacloprid and nematodes improves the feasibility of combining these agents for curative white grub control. The positive effect of imidacloprid on nematode reproduction after combined application may increase the likelihood of infection of white grubs by subsequent generations of nematodes, thereby improving their field persistence and biological control potential.     

Effects of Microbial and Other Antagonistic Organism and Competition on Entomopathogenic Nematodes

Antagonistic factors, broadly identified as antibiosis, competition and natural enemies, impact on entomopathogenic nematodes. Antibiosis can occur through the release of plant chemicals from the roots into the soil, which may adversely affect the host-finding behavior of the infective stage nematode, or the presence of these chemicals in the host insect may negatively affect nematode reproduction. In laboratory studies, intra-specific and inter-specific competition reduces nematode fitness, and inter-specific competition can cause local extinction of a nematode species. For example, after concomitant infection of a host, a steinernematid species usually excludes a heterorhabditid species. The mechanism for the steinernematid superiority has been postulated to be a bacteriocin(s) produced by Xenorhabdus, the symbiotic bacterium of the steinernematid, which prevents Photorhabdus, the symbiotic bacterium of the heterorhabditid, from multiplying. Inter-specific competition between two steinernematid species shows that both can co-exist in a host, but one species will eventually prevail in the environment. By having different foraging strategies, however, both steinermatid species may co-exist in the same habitat. An important issue is whether the introduction of an exotic entomopathogenic nematode species will competitively displace an indigenous nematode species. Although the environmental risks are small, the recommended policy is that the introduction of exotic nematodes be regulated. With other pathogens, entomopathogenic nematodes can out-compete entomopathogenic fungi, but not Bacillus thuringiensis, for the same host individual when both the nematode and entomopathogen are applied simultaneously. The best studied natural enemy is the nematophagous fungus, Hirsutella rhossiliensis, which causes higher mortality in Steinernema glaseri compared with Heterorhabditis bacteriorphora. Differential susceptibility to the fungus may be associated with the retention of the second-stage cuticle by H. bacteriophora. Invertebrate predators including mites and collembolans feed on entomopathogenic nematodes. Although a number of studies have been conducted with antagonists, there is a dearth of field data. We suggest that long-term research plots be established where natural populations of entomopathogenic nematodes occur and include antagonists as a component of such studies.     

Parameters affecting plant defense pathway mediated recruitment of entomopathogenic nematodes

Entomopathogenic nematodes are natural enemies and effective biological control agents of subterranean insect herbivores. Interactions between herbivores, plants, and entomopathogenic nematodes are mediated by plant defense pathways. These pathways can induce release of volatiles and recruit entomopathogenic nematodes. Stimulation of these plant defense pathways for induced defense against belowground herbivory may enhance biological control in the field. Knowledge of the factors affecting entomopathogenic nematode behaviour belowground is needed to effectively implement such strategies. To that end, we explore the effect of elicitor, elicitor dose, mechanical damage, and entomopathogenic nematode release distance on recruitment of entomopathogenic nematode infective juveniles to corn seedlings. Increasing doses of methyl jasmonate and methyl salicylate elicitors recruited more entomopathogenic nematodes as did mechanical damage. Recruitment of entomopathogenic nematodes was higher at greater release distances. These results suggest entomopathogenic nematodes are highly tuned to plant status and present a strategy for enhancing biological control using elicitor-stimulated recruitment of entomopathogenic nematodes.     

Anfälligkeit von entomopathogenen Nematoden gegenüber nematodenfangenden und endoparasitären Pilzen

Abstract Susceptibility of entomopathogenic nematodes to nematode-trapping and endoparasitic fungi
Laboratory tests in petri dishes demonstrated that the nematode-trapping fungi Arthrobotrys superba and A. robusta preyed upon the entomopathogenic nematodes Steinernema bibionis, S. feltiae and a species of Heterohabditis. Up to 100% of the nematodes were trapped 48 h after application. Large differences existed in the time required for trapping as well as for mortality after trapping and was dependent on the fungal or nematode species studied. Arthrobotrys robusta , reduced S. bibionis densities in sand-block bioassay chambers. The prior introduction of a microphagous nematode, Panagrellus redivivus , to induce trap formation caused a 52% reduction in S. bibionis levels.
The endoparasitic fungi Harposporium anguillulae and Drechmeria coniospora did not parasitize the entomopathogenic nematodes in petri dish tests. However, Verticillium balanoides , was shown to be a parasite of S. bibionis. Predivivus and a mycophagous species of Ditylenchus were more quickly parasitized than S. bibionis , with parasitism reaching 100, 90 and 30% after 42 days, respectively. In the sand-block chambers V. balanoides did not reduce the, number of recovered S. bibionis juveniles after 14 or 28 days.     

Spatial and temporal distribution of endemic entomopathogenic nematodes in a heterogeneous vegetable production landscape

Entomopathogenic nematodes within the families Steinernematidae and Heterorhabditidae (Order: Rhabditida) are potential biological control agents for many soil-dwelling vegetable pests. However, their low persistence and efficacy after field releases have resulted in limited use in pest management programs. Understanding the factors regulating natural populations of entomopathogenic nematodes may provide insight into practices to conserve populations within production systems. A series of investigations were conducted within a vegetable production area in Willard, Ohio during 2000–2003 to gain insight into the population ecology of endemic populations of entomopathogenic nematodes. A total of 440 sites across four habitats associated with the production landscape were sampled to ascertain the natural occurrence of these beneficial nematodes. Habitats included cultivated areas, grassy banks adjacent to cultivated areas, undisturbed shrub lands and forests. Twelve sites along grassy banks were monitored over a growing season to estimate associations between abiotic and biotic factors and endemic populations. Entomopathogenic nematodes were only detected along grassy banks adjacent to the cultivated areas; nematodes were recovered from 15 to 30% of sites sampled in 2001 and 2002, respectively. Two species of nematodes were isolated, Heterorhabditis bacteriophora Poinar and Steinernema feltiae Filipjev. H. bacteriophora was the most prevalent nematode species and was recovered from 60% of positive samples. Nematode populations varied temporally and spatially along grassy banks; mean population density over the growing period was 1313 infective juveniles/m². Neither macro- nor microarthropod communities nor soil temperature differed between sites at which nematodes were detected and those at which nematodes were not detected. Soil moisture, however, was associated with the occurrence and persistence of nematodes along grassy banks; mean soil moisture at sites at which nematodes were detected and those sites at which nematodes were not detected was 37.3 and 26.8%, respectively. Water management is an important component of vegetable production and our results suggest that soil moisture manipulation would be important in the establishment and sustained presence of entomopathogenic nematode populations within cultivated areas over the growing season.     

Infectivity and reproductive potential of the Oswego strain of Heterorhabditis bacteriophora associated with life stages of the clover root curculio,Sitona hispidulus

The infectivity and reproductive potential of the entomopathogenic nematode Heterorhabditis bacteriophora (Oswego strain), at different concentrations, was studied. Seventy to 80.0% mortality to late instar larvae of the clover root curculio, Sitona hispidulus, and 40.0-76.0% mortality to pupae, was observed at concentrations of 15-100 infective juveniles. There were no significant differences in mortality among nematode concentrations. LC(50) levels of 4.0 and 21.4 nematodes were determined for clover root curculio larvae and pupae, respectively. Nematodes did not cause significant mortality to adult or first instar clover root curculio. H. bacteriophora was able to complete its development and reproduce in 74.0-95.0% of clover root curculio late instar larvae and pupae. Reproductive potential in curculio larvae and pupae ranged from 0 to 7040 infective juveniles per host. Larvae exposed to 100 nematodes had a reproductive potential significantly higher than in those larvae exposed to 15 and 50 nematodes. Reproductive potential in pupae decreased with an increased nematode dose, indicating potential crowding effects. Host larval and pupal mass were positively correlated with nematode progeny production.     

Evaluating mulches together with Heterorhabditis zealandica (Rhabditida: Heterorhabditidae) for the control of diapausing codling moth larvae,Cydia pomonella (L.) (Lepidoptera: Tortricidae)

The potential of using an entomopathogenic nematode, Heterorhabditis zealandica Poinar, together with different test mulches (pine chips, wheat straw, pine wood shavings, blackwood and apple wood chips) to control diapausing codling moth, Cydia pomonella (L.) larvae was evaluated. Mesh cages were identified as a suitable larval-containment method. High levels of codling moth mortality were obtained when using pine wood shavings as mulch (88%) compared to pine chips, wheat straw, blackwood and apple wood chips (41–88%). Humidity (>95% RH) has to be maintained for at least 3 days to ensure nematode survival. It was also proven that nematodes had the ability to move out of infected soil into moist mulch, to infect the codling moth larvae residing at heights of up to 10 cm. Field experiments showed the importance of climatic conditions on nematode performance. Low temperatures (<15°C) recorded during the first trial resulted in low levels of control (48%), as opposed to the 67% mortality recorded during the second trial (temperatures ranged between 20 and 25°C). Low levels of persistence (<10%) were recorded in the mulches post-application. The study conclusively illustrated some of the baseline requirements fundamental to the success of entomopathogenic nematodes together with mulches for the control of codling moth.     

Effect of humidity and a superabsorbent polymer formulation on the efficacy of Heterorhabditis zealandica (Rhabditida: Heterorhabditidae) to control codling moth,Cydia pomonella (L.) (Lepidoptera: Tortricidae)

Adequate moisture levels are required for nematode survival and subsequent efficacy as entomopathogens. Formulation of nematodes aimed at aboveground applications may assist in maintaining such moisture levels. In this study, we report the effects of a superabsorbent polymer formulation, Zeba® on the performance of an entomopathogenic nematode, Heterorhabditis zealandica Poinar, for controlling diapausing codling moth, Cydia pomonella (L.) larvae in cryptic habitats on trees. Water activity (a_w-value) on bark was considered to be an indication of moisture levels on trees in cryptic habitats where codling moth larvae are known to occur, thereby influencing nematode efficacy. H. zealandica was only able to infect codling moth larvae at a_w≥0.92, with a_w50=0.94 and a_w90=0.96. Laboratory experiments in which nematode concentration was investigated indicated a positive linear relationship between the concentration of nematodes applied and the level of control obtained, with the highest level of mortality recorded at 80 IJs/larva, requiring at least 4 h of conditions conducive to nematode activity to ensure infectivity and subsequent efficacy. Further experimentation showed that the use of the Zeba formulation, together with the nematodes, improved the level of control obtained at 60% and 80% RH in the laboratory and that it also enhanced the survival and infection-ability of the nematodes in the field. The study conclusively illustrates that the tested formulation assisted in maintaining adequate moisture levels on the application substratum, as required for nematode survival and subsequent efficacy.     

Effects of Entomopathogenic Nematodes on Control of a Mushroom-infesting Sciarid Fly and on Mushroom Production

The entomopathogenic nematodes Steinernema feltiae (Biosys strain #27) and Heterorhabditis heliothidis were evaluated for the larval control of a mushroom-infesting sciarid, Lycoriella mali, and for the effects of these nematodes on mushroom (Agaricus bisporus) production. In a series of small-scale mushroom crops, infective-stage H. heliothidis and S. feltiae were applied to the mushroom casing surface in the irrigation water or incorporated into the casing material at densities ranging from 28 to 1120 and 11 to 1120 nematodes cm-2 of casing surface respectively. The mortality of L. mali larvae ranged from 52 to 100% for H. heliothidis and 38 to 100% for S. feltiae. Both nematode species reduced mycelial coverage on the casing surface at primordia initiation. Neither mushroom strain (off-white or white hybrid) or method of application (incorporation into or irrigation onto the casing surface) altered the effect on mycelial coverage. The nematodes's negative effect on mycelial growth confounded the benefit of fly control. At high nematode densities (up to 1120 nematodes cm-2), damage-free mushroom yields for the first week of harvest were less than those from the untreated control. However, at lower nematode densities, at or below 140 cm-2, the nematodes had less effect on mushroom growth, and consequently, damage-free mushroom yields for the first week of harvest were frequently greater than those from the untreated control. In the absence of flies, the first-week mushroom yield generally declined with increasing nematode densities for both white and off-white mushroom hybrids. After 4 weeks of harvest, accumulated mushroom yields had nearly recovered from the earlier decline.     

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.     

Effects of Adjuvants on Entomopathogenic Nematode Persistence and Efficacy Against Plutella xylostella

The effects of spray additives on entomopathogenic nematode persistence and efficacy against Plutella xylostella (L . ) were studied . Several adjuvants were toxic to radish seedlings ( Raphanus sativus var . capitata L . ) but none was toxic to the nematodes or P. xylostella. In the laboratory , the adjuvants that provided the best antidesiccant activity based on a rank score were TX7719 , Rodspray oil and Nufilm P . Those providing less protection but better than the remaining adjuvants were 38 - F , dextrose and Pluronic F - 127 . In greenhouse trials , TX7719 and Rodspray oil were more effective than the other adjuvants tested . The stilbene brightener , Blankophor BBH , did not increase nematode efficacy consistently in greenhouse trials probably because the concentration used was too low . In field trials , the combination of TX7719 plus Blankophor BBH increased nematode persistence on watercress leaves ( Nasturium officinale R . Br . ) and efficacy against P. xylostella significantly . In vitro- pro duced nematodes benefited more from additives than in vivo- produced nematodes in the laboratory , but that difference was lost in the field . Overall , it was found that additives generally improved nematode persistence and efficacy , but the improvement was probably not sufficient to increase the feasibility of foliar applications of nematodes against P. xylostella. However , further evaluation of adjuvants is warranted for applications of nematodes to watercress for the control of P. xylostella.     

Selective breeding of Steinernema feltiae (Filipjev) (Nematoda: Steinernematidae) for improved efficacy in control of a mushroom fly,Lycoriella solani Winnertz (Diptera: Sciaridae)

A method of selecting a Steinernema feltiae strain that is effective against a mushroom fly, Lycoriella solani, is described in detail. The pest control efficacy of the selected nematode strain was evaluated and compared with the efficacy of two unselected strains. The selection procedure was designed to give preference to nematode individuals with the greatest ability (1) to search effectively for the target insect larvae in their natural habitat, (2) to infect them shortly after application and (3) to reproduce in their haemocoel. Thirty‐four rounds of selection achieved a 4‐fold improvement in nematode ability to find and parasitize third‐ and fourth‐instar larvae of the pest in the mushroom substrate. In 24‐h laboratory experiments, mortality of the insect caused by nematode juveniles rose from 22.5%, recorded for the original unselected isolate, to 92.5% for the selected strain. In a 51‐day experiment conducted on a mixed age mushroom house population of L. solani, the enhanced pest control ability of the selected strain was detected shortly after nematode application and remained high throughout the experimental period. During the first 4 weeks of the trial the selected nematode strain was significantly better than both unselected strains and caused 91.1–92.7% reduction of the fly emergence from the mushroom substrate. No difference was observed between the efficacy of the selected nematodes applied at 1 × 10⁶ and 3 ×10⁶ infective juveniles per m², while the unselected strains performed significantly better at the higher concentration. All the nematodes examined showed good persistence in the mushroom casing apparently due to recycling in the insect host.     

昆虫病原线虫资源概况和分类技术进展

昆虫病原线虫是具有重要潜在应用价值的害虫生物防治资源,主要包括斯氏线虫科（Steinernematidae）的斯氏线虫属Steinernema与新斯氏线虫属Neosteinernema线虫和异小杆线虫科（Heterorhabditidae）的异小杆线虫属Heterorhabditis线虫。近10年来,分子生物学方法与传统的形态学方法相结合应用到线虫的鉴定与分类,昆虫病原线虫的分类进入稳定与发展时期,越来越多的新种或品系被发现及应用于生物防治。目前已描述的昆虫病原线虫种类达65种,其中斯氏线虫属52种,新斯氏线虫属1种,异小杆线虫属12种。本文整理列出了迄今报道的昆虫病原线虫种类及其来源,并综述了昆虫病原线虫分类现状以及鉴定与分类方法上的研究进展,重点阐述了分子生物学技术在昆虫病原线虫鉴定与分类的应用状况。     

Evaluation of Entomopathogenic Nematodes for the Control of Mediterranean Fruit Fly (Diptera: Tephritidae)

The virulence of various entomopathogenic nematode (EPN) strains was evaluated against the Mediterranean fruit fly, C. capitata . The selected nematodes were assessed for their infectivity for the final larval stage of the insect host and under varying environmental conditions. Among 12 EPN strains tested, Steinernema riobrave Texas ( Sr TX) and Heterorhabditis sp. IS-5 (H IS-5), showed high activity and induced >80% mortality. Six EPN strains showed limited activity (>30% mortality), and four strains had no effect (<20% mortality). Sr TX was more effective than H IS-5. Mature C. capitata larvae were most susceptible to nematode infection during the first 4h after they began to emerge from their diet to pupate. Activity of the two nematode strains at a constant inoculation rate was dependent on insect larval density. The highest activity was recorded at 1.88 larvae cm -2 and decreased at higher larval densities. EPN activity was also directly related to nematode density. Maximal activity was shown at a density of 150 infective juveniles cm -2 . A similar activity pattern was also recorded with Sr TX in four different soil types. The persistence of this EPN in the soil extended over 5 days but there was no activity after 14 days. Except for a lower activity under cool conditions (17°C), temperatures ranging between 22 and 41°C, or moisture levels in the treated soil ranging between 3 and 20%, had no significant effect on nematode activity. Our results suggest that application of Sr TX against C. capitata may have potential for controlling C. capitata .     

Insect protein digestion improves purity of Steinernema carpocapsae in vitro culture and reduces culture period

Insect protein, used for in vitro culture media for entomopathogenic nematode, produces nematodes of high quality. However, the time-consuming culture and poor purity of nematodes hinder the commercial application of insect protein media. We show that hydrolyzed insect protein improves nematode purity in in vitro culture. The results revealed that nematode purity was increased by more than 90 %, and the culture period was reduced by 6 days. Estimated economic efficiency of using hydrolyzed insect protein medium was increased by 44.25 % over that obtained with non-hydrolyzed insect medium.     

Effectiveness of the entomopathogenic nematode <Emphasis Type="Italic">Steinernema feltiae</Emphasis> in agar gel formulations against larvae of the Colarado potato beetle, <Emphasis Type="Italic">Leptinotarsa decemlineata</Emphasis> (Say.) (Coleoptera: Chrysomelidae)

The entomopathogenic nematode Steinernema feltiae strain Ustinov Russia was used on potato foliage to control larvae of the Colorado potato beetle, Leptinotarsa decemlineata (Say.) (Coleoptera: Chrysomelidae). Nematodes were applied in formulations of agar at 4%, 2%, 1% and 0.5% concentrations and compared to a control application of nematodes in water for nematode survival. Agar formulation significantly improved efficacy by increasing nematode survival through reduction in desiccation when compared to water-based formulation. More than 70% of infective juvenile nematodes (IJs) died after being incubated in the highest concentration of agar for 12 h, while only 8% mortality was recorded at the 1% concentration. Suspension of nematodes in 1% agar gel was shown to be efficacious in both laboratory and greenhouse tests for extension of the nematodes’ survival. Agar formulation droplets dried slower than control droplets by 127.8 min. Leptinotarsa decemlineata mortality significantly increased when insects were exposed to infective juvenile nematodes for four hours after application. In conclusion, the agar formulation enhanced nematode survival by providing a suitable environment thereby delaying dryingand increasing the possibility for nematodes to invade their host on the foliage.