Left: Juvenile nematodes emerging from a beet armyworm pupa. Note the large adult female (center left). A.Hara Center: An infective juvenile nematode. P.Timper Right: Steinernema carpocapsae infective juvenile nematodes are "ambushers" that may assume an upright or nictation position when seeking insect hosts. J.Ogrodnick

Nematodes
(Rhabditida: Steinernematidae & Heterorhabditidae)

Nematodes are simple roundworms. Colorless, unsegmented, and lacking appendages, nematodes may be free-living, predaceous, or parasitic. Many of the parasitic species cause important diseases of plants, animals, and humans. Other species are beneficial in attacking insect pests, mostly sterilizing or otherwise debilitating their hosts. A very few cause insect death but these species tend to be difficult (e.g., tetradomatids) or expensive (e.g. mermithids) to mass produce, have narrow host specificity against pests of minor economic importance, possess modest virulence (e.g., sphaeruliids) or are otherwise poorly suited to exploit for pest control purposes. The only insect-parasitic nematodes possessing an optimal balance of biological control attributes are entomopathogenic or insecticidal nematodes in the genera Steinernema and Heterorhabditis. These multi-cellular metazoans occupy a biocontrol middle ground between microbial pathogens and predators/parasitoids, and are invariably lumped with pathogens, presumably because of their symbiotic relationship with bacteria. 

Entomopathogenic nematodes are extraordinarily lethal to many important soil insect pests, yet are safe for plants and animals. This high degree of safety means that unlike chemicals, or even Bacillus thuringiensis, nematode applications do not require masks or other safety equipment; and re-entry time, residues, groundwater contamination, chemical trespass, and pollinators are not issues. Most biologicals require days or weeks to kill, yet nematodes, working with their symbiotic bacteria, kill insects in 24-48 hr. Dozens of different insect pests are susceptible to infection, yet no adverse effects have been shown against nontargets in field studies (Georgis et al., 1991). Nematode production is easily accomplished for some species using standard fermentation in tanks up to 150,000 liters. Nematodes do not require specialized application equipment as they are compatible with standard agrochemical equipment including pressurized, mist, electrostatic, fan, and aerial sprayers. Application through irrigation systems has improved grower acceptance. Insecticidal nematodes are virtually without competition from other biological agents for control of soil-inhabiting and plant-boring insects. 

Hundreds of researchers representing more than forty countries are working to develop nematodes as biological insecticides. Nematodes are sold in the U.S., Europe, Japan, and China for control of insect pests in high-value horticulture, agriculture, home and garden niche markets.

Life Cycle

Diagram courtesy of H. Kaya

Steinernematids and heterorhabditids have similar life histories. The non-feeding developmentally arrested infective juvenile seeks out insect hosts and initiates infections. When a host has been located, the nematodes penetrate into the insect body cavity, usually via natural body openings (mouth, anus, spiracles) or areas of thin cuticle. Once in the body cavity, a symbiotic bacterium (Xenorhabdus for steinernematids, Photorhabdus for heterorhabditids) is released from the nematode gut, which multiplies rapidly and causes rapid insect death. The nematodes feed upon the bacteria and liquefying host, and mature into adults. Steinernematid infective juveniles may become males or females, where as heterorhabditids develop into self-fertilizing hermaphrodites although subsequent generations within a host produce males and females as well. The life cycle is completed in a few days, and hundreds of thousands of new infective juveniles emerge in search of fresh hosts. 

Thus, entomopathogenic nematodes are a nematode-bacterium complex. The nematode may appear as little more than a biological syringe for its bacterial partner, yet the relationship between these organisms of one of classic mutualism. Nematode growth and reproduction depend upon conditions established in the host cadaver by the bacterium. The bacterium further contributes anti-immune proteins to assist the nematode in overcoming host defenses, and anti-microbials that suppress colonization of the cadaver by competing secondary invaders. Conversely, the bacterium lacks invasive powers and is dependent upon the nematode to locate and penetrate suitable hosts. 

Relative Effectiveness

Growers will not adopt biological agents that do not provide efficacy comparable with standard chemical insecticides. Technological advances in nematode production, formulation, quality control, application timing and delivery, and particularly in selecting optimal target habitats and target pests, have narrowed the efficacy gap between chemical and nematode agents. Nematodes have consequently demonstrated efficacy in an number of agricultural and horticultural market segments. 

Entomopathogenic nematodes are remarkably versatile in being useful against many soil and cryptic insect pests in diverse cropping systems, yet are clearly underutilized. Like other biological control agents, nematodes are constrained by being living organisms that require specific conditions to be effective. Thus, desiccation or ultraviolet light rapidly inactivates insecticidal nematodes; chemical insecticides are less constrained. Similarly, nematodes are effective within a narrower temperature range than chemicals, and are more impacted by suboptimal soil type, thatch depth, and irrigation frequency (Georgis and Gaugler, 1991). Nematode-based insecticides may be inactivated if stored in hot vehicles, cannot be left in spray tanks for long periods, and are incompatible with several agricultural chemicals. Certain species cannot be applied with high-pressure application equipment; unused nematodes cannot be applied the following year; different species require different screen sizes. Chemicals also have problems (e.g., mammalian toxicity, resistance, groundwater pollution, etc.) but a large knowledge base has been developed to support their use. Accelerated implementation of nematodes into IPM systems will require users to be more knowledgeable about how to use them effectively. 

Appearance

Nematodes are formulated and applied as infective juveniles, the only free-living and therefore environmentally tolerant stage. Infective juveniles range from 0.4 to 1.1 mm in length and can be observed with a hand lens or microscope after separation from formulation materials. Disturbed nematodes move actively, however sedentary ambusher species (e.g. Steinernema carpocapsae, S. scapterisci) in water soon revert to a characteristic "J"-shaped resting position. Low temperature or oxygen levels will inhibit movement of even active cruiser species (e.g., S. glaseri, Heterorhabditis bacteriophora). In short, lack of movement is not always a sign of mortality; nematodes may have to be stimulated (e.g., probes, acetic acid, gentle heat) to move before assessing viability. Good quality nematodes tend to possess high lipid levels that provide a dense appearance, whereas nearly transparent nematodes are often active but possess low powers of infection. 

Insects killed by most steinernematid nematodes become brownish-yellow, whereas insects killed by heterorhabditids become red and the tissue assumes a gummy consistency. A dim luminescence given off by insects freshly killed by heterorhabditids is a foolproof diagnostic for this genus (the symbiotic bacteria provide the luminescence). Black cadavers with associated putrefaction indicate that the host was not killed by entomopathogenic species. Nematodes found within such cadavers tend to be free-living soil saprophages. 

Habitat

Steinernematid and heterorhabditid nematodes are exclusively soil organisms. They are ubiquitous, having been isolated from every inhabited continent from a wide range of ecologically diverse soil habitats including cultivated fields, forests, grasslands, deserts, and even ocean beaches. In New Jersey, entomopathogenic nematodes were recovered from 22% of sites sampled (Gaugler, et al., 1992). 

Pests Attacked

Because the symbiotic bacterium kills insects so quickly, there is no intimate host-parasite relationship as is characteristic for other insect-parasitic nematodes. Consequently, entomopathogenic nematodes are lethal to an extraordinarily broad range of insect pests in the laboratory. Field host range is considerably more restricted, with some species being quite narrow in host specificity. When considered as a group of nearly 30 species, however, entomopathogenic nematodes are useful against a large number of insect pests, many of which are listed in the table below. As field research progresses and improved insect-nematode matches are made, this list is certain to expand. Regrettably, nematodes have yet to be found which are effective against several of the most important soil insects, including wireworms, grape phylloxera, fire ants, or corn rootworms.

Characteristics of Key Species

Steinernema carpocapsae: The most studied, available, and versatile of all entomopathogenic nematodes. Important attributes include ease of mass production and ability to formulate in a partially desiccated state that provides several months of room-temperature shelf-life. Particularly effective against lepidopterous larvae, including various webworms, cutworms, armyworms, girdlers, and wood-borers. This species is a classic sit-and-wait or "ambush" forager, standing on its tail in an upright position near the soil surface and attaching to passing hosts. Consequently, S. carpocapsae tends to be most effective when applied against highly mobile surface-adapted insects. Highly responsive to carbon dioxide once a host has been contacted, the spiracles are a key portal of host entry. It is most effective at temperatures ranging from 22 to 28°C. 

Steinernema feltiae: Attacks primarily immature dipterous insects, including mushroom flies, fungus gnats, and tipulids. This nematode is unique in maintaining infectivity at soil temperatures below 10°C. S. feltiae offers lower stability than other steinernematids. 

Steinernema glaseri: The largest entomopathogenic nematode at twice the length but eight times the volume of S. carpocapsae infective juveniles. Attacks coleopterous larvae, particularly scarabs. This species is a cruise forager, neither nictating nor attaching well to passing hosts, but highly mobile and responsive to long-range host volatiles. Thus, this nematode is best adapted to parasitize hosts possessing low mobility and residing within the soil profile. Field trials, particularly in Japan, have shown that S. glaseri can provide control of several scarab species. Large size, however, reduces yield, making this species significantly more expensive to produce than other species. A tendency to occasionally "lose" its bacterial symbiote is bothersome. Moreover, the highly active and robust infective juveniles are difficult to contain within formulations that rely on partial nematode dehydration. In short, additional technological advances are needed before this nematode is likely to see substantial use. 

Steinernema kushidai: Only isolated so far from Japan and only known to parasitize scarab larvae. Although still in development as of this writing, early field trials against scarab larvae in Japan and the U.S. have been highly promising. Good laboratory stability and field persistence have been demonstrated. This small nematode is likely be see rapid implementation if quirky problems encountered in mass rearing can be resolved. 

Steinernema riobravis: This novel and highly pathogenic species, isolated to date only from the Rio Grande Valley of Texas, possesses several novel features. Its effective host range runs across multiple insect orders. This versatility is likely due in part to its ability to exploit aspects of both ambusher and cruiser means of finding hosts. Trials have demonstrated its effectiveness against corn earworm and mole crickets. In Florida, 60,000 acres of citrus are treated annually for control of citrus root weevil with impressive results. This is a high temperature nematode, effective at killing insects at soil temperatures above 35°C. Persistence is excellent even under semi-arid conditions, a feature no doubt enhanced by the uniquely high lipid levels found in infective juveniles. Its small size provides high yields whether using in vivo (up to 375,000 infective juveniles per wax moth larvae) or in vitro methods. Only formulation improvements that impart increased stability are needed for this parasite to achieve its full potential. 

It must also be noted that S. riobravis is currently being marketed for suppression of plant parasitic nematodes infesting turfgrass. There is substantial correlative data suggesting that some entomopathogenic nematodes can suppress plant species. Some skepticism may be healthy until this puzzling assertion can be fully confirmed by rigorously designed, multiple field experiments 

Steinernema scapterisci: The only entomopathogenic nematode to be used in a classical biological control program, S. scapterisci was isolated from Uruguay and first released in Florida in 1985 to suppress an introduced pest, mole crickets. The nematode become established and presently contributes to control. Steinernema scapterisci is highly specific to adult mole crickets. Its ambusher approach to finding insects is ideally suited to the turfgrass tunneling habits of its host. Commercially available since 1993, this nematode is also sold as a biological insecticide, where its excellent ability to persist and provide long-term control contributes to overall efficacy. Availability is severely restricted due to the small market niche this nematode occupies. This is aggravated by the specificity of this nematode for a host that is very difficult to rear, precluding in vivo production. 

Heterorhabditis bacteriophora: Among the most important entomopathogenic nematodes, H. bacteriophora possesses considerable versatility, attacking lepidopterous and coleopterous insect larvae among other insects. This cruiser species appears most useful against root weevils, particularly black vine weevil where it has provided consistently excellent results in containerized soil. A warm temperature nematode, H. bacteriophora shows reduced efficacy when soil drops below 20°C. Characteristic poor stability has limited the usefulness of this interesting nematode: shelf-life is problematic and most infective juveniles persist only a few days following field release. 

Heterorhabditis megidis: First isolated in Ohio, this nematode is marketed in western Europe for control of black vine weevil and various other soil insects. Its large size, characteristic heterorhabditid instability, and dearth of field efficacy data limit its utility at present. 

Conservation

Conservation strategies are poorly developed and largely limited to avoiding applications onto sites where the nematodes are ill-adapted; for example, where immediate mortality is likely (e.g., exposed foliage) or where they are completely ineffective (e.g., aquatic habitats) (Lewis et al., 1997). Minimizing deleterious effects of the aboveground environment with a post-application rinse that washes infective juveniles into the soil is also a useful approach to increasing persistence and efficacy. 

Native populations are highly prevalent, but other than scattered reports of epizootics their impact on hosts populations is not well documented. This is largely attributable to the cryptic nature of soil insects. Consequently, guidelines for conserving native entomopathogenic nematodes have not been advanced. 

Pesticide Susceptibility

Infective juveniles are compatible with most but not all agricultural chemicals under field conditions. Heterorhabditis bacteriophora is the most sensitive among entomopathogenic nematodes to physical stress, yet even this species was tolerant to three days of laboratory immersion in 58 of 75 fungicides, herbicides, insecticides, and nematicides assayed (Rovesti et al., 1988). Moreover, many of the chemicals noted to be toxic had a transient effect only, as the nematodes recovered quickly when the exposure ended. Thus, tank mixes and simultaneous application of nematodes with most pesticides and fertilizers should be practical when exposures are short. Chemicals to be used with care or avoided include the insecticides bendiocarb, chlorpyrifos, ethoprop, and isazophos, the fungicides anilazine, dimethyl benzyl ammonium chloride, fenarimol, and mercurous chloride, the herbicides 2,4-D and trichlopyr, and the nematicide fenamiphos. 

Commercial Availability

Of the nearly thirty steinernematid and heterorhabditid nematodes identified to date, seven species are commercially available. The list of nematode suppliers provided below emphasizes U.S. suppliers and has been adapted from Dunn's (1997) Beneficial Nematodes, which was based on Hunter's (1994) Suppliers of Beneficial Organisms in North America. Comparison-shopping is recommended as prices vary greatly among suppliers. Caution is further advised with regard to application rates. One billion nematodes per acre (250,000 per m²) is the rule-of-thumb against most soil insects (containerized and greenhouse soils tend to be treated at higher rates). Broadcast application rates suggested by some suppliers of 100 million per acre or less are not credible and not supported by controlled studies. A final caveat is that, just as one must select the appropriate insecticide to control a target insect, so must one choose the appropriate nematode species or strain. Ask suppliers about field tests supporting their recommended matching of insect target and nematode.

References

Dunn, R. A. and G. C. Smart, Jr. 1977. Suppliers of beneficial nematodes. Nematology Pointer No. 45 (SS-ENY-27), Fl. Coop. Ext. Serv., IFAS, Univ. Florida. 

Georgis, R. and R. Gaugler. 1991. Predictability in biological control using entomopathogenic nematodes. J. Econ. Entomol. [Forum] 84:713-20. 

Gaugler, R., J. Campbell, M. Selvan & E. Lewis. 1992. Large-scale inoculative releases of the entomopathogen Steinernema glaseri: assessment 50 years later. Biol. Control 2:181-7. 

Georgis, R., H. Kaya and R. Gaugler. 1991. Effect of steinernematid and heterorhabditid nematodes on nontarget arthropods. Environ. Entomol. 20:815-22. 

Hunter, C. D. 1994. Suppliers of Beneficial Organisms in North America. California Environmental Protection Agency, Dept. Pesticide Regulation, Environmental Monitoring and Pest Management Branch. PM 94-03. 30 pp. 

Lewis, E., J. Campbell and R. Gaugler. 1997. A conservation approach to using entomopathogenic nematodes in turf and landscapes. In "Perspectives on the Conservation of Natural Enemies of Pest Species" (P. Barbosa, Ed.), Academic Press (in press). 

Rovesti, L., E.W. Heinzpeter, F. Tagliente and K.V. Deseo. 1988. Compatibility of pesticides with the entomopathogenic nematode Heterorhabditis bacteriophora. Nematologica 34:462-476. 

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