entomopathogenic nematodes - a review - aws

AGRICULTURAL RESEARCH COMMUNICATION CENTRE

www.arccjournals.com / indianjournals.comAgri. Reviews, 34 (3) : 163-175, 2013DOI- 10.5958/j.0976-0741.34.3.001

ENTOMOPATHOGENIC NEMATODES - A REVIEW

Sumit Vashisth* , Y.S. Chandel and P. K. Sharma

Department of Entomology,Himachal Pradesh Krishi Vishvavidyalaya, Palampur, 176 062, India

Received: 20-02-2012 Accepted: 19-10-2012

ABSTRACTThe entomopathogenic nematodes possessing balanced biological control attributes belong to

genera Steinernema and Heterorhabditis and are having mutualistic association with bacteria of thegenus Xenorhabdus for Steinernematidae and Photorhabdus for H eterorhabditidae.Entomopathogenic nematodes being highly lethal to many important insect-pests, are safe to non-target organisms and working with their symbiotic bacteria kill the insects within 24-28 hours ascompared to days and weeks required for insect killing in other biological control agents. Theirinfective juveniles (IJs) have been reported to tolerate short-term exposure to many chemical andbiological insecticides, fungicides, herbicides, fertilizers and growth regulators, hence providing anopportunity of tank-mixing and application together. Entomopathogenic nematodes are also reportedto be compatible with a number of agrochemicals and their use can offer a cost-effective alternativeto pest control. Only twelve species out of nearly eighty identified Steinernematids and Heterorhabditisnematode species have been commercialized. The EPNs have the great potential to be used inintegrated pest management systems and work done have been reviewed in this article to facilitatethe students and researchers to have an overview of the work done and proceed further to undertakethe advanced research in different aspects related to entomopathogenic nematodes.

Keywords: Entomopathogenic nematodes (EPN), Heterorhabditis, Photorhabdus,Steinernema, Xenorhabdus.

*Corresponding author’s e-mail: [email protected]

Nematodes are simple roundworms,colorless, unsegmented, and lacking appendages,may be free-living, predaceous, or parasitic. Manyof the parasitic species cause important diseases ofhumans plants and animals. The term ‘Entomophilicnematodes’ includes all relationships of insects andnematodes ranging from phoresis to parasitism andpathogenesis. ‘Entomogenous nematodes’ are thosethat have a facultative or obligate parasiticassociations with insects. Entomogenous nematodeshave several deleterious effects on their hostsincluding sterility, reduced fecundity, longevity andflight activity, delayed development, or otherbehavioral, physiological and morphologicalaberrations and in some cases, rapid mortality.Parasitic associations with insects have beendescribed from 23 nematode families. Seven of thesefamilies contain species that have potential forbiological control of insects (Koppenhofer and Kaya,2001). A very few cause insect death but these

species are difficult (e.g., tetradomatids) or expensive(e.g. mermithids) to mass produce, have narrow hostspecificity against pests of minor economicimportance, possess modest virulence (e.g.,sphaeruliids) or are otherwise poorly suited toexploit for pest control purposes. The only insect-parasi tic nematodes possessing an optimalbalance of biological control attributes areentomopathogenic or insecticidal nematodes in thegenera Steinernema and Heterorhabditis because oftheir ability to kill hosts quickly (1-4 d depending onnematode and host species). Mutualistic associationof these nematodes with bacteria of the genusXenorhabdus for Steinernematidae andPhotorhabdus for Heterorhabditidae have also beenreported by Boemare et al. (1993). However,although both the families belong to the same order,but are not closely related and have distinctlydifferent reproductive strategies (Blaxter et al. 1998).

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Advantages:Entomopathogenic nematodes areextraordinarily lethal to many important insect pests,yet are safe for non target organisms (Georgis et al.,1991). This high degree of safety means that unlikechemicals, or even Bacillus thuringiensis, nematodeapplications do not require masks or other safetyequipment; and re-entry time, residues, groundwatercontamination, chemical trespass, and pollinatorsare not issues. Most biologicals require days or weeksto kill, yet nematodes, working with their symbioticbacteria, can kill insects within 24-48 hours. Dozensof different insect pests are susceptible to infection,yet no adverse effects have been shown againstbeneficial insects or other nontargets in field studies(Georgis et al., 1991; Akhurst and Smith, 2002).Nematodes are amenable to mass production anddo not require specialized application equipment asthey are compatible with standard agrochemicalequipments, including various sprayers (e.g.,backpack, pressurized, mist, electrostatic, fan, andaerial) and irrigation systems.

Life cycle: The entomopathogenic nematodes underSteinernematids and heterorhabditids have similarli fe histories. The free-l iving, non-feedingdevelopmentally arrested infective juveniles of thesenematode species have attributes of both insectparasitoids or predators, they have chemoreceptorsand are motile, like pathogens, highly virulent, killingtheir hosts quickly, and can be cultured easily andhave a high reproductive potential. When a host hasbeen located, the nematodes penetrate into the insectbody cavity, usually via natural body openings(mouth, anus, spiracles) or areas of thin cuticle.Wang and Gaugler (1999) compared the penetrationbehavior of S. glaseri and H. bacteriophora intoPopillia japonica larvae and found that S. glaseripenetrated primarily through the gut. H.bacteriophora was not efficient at penetrating thegut, presumably because of the thick peritrophicmembrane, but penetration through theintersegmental membranes of the cuticle. Once inthe body cavity, a symbiotic bacterium (Xenorhabdusfor steinernematids, Photorhabdus forheterorhabditids), that are motile, Gram-negative,faculat ively anaerobic rods in the fami lyEnterobacteriaceae released from the nematode gut,which multiplies rapidly and causes rapid insectdeath due to septicemia. Presently, eight species arerecognized but most isolates from recently described

entomopathogenic nematodes species have yet tobe determined (Hazir et al. 2003). The nematodesfeed upon the bacteria and liquefying host, andmature into adults. Steinernematid infective juvenilesmay become males or females, where asheterorhabditids develop into self-ferti lizinghermaphrodites although subsequent generationswithin a host produce males and females as well(Fig. 1). The life cycle is completed in a few days,and hundreds and thousands of new infectivejuveniles emerge in search of fresh hosts.

Mode of action:Entomopathogenic nematodes area nematode-bacterium complex. In this association,the nematode is dependent upon the bacterium forquickly killing its insect hosts, creating a suitableenvironment for its development by producingantibiotics that suppress competing secondarymicroorganisms and transforming the host tissuesinto a food source. The bacterium requires nematodefor protection from the external environment,penetration into the hosts haemocoel and inhibitionof the host’s antibacterial proteins.

Edaphic factors: Soil texture has an effect oninfective juvenile survival with survival being lowestin clay soils. The poor survival rate in clay soils isprobably due to lower oxygen levels in the smallersoil pores. Oxygen may also become a limiting factorin water-saturated soils and soils with contents oforganic matter. Soil pH does not have a strong effecton infective juvenile survival. Thus, infective juvenilesurvival at pH values between 4 and 8 does not vary,but survival declines at pH 10. Similarly, soil salinityhas only l imited negative effects onentomopathogenic nematode survival even atsalinity above the tolerance levels of most crop plants(Thurston et al., 1994). Sea water has no negativeeffects on survival of several Heterorhabditis species/strains (Griffin et al., 1994) and they have beenfrequently isolated from soils near the sea shore.However, high salinity (seawater) reduced the abilityto Heterorhabdistis to infect hosts but did improvetheir tolerance to high temperatures (Finnegan et al.,1999).

Production and storage technologyEntomopathogenic nematodes are mass

produced as biopesticides using in vivo or in vitromethods (Shapiro-Ilan and Gaugler 2002).

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In vivo production: Production methods forculturing entomopathogenic nematodes in insecthosts have been reported by various authors (Dutkyet al., 1964, Poinar 1979 and Kaya and Stock,1997). These references essentially describe systemsbased on the White trap (White, 1927), which takesadvantage of infective juveniles (IJ) naturalmigration away from the host cadaver uponemergence. Yield is affected by choice of nematodeand host species. Among nematode species, yield isgenerally proportional to host size (Blinova andIvanova, 1987 and Flanders et al., 1996) but yieldper milligram insect (within host species) andsusceptibility to infection is usually inverselyproportional to host size or age (Dutky et al., 1964and Blinova and Ivanova, 1987). In vivo productionrequires the least capital outlay and technicalexpertise (Friedman, 1990; Gaugler and Han, 2002).The most common insect host used for in vivolaboratory and commercial entomopathogenicnematode production is the last instar of the greaterwax moth, Galleria mellonella, because of its highsusceptibility to most nematodes, ease of rearing,wide availability and ability to produce high yields(Woodring and Kaya, 1988).

In vitro production: Production of entomo-pathogenic nematodes through in vitro requires athorough knowledge of biology and behavior of thenematode species produced. The only stage that canbe commercially used is the dauer juvenile (DJ), amorphologically distinct juvenile, formed as aresponse of depleting food sources and adverseenvironmental conditions. Mass production ofentomopathogenic nematodes has evolved from thefirst large scale in vitro soild media production byGlaser et al. (1940) to the three dimensional solidmedia in vitro process (Bedding, 1981 and 1984)and to the in vitro liquid fermentation productionmethod (Friedman, 1990). Currently, commercialnematodes are produced monoxenically using thesolid-media process developed by Bedding (1981and 1984) or the liquid-fermentation method. Thesolid media process has successfully producedpathogenic steinernematids and heterorhabditids,but the high labour cost is the main constraint(Bedding, 1990). The liquid-fermentation process,is highly efficient for several steinernematid speciesbut not for heterorhabditids (Gaugler and Georgis,1991).

Formulation of nematodes into a stableproduct has played a signi ficant role incommercialization of these biological-control agents.Active nematodes must be immobilized to preventdepletion of their lipid and glycogen reserves. Avariety of formulations have been developed tofacilitate nematode storage and application includingactivated charcoal, alginate and polyacrylamide gels,baits, clay, paste, peat, polyurethane sponge,vermiculite, and water-dispersible granules.Depending on the formulation and nematodespecies, successful storage under refrigeration rangesfrom one to seven months. Low temperature (2-50C)generally reduce metabolic activity and can thereforeenhance their shelf-life, some warm adapted speciessuch as H. indica and S. riobrave do not store wellattemperature below 100C (Strauch et al., 2000 andGrewal 2002)

Relative effectiveness and applicationparameters: Entomopathogenic nematodes arecertainly more specific and have no threat to theenvironment than chemical insecticides (Ehlers andPeters, 1995). In some instances, entomopathogenicnematodes have proven to be safe and effectivealternatives to chemical pesticides, but in numerousother cases they have failed to compete successfully(Georgis et al., 2006). Technological advances innematode production, formulation, quality control,application timing and delivery, and particularly inselecting optimal target habitats and target pests,have narrowed the efficacy gap between chemicaland nematode agents. S. feltiae is an efficacious andeconomical replacement for chemical insecticidesin the floriculture industry in the Netherlands,England and Germany (Jagdale et al., 2004)Entomopathogenic nematodes are remarkablyversatile in being useful against many soil and crypticinsect pests in diverse cropping systems, yet areclearly underutilized. Like other biological controlagents, nematodes are constrained by being livingorganisms that require specific conditions to beeffective. Desiccation and temperature extremes arethe most important abiotic factors affecting survivalof entomopathogenic nematodes (Glazer, 2002) andin soilinfective juveniles are subject to attack by avariety of microbial and invertebrate antagonists(Kaya, 2002). Thus, desiccation or ultraviolet lightrapidly inactivates nematodes in comparison tochemical insecticides. Similarly, nematodes are


TABLE 1: List of insect pests targeted with entomopathogenic nematodes.

Order Famil y

Scientifi c and/or common name

Commodity

Country Nematode sp.a

References

Lepidoptera Carposinidae

Carposina niponensis Walsingham

Apple China Sc Wang (1993), Yang et al. (2000)

Gelechiidae Phthorimaea operculella (Zeller)

Potato India Sb, Hi Hussaini (2003)

Pyralidae Chilo suppressalis (Walker) Rice Korea Sc, Sg, Hb Choo et al. (1991), Choo et al. (1995)

C. zonellus Swinhoe Maize India Sc Mathur et al. (1966), Rao et al. (1971)

Maize India Sb, Hi Hussaini (2003) Tryporyza i ncertulas (Walker) Rice India Sc Rao et al. (1971) Noctuidae Spodoptera littoralis Cabbage Egypt Hb, H i, Sc,

Sa, Sr Salem et al. (2007)

Spo doptera litura (F.) Fo liar crops

India H i, Sg Saravanapriya and Subramanian (2007)

Glasshouse and

nursery crops

India H i Divya et al. (2010)

Crucifers Korea Sc, Hb, Sm, Sl, Sg

Park et al. (2001)

Tobacco India Sf Narayanan and Gopalakrishnan (1987)

Tobacco India Sb, Hi Hussaini (2003) India Sf Narayanan and

Gopalakrishnan (1987) Spodoptera furgiperda Corn USA S sp. Raulston et al. (1992) Helicoverpa armigera Fo liar

crops India H i, Sg Saravanapriya and

Subramanian (2007) Glasshous

e and nursery crops

India H i Divya et al. (2010)

India Sg, Sc, St, Sm, Ss

Ali et al. (2008)

India H i Prabhuraj et al. (2006) Helicoverpa zea Corn USA Sr Cabanillas and Raulston

(1995) USA S sp. Raulston et al. (1992) Cnaphalocrosis medinalis Fo liar

crops India H i, Sg Saravanapriya and

Subramanian (2007) Pieris rapae Cabbage Egypt Hb, H i, Sc,

Sa, Sr Salem et al. (2007)

Pieris brassicae L innaeus Crucifers India H i, Sc, St Lalramliana and Yadav (2010)

Agrotis ipsilon (Hufnagel) Potato India Sf Singh (1993) India Sb, Sc, H i Hussaini et al. (2000) India Sr Mathasoliya et al. (2004) A. segtum (Denis et

SchiVermueller) Potato India Sf Singh (1993)

India Sb, Sc, H i Hussaini et al. (2000) Turfgrass Korea Sc, Sg, Hb,

Sl,. Lee et al. (1997)

Sm, H sp. Kang et al. (2004) Palpi ta indi ca (Saunder) Vegetables Korea Sc, Sg, Sl,

Hb Kim et al. (2001b)

a Sb, Steinernema bicornutum; Sc, S. carpocapsae; Sf, S. feltiae; Sg, S. glaseri; Sl, S. longicaudum; Sm, S. monticolum; St, S.thermophilum; S sp,. Steinernema species; H sp., Heterorhabditis species; Hb, H. bacteiophora; Hi, H. indica; Hm, H. marelatus

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effective within a narrower temperature range(generally between 20 °C and 30 °C) than chemicals,and are more impacted by suboptimal soil type,thatch depth, and irrigation frequency (Georgis andGaugler, 1991; Shapiro-Ilan et al., 2006).Maintenance of high viability and virulence duringproduction, formulation and storage forms thebackbone of an effective quality control strategy.Effect of ageing of infective juveniles onpathogenicity was evaluated and compared byHussaini et al., 2005 with freshly hatched infectivejuveniles of the genera Steinernema andHeterorhabditis. S. tami and S. carpocapsaesurvived up to 40 days without mortality and loss ofinfectivity and S. abbasi with 2% mortality.Heterorhabditis spp. survived up to 20 days withoutmortality and declined to 60% and 20% after 40and 60 days of storage, respectively.

Therefore, based on the nematodes’ biology,applications should be made in a manner thatavoids direct sunlight, e.g., early morning or eveningapplications are often preferable. Soil in the treatedarea should be kept moist for at least two weeksafter applications (Klein, 1993). Application toaboveground target areas is difficult due to thenematode’s sensitivity to desiccation and UVradiation; however, improved formulations haveenhanced the efficacy of nematodes against aboveground target pests (e.g., Shapiro-Ilan et al., 2010).The most commonly used application method forentomopathogenic nematodes is spraying directlyon to the soil (or other) surface. Nematodes can beapplied with most commercially available sprayequipment including hand or ground sprayers, mistblowers etc. (Georgis et al., 1995). The infectivejuveniles can withstand pressure up to 1068 kPa andpass through all common nozzle type sprayers withopenings of about 100 µm (screen in nozzles shouldbe removed) in diameter. Nematodes can also bedelivered via irrigation systems including microjet,drip, sprinkler, etc. (Georgis et al., 1995 andCabanillas and Raulston, 1996). In all cases, thenematodes must be applied at a rate that is sufficientto kill the target pest; generally, 25 infective juvenilesper cm2 of treated area is required (rate can beincreased or decreased according to target pest andconditions) (Shapiro-Ilan et al., 2002). Selectivityof nematode species should be according to targetpest.

Appearance: Nematodes are formulated andapplied as infective juveni les as they aremorphologically, physiologically and behaviorallyadapted to act as the active ingradient of a biologicalpesticide. Infective juveniles range from 0.4 to 1.5mm in length. The location of the infective juvenilewithin the soil profile is one of the most importantfactor (Lewis, 2002). Formulation in water-dispersable granules is very successful with theambush forager S. carpocapsae, while the cruiseforaging S. feltiae and S. riobrave rapidly migrateout of the granules (Grewal, 2002). Disturbednematodes move actively, however sedentaryambusher species (e.g. S. carpocapsae, S.scapterisci) in water soon revert to a characteristic“J”-shaped resting position. Ambushing nematodespecies are usually associated with highly mobilesurface dwelling hosts. Entomopathogenicnematodes disperse horizontally and vertically afterapplication. S. carpocapsae infective juveniles moveupwards in soil column (Georgis and Poinar, 1983;Schroeder and Beavers, 1987) where as S. glaseriand H. bacteriophora move downwards, but theyalso disperse throughout the soil column. Lowtemperature or low oxygen levels will inhibitmovement of even active cruiser species (e.g., S.glaseri , H. bacteriophora). In short, lack ofmovement is not always a sign of mortality;nematodes may have to be stimulated (e.g., probes,acetic acid, gentle heat) to move before assessingviability. The infective juveniles do not feed, butrelies on stored energy reserves. Lipids (especiallytriglycerides) constitute up to 40% of the body weight(Selvan te al., 1993, Fitters et al., 1999) and aremost important energy reserve, though protein andthe carbohydrates, glycogen and trehalose, also yieldenergy (Qiu and Bedding, 2000). Good qualitynematodes tend to possess high lipid levels thatprovide a dense appearance, whereas nearlytransparent nematodes are often active but possesslow powers of infection.

Insects killed by most steinernematidnematodes become brown or tan, whereas insectskilled by heterorhabditids become red and the tissuesassume a gummy consistency. A dim luminescencegiven off by insects freshly killed by heterorhabditidsis a foolproof diagnostic for the genusHeterorhabdit is (the symbiotic bacteriaPhotorhabdus provide the luminescence). Black


Nematode species Product formulation Country

Steinernema carpocapsae ORTHO Biosafe USA Bio Vector USA

Exhibit USA Sanoplant

Boden Nutz;li nge Helix

X-GNAT Vector TL

USA USA USA

Switzerland Germany Canada

USA USA

S. feltiae Magent Nemasys Stealth

Entonem

USA UK UK

USA S. riobrave Vector MG

Bio Vector USA USA

S. scapteri sci USA Heterorhabditis bacteriophora Otinem USA H. megidis Nemasys UK S. carpocapsae Green commandos

Soil commandos India

TABLE 2: Commercial products available in international market.

cadavers with associated putrefaction indicate thatthe host was not killed by entomopathogenic species.Nematodes found within such cadavers tend to befree-living soil saprophages.

H abitat:Steinernematid and heterorhabditidnematodes are exclusively soil organisms. They areubiquitous, having been isolated from everyinhabited continent from a wide range of ecologicallydiverse soil habitats including cultivated fields,forests, grasslands, deserts, and even ocean beaches.When surveyed, entomopathogenic nematodes arerecovered from 2% to 45% of sites sampled(Hominick, 2002; Lalramliana and Yadav, 2010;Khatri-Chhetri et al. 2010).

Host range: Entomopathogenic nematodes havebeen tested against a large number of insect pestspecies with results varying from no effects toexcellent control. Because the symbiotic bacteriumkills insects so quickly, there is no intimate host-parasite relationship as is characteristic for otherinsect-parasitic nematodes. Consequently,entomopathogenic nematodes attacks a far widerspectrum of insects in the laboratory where the hostcontact is assured, environmental conditions areoptimal and no ecological or behavioral barriers toinfection exists(Gaugler 1981 and 1988) . Field hostrange is considerably more restricted, with somespecies being quite narrow in host specificity.Matching biology and ecology of both the nematode

and target pest are of great importance if nematodeapplications are to result in significant pestreductions. Nonetheless, when considered as a groupof nearly 80 species, entomopathogenic nematodesare useful against a large number of insect pests(Grewal et al., 2005). Although in field infections,impact on target populations has been frequentlyjudged modest or negligible, even whenconcentrations equivalent to billion of nematodesper hectare are applied (Finney and Walker, 1979).It would be premature to conclude that nematodesare safe for non target invertebrates in all habitats(Akhurst, 1990). Nevertheless, considerable evidenceindicates that nematodes possess a restricted hostrange in natural systems and, therefore, are not nearlyas threatening as their experimental host range mightsuggest (Kaya and Gaugler, 1993). Smitley et al.,(1992) were the fi rst to test the impact ofentomopathogenic nematodes on plant parasiticnematodes in a field trial and presently,entomopathogenic nematodes have been marketedfor control of certain plant parasitic nematodes,though efficacy has been variable depending onspecies (Lewis and Grewal, 2005).

Conservation:Reports and studies of naturaloccurrence and ecology of entomopathogenicnematodes are relatively uncommon. Informationfrom the manipulative experiments, however, canhelp to assess the effects of environmental factors,

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FIG. 1: Generalized life cycle of steinernematids andheterorhabditids. IJ = infective juvenile.

point out the gaps in our knowledge and help us tounderstand how this important group of naturalenemies of insects can be conserved (Lewis et al.,1998). The reasons for success or lack of success incontrolling insect pests, particularly in the soilenvironment, often remain unknown, underscoringthe need to obtain basic information on the biology,behavior, ecology and genetics of these nematodes.Research in behavioral ecology has clearlydemonstrated that these entomopathogenicnematodes are not generalist pathogens. Althoughmost of them have a broad host range in thelaboratory, in the field they are adapted to hosts in aparticular environment depending on their foragingstrategy. Their foraging strategy restricts much of theiractivity to certain soil stratum eliminating manyinsects from infection. Native populations are highlyprevalent, but, other than scattered reports ofepizootics, their impact on host populations isgenerally not well documented (Stuart et al., 2006).Epizootics of entomopathogenic nematode diseasesprobably occur regularly in soil, but they are difficultto detect and often go unrecorded (Kaya, 1987). Twoepizootics, both from Australia involving undescribedHeterorhabditis species, have been documented(Akhurst et al., 1992; Sexton and Williams 1981).In the first case, a significant reduction of thewhitefringed beetle, Graphagnathus leucoloma,larvae and adults was observed in lucerne field(Sexton and Williams 1981). In the second, twoundescribed species of Heterorhabditis were found

infecting four scarabaeid species in three adjoiningsugarcane fields (Akhurst et al., 1992). Underlaboratory conditions, the scarabaeid larvae are notsusceptible to the nematodes, but in the field infectedlarvae were readily recovered. This is largelyattributable to the mysterious nature of soil insects.Consequently, research and guidelines for conservingnative entomopathogenic nematodes are in need ofadvancement.

Compatibility: EPN infective juveniles (IJs) cantolerate short-term exposures (2-24 h) to manychemical and biological insecticides, fungicides,herbicides, fertilizers and growth regulators, and canthus be tank-mixed and applied together. This offersa cost-effective alternative to pest control, andfacilitates the use of nematodes in integrated pestmanagement systems. However, the actualconcentration of the chemical to which thenematodes will be exposed will vary depending uponthe application volume and system used (Alumai andGrewal, 2004). Compatibility has been tested withwell over 100 different chemical pesticides.Entomopathogenic nematodes are compatible (e.g.,may be tank-mixed) with most chemical herbicidesand fungicides as well as many insecticides (suchas bacterial or fungal products) (Koppenhöfer andGrewal, 2005). In fact, in some cases, combinationsof chemical agents with nematodes results insynergistic levels of insect mortality. Some chemicalsto be used with care or should be avoided likealdicarb, carbofuran, diazinon, dodine, methomyl,and various nematicides as they do not have anycompatible interact ion. However, specif icinteractions can vary based on the nematode andhost species and applicat ion rates. Theorganophosphate oxamyl increased S. carpocapsaeefficacy against Agrotis segetum synergistically, butonly in fumigated soil, probably by enhancing thenematodes nictation behavior (Ishibashi, 1993).Furthermore, even when a specific chemicalpesticide is not considered compatible, use of bothagents (chemical and nematode) can beimplemented by maintaining an appropriate intervalbetween applications (e.g., 1 – 2 weeks). Prior touse, compatibility and potential for tank-mixingshould be based on manufacturer recommendations.Combination of two nematode species may provideeffective control of two pests (Kaya et al., 1993) and


the nematode species may coexist (Koppenhofer andKaya, 1996) if the target differs in susceptibilityagainst the search strategies and/or pathogenicityof the two nematodes. Similarly, entomopathogenicnematodes are also compatible with many pathogens(Koppenhöfer and Grewal, 2005) like combinationof Bacillus thuringiensis (Bt) against severallepidopterous pests resulted in additive andantagonistic interactions. Interactions range fromantagonism to additivity or synergism depending onthe specific combination of control agents, targetpest, and rates and timing of application.Koppenhofer et al., (2000) documented synergismbetween Heterrhabditis spp. and S. glaseri andneonicotinoid imidacloprid in scarab larvae. Asimidacloprid reduces the grub’s defensive behaviorresulting in increased nematode attachment andpenetration. Nematodes are generally compatiblewith chemical fertilizers as well as composted manurethough fresh manure can be detrimental.

Commercial availability: Of the nearly eightysteinernematid and heterorhabditid nematodesidentified to date, at least twelve species have beencommercialized. A list of some commerciallyavailable entomopathogenic nematode products ininternational market are given in table 2 (Siddiquiet al., 2010). One billion nematodes per acre(250,000 per m2) is the rule-of-thumb against mostsoil insects (greenhouse soils tend to be treated athigher rates).

Conclusions: Entomopathogenic nematology hasa relatively short history dating back to the pioneeringresearch of R.W. Glaser and his coworkers in the1930s and 1940s (Gaugler R and Kaya HK, 1990).The primary emphasis of their research and of othersfollowing them focused on developing and usingthese nematodes as biological insecticides. Thereasons for success or lack of success in controllinginsect pests, particularly in the soil environment, oftenremains unknown, underscoring the need to obtainbasic information on the biology, behavior, ecology

and genetics of these nematodes. Recently advancesmade in nematode behavior and ecology clearlydemonstrates that they are not generalist pathogens;their behavior, for example, restricts much of theiractivity to certain soil stratum, eliminating manyinsects from infection. Understanding thesebehavioral patterns and their genetics will enhancethe use and production of the most adapted speciesfor insect control in the field.

Entomopathogenic nematodes (Rhabditida:Steinernematidae and Heterorhabditidae) inparticular have emerged as excellent biocontrolagents of soil-dwelling insect pests and henceattracted widespread commercial interest. Thesebiological control agents are endowed with manyadvantages including host seeking capability, highvirulence, ease of production, ease of application,mammalian safety and exception from registrationin many countries. They also possess a broad hostrange, are compatible with many other controlagents, are widespread in distribution and can beformulated and stored for reasonable length of time.However, the efficacy of these nematodes againstinsect pests varied, particularly in soil environment,are often unknown, emphasizing the need to obtainbasic information on the biology, behavioral ecologyand genetics of these nematodes. Indeed, there hasbeen a surge in research to obtain more basicinformation and in exploration to discover newnematode isolates. These nematodes are ubiquitousin nature, but their populations usually are notsufficiently high to cause epizootic and reduce pestpopulation. Research is needed to better understandthe factors that regulate their population and on howtheir population can be manipulated to initiateepizootic in insect-pest populations. Finally, thesefascinating animals may contribute more to sciencethan their use solely as biological control agents. Tobegin with, they may be useful tools in understandingthe evolution of parasitism and symbiosis andmechanisms by insect resistance to infection.

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Microbial pp. 234-238. Boca Raton, FL: CRC 259 pp.Akhurst, R.J., Bedding, R.A., Bull, R.M. and Smith, D.R.J. (1992). An epizootic of Heterorhabditis spp. (Heterorhabditidae:

Nematoda) in sugarcane scarabaeids (Coleoptera). Fund Appl Nematol., 15: 71-73.

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Alumai, A. and Grewal, P.S. (2004). Tankmix-comaptibility of the entomopathogenic nematodes, Heterorhabditisbacteriophora and Steinnernema carpocapsae, with selected chemical pesticides using in turfgrass. BiocontrolSci Tech., 14: 725-730.

Bedding, R.A. (1981). Low cost in vitro production of Neoplectana and Heterorhabditis species (Nematoda) for fieldcontrol of insect pests. Nematologica., 27: 109-114

Bedding, R.A. (1984). Large scale production, storage and transport of the insect-parasitic nematodes Neoaplectanaspp. and Heterorhabditis spp. Ann Appl Bio., 104: 117-120.

Bedding, R.A. (1990). Logistics and strategies for introducing entomopathogenic nematodes technology into developingcountries. In: Gaugler, R. and Kaya, H.K. (eds) EPN. Biol. Control., CRC Press, Boca Raton, Florida, pp-233-248.

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