This article was downloaded by: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] On: 10 June 2011 Access details: Access Details: [subscription number 923464531] Publisher Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 3741 Mortimer Street, London W1T 3JH, UK
Archives Of Phytopathology And Plant Protection
Publication details, including instructions for authors and subscription information: http://www.informaworld.com/smpp/title~content=t713454295
Entomopathogenic nematodes, a potential microbial biopesticide: mass production and commercialisation status - a mini review Mahaveer P. Sharmaa; Amar N. Sharmaa; Syed S. Hussainib a Directorate of Soybean Research (DSR-ICAR), Indore, India b National Bureau of Agriculturally Important Insects (formerly Project Directorate of Biological Control), Bangalore, India Online publication date: 25 May 2011
To cite this Article Sharma, Mahaveer P. , Sharma, Amar N. and Hussaini, Syed S.(2011) 'Entomopathogenic nematodes, a
potential microbial biopesticide: mass production and commercialisation status - a mini review', Archives Of Phytopathology And Plant Protection, 44: 9, 855 — 870 To link to this Article: DOI: 10.1080/03235400903345315 URL: http://dx.doi.org/10.1080/03235400903345315
PLEASE SCROLL DOWN FOR ARTICLE Full terms and conditions of use: http://www.informaworld.com/terms-and-conditions-of-access.pdf This article may be used for research, teaching and private study purposes. Any substantial or systematic reproduction, re-distribution, re-selling, loan or sub-licensing, systematic supply or distribution in any form to anyone is expressly forbidden. The publisher does not give any warranty express or implied or make any representation that the contents will be complete or accurate or up to date. The accuracy of any instructions, formulae and drug doses should be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims, proceedings, demand or costs or damages whatsoever or howsoever caused arising directly or indirectly in connection with or arising out of the use of this material.
Archives of Phytopathology and Plant Protection Vol. 44, No. 9, May 2011, 855–870
Entomopathogenic nematodes, a potential microbial biopesticide: mass production and commercialisation status – a mini review Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Mahaveer P. Sharmaa*, Amar N. Sharmaa and Syed S. Hussainib a
Directorate of Soybean Research (DSR-ICAR), Khandwa Road, Indore 452001, India; National Bureau of Agriculturally Important Insects (formerly Project Directorate of Biological Control), PB No 2491, HA Farm Post, Bellary Road, Bangalore 560024, India b
(Received 9 August 2009; final version received 1 September 2009) Parasitic nematodes have several important attributes that make them excellent candidates for biological control of soil insects. These nematodes can be produced by in vivo by baiting technique on insects and commercially by in vitro solid/liquid culturing. Numerous insect pests on many different crops are being controlled by these insect parasitic nematodes, including root weevils, flea beetles, mint root borer, colorado potato beetle, white grubs, caterpillars and plant parasitic root nematode, e.g. root-knot nematodes. Utilisation of entomopathogenic nematodes (EPN) has raised intense interest and has been a growing concern globally mainly because of its potential efficiency, exemption from registration and other impressive attributes for utilising against the control of soil dwelling pests. This review highlights the mass production, commercialisation and utilisation of EPN as microbial biopesticide in bio-intensive pest management programmes. Keywords: Entomopathogenic production techniques
nematodes;
Steinernema;
Hetrerhabditis;
Introduction Indiscriminate pesticide use has raised many environmental concerns in respect to ground water contamination, residues on food, resistance development and wild life kills and has resulted in prohibitive legislation (Zimmerman and Cranshaw 1990) thus raised interest for safer alternatives. Biological control exploits insects, fungi, bacteria, viruses and nematodes as biological insecticides. Nematodes are the most numerous multicellular animals on earth. Nematodes are simple roundworms, colourless, 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 (Gaugler 2002). A handful of soil will contain thousands of the microscopic worms, many of them parasites of insects, plants or animals. There are nearly 20,000 described species classified in the phylum Nematoda. It has been estimated that half table spoon of soil will have a numbers of organisms – bacteria 108–9, actinomycetes 105–8, fungi 105–6, micro-algae 103–6, protozoa 103–5, nematodes 101–2, other invertebrates 103–5 (Dindal 1990). Free-living species are abundant, including nematodes that feed on insects, bacteria, fungi and other
*Corresponding author. Email:
[email protected] ISSN 0323-5408 print/ISSN 1477-2906 online Ó 2011 Taylor & Francis DOI: 10.1080/03235400903345315 http://www.informaworld.com
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
856
M.P. Sharma et al.
nematodes, yet the vast majority of species encountered are poorly understood biologically. A certain group of nematodes that parasitise insects are called insect pathogenic or entomopathogenic nematodes (EPN) (Poinar 1990). EPN comprises two words, i.e. Entomo-‘ ¼insect or pathogenic’ ¼ producing disease. Nematodes associated with insects are referred to as entomophilic, entomogenous, or ‘entomopathogenic’ are known to parasitise, cause disease and kill the insects. EPNs are nematodes capable of infecting and killing insects with the aid of symbiotic bacteria (Poinar 1990). EPN in the families Steinernematidae and Heterorhabditidae are potential nematodes as they are highly virulent because of their symbiotic association with bacteria Xenorhabdus and Photorhabdus sp (Akhurst and Boemare 1990). Within these families, the genera Steinernema and Heterorhabditis are important and have gained commercial application (Peters and Ehlers 1994; Georgis and Manweiler 1994; Gaugler et al. 2000). Nematode unique to EPN is their close association with specific bacteria of the genera Xenorhabdus with Steinernema and Photorhabdus sp. with Heterorhabditis (Boemore et al. 1993). These bacterial symbionts belong to the Enterobacteriaceae within the gamma sub division of the purple bacteria (Ehlers et al. 1998). The EPNs are excellent biological control agents for soil-dwelling stages of many insect pests and are fast acting, killing target insect pests in 24–48 h (Kaya et al. 1999). In comparison, many other biological control agents take days or weeks to kill the target insect pest. EPN are safe to most non-target organisms and the environment, are easy to apply, and are compatible with most agricultural chemicals. They also have a broad host range, ability to search for pests, and a potential to reproduce after application (Kaya and Gaugler 1993). Large-scale application and demonstration of EPN has been dominated in the western countries covering thousands hectares of land. So far, more than 30 species of these EPN have been described (Hominick et al. 1997) out of these nine species of Steinernema and three of Heterorhabditis (seven alone in US) have been commercially exploited (Shapiro-llan et al. 2002) using in vivo and in vitro techniques (Kaya 2002). Many pests of horticulture, agriculture, home and garden besides public health have reached the second position after Bacillus thuringiensis-based products. In India, extensive and systematic surveys undertaken by Project Directorate of Biological Control, Bangalore for determining the existence of several potential native isolates of Steinernema and Heterorhabditis. Significant amount of work on EPN research on selection of improved strains, mass production and developing their simple ready-to-use formulations is being carried out at PDBC, Bangalore, India. In the world, the most dominant work on EPN related to mass production, commercialisation and application has been carried out in US, Israel, Germany, UK and other part of Europe. A large number of companies are in the business of commercial production of EPN (Table 1). Attempts have been made in this article to appraise the status of EPN as microbial bioagents, their commercial potential and future directions for enhanced use of Sternernema sp. and Heterorhabditis sp. as microbial biopesticide. Production, commercialisation and application The symbiotic association of EPN with specific bacteria facilitates rapid production of nematodes (bacteria serve as food) and successful pathogenicity. Although axenic
Archives of Phytopathology and Plant Protection
857
Table 1. Some available commercial products containing Steinernema and Heterorhabditis nematodes (Grewal and Georgis 1988). Formulation
Nematode species
Alginate gel
S. carpocapsae S. carpocapsae S. carpocapsae S. feltiae S. feltiae H. megidis H. megidis S. feltiae S. feltae S. carpocapsae S. carpocapsae S. carpocapsae S. carpocapsae S. carpocapsae S. carpocapsae S. feltae S. riobravis
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Clay
Water dispersible granules
Product
Company
Microplant Boden-Nutzlinge Biosafe Exhibit Stealth Nemasys-H Larva Nem NemaSys Entonem Proactant Ss Biosafe Biosafe-N BioVector Vector TL Helix X-GNAT Vector MC
Novartis, Vienna, Australia Rhone-Poulenc, Germany SDS Biotech, Japan Novaritis, Switzerland Novaritis, UK Micro Bio, UK Koppert BV, Netherlands Micro Bio Koppert BV, Netherlands Biocontrol, FL, USA Thermo Trilogy, Columbia, MD Thermo Trilogy, Columbia, MD Thermo Trilogy, Columbia, MD Lesco, Lansing, MI Novaritis, Canada E.C. Geiger, USA Lesco, UK
nematodes (nematodes without bacteria) requires great deal of work to ensure host survival and reproducibility (Grewal and Georgis 1988). Furthermore, bacteria alone are not capable of penetrating the gut and cannot independently gain entry to the host’s hemocoel. Thus nematode acts as vector to transport bacteria into host within which they can proliferate, and bacteria create conditions necessary survival and reproduction within the insect cadaver (Grewal and Georgis 1988). A key factors in the success of EPN, as biopesticides is their amenability to mass production. EPNs can be mass-produced by in vivo or in vitro methods (Friedman 1990; Ehlers 2001; Gaugler and Han 2002; Hussaini 2002). These nematodes were first cultured more than 70 years ago and currently they are commercially produced using these culture methods: in vivo and in vitro solid and liquid culture. In the in vivo process, an insect serves as a bioreactor; in the in vitro process, artificial media are used. In vivo culture method In vivo production is a cottage industry of low volume producers (Gaugler and Han 2002) and is based on adaptation of the white trap (White 1927), although, with some modifications (Lindegren et al. 1993) in which nematode killed hosts are placed above a water reservoir. With limited capital for in vitro culture the default strategy is to rear nematodes in insect hosts. In vivo mass production depends on the availability of a highly reliable, highly susceptible and inexpensive supply of hosts (Gaugler and Han 2002). In vivo culture is two-dimensional system that relies on production in trays and shelves. Production methods for culturing EPN in insect hosts using greater wax moth (Galleria mellonella) have already been reported (Gaugler and Brown 2001). All of these describe (with some variation) a system based on the White trap, which take advantage of the infective juveniles (IJs) natural migration away from the host
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
858
M.P. Sharma et al.
cadaver upon emergence. First scalable system for in vivo nematode mass production was developed by Gaugler group (Gaugler et al. 2002) where unlike white trap, there is no requirement for nematode migration to a water reservoir. The LOTEK system of tools and procedures provides process technology for low-cost high efficiency mass production of EPN. The LOTEK system consists of perforated trays to secure insect hosts, an automated self cleaning harvester with misting nozzles that trigger IJ emergence and rinse nematode through the holding trays to a central bulk storage tank and a continuous deflection separator for washing and concentrating nematodes (Gaugler et al. 2002). The methods described consist of inoculation, harvest, concentration and if necessary decontamination. Insects are inoculated with nematodes on a dish or tray lined with absorbent paper. After 2–5 days, infected insects are transferred to White traps; if infection is allowed to progress too long before transfer harm to nematode reproductive stages may occur and the cadavers are more likely to rupture. White traps consist of a dish on which the cadavers rest surrounded by water which is contained by a larger dish or tray. The central dish (containing the cadavers) provides a moist substrate for the nematodes to move upon, e. g. inverted Petri dish lid lined with filter paper. In the long run, the in vivo system would lack economy of scale; labour, equipment and insect’s costs increase as a linear function of production capacity (Grewal and Georgis 1988). Perhaps even more important is the lack of improved quality while increasing scale. The in vivo nematode production is increasingly sensitive to biological variation and catastrophes as scale increases (Friedman 1990). In vitro solid culture method EPN were first grown in vitro on a solid medium axenically (Glaser 1932). Thereafter, it was realised that growth increased with presence of bacteria. The importance of the natural symbiont was recognised (Poinar 1966). Therefore, the first successful commercial scale monoxenic culture was developed by Bedding, and has known to be as solid culture (Bedding 1981, 1984) using chicken offal or another protein rich medium soaked in an inert carrier (sponge, polyurethane). In this method, nematodes are cultured on a crumbed polyether polyurethane sponge impregnated with emulsified beef fat and pig’s kidney along with symbiotic bacteria. Using this method, approx. 6 6 105–10 6 105 IJs/g of medium was achieved (Bedding 1984). Today the need for monoxenicity is universally recognised as one of the cornerstone of nematode in vitro culture (Poinar and Thomas 1966). The solid media process has successfully produced pathogenic steinernematids and heterorhabditids but the high labour cost limits economics of scale. This technology is most suitable for countries where labour costs are minimal (Bedding 1990). In vitro solid culture advanced considerably with the invention of a threedimensional rearing system involving nematode culture on crumb polyether polyurethane foam (Bedding 1981). A liquid medium is mixed with foam and autoclaved. Bacteria are inoculated first followed by nematode 3 days later. Nematodes can be harvested within 2–5 weeks by placing the foam onto sieves, which are immersed in water. IJs migrate out of the foam, settle downward, and are pumped to collection tank; the product is cleaned through repeated washings with water, i.e. sedimentation and decanting. As in Petri dishes, media for this approach
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Archives of Phytopathology and Plant Protection
859
were animal product based but was later improved for cost and consistency and may include various ingredients including peptone, yeast extract, eggs, soy flour and lard. The bacteria can be inoculated first followed by the nematode several days later. It was realised the two organisms could be added simultaneously if a large concentration of bacteria is used. The potential for scale up production was further advanced through several measures including using bags with a gas permeable Tyvac strip for ventilation automated mixing and autoclaving, and harvest through centrifugal sifters. These all developments take place in the phases of scaling up at Biotech Australia and then Ecogen-Sylvan, USA (Gaugler and Han 2002). In vitro liquid culture method Glaser group (Glaser 1940) developed the first liquid medium, based on kidney extract, for axenic culture of Steinernema glaseri. Then, EPN was grown in liquid culture axenically (Stoll 1952). He used a liquid medium containing raw liver extract and incubated the cultures on a shaker. The nematode developed and produced offspring reaching approximately 400-dauer juveniles/ml. A significant step forward was achieved in this direction (Buecher and Hansen 1971). By simply supplying sterile air through liquid media bottles, they showed that bubbling is an acceptable means of supplying aeration to nematodes without affecting shear effect due to forced aeration. However, nematodes thus producing axenically could not be used for biocontrol purposes due to low yields, high cost on media and more important absence of symbiotic bacteria in the culture (Ehlers et al. 1997). It was also showed that even gently agitation (shear effect) of solid cultures suppressed nematode development (Bedding 1984). Various ingredients for liquid culture have been reported including soy flour, milk powder, yeast extract, corn oil, casein peptone, thistle oil, egg yolk, liver extract and cholesterol (Buecher and Popiel 1989; Friedman et al. 1989). Culture times, which can vary depending on media and species, may be as long as 3 weeks, but many species reach max production in 2 weeks or less. Once the culture is completed, nematodes can be removed from the medium through centrifugation. Lipids have received more attention that other nutritional components because 60% of the total energy for the non-feeding IJs is derived from metabolising lipids (Hatab and Gaugler 1997). It was also concluded that lipid sources with high proportions of saturated fatty acids result in suboptimal yield (Hatab and Gaugler 2001). Further a medium was tested for Heterorhabditis bacteriophora rich in canola oil to enhance lipids rich in mono-saturated fatty acids (Yoo et al. 2000). Pace group (Pace et al. 1986) cultured nematodes in a standard 10-l bioreactor/ fermenter but due to shear effect adult females got disrupted. Therefore, to increase the recovery/yield and reduce losses caused by shear stress there is need to replace flat blade impeller with a paddle stirrer. In 1992, Biosys (Palo Alto, Calif) attempted liquid culture on large scale for the production of Steinernema carpocapsae and production was scaled-up to volumes of 80,000 l. Currently, the in vitro liquid culture method is a commercially viable method wherever expertise and initial capital is available. Companies that include Microbio, USA, E-Nema GmbH, Germany and SDS Biotech have adopted this system (Ehlers 2001; Gaugler and Han 2002).
860
M.P. Sharma et al.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Formulations, storage and field application Formulation is intended to improve activity, absorption, delivery, ease-of-use or storage stability of an active ingredient. Typical examples of pesticide formulation ingredients (additives) include absorbents, adsorbents, anti-microbial agents, antioxidants, binders, carriers, dispersants, preservatives, solvents, surfactants and UV absorbers (Grewal 2002). Although the overall concept for EPN formulations is similar to pesticide formulations only different is nematode presence that is unique and has challenges because of high demand for oxygen, moisture and sensitive to solar radiation and extreme temperature (Grewal 2002). The success of EPN infectivity depends on formulated stable product, which is important for commercialisation. Large scale produced nematodes can be stored in suspension for several days/weeks in a refrigerated bubbled tank, though with a risk of contamination and high oxygen demand (Mracek 2002). But this method is not commercial feasible. Nematode metabolism is temperature driven and warm temperature (20–308C) increase metabolic activities, reducing nematode viability (Georgis 1990a). Most common formulations are made in alginate, polyacrylamide gels, clay, vermiculite and activated charcoal suitable for 6 months at refrigerated temperature and 3 months at room temperature (Georgis 1990a). Formulation stability is also achieved by partially desiccating and immobilising IJs in specific carriers. Immobilisation has been accomplished by maintenance in aqueous suspension at low temperature (5–158C) (Hussaini 2002). Steinernematids can be stored at 4–108C for 6–12 months without much loss of activity whereas heterorhabditids do not store well and 2–4 months of storage at 4–108C is considered good. General range storage range for steinernematids is 5–108C and 10– 158C for heterorhabditids (Georgis 1990a). Encapsulation of EPN with calcium alginate perhaps the first report (Kaya and Nelson 1985). The alginate-based S. carpocapsae products were the first to possess root temperature shelf life of about 3– 4 months (Grewal 2002) and led to an increased acceptability of EPNs in local markets. However, while scaling-up on mass scale this formulation was found unsuitable for large-scale application (Grewal and Georgis 1988). Although EPN have been successfully formulated in gel-forming polyacrilamides (Georgis 1990a), flowable gels (Georgis and Manweiler 1994), attapulgite clay (Bedding 1988), all these were having low shelf life compared to alginate gels. A significant advancement in formulation and storage has taken place in recent years (Grewal 2002) (Table 2) with the advent of water-dispersible granules (WDG). WDG in which IJs are encased in 10–20 mm diameter granules consisting of mixture of various types of silica, clays, cellulose, lignin and starches (Silver et al.1995). Induction of partial anhydrobiosis has been achieved successfully by controlling water activity of the formulations (Silver et al. 1995; Grewal 2000). The water activity is a measure of how tightly water is bound, structurally or chemically to the nematodes. The WDG extended nematode storage stability to several months at 15–258C, enhanced nematode tolerance to temperature extremes, enabling easier and less expensive transport and labour intensive preparation steps and proved overall economic (Grewal 2000, 2002). Very recently a method (Chen and Glazer 2005) for long term storage of EPN juveniles at 238C in distilled water containing osmotic solution of 18% glycerol and 2% sodium alginate was dropped in 0.5% CaCo3.2H2O, which formed calcium alginate granules. The granules were placed in 5 cm Petri dishes and transformed into sealed 2.5-l dessicator under 85%
Archives of Phytopathology and Plant Protection Table 2. 2002).
Expected shelf life Steinernema and Heterorhabditis spp. in formulations (Grewal
Formulation Actively moving nematodes Spongea Vermiculitea Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
861
Reduced mobility nematodes Alginate gels Flowable gels Liquid concentratea Anhydrobiotic nematodes Wettable powder Water dispersible granulesa
Nematode species
Strain
Shelf-life (months)
S. carpocapsae H. bacteriophora S. carpocapsae S. feltiae H. megidis
All HP88 All UK UK
22–258C 0.03–0.1 0 0.1–0.2 0.03–0.1 0
2–108C 2.0–3.0 1.0–2.0 5.0–6.0 4.0–5.0 2.0–3.0
S. S. S. S. S. S.
carpocapsae feltiae carpocapsae glaseri carpocapsae riobrave
All SN All NJ43 All RGV
3.0–4.0 0.5–1.0 1.0–1.5 0.03–0.06 0.16–0.2 0.1–0.13
6.0–9.0 4.0–5.0 3.0–5.0 1.0–1.5 0.4–0.5 0.23–0.3
S. carpocapsae S. feltiae H. megidis S. carpocapsae S. feltiae S. riobrave
All UK UK All SN RGV
2.0–3.5 2.5–3.0 2.0–3.0 4.0–5.0 1.5–2.0 2.0–3.0
6.0–8.0 5.0–6.0 4.0–5.0 9.0–12.0 5.0–7.0 4.0–5.0
a
commercially available formulation.
relative humidity, or sealed into plastic boxes in which a high RH was maintained by the presence of a piece of distilled water-soaked sponge. They concluded that EPN survived (99%) upto 6 months in granules containing osmotic solution in presence of 100% RH in dessicator. A non-viscous, non-adhesive and non-toxic liquid formulation has developed for EPN storage and transport based on neutral density colloidal silica suspensions (Wilson and Ivanova 2004). Survival and virulence of stored nematodes in this formulation without aeration was found superior than stored in aerated quarter strength Ringer’s solution. Shapiro-Ilan group (Shapiro-llan et al. 2002) has attempted the feasibility of formulating nematode-infected insect cadavers to overcome the storage and application hindrances. The formulated cadavers (starches, clays, etc.) were more resistant to rupturing and sticking during agitation than non-formulated cadavers. Lewis and Shapiro-Ilan (2002) also showed that nematode-infected insect cadavers were found to be resistant when exposed to freezing. The quality of nematodes that survive the manufacturing process is analysed by determining their shelf life and virulence potential. Shelf life is predicted from storage energy reserves of IJs, whereas virulence potential is measured using 1:1 bioassay against wax moth (Miller 1989; Grewal et al. 1998). For effective application EPN have been applied in various formulations, e.g. IJs through water, desiccated cadavers, baits and capsules (Grewal 2002; Georgis 1990b), etc. Water dispersible granular formulation was found to be the most appropriate, had longer shelf life, minimum space for storage and ease of application (Grewal 2000, 2002). EPN have been most efficacious for insects that reside in soil or cryptic habitats where there is protection from desiccation, UV radiation and high temperature. Moreover, best results are usually obtained when nematodes are
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
862
M.P. Sharma et al.
applied to moist soil in the evening or early morning (Grewal and Georgis 1988). Spraying IJs directly onto the soil surface is the most common method of application. A spray volume between 750 and 1900 l/ha is usually sufficient for most nematode species to reach the target insects in soil (Grewal 2002). Post application irrigation improves efficacy and rinses the nematodes from the foliage and into their natural reservoir, the soil (Selvan et al. 1994). The spraying can be done with any commercially available ground or aerial spray equipments, which include smallpressurised sprayers, mist blowers and electrostatic sprayers as well as traditional sprayers used in aerial application via helicopters (Georgis 1990b). To increase the efficacy, several considerations have been taken in account. Antidesiccants have been used successfully to retard evaporation of the nematode suspension in foliar applications and thus reduce desiccation of nematodes (Glazer and Navon 1990). Recently, PDBC (Hussaini et al. 2001) have been reported good UV protectants for PDBC isolates of Steinernema sp. and Heterorhabditis indica. Registration regulations So far EPN has been exempted from registration process in many countries, e.g. in USA by the Environmental Protection Agency (EPA) but some countries have restrictions for exotic or non-indigenous species/strains which require registration. In UK the indigenous and genetically unmodified EPNs and their bacterial symbionts do not have obligations for registration regulations. The European Commission is regulating and streamlining the procedures for the authorisation of the plant protection products including EPN under the provision of Council Directive 91/414/ EEC (Richardson 1996). Other European country either has the mandatory regulation based on Council Directive or no registration is required. EPN: A tool for integrated and bio-intensive pest management EPNs in the genera Steinernema spp. and Heterorhabditis spp. are found to be potential agents for control of insect pests mainly belonging to order Diptera, Coleoptera, Lepidoptera and Orthoptera. Many workers have tried the use of EPN for the control of target insects. The efficacy of EPN has been found to influence by nematode species, strain, production and storage conditions, and persistence in the habitat and susceptibility of target insect pests. EPN potential has been realised for controlling insect pests, e.g. beetles, fly hosts and butterfly hosts mainly from cryptic and foliar habitats. Today, EPN are mainly used in environments where chemical pesticides fail, i.e. in the soil, in the galleries of boring insects, or in cases where resistance to insecticides has developed (Ehlers 2001) (Table 3). Very recently a mechanism of insect hosts attracting EPN to control root pests has been identified by a Swiss group comparing two different germplasm of maize infecting Diabrotica virgifera for validating the basis for EPN attraction. They found b-caryophyllene compound secretions from European maize roots infected with Diabrotica virgifera had lower infestation due to attraction of EPN whereas an American maize variety did not secrete b-caryophyllene had more insect attack due to no attraction of EPN (Rasmann et al. 2005). In India, various workers have used EPN against cutworms, ragi pink borer, stem borer, white grubs, etc. in laboratory and field conditions (Singh 1977; Sivakumar et al. 1989; Hussaini et al. 2003; Sitaramaiah et al. 2003). In USA and Europe, Steinernema carpocapsae, S. riobrave and Heterorhabditis sp. were
Archives of Phytopathology and Plant Protection Table 3.
Commercial use of entomopathogenic nematodes.
Common name Fungus gnats
Scientific name
Order
Mushrooms
Diptera
Ornamentals
Diptera Diptera Diptera Coleoptera
Cabbage Turf Stables Ornamentals, vegetables Strawberry, ornamentals Strawberry, cranberry Hop Sugarbeet Citrus Turf
Cabbage root flym March flies House flym Leafminer Black vine weevil
Otiorhynchus sulcatus
Coleoptera
Strawberry weevil
O. ovatus
Coleoptera
Hop weevil Sugarbeet weevil Citrus root weevil White grubs
Coleoptera Coleoptera Coleoptera Coleoptera
Peanut white grubm
O.ligustici Temborhinus mendicus Diaprepes abbreviatus Popillia japonica, Anomala spp., Phyllopertha horticola, etc. Maladera matrida
Cutworms Banana moth Ghost moth
Agrotis ipsilon, etc. Opogona sacchari Hepialus spp.
Lepidoptera Lepidoptera Lepidoptera
Cockroachm
Blattaria
Mole cricket Western flower thripsm
Periplaneta americana, Blatta spp., Scapteriscus spp., Frankliniella occidentalis
Orthoptera Tysanoptera
Cat flea
Ctenocephalides felis
Siphonaptera
Application against insects pest marked 2001).
m
Culture
Diptera
Lycoriella solani, L. auripila, Lycoriella spp., Bradysia coprophila, Bradysia spp. Delia radicum Bibio hortulans Musca domestica Liriomyza spp.
Fungus gnats
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
863
Coleoptera
Sweet potatoes, peanuts Various Ornamentals Chives, ornamentals Household Turf Ornamentals, vegetables House gardens
is currently under development for market introduction (Ehlers
used against large number of pests those of white grubs (Koppenho¨fer and Fuzy 2003; Grewal et al. 2004) mole crickets, fungus gnats, root weevils (Miklasiewicz et al. 2002; Stuart et al. 2004), bugs borers on variety of crop plants, turfs, black vine weevil, strawberry root weevil (Table 4). EPN has been found successful in reducing damage to turf grass by mole cricket (Hudson et al. 1988). The performance of EPN has got more success to control soil pests when compared to foliar pests. The major reason for lack of success of foliar application of EPN is the intolerance of juveniles to extreme of desiccation (Lello et al. 1996), temperature (Grewal et al. 1994) and ultraviolet radiation (Gaugler et al. 1992). Very recently, Schroer and Ehlers (2005) used S. carpocapsae on cabbage against foliar insect Plutella xylostella formulated with surfactant Rimulgan and polymer xanthan to provide optimal conditions that support nematode invasion on the foliage surface. In agro-ecosystem soil moisture, temperature and sunlight (UV radiation) are the three most important factors that influence EPN efficacy (Richardson and Grewal 1991) and field efficacy has been reviewed recently (Hussaini 2002; Mracek 2002) and presented in Table 4. EPN
Cydia pomonella Hoplocampa testudinea Diaprepes abbreviatus Otiorhynchus sulcatus Comoritis albicapilla Popillia japonica Popillia japonica Cyclocephala borealis Popillia japonica
Apple
Listronotus oregonensis Maladera matrida Maladera insanabilis
Plutella xylostella
Liriomyza huidobrensis
L. trifolii Ceratitis capitata Lycoriella auripila, L. solani
Carrot Peanut Field and plantation crops Cabbage
Lettuce
Bean Soil Mushroom Sc Sf Sf Sf
Sf
Sc
Hb Hb Hb, Sf, Sg
Native Sr Sc Hb, Sc, Sg, Sf Sg Hb Hb Hb, Sg with Neonicotinoides Sc Hb
Sc
EPN species
9 6 109/ha 150–5000/cm2 3 6 106/tray 1–3 6 106/m2
1–2 6 1011/ha
75 IJ/cm2
Above 65 76–95 8 and 11% increased weight and number 91–93% fly emergence reduction 83%
80% in surfactantpolymer formulation 82
Significant reduction 50–90 Significant reduction
100 40–83
2 6 105/m2 0.5–1.5 6 106 – 0.25 6 106/m2 –
83 100 65–80 50–65 14–92 44–66 34–98 47–83 75
Efficacy (%)
5 b/ha 1 6 105/500 cm long branch Soil baiting technique 7.6 6 109/ha – 5 6 109/ha 2.5 6 109/ha 2.5 6 109/ha –
Dosage (IJs)
Sc, Steinernema carpocapsae; Sf, S. feltae; Sr, S. riobrave; Sg, S. glaseri; Hb, Heterorhabditis bacteriophora; Hm, H. megidis.
Scapteriscus spp. Phylloperta horticola
Golfcourse
Citrus Strawberry Litchi Turf field
Pest
Some recent examples of biological control by EPN against various insect pests.
Crop
Table 4.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Williams and Walters 2000 Hara et al. 1993 Lindegren et al. 1990 Grewal et al. 1993; Tomalak 1994
Schroer and Ehlers 2005
Parkman et al. 1994 Sulistyanto and Ehlers 1996 Miklasiewicz et al. 2002 Glazer and Gol’Berg 1993 Bhatnagar et al. 2004
Grewal et al. 2004 Koppenho¨fer et al. 2002
Lacey and Unruh 1998 Belair et al. 1998 Stuart et al. 2004 Kakouli-Duarte et al. 1997 XuJie et al. 2000 Selvan et al. 1994
References
864 M.P. Sharma et al.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Archives of Phytopathology and Plant Protection
865
efficacy was also found be compatible with chemical insecticides, fungicides and acaricides (Ishibashi 1993) and therefore can often be tank mixed and applied with other pesticides as integrated pest management tool. Some pesticides, such as Imidacloprid (Koppenho¨fer et al. 2000), tefluthrin (Nishimatsu and Jackson 1998), neonicotinoid (Koppenho¨fer et al. 2002) and Bacillus thuringiensis (Koppenho¨fer and Kaya 1997) were found to be synergistic with EPN. There are few pesticides that can reduce EPN efficacy and survival (Grewal et al. 1997; Hussaini et al. 2000). Therefore, formulation and integration of EPN with pesticides and surfactants should be properly evaluated before it is released into the field. EPN: A tool for integrated disease management of plant parasitic nematodes Non-availability of potential nematicides, expensive field application and the present day trend for eco-friendly approaches for pest and disease control are the compelling reasons as nematode control is considered more of an emergency. Today’s era of globalisation discourages the use of nematicides. The use of nematicides is particularly prohibited in organic farming and is thus likely become unavailable in future. In India, large number of crops such as vegetables, fruits and pulses are affected by root-knot nematode, which cause significant damage to these crops. Since, EPN has already established to control insect pests, it would be highly economical if the same were used for the control of plant parasitic nematodes (PPN). There are numerous reports available which show the application of EPN in the suppression of PPN (Grewal et al. 1997; Perez and Lewis 2002; Perez and Lewis 2004, Hussaini et al. 2009) EPN in the genera Steinernema and Heterorhabiditis and their associated bacteria Xenorhabdus spp. and Photorhabdus spp. respectively have suppressed selected species of PPN including root-knot nematode in green house experiments (Ishibashi and Choi 1991; Perez and Lewis 2004) and in the field (Grewal et al. 1997). Root-knot nematode IJs penetrate plant roots usually near the tips and the carbon dioxide secreted by actively dividing cells are known to attract the IJs of root knot as well as Steinernema /Heterorhabditis spp. Second-stage juveniles of the EPN either infect a new host or infect their original host insect in the soil (Hussey 1985). However, the existing literature has limited information on what stage(s) of plant-parasitic nematodes are affected by EPN applications. Tests conducted in the field (Grewal et al. 1997) have demonstrated plant-parasitic nematode population suppression for up to 8 weeks after application of EPN products. However, suppression cannot yet be attributed to any specific effect of EPN on plant-parasitic nematodes and so may vary significantly within the life stage of the plant-parasitic nematode exposed to EPN and with the dose applied. Moreover, EPN application rates and methods varied among experiments. M. javanica was suppressed on tomatoes when one application of 5 6 106 or 10 daily application of 5 6 105 S. glaseri (Steiner) IJs/ potted plant was applied (Bird and Bird 1986). In turf, H. bacteriophora Poinar used a mix of 2.47 6 109 and 2.08 6 109 S. carpocapsae IJs/ha (Smitley et al. 1992), whereas a single application of 2.47 6 109 S. riobrave Cabanillas, Poinar and Raulston IJs/ha used and suppressed plant-parasitic nematodes in field experiments (Grewal et al. 1997). These reports along with findings of others (Tsai and Yeh 1995) indicated that host-plant, application rate, species of PPN, and species of EPN impact the level of PPN suppression by EPN.
866
M.P. Sharma et al.
Recently it has been reported that rate, application time and EPN species affected M. incognita suppression on tomato (Perez and Lewis 2002). They showed that 25 IJs/cm2 (2.5 billion/ha) of S. feltiae (Filipjev) were suppressive when applied at the time of M. incognita infestation but, when EPN application time was delayed by 2 weeks, a five-fold rate increase was required to achieve the same level of suppression.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Conclusion and future prospects The use of EPNs belonging to genera Steinernema and Heterorhabditis as biological control agents against insect pests has grown rapidly in recent years. It has been established that these EPN effectively kill and multiply within the host. These nematodes are safe to apply and had no adverse effects on humans and non-target organisms and exempted from registration. The large scale mass production, formulation; storage shelf life for IJs of EPN is need of the hour for commercialising EPN as microbial biopesticide against wide spectrum of soil inhabiting (including cryptic habitats) insect pests. The commercial potential of in vitro mass production need critical evaluation for increased proven shelf life and formulations for ease application, without affecting nematode virulence and metabolism yet producing at much lower costs. However, significant progress has been made on water dispersible granules in which S. carpocapsae can be stored for 5–6 months at 258C. These developments has led to the use of EPN for the control of white grubs, mole crickets and bugs in turfs, root weevils in citrus and strawberries, foliar insets in vegetables and fungus gnats in mushroom. However, EPN has given different levels of efficacy against the mortality of variety of soil pests. Moreover, EPN was also found to be compatible with chemical pesticides, which opens up its large-scale application in IPM programme. Despite this progress, EPN has not reached the reliance level where chemical pesticides use can be reduced. Thus, to enhance target pest mortality, the future goal should be EPN optimisation more closely into different pest management systems, application optimisation for foliar pests, selection and discovery of potential EPN strains against target pests. EPN has also prospects in soil and cryptic habitats that provide the most congenial conditions where IJs may survive, persist, establish, re-cycle and develop a long-term regulation of insect populations. Research should also be focussed on the mechanisms of suppression of PPN by EPN, which will emphasise the need on the integration of EPN in integrated disease and pest management systems. Acknowledgements This review was compiled at TERI, New Delhi by MPS under an EPN project funded by the Department of Biotechnology, Government of India. Thanks are due to Dr R.K. Pachauri, Director General, TERI and Dr. Alok Adholeya, Director, Management of Bioresources Division, TERI, New Delhi for providing necessary facilities for preparing this MS.
References Akhurst R, Boemare NE. 1990. Biology and taxonomy of Xenorhabdus. In: Gaugler R, Kaya HK, editors. Entomopathogenic nematodes in biological control. Boca Raton (FL): CRC Press. p. 75–87. Bedding RA. 1981. Low cost-invitro mass production of Neopectana and Heterorhabditis sp. for field control of insects pests. Nematologica. 27:109–114.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Archives of Phytopathology and Plant Protection
867
Bedding RA. 1984. Large scale production, storage, and transport of insect parasitic nematodes (Neopectana sp. and Heterorhabditis sp.). Annl Appl Biol. 104:117–120. Bedding RA. 1988. Storage of insecticidal nematodes. World Patent No. WO 88/08668. Bedding RA. 1990. Logistics and strategies for introducing entomopathogenic nematode technology into developing countries. In: Gaugler R, Kaya HK, editors. Entomopathogenic nematodes in biological control. Boca Raton (FL): CRC Press. p. 233–248. Belair G, Vincent C, Chouinard G. 1998. Foliar spray with Steinernema carpocapsae against early season apple pests. J Nematol. 30:559–606. Bhatnagar A, Shinde V, Bareth SS. 2004. Evaluation of entomopathogenic nematodes against wegrub, Maladera insanabilis Brenske. Int J Pest Mngt. 50:285–289. Bird AF, Bird J. 1986. Observations on the use of insect parasitic nematodes as means of biological control of root-knot nematodes. Int J Parasitol. 16:511–516. Boemore NE, Boyer-Giglio MH, Thaler JO, Akhurst RJ. 1993. The phages and bacteriocins of Xenorhabdus spp. symbiont of the nematode Steinernema spp. and Heterorhabditis spp. In: Bedding R, Akhurst R, Kaya H, editors. Nematodes and the biological control of insect pests. East Melbourne (VIC). Australia: CSIRO publications. p. 137–145. Buecher EJ, Popiel I. 1989. Liquid cultures of the entomogenous nematode, Steinernema feltiae with its bacterium symbiont. J Nematol. 21:500–504. Buecher EJ, Hansen EL. 1971. Mass culture of axenic nematodes using continuous aeration. J Nematol. 3:199–200. Chen S, Glazer I. 2005. A novel method for long-term storage of the entomopathogenic nematode Steinernema feltiae at room temperature. Biol Cont. 32:104–110. Dindal DL. 1990. Soil biology guide. New York: John Wiley & Sons. p. 1349. Ehlers RU. 2001. Mass production of entomopathogenic nematodes for plant protection. Appl Microbiol Biotechnol. 56:623–633. Ehlers RU, Lunau S, Krasomil-Osterfeld KC, Osterfeld KH. 1998. Liquid culture of the entomopathogenic nematode bacterium complex Heterorhabdtis megidis/Photorhabdus luminescens. Biocontrol. 43:77–86. Ehlers RU, Wulff A, Peters A. 1997. Pathogenicity of axenic Steinernema feltiae, Xenorhabdus bovienii and bactohelminthic complex to larvae of Tipula oleracea and Galleria mellonella. J Invertebrate Pathol. 69:212–217. Friedman MJ. 1990. Commercial production and development. In Gaugler R. Kaya H. editors. Entomopathogenic nematodes in biological control. Boca Raton (FL): CRC Press. p. 153–172. Friedman M, Langston S, Pollitt S. 1989. Mass production in liquid culture of insect killing nematodes. International Patent no. W0 89/04602, US Patent 5,023,183. Gaugler R. 2002. Entomopathogenic nematology. UK: CABI Publications. p. 388. Gaugler R, Bednark A, Campbell JF. 1992. Ultraviolet inactivation of hererorhabditids and Steinernematids. J Invertebr Pathol. 59:155–160. Gaugler R, Brown I. 2001. LOTEK: an automated apparatus for production of insecticidal nematodes. USSN 09/845, 816. Gaugler R, Brown I, David Shapiro-Ilan D, Atwac A. 2002. Automated technology for in vivo mass production of entomopathogenic nematodes. Biol Cont. 24:199–206. Gaugler R, Grewal PS, Kaya HK, Smith-Fiola D. 2000. Quality assessment of commercially produced entomopathogenic nematodes. Biol Cont. 17:100–109. Gaugler R, Han R. 2002. Production technology. In: Gaugler R, editor. Entomopathogenic nematology. USA: CABI Publications. p. 289–310. Georgis R. 1990a. Commercialization of steinernematid and heterorhabditid entomopathogenic nematodes. Proceedings of Brighton Crop Protection Conference Insecticides Fungicides. Vol. 1, Boca Raton (FL): CRC Press. p. 275–280. Georgis R. 1990b. Formulation and application technology. In: Gaugler R, Kaya HK, editors. Entomopathogenic nematodes in biological control. Boca Raton (FL): CRC Press. p. 352. Georgis R, Manweiler S. 1994. Entomopathogenic nematodes: a developing biological control technology. Agri Zool Rev. 6:63–94. Glaser RW. 1932. A pathogenic nematode of the Japanese beetle (Popillia japonica). J Parasitol. 18:119. Glaser RW. 1940. Continued culture of a nematode parasitic in the Japanese beetle. J Parasitol Zool. 84:1.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
868
M.P. Sharma et al.
Glazer I, Gol’Berg A. 1993. Field efficacy of entomopathogenic nematodes against the beetle Maladera matrida (Coleoptera:Scarabaeidae). Biocont Sci Technol. 3:367–376. Glazer I, Navon A. 1990. Activity and persistence of entomopathogenic nematodes tested against Heliothidis armigera. J Econ Entomol. 83:1795–1800. Grewal PS. 2000. Enhanced ambient storage stability of an entomopathogenic nematode through anhydrobiosis. Pest Mangt Sci. 56:401–406. Grewal PS. 2002. Formulation and application technology. In: Gaugler R, editor. Entomopathogenic nematology. USA: CABI Publications. p. 265–287. Grewal PS, Converse V, Georgis R. 1998. Influence of production and bioassay methods on infectivity of two ambush foragers (Nematosa: Steinernematidae). J Invertebr Pathol. 6:262–269. Grewal PS, Georgis R. 1988. Entomopathogenic nematodes. In: Hall FR, Menn JJ, editors. Methods in biotechnology, Vol. 5, Biopesticides: use and delivery. Totowa (NJ): Humana Press Inc. p. 271–297. Grewal PS, Martin WR, Miller RW, Lewis EE. 1997. Suppression of plant-parasitic nematode populations in turfgrass by application of entomopathogenic nematodes. Biocontrol Sci Technol. 7:393–399. Grewal PS, Power KT, Grewal SK, Suggars A, Haupricht S. 2004. Enhanced predictability in biological control of white grubs with new strains of entomopathogenic nematodes. Biol Cont. 30:73–82. Grewal PS, Selvan S, Gaugler R. 1994. Thermal adaptation of entomopathogenic nematodes, niche breadth for infection, establishment, and reproduction. J Thermal Biol. 19:245–253. Grewal PS, Tomolak M, Kiel CBO, Gaugler R. 1993. Evaluation of genetically selected strain of Steinernema feltiae against the mushroom sciarid Lycoriella mali. Ann Appl Biol. 123:695–702. Hara AH, Kaya HK, Gaugler R, Lebeck LM, Mello CL. 1993. Entomopathogenic nematodes for biological control of the leafminor, Liriomyza trifoli. Entomophaga. 38:359–369. Hatab M, Gaugler R. 1997. Growth-mediated variations in fatty acids of Xenorhabdus sp. J Appl Microbiol. 82:351–358. Hatab M, Gaugler R. 2001. Diet composition and lipids of in vitro-produced Heterorhabditis bacteriophora. Biol Contr. 20:1–7. Hominick WM, Briscoe BR, Pino FG del, Heng J, Hunt DJ, Kozodoy E, Mracek Z, Nguyen KB, Reid AP, Spiridonov S, et al. 1997. Biosystematics of entomopathogenic nematodes, current status, protocols and definitions. J Helminthol. 71:271–298. Hudson WG, Frank JH, Castner JL. 1988. Biological control of Scapteriscus spp. mole crickets in Florida. Bull Entomol Soc. 34:192–198. Hussaini SS. 2002. Entomopathogenic nematodes for the control of crop pests. In: Upadhyay RK, editor. Advances in microbial control of insect pests. New York: Kluwer Academic/ Plenum Publishers. p. 265–296. Hussaini SS, Kavitha J, Satya, Hussain MA. 2003. Survival and pathogenicity of indigenous entomopathogenic nematodes in different UV protectants. Indian J Plant Protect. 31: 12–18. Hussaini SS, Kiran Kumar KC, Adholeya A, Shakeela V. 2009. Interaction Effect of Steinernema spp. and Arbuscular Mycorhizal Fungus, Glomus intraradices on the development and reproduction of Meloidogyne incognita (Kofoid and White) in Tomato. Trends in Biosciences 2(1):33–36. Hussaini SS, Singh SP, Parthasarathy R, Shakeela V. 2001. Survival and pathogenicity of ingineous EPN in different UV protectants. National congress on Centenary of Nematology in India – Appraisal and future plan, 5th December. India: IARI; 2001. p. 120–121. Hussaini SS, Singh SP, Shakeela V. 2000. Compatibility of entomopathogenic nematodes (Steinernematidae, Heterorhabditidae: Rhabiditoidea) with selected pesticides and their influence on some biological traits. Entomon. 26:37–44. Hussey RS. 1985. Host-parasite relationships and associated physiological changes. In: Sasser JN, Carter CC, editors. An advanced treatise on meloidogyne. Vol. 1: Biology and control. Raleigh (NC): North Carolina State University. p. 143–154. Ishibashi N. 1993. Integrated control of insects pest by Steinernema carpocapsae. In: Bedding R, Akhurst R, Kaya HK, editors. Nematodes and biological control of insects. East Melbourne, Australia: CSIRO. p. 105–113.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
Archives of Phytopathology and Plant Protection
869
Ishibashi N, Choi DR. 1991. Biological control of soil pests by mixed application of entomopathogenic and fungivorous nematodes. J Nematol. 23:175–181. Kakouli-Duarte T, Labuschagne L, Hague NGM. 1997. Biological control of black vine weevil Otiorhynchus sulcatus with entomopathogenic nematodes. Ann Appl Biol. 131:11–27. Kaya HK. 2002. Natural enemies and other antagonists. In: Gaugler R, editor. Entomopathogenic nematology. Wallingford, UK: CAB International. p. 189–203. Kaya H, Gaugler R. 1993. Entomopathogenic nematodes. Ann Rev Entomol. 38:181–206. Kaya HK, Koppenho¨fer AM, Johnson M. 1999. Natural enemies of entomopathogenic nematodes. Japanese J Nematol. 28:13–21. Kaya HK, Nelson CE. 1985. Encapsulation of Steinernematids and Heterorhabditids nematode with calcium alginate, a new approach for insect control and other applications. Environ Entmol. 14:572–574. Koppenho¨fer AM, Cowles RS, Cowles EA, Fuzy EM, Baumgartner L. 2002. Comparison of neonicotinoid insecticides as synergists for entomopathogenic nematodes. Biol Cont. 24:90–97. Koppenho¨fer AM, Fuzy EM. 2003. Steinernema scarabaei for the control of white grubs. Biol Cont. 28:47–59. Koppenho¨fer AM, Grewal PS, Kaya HK. 2000. Synergism of imidacloprid and entomopathogenic nematodes against white grub: the mechanism. Entomol Exp Et Appl. 94:283– 294. Koppenho¨fer AM, Kaya HK. 1997. Additive and synergistic interaction between entomopathogenic nematodes and Bacillus thuringiensis for scarab grub control. Biol Cont. 8:131– 137. Lacey LA, Unruh TR. 1998. Entomopathogenic nemtodes for control of codling moth, Cydia pomonella: effect of nematode species, concentration, temperature, and humidity. Biol Cont. 13:1–8. Lello ER, Patel MN, Mathews GA, Wright DJ. 1996. Application technology for entomopathogenic nematodes against foliar pests. Crop Prot. 15:567–574. Lewis EE, Shapiro-Ilan DI. 2002. Host cadavers protect entomopathogenic nematodes during freezing. J Invertebr Pathol. 81:25–32. Lindegren JE, Valero KA, Mackey BE. 1993. Simple in vivo production and storage methods for Steinernema carpocapsae infective juveniles. J Nematol. 25:193–197. Lindegren JE, Wong TT, Mclnnis DO. 1990. Response of Mediterranean fruit fly to the entomogenous nematode Steinernema feltiae in fields test in Hawaii. Environ Entomol. 19:383–386. Miklasiewicz TJ, Grewal PS, Hoy CW, Malik VS. 2002. Evaluation of entomopathogenic nematodes for suppression of carrot weevil. Biocontrol. 47:545–561. Miller RW. 1989. Novel pathogenicity assessment technique for Steinernema and Heterorhabditis entomopathogenic nematodes. J Nematol. 21:574. Mracek Z. 2002. Use of entomoparasitic nematodes (EPANS) in biological control. In: Upadhyay RK, editor. Advances in microbial control of insect pests. (NY): Kluwer Academic/Plenum Publishers. p. 235–264. Nishimatsu T, Jackson JJ. 1998. Interaction of insecticides, entomopathogenic nematodes, and larvae of the western corn rootworm. J Econ Entomol. 91:410–418. Pace GW, Grote W, Pitt DE, Pitt JM. 1986. Liquid cultures of nematodes. International Patent no. W0 86/011074. Parkman JP, Frank JH, Nguyen KB, Smart GC, Jr. 1994. Inoculative release of Steinernema scapterisci to suppress pest mole crickets on golf courses. Environ Entomol. 23:1331–1337. Perez EE, Lewis EE. 2002. Use of entomopathogenic nematodes to suppress Meloidogyne incognita on greenhouse tomatoes. J Nematol. 34:171–174. Perez EE, Lewis EE. 2004. Suppression of Meloidogyne incognita and Meloidogyne hapla with entomopathogenic nematodes on greenhouse peanuts and tomatoes. Biol Cont. 30:336– 341. Peters A, Ehlers RU. 1994. Susceptibility of leatherjackets (Tipula paludosa and T. oleracea) to the entomopathogenic nematode, Steinernema feltiae. J Invertebra Pathol. 63:163–171. Poinar GO, Jr. 1966. The presence of Achromobactor nematophilus in the infective stage of Neoaplectana sp. Nematologica. 12:105–108.
Downloaded By: [Sharma, Mahaveer P.][Consortium for e-Resources in Agriculture] At: 09:04 10 June 2011
870
M.P. Sharma et al.
Poinar GO, Jr. 1990. Biology and taxonomy of Steinernematidae and Heterorhabditidae. In: Gaugler R, Kaya HK, editors. Entomopathogenic nematodes in biological control. Boca Raton (FL): CRC Press. p. 23–61. Poinar GO, Jr, Thomas GM. 1966. Significance of Achromobactor nematophilus in the development of the nematode, DD-136 (Neoplectana sp. Steinernematidae). Parasitol. 56:386. Rasmann S, Ko¨llner TG, Degenhardt J, Hiltpold I, Toepfer S, Kuhlmann U, Gershenzon J, Turlings TCJ. 2005. Recruitment of entomopathogenic nematodes by insect-damaged maize roots. Nature. 434:732–737. Richardson PN. 1996. British and European legislation regulating rhabditid nematodes. Biocontr Sci Technol. 6:449–463. Richardson PN, Grewal PS. 1991. Comparative assessment of biological and chemical methods of control for the mushroom fly Lycoriella auripila. Biocontrol Sci. 1:217–228. Schroer S, Ehlers RU. 2005. Foliar application of the entomopathogenic nematode Steinernema carpocapsae for biological control of diamondback moth larvae (Plutella xylostella). Biol Cont. 3:81–86. Selvan SP, Grewal PS, Gaugler R, Tomalak M. 1994. Evaluation of steinernematid nematodes against Popillia japonica: species, strain and rinse after application. J Econ Entomol. 89:605–609. Shapiro-llan D, Gouge D, Koppenhofer A. 2002. Factors affecting commercial success: case studies in cotton, turf, and citrus. In: Gaugler R, editor. Entomopathogenic nematology. CABI Publications. p. 333–355. Silver SC, Dunlop DB, Grove DI. 1995. Granular formulations of biological entities with improved storage stability. World Patent No. WO 95/0577. Singh SP. 1977. Isolation of an entomophilic nematode from potato cutworms. Curr Sci. 46:454–455. Sitaramaiah S, Gunneswara RS, Hussaini SS, Venkateswarlu P, Nageswara Rao S. 2003. Use of entomopathogenic nematode Steinernema carpocapsae against Spodoptera. In: Tandon PL, Balal CR, Jalali SK, Rabindra RJ, editors. Biocontrol of lepidopterous pests. Bangalore, India: Project Directorate of Biological Control. p. 211–213. Sivakumar CV, Jayaraj S, Subramaniam S. 1989. Observations on Indian population of the entomopathogenic nematode Heterorhabditis bacteriophora. J Biol Cont. 2:112–113. Smitley DR, Warner FW, Bird GW. 1992. Influence of irrigation and Heterorhabiditis bacteriophora on plant-parasitic nematodes in turf. J Nematol. 24:637–641. Stoll NR. 1952. Axenic cultivation of the parasitic nematode, Neoaplectana glaseri, in a fluid medium containing raw liver extract. J Parasitol. 39:422–444. Stuart RJ, Shapiro-Ilan DI, James RR, Nguyen KB, McCoy CW. 2004. Virulence of new and mixed strains of the entomopathogenic nematode Steinernema riobrave to larvae of the citrus root weevil Diaprepes abbreviatus. Biol Cont. 30:439–445. Sulistyanto D, Ehlers RU. 1996. Efficay of the entomopathogenic nematodes Heterorhabtidis magidis and H. bacteriophora for the control of grubs, Phyllopertha horticola and Aphodius contaminatus in golf course turf. Biocont Sci Technol. 6:247–250. Tomalak M. 1994. Selective breeding of Steinernema feltiae for improved efficacy in control of a mushroom fly Lycoriella solani Winn. BioControl. 4:187–198. Tsai BY, Yeh H. 1995. Effect of Steinernema carpocapsae Weiser on the infectivity of Pratylenchus coffeae (Zimmermann) Filipjev and Schuurmans Stekhoven and Meloidogyne javanica (Treub) Chitwood. Plant Pathol Bull. 4:106–110. White GF. 1927. A method for obtaining infective nematode larvae from cultures. Science. 66:302–303. Williams EC, Walters KFA. 2000. Foliar application of the entomopathogenic nematodes Steinernema feltiae against leafminers on vegetables. Bicontrol Sci Technol. 10:61–70. Wilson M, Ivanova E. 2004. Neutral density liquid formulations for nematode-based biopesticides. Biotech Lett. 26:1167–1171. XuJie, Lian Ping Y, Xie RC. 2000. Studies on the applications of some entomopathogenic nematodes against litchi lyponomeutid Comoritis albicapilla M. Acta Phytophol. 27:27–31. Yoo S, Brown I, Gaugler R. 2000. Liquid media development for Heterorhabditis bacteriophora: lipid source and concentration. Appl Microbiol Biotechnol. 54:759–763. Zimmerman RJ, Cranshaw WS. 1990. Compatibility of three entomogenous nematodes in aqueous solutions of pesticides used in turfgrass maintenance. J Econ Entomol. 83:97–100.
View publication stats
