Critical factors required by the nematode Steinernema feltiae for the control of the leafminers Liriomyza huidobrensis, Liriomyza bryoniae and Chromatomyia syngenesiae

The critical factors required by entomopathogenic nematodes for the control of dipteran leafminers were investigated and compared. The foliar application of Steinernema feltiae and Heterorhabditis sp. (strain UK 211) caused a significant reduction in the numbers of dipteran leafminer larvae. There was no significant difference in efficacy between these nematode species against Liriomyza huidobrensis at 20°C and 85–90% r.h. Application of S. feltiae to all three larval instars of L. huidobrensis significantly reduced larval survival, with application to the second instar being the most effective at 20 ± 2°C and 80 ± 10% r.h. Humidities of >90% significantly enhanced the efficacy of S. feltiae. When S. feltiae was applied to second instar larvae, it was equally effective throughout the temperature range 10–30°C. At 20°C and >90% r.h., sufficient nematodes were able to enter the leaf tissues within the first 12 h after application to reduce larval survival to c. 15%. A comparison between L. huidobrensis, L. bryoniue and Chromatomyia syngenesiue indicated that the efficacy of S. feltiae was affected by the same critical factors for all three species.     

Controlling the sugar beet fly Pegomyia mixta Vill. with entomopathogenic nematodes

Sugar beet, Beta vulgaris L. is a strategic crop of sugar industry in Egypt. It is threatened by several insect pests among most important of them is the beet fly Pegomyia mixta. This work deals with the biological control of this insect using four entomopathogenic nematodes (EPNs). The nematodes included Steinernema carpocapsae S2, Steinernema feltiae, Heterorhabditis bacteriophora (HB1-3) and Heterorhabditis bacteriophora S1. Daily mortality of larvae and pupae of P. mixta were recorded after treatment with serial concentrations (500, 1000, 2000 and 4000 infective juveniles (IJs)/ml) of each of four studied EPNs. In the laboratory all tested nematodes killed the larvae inside their mines in the sugar beet leaves and developed in their bodies in different extends. They also killed the insect pupae in the soil and developed in their bodies. Young larvae were more susceptible than old ones. New pupae were more susceptible than old ones. In the field a single spray of S. feltiae or H. bacteriophora caused 81.3 or 75.9% reduction in the larval population of the in sugar beet leaves.     

Multiple-species natural enemy approach for biological control of alfalfa snout beetle (Coleoptera: Curculionidae) using entomopathogenic nematodes

Multiple-species natural enemy approach for the biological control of the alfalfa snout beetle, Otiorhynchus ligustici (L.) (Coleoptera: Curculionidae), was compared with using single-species of natural enemies in the alfalfa ecosystem by using entomopathogenic nematodes with different dispersal and foraging behaviors. Steinernema carpocapsae NY001 (ambush nematode), Heterorhabditis bacteriophora Oswego (cruiser nematode), and Steinernema feltiae Valko (intermediate nematode) were applied in single-species, two-species combinations, and one three-species combination treatments at 2.5 x 10(9) infective juveniles per hectare. All nematode species persisted for a full year (357 d). S. carpocapsae NY001 protected the plants from root-feeding damage better than H. bacteriophora Oswego but allowed for higher larval survival than all other nematode treatments. S. feltiae Valko protected the plants better than H. bacteriophora Oswego and controlled alfalfa snout beetle larvae better than S. carpocapsae NY001. H. bacteriophora Oswego allowed for similar root damage compared with control plots but reduced larval populations better than S. carpocapsae NY001. The combination of S. carpocapsae NY001 and H. bacteriophora Oswego provided significantly better protection for the plants than the control (unlike H. bacteriophora Oswego alone) and reduced host larva survival more than S. carpocapsae NY001 alone. The combination S. feltiae Valko and H. bacteriophora Oswego could not be statistically separated from the performance of S. feltiae Valko applied alone.     

Interaction of microbial populations in Steinernema (Steinernematidae,Nematoda) infected Galleria mellonella larvae

Infection of Galleria mellonella larvae with the entomopathogenic nematodes Steinernema feltiae (A21 and R strains) and Steinernema glaseri (Dongrae) resulted in several species of bacteria, including the respective bacterial symbiont, Xenorhabdus spp., growing in the infected insect cadavers. These other bacteria were Enterococcus in all three nematode infections studied and Acinetobacter in the S. feltiae infections. The respective populations of these bacteria changed with time. Following infection of G. mellonella larvae with any one of the Steinernema sp., only Enterococcus bacteria were detected initially in the dead larvae. Between 30 and 50h post-infection Xenorhabdus bacteria were detected and concurrent with this Enterococcus population declined to zero. This was probably due to secondary metabolites with antibacterial properties that were produced by Xenorhabdus. In the S. feltiae (both R and A21 strains) infections a third bacterium, Acinetobacter, appeared at about 130h (in S. feltiae A21 infections) or 100h (in S. feltiae R infections) and increased in population size to approximately that of Xenorhabdus. It was demonstrated that Enterococcus, orginating from the G. mellonella digestive tract, was sensitive to the organically soluble antimicrobials produced by Xenorhabdus but Acinetobacter, which was carried by the nematode, was not.     

Laboratory Evaluation of Seven Pakistani Strains of Entomopathogenic Nematodes Against a Stored Grain Insect Pest, Pulse beetle Callosobruchus chinensis (L.)

Seven Pakistani strains of entomopathogenic nematodes belonging to the genera Steinernema and Heterorhabditis were tested against last instar and adult stages of the pulse beetle, Callosobruchus chinensis (L.). These nematodes included Steinernema pakistanense Shahina, Anis, Reid and Maqbool (Ham 10 strain); S. asiaticum Anis, Shahina, Reid and Rowe (211 strain); S. abbasi Elawad, Ahmad and Reid (507 strain); S. siamkayai Stock, Somsook and Reid (157 strain); S. feltiae Filipjev (A05 strains); Heterorhabditis bacteriophora Poinar (1743 strain); and H. indica Poinar, Karunakar and David (HAM-64 strain). Activity of all strains was determined at four different nematode densities in Petri dishes and in concrete containers. A significant nematode density effect was detected for all nematode species tested. Overall, Heterorhabditis bacteriophora, S. siamkayai, and S. pakistanense were among those that showed the highest virulence to pulse beetle larvae and adults. For all nematode species, the last larval stage of the pulse beetle seems to be more susceptible than the adult. LC(50) values in Petri dish and concrete containers were 14-340 IJs/larvae and 41-441 IJs/larvae, respectively, and 59-1376 IJs/adult and 170-684/adult, respectively.     

Susceptibility of Sap Beetles (Coleoptera: Nitidulidae) to Entomopathogenic Nematodes

Nitidulid beetles (Coleoptera) are considered to be serious pests of date palms throughout the world. They attack ripe fruit, causing it to rot, and damage is reflected in both reduced yield and lower fruit quality. The present study was aimed at an evaluation of the susceptibility to different sap beetles to entomopathogenic nematodes. We further tested nematode efficacy in pots filled with soil infested by third instar larvae of the two beetle species. In Petri dish assay, mortality levels of Carpophilus humeralis and C. hemipterus exposed to Heterorhabditis sp. IS-5 strain indicated that the latter is less susceptible to nematode infection. Exposure of both sap beetle species to different nematode strains gave moderate levels of mortality (35-65%) with the heterorhabditid strains HP88, IS-5 and IS-25. The IS-12 strain of Heterorhabditis sp. showed poor virulence (<35% mortality) against larvae of C. humeralis as well as larvae and pupae of C. humipterus. The nematode species S. riobrave showed moderate virulence (35-65%) mortality to larvae and pupae of S. humeralis as well as to larvae of C. hemipterus . Exposure of C. hemeralis to different concentrations of Heterorhabditis sp. IS-5 in pots containing soil resulted in high mortality (>65%). In contrast, the lower concentrations (500 and 1000 nematodes/pot) caused low mortality (35%) of C. hemipterus . Other heterorhabditid strains caused 95-100% mortality of C. humeralis in pot assay. The HP88 strain of H. bacteriophora and the Tx strain of Steinernema riobrave showed poor effectiveness. Incubation of different nematode strains with the C. humeralis larvae at high temperature (32 C) resulted in an increase in insect mortality with the IS-12 and IS-21 strains. Reduced mortality was recorded with the HP88 strain treatment at the higher temperature. The IS-5 and IS-12 strains were equally effective in all three soil types tested, whereas the IS-19 strain was more effective in the Almog type soil than in the others.     

Screening of entomopathogenic nematodes for virulence against the invasive western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae) in Europe

Entomopathogenic nematode species available in Europe were screened for their efficacy against both the root-feeding larvae and silk-feeding adults of the western corn rootworm, Diabrotica virgifera virgifera LeConte. Laboratory screening tests were aimed at the selection of candidate biological control agents for the management of this invasive alien pest in Europe. Steinernema glaseri, S. arenarium, S. abassi, S. bicornutum, S. feltiae, S. kraussei, S. carpocapsae and Heterorhabditis bacteriophora were studied to determine their virulence against third instar larvae and adults of D. v. virgifera in small-volume arenas (using nematode concentrations of 0.5, 0.8, 7.9 and 15.9 infective juveniles cm-2). All nematode species were able to invade and propagate in D. v. virgifera larvae, but adults were rarely infected. At concentrations of 7.9 and 15.9 cm-2, S. glaseri, S. arenarium, S. abassi and H. bacteriophora caused the highest larval mortality of up to 77%. Steinernema bicornutum, S. abassi, S. carpocapsae and H. bacteriophora appeared to have a high propagation level, producing 5970+/-779, 5595+/-811, 5341+/-1177 and 4039+/-1025 infective juveniles per larva, respectively. Steinernema glaseri, S. arenarium, S. feltiae, S. kraussei and H. bacteriophora were further screened at a concentration of 16.7 nematodes cm-2 against third instar larvae in medium-volume arenas (sand-filled trays with maize plants). Heterorhabditis bacteriophora, S. arenarium and S. feltiae caused the highest larval mortality with 77+/-16.6%, 67+/-3.5%, and 57+/-17.1%, respectively. In a next step, criteria for rating the entomopathogenic nematode species were applied based on results obtained for virulence and propagation, and for current production costs and availability in Europe. These criteria were then rated to determine the potential of the nematodes for further field testing. Results showed the highest potential in H. bacteriophora, followed by S. arenarium and S. feltiae, for further testing as candidate biological control agents.     

Foliar Application of the Entomopathogenic Nematode Steinernema feltiae Against Leafminers on Vegetables

Data from a comparative study of the efficacy of Steinernema feltiae for the control of three species of leafminer formed the basis of an application schedule which successfully suppressed an outbreak of the statutory leafminer pest, Liriomyza huidobrensis . All three instars of Liriomyza bryoniae and L. huidobrensis were similarly susceptible to S. feltiae at 20 o C and > 90% relative humidity (RH). Although all larval instars of Chromatomyia syngenesiae were susceptible to S. feltiae , mortality was lower than for L. bryoniae . Repeat applications of S. feltiae to L. bryoniae and C. syngenesiae indicated that a nematode treatment to the second/early third instar larvae was more effective than applying higher rates of nematodes when humidities were less than 90% RH. In a trial on lettuce at a commercial glasshouse a mean L. huidobrensis mortality of 82 +/- 5% was recorded after an S. feltiae application, significantly higher than the chemical treatment, heptenophos.     

Host range, specificity, and virulence of Steinernema feltiae, Steinernema rarum, and Heterorhabditis bacteriophora (Steinernematidae and Heterorhabditidae) from Argentina

Infections were carried out in the laboratory to determine the host range, specificity, and virulence of Steinernema rarum, S. feltiae, and Heterorhabditis bacteriophora that were isolated from different regions of Argentina. All insect orders showed a remarkable susceptibility to the three nematode species, showing mortality values higher than 66%, except for Anoplura. The mortality of the insects of agronomic interest was more than 90%. As for insects of sanitary importance, S. feltiae and S. rarum caused 60% of mortality, whereas H. bacteriophora caused 80%. The results fluctuated when considering the buccal apparatus function of the insects. Nematodes completed their cycle in anoplurans, orthopterans, lepidopterans, and hymenopterans, while the development varied in coleopterans, homopterans, hemipterans, and dipterans. S. rarum developed completely both in immature and adult, while S. feltiae and H. bacteriophora developed preferably in immature. The three isolates are capable of parasitizing a wide host range under laboratory conditions; H. bacteriophora is more virulent than the evaluated Steinernema spp. against Galleria mellonella larvae; the pathogenicity and specificity depend on the bioecological characteristics of nematodes and hosts.     

病原线虫对桔小实蝇种群的控制作用

通过室内和田间实验研究了昆虫病原线虫对桔小实蝇Bactrocera (Bactrocera) dorsalis (Hendel)的控制作用。室内实验结果表明,供试的3种线虫的4个品系（小卷蛾斯氏线虫Steinernema carpocapsae All品系与A24品系,夜蛾斯氏线虫Steinernema feltiae SN品系和嗜菌异小杆线虫Heterorhabditis bacteriophora H06品系）,以小卷蛾斯氏线虫All品系对桔小实蝇的侵染力最强,其3天的LD₅₀和LD₉₅分别为35.0和257.1条/cm²土壤。按300条/cm²土壤的量施用,小卷蛾斯氏线虫All品系对当代桔小实蝇的控制效果为86.3%。用以虫期作用因子组建的生命表方法评价了小卷蛾斯氏线虫All品系对田间桔小实蝇下代种群的控制作用,结果表明,按300条/cm²土壤的量施用线虫,对照杨桃园的桔小实蝇种群趋势指数为105.9,而处理杨桃园的桔小实蝇种群趋势指数下降为15.5;小卷蛾斯氏线虫All品系对桔小实蝇的干扰控制指数为0.146,即线虫处理果园的下代种群密度仅为对照果园的14.6%。     

Effectiveness of different species of entomopathogenic nematodes for biocontrol of the Mediterranean flatheaded rootborer, Capnodis tenebrionis (Linné) (Coleoptera: Buprestidae) in potted peach tree

The susceptibility of larvae of the Mediterranean flatheaded rootborer (Capnodis tenebrionis) to 13 isolates of entomopathogenic nematodes was examined using GF-677 potted trees (peachxalmond hybrid) as the host plant. The nematode strains tested included nine Steinernema feltiae, one S. affine, one S. carpocapsae and two Heterorhabditis bacteriophora. Nematodes showed the ability to locate and kill larvae of C. tenebrionis just after they enter into the roots of the tree. S. feltiae strains provided an efficacy ranging from 79.68% to 88.24%. H. bacteriophora strains resulted in control of 71.66-76.47%. S. carpocapsae (B14) and S. affine (Gspe3) caused lower control of C. tenebrionis larvae (62.03% and 34.76%, respectively). The influence of foraging strategy and the use of autochthonous nematodes to control C. tenebrionis larvae inside the roots is discussed.     

Assessment of temperature effects on the development and fecundity of Pullus mediterraneus (Col., Coccinellidae) and consumption of Saissetia oleae eggs (Hom., Coccoida)

Eggs, larval and nymphal periods and fecundity of Pullus mediterraneus were examined under 16 h light : 8 h dark combined with six constant temperatures: 15, 20, 25, 30, 35 and 40°C. Eggs of Saissetia oleae were used as prey. The developmental time at 15, 20, 25, 30 and 35°C was 17.23, 4.5, 2.64, 1.67, 1.28 days for eggs and 98.47, 68.88, 53.94, 28.96, 36.51 days for larval–pupal duration, respectively. At 7°C no eggs hatched, and at 40°C all the stages died after 36 h of maximum exposure except the three last stages. The fecundity of females rearing at different temperatures ranged between 1.7 eggs at 15°C and 601.86 eggs at 30°C. The pre-oviposition period ranged between 23.75 days at 15°C and 3.47 days at 35°C. The consumption of S. oleae eggs by the larvae reached 597.69 eggs during the pre-imaginal development. Females attacked more eggs than males averaging 77.69 ± 22.34 eggs per 4 day period compared with 46.97 ± 10.12 eggs per 4 day period for males.     

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

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

Impact of Entomopathogenic Nematodes on Different Soil-dwelling Stages of Western Flower Thrips, Frankliniella occidentalis (Thysanoptera: Thripidae), in the Laboratory and Under Semi-field Conditions

The impact of entomopathogenic nematodes (EPN) on mortality of soil-dwelling stages of western flower thrips (WFT), Frankliniella occidentalis (Thysanoptera: Thripidae) with different insect stage combinations was studied in the laboratory and under semi-field conditions. In laboratory experiments, the efficacy of Steinernema feltiae strain Sylt (Rhabditida: Steinernematidae) at a concentration of 400 infective juveniles (IJs) cm -2 was tested against different proportions of soil-dwelling stages of WFT, i.e. late second instar larvae (L2), prepupae and pupae. Soil was used as the testing medium. S. feltiae significantly affected the mortality of all soil-dwelling life stages of WFT at all tested insect stage combinations. The proportion of late L2 in the population negatively correlated to EPN-induced mortality. WFT prepupa and pupa were similarly susceptible to S. feltiae and their proportion in the population did not affect the EPN-induced mortality under laboratory conditions. The highest mortality (80%) was recorded when the population consisted only of prepupae and/or pupae. In the semi-field study, the impact of S. feltiae , S. carpocapsae strain DD136 and Heterorhabditis bacteriophora strain HK3 (Rhabditida: Heterorhabditidae) ( H. bacteriophora ) at concentrations of 400 and 1000 IJs cm -2 was evaluated against WFT reared on green beans, Phaseolus vulgaris L., as host plant in pot experiments in a controlled climate chamber. All tested EPN strains at both dose rates significantly reduced the WFT populations. Up to 70% reduction of the WFT population was obtained at the higher EPN concentration.     

Cold shock and cold tolerance in larvae and pupae of the blow fly, Lucilia sericata

Abstract.  Understanding the effects of low winter temperatures on mortality is essential in the development of a full understanding of the long-term population dynamics of any insect. The present study aims to examine the survival of pupae and larvae of the blow fly, Lucilia sericata , at overwintering temperatures. Groups of pupae and diapausing and nondiapausing third-stage larvae of L. sericata are maintained in cooled incubators at either 3 °C and 6 °C. Groups are removed from the incubators at 3–4-day intervals and transferred either to−8 °C or to 25 °C. After 1 h in the freezer, the larvae and pupae exposed to this cold-shock are also transferred to 25 °C. Larvae and pupae are then allowed to continue development and the number of adults emerging from each group is counted. The results demonstrate that survival decreases linearly with the period of exposure at both 3 °C and 6 °C. Mortality is higher at 3 °C than at 6 °C and, in groups that receive the cold shock, cold-shock reduces emergence by over 50%. However, there is no consistent tendency for diapausing larvae to survive prolonged cold or cold shock better than other life-cycle stages. The results suggest that the facultative development of an overwintering diapause stage in L. sericata does not appear to be an adaptation to enhance cold tolerance or resistance to cold shock. It is concluded that the survival of overwintering L. sericata is likely to be relatively less affected by low temperatures than it is by, for example, biotic factors, particularly given the buffered soil environment and short time-scales over which periods of cold act.     

Efficacy of entomopathogenic nematode Steinernema feltiae (Rhabditida: Steinernematidae) as influenced by Frankliniella occidentalis (Thysanoptera: Thripidae) developmental stage and host plant stage

Entomopathogenic nematodes were investigated as an alternative biological control strategy for western flower thrips, Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), in ornamental greenhouse crops, by using potted chrysanthemum as a model crop. The susceptibility of various life stages of F. occidentalis to different concentrations of the nematode Steinernema feltiae (Filipjev) (Rhabditida: Steinernematidae) was investigated in petri dish bioassays. This was followed with trials using potted chrysanthemums comparing the efficacy of nematode application to plants in vegetative (exposed habitat) versus flowering (cryptic habitat) stages. In both trials, the effect of the wetting agent Agral 90 (nonylphenoxy polyethoxyethanol), which is used in combination with the nematode spray, on F. occidentalis mortality was assessed. In petri dish trials, the prepupae and pupae were the most susceptible developmental stages of F. occidentalis to infection by S. feltiae. First and second instars were killed by very high rates of nematodes (> or =20,000 infective juveniles per milliliter), but corrected mortality was only approximately 28-37%. No significant mortality was observed for adult thrips. Results from the petri dish trials were confirmed on chrysanthemum plants. Foliar application of S. feltiae did not result in significant mortality in larvae or adults. No significant differences in efficacy were detected by application of nematodes on vegetative versus flowering chrysanthemum. Agral 90 had a significant impact on mortality on the first stage larvae and prepupae in the petri dish trials but not in the plant trials. Thrips control by S. feltiae in greenhouses may be partly or completely due to prepupal and pupal mortality.     

Comparison of Bioassays to Measure Virulence of Different Entomopathogenic Nematodes

Five bioassays were compared for their usefulness to determine the virulence of four nematode strains. The objective of this study was to develop standard assays for particular nematode species. In all assays, the nematodes Steinernema feltiae (strain UK), S. riobravis, S. scapterisci Argentina and Heterorhabditis bacteriophora HP88 were exposed to Galleria mellonella larvae. All bioassays except the sand column assay were conducted in multi-well plastic dishes. In the penetration rate assay, the number of individual nematodes invading the insect was determined after a 48-h exposure to 200 infective juveniles (IJs). In the one-on-one assay, each larva was exposed to an individual nematode for 72 h before insect mortality was recorded. In the exposure time assay, insect mortality was recorded after exposure to 200 IJs for variable time periods. The dose-response assay involved exposing larvae to different nematode concentrations over the range 1-200 IJs/insect and recording mortality every 24 h for a 96-h period. In the sand columns assay, insects were placed in the bottom of a plastic cylinder filled with sand. Nematodes were applied on top of the sand and insect mortality was determined after IJs had migrated through the cylinder. The highest mortality level in the sand column assay was obtained with IJs of S. feltiae followed by H. bacteriophora; treatments with S. riobravis and S. scapterisci produced low levels of insect mortality. In the other four assays, S riobravis was the most virulent, followed by S. feltiae, H. bacteriophora and S. scapterisci. In the exposure time assay, rapid mortality was achieved when the insects were exposed to S. feltiae and S. riobravis. For these nematode species, a gradual increase in the number of individuals which penetrated into cadavers was recorded. Conversely, the number of nematodes in the cadavers of insects infected by H. bacteriophora and S. scapterisci remained low during the entire exposure period. In this assay, exposing the insects to these nematodes resulted in a gradual increase in mortality. In the dose-response assay, complete separation among nematode species was obtained only after 48 h of incubation at a concentration of 15 IJs/insect. LD and LD values were calculated from 50 90 dose-response assay data. However, these values did not indicate differences among the different nematode species. The present study demonstrated the variation in entomopathogenic nematode performance in different bioassays and supports the notion that one common bioassay cannot be used as a universal measure of virulence for all species and strains because nematodes differ in their behavior. Furthermore, particular assays should be used for different purposes. To select a specific population for use against a particular insect, assays that are more laborious but which simulate natural environmental conditions (e.g. the sand column assay) or invasion by the nematode (e.g. the penetration rate assay) should be considered. In cases where commercial production batches of the same nematode strains are compared, simple and fast assays are needed (e.g. the one-on-one and exposure time assays). Further studies are needed to determine the relationships between data obtained in each assay and nematode efficacy in the field.     

Factors affecting entomopathogenic nematodes (Steinernematidae) for control of overwintering codling moth (Lepidoptera: Tortricidae) in fruit bins

Fruit bins infested with diapausing codling moth larvae, Cydia pomonella (L.), are a potential source of reinfestation of orchards and may jeopardize the success of mating disruption programs and other control strategies. Entomopathogenic nematodes (EPNs) were tested as a potential means of control that could be applied at the time bins are submerged in dump tanks. Diapausing cocooned codling moth larvae in miniature fruit bins were highly susceptible to infective juveniles (IJs) of Steinernema carpocapsae (Weiser) and Steinernema feltiae (Filipjev) in a series of experiments. Cocooned larvae are significantly more susceptible to infection than are pupae. Experimental treatment of bins in suspensions of laboratory produced S. feltiae ranging from 10 to 100 IJs/ml of water with wetting agent (Silwet L77) resulted in 51-92% mortality. The use of adjuvants to increase penetration of hibernacula and retard desiccation of S. feltiae in fruit bins resulted in improved efficacy. The combination of a wetting agent (Silwet L77) and humectant (Stockosorb) with 10 S. feltiae IJs/ml in low and high humidity resulted in 92-95% mortality of cocooned codling moth larvae versus 46-57% mortality at the same IJ concentration without adjuvants. Immersion of infested bins in suspensions of commercially produced nematodes ranging from 10 to 50 IJs/ml water with wetting agent in an experimental packing line resulted in mortality in cocooned codling moth larvae of 45-87 and 56 - 85% for S. feltiae and S. carpocapsae, respectively. Our results indicate that EPNs provide an alternative nonchemical means of control that could be applied at the time bins are submerged in dump tanks at the packing house for flotation of fruit.     

The influence of humidity on the effect of Steinernema feltiae against diapausing codling moth larvae (Cydia pomonella L.) (Lepidoptera: Tortricidae)

Codling moth (CM) is a serious and global pest of pome fruit. It overwinters in cryptic habitats as cocooned diapausing larvae. Field trials with the entomopathogenic nematode Steinernema feltiae (Rhabditida: Steinernematidae) report control of diapausing CM of up to 70%, but results are variable. The objective of this study was to define environmental conditions favouring the performance of the nematodes. Cocooned larvae were more susceptible than non-cocooned larvae. S. feltiae was unable to infect CM at a water activity (aw-values) < or = 0.9. Mortality of cocooned larvae was reported at lower aw-values than of non-cocooned larvae. Exposure time and impact of external relative humidity (RH) was studied. Mortality of cocooned larvae did not further increase after half an hour of exposure, whereas the mortality increased with increasing exposure time in non-cocooned larvae. LC50 and LC90 considerably decreased with increasing RH. The influence of the relative humidity was less pronounced when surpassing 80% than the effect of the volume of applied water. When S. feltiae was formulated in a surfactant-polymer-formulation (SPF), mortality significantly increased when compared to application in water.     

Biocontrol Potential of Entomopathogenic Nematodes Against Winter Moths (Operophtera brumata and O. fagata) (Lepidoptera: Geometridae) Infesting Urban Trees

Infectivity and biocontrol potential of entomopathogenic nematodes against winter moths (Operophtera brumata and O. fagata)pupating in the soil were examined in laboratory, semi-field and field conditions. A pilot experiment conducted in the field showed that Steinernema feltiae was completely ineffective against pupae of these moths in the soil. Subsequent laboratory tests revealed that none of the tested species (i.e. S. feltiae, S. affinae, S. carpocapsae, Heterorhabditis megidis and H. bacteriophora) could colonise the pupae, while mature larvae descending to the soil for pupation and prepupae were highly susceptible to nematode infection. No differences were observed between O. brumata and O. fagata in susceptibility to nematodes. In laboratory experiments H. megidis applied at 1.5×10⁵infective juveniles (IJ) m^-2infected almost 100% of insects exposed for 6 days in the soil. It was significantly more infective than H. bacteriophora (73-77%) and Steinernema species (29-50%). H. megidis was also highly effective in semi-field conditions when applied at an even lower dose, i.e. 10⁵IJ m^-2. After a 45-day experiment, only 3% of insects descending for pupation survived in the soil pre-treated with this species. This was significantly less than in soil with S. feltiae (43%) and control treated with water only (59%). Very high efficacy of H. megidis and a relatively easy method for its field application through ground spraying gives some promise for environmentally safe and successful biological control of winter moths during their pupation in the soil. The low application rate required and recycling in the host could be additional advantages for economic and long lasting protection of high value trees, particularly those in urban parks and forests.