Use of Cellulose Acetate Electrophoresis in the Taxonomy of Steinernematids (Rhabditida,Nematoda)

A steinernematid nematode was isolated from soil samples collected near St. John''s, Newfoundland, Canada. On the basis of its morphometry and RFLPs in ribosomal DNA spacer, it was designated as a new strain, NF, of Steinernema feltiae. Cellulose acetate electrophoresis was used to separate isozymes of eight enzymes in infective juveniles of S. feltiae NF as well as four other isolates: S. feltiae Umeå strain, S. feltiae L1C strain, Steinernema carpocapsae All strain, and Steinernema riobravis TX strain. Based on comparisons of the relative electrophoretic mobilities (μ) of the isozymes, one of the eight enzymes (arginine kinase) yielded zymograms that were distinctive for each of the isolates, except for the Umeå and NF strains of S. feltiae, which had identical banding patterns. Four enzymes (fumarate hydratase, phosphoglucoisomerase, phosphoglucomutase, and 6-phosphogluconate dehydrogenase) yielded isozyme banding patterns that were characteristic for all isolates, except for the L1C and NF strains of S. feltiae, which were identical. Two enzymes (aspartate amino transferase and glycerol-3-phosphate dehydrogenase) yielded zymograms that permitted S. carpocapsae All strain to be discriminated from the other four isolates, while the remaining enzyme (mannose-6-phosphate isomerase) was discriminatory for S. riobravis TX strain. Except for one enzyme, the isozyme banding pattern of the NF isolate of S. feltiae was the same as in the L1C strain, isolated 13 years previously from Newfoundland. Cellulose acetate electrophoresis could prove invaluable for taxonomic identification of isolates of steinernematids, provided that a combination of enzymes is used.     

Variations in Immune Response of Popillia japonica and Acheta domesticus to Heterorhabditis bacteriophora and Steinernema Species

The infectivities of Steinernema carpocapsae, S. glaseri, S. scapterisci, and Heterorhabditis bacteriophora to Japanese beetle larvae, Popillia japonica, and house cricket adults, Acheta domesticus, were compared using external exposure and hemocoelic injection. Only H. bacteriophora and S. glaseri caused high P. japonica mortality after external exposure. When nematodes were injected, P. japonica had a strong encapsulation and melanization response to all species except S. glaseri. Heterorhabditis bacteriophora and S. carpocapsae were able to overcome the immune response, but S. scapterisci was not. All species except S. scapterisci were able to kill and reproduce within the host. Only S. scapterisci and S. carpocapsae caused A. domesticus mortality after external exposure. When nematodes were injected, A. domesticus had a strong immune response to all species except S. scapterisci. Steinernema carpocapsae effectively overcame the strong immune response and caused high host mortality, but S. glaseri and H. bacteriophora did not. Steinernema scapterisci caused high host mortality and reproduced, S. glaseri and H. bacteriophora caused low host mortality but only S. glaseri reproduced, and S. carpocapsae was able to kill the host but reproduced poorly. Most (ca. 90%) of the S. carpocapsae in the hemocoel of P. japonica became encapsulated and melanized within 8 hours postinjection. The symbiotic bacterium, Xenorhabduf nematophilus, was often released before this encapsulation and melanization.     

Effects of Temperature and Dietary Lipids on Phospholipid Fatty Acids and Membrane Fluidity in Steinernema carpocapsae

The phospholipid composition of Steinernema carpocapsae was studied in relation to diet and culture temperature. When reared at 18 and 27.5 C on Galleria mellonella or on an artificial diet supplemented with lard, linseed oil, or fish oil as lipid sources, nematode phospholipids contained an abundance of 20-carbon polyunsaturated fatty acids, with eicosapentaenoic acid (20:5(n - 3)) predominant, regardless of the fatty acid composition of the diet. Because the level of linolenic acid (18:3(n - 3)) in nematode phospholipids was very low and because eicosapentaenoic acid was present even when its precursor (linolenic acid) was undetectable in the diet, S. carpocapsae likely produces n - 3 polyunsaturated fatty acids by de novo biosynthesis, a pathway seldom reported in eukaryotic animals. Reduction of growth temperature from 25 to 18 C increased the proportion of 20:5(n - 3) but not other polyunsaturated fatty acids. A fluorescence polarization technique revealed that vesicles produced from phospholipids of nematodes reared at 18 C were less ordered than those from nematodes reared at 27.5 C, especially in the outermost region of the bilayer. Dietary fish oil increased fluidity in the outermost region but increased rigidity in deeper regions. Therefore, S. carpocapsae appears to modify its membrane physical state in response to temperature, and eicosapentaenoic acid may be involved in this response. The results also indicate that nematode membrane physical state can be modified dietarily, possibly to the benefit of host-finding or survival of S. carpocapsae at low temperatures.     

Comparative effects of Steinernema feltiae (Nematoda: Steinernematidae) and insecticides on yield and cropping of the mushroom Agaricus bisporus

In a mushroom crop (Agaricus bisporus) affected by a very low level of sciarid fly (Lycoriella auripila) infestation, the effects of an indigenous isolate of insect-parasitic nematode (Steinernema feltiae) and of two commonly used insecticides (diazinon and diflubenzuron) were studied. When compared with untreated plots, nematodes applied to the casing had no adverse effects on mushroom yields whereas insecticides decreased yields. At a rate of 3 × 10⁶ infective juveniles per tray (surface area = 0.56 m²), S. feltiae elicited increases of 28.5% and 19% in the mean total numbers and weights of mushrooms respectively. Treatment only with diflubenzuron resulted in 14.6% and 6% reductions in mean total numbers and weights of mushrooms, respectively; treatment with both diazinon and diflubenzuron caused 18.5% and 9.4% losses. Application of nematodes generally reduced the mean weight per mushroom whereas insecticides increased it; nematodes delayed the onset of mushroom production (first flush) whereas diflubenzuron delayed the third and fourth flushes. Nematode contamination of sporophores was minimal when S. feltiae was applied at casing. Although their numbers declined with time, the nematodes persisted, in the casing layer, throughout the cropping period of seven weeks. It is concluded that yield benefits associated with nematode application can result mainly from nematode effects on A. bisporus and not solely from suppression of a damaging pest population.     

Dispersal Behaviour and Transmission Strategies of the Entomopathogenic Nematodes Heterorhabditis and Steinernema

Entomopathogenic nematodes of the Heterorhabditidae and Steinernematidae appear to be capable of long-distance dispersal and local migration. Their transmission strategies include both highly active seek-and-destroy behaviours and ambusher strategies, and they may be sensitive to sex-related factors in their own populations. Their host-finding abilities are poorly understood, despite the fact that these abilities are fundamental to their success as biocontrol agents in soil. Like the vast numbers of exotic hymenopterans and other natural enemies that have been released for biological control over the past century, they may be used in their ecologically competent wild-type form. On the other hand, because they are applied inundatively, they may be tailored, by breeding or transformation, to their intended purpose and to ecological incompetence, improving both their efficacy and their ecological safety.     

Compatibility of Soil Amendments with Entomopathogenic Nematodes

The impact of inorganic and organic fertilizers on the infectivity, reproduction, and population dynamics of entomopathogenic nematodes was investigated. Prolonged (10- to 20-day) laboratory exposure to high inorganic fertilizer concentrations inhibited nematode infectivity and reproduction, whereas short (1-day) exposures increased infectivity. Heterorhabditis bacteriophora was more sensitive to adverse effects than were two species of Steinernema. In field studies, organic manure resulted in increased densities of a native population of Steinernema feltiae, whereas NPK fertilizer suppressed nematode densities regardless of manure applications. Inorganic fertilizers are likely to be compatible with nematodes in tank mixes and should not reduce the effectiveness of nematodes used for short-term control as biological insecticides, but may interfere with attempts to use nematodes as inoculative agents for long-term control. Organic manure used as fertilizer may encourage nematode establishment and recycling.     

Evaluation of Steinernema carpocapsae Weiser for Control of Carrot Weevil Adults, Listronotus oregonensis (LeConte) (Coleoptera: Curculionidae), in Organically Grown Carrots

Under controlled conditions, application of Steinernema carpocapsae using a baiting method consistently reduced the oviposition of Listronotus oregonensis adults at all moisture regimens. At 80 and 95% relative humidity, soil applications at 10 4 infective juveniles (IJs) cm-2 and 10 5 infective stages (400 IJs cm-2) per pot reduced the survival and oviposition of L. oregonensis adults; soil applications were ineffective at relative humidity lower than 80%. When timed correctly, soil application at the rate of 2 105 IJs/linear meter of row (4.4 109 IJs ha-1) reduced damage by up to 59%.     

Competitive interactions between entomopathogenic nematodes and parasitoid venom

Entomopathogenic nematodes and parasitoid larvae of some wasps play important roles in the natural control of the pest insects. However, it has not been excluded that competition between nematodes and wasps may in some cases reduce their efficacy in the pest control. Using caterpillars of Spodoptera littoralis, we examined interactions between the nematode Steinernema carpocapsae and the venom of the parasitoid Habrobracon hebetor. The survival of S. littoralis caterpillars was reduced in a dose-dependent manner when 5 to 500 nematodes or 0.005–0.1 venom units were applied to single caterpillars. High doses of either nematodes or the venom caused death within 1–3 days in all treated hosts. The low doses of nematodes killed caterpillars within a week, in some cases when they attempted to pupate. Caterpillars receiving low venom doses were characterized by extended survival time terminated with death due to starvation. Combined treatment of nematodes and the venom were mutually synergistic and elicited severe lethal effects. The nematodes were fully resistant to the venom and can feed and grow on the symbiotic bacteria in vitro. The venom impairs food processing and causes death of caterpillars due to starvation. Disruption of the hormonal regulation of metamorphosis by ecdysteroids and juvenile hormone could be responsible for defective moults block at different stages of the moulting process, regionally restricted moulting, moults to “intermediates” combining regions of newly secreted larval and pupal cuticles.     

Survey of entomopathogenic nematodes and associate bacteria in Thailand and their potential to control Aedes aegypti

Aedes aegypti is an insect vector that transmits several viruses affecting humans worldwide. Entomopathogenic nematodes (EPNs) and their symbiotic bacteria are organisms with the potential to control many insects. In this study, we did a survey aimed to identify EPNs and their symbiotic bacteria and evaluate the larvicidal activity of bacteria against Ae. aegypti. We collected 540 soil samples from 108 sites in Phitsanulok Province, lower northern Thailand. Baiting techniques and White traps were used to isolate EPNs from soil samples. By sequencing of 28S rDNA and internal transcribed spacer regions, 51 EPN isolates were identified as Steinernema surkhetense (35 isolates), Heterorhabditis indica (14 isolates) and Heterorhabditis sp. SGmg3 (two isolates). Based on sequencing of a partial region of the recA gene, 35 isolates of Xenorhabdus were identified as Xenorhabdus stockiae, and 20 Photorhabdus isolates were identified as Photorhabdus luminescens subsp. akhurstii (10 isolates), P. luminescens subsp. hainanensis (seven isolates) and P. asymbiotica subsp. australis (three isolates). Screening for larvicidal activity of bacteria against Ae. aegypti was performed in the laboratory. Xenorhabdus WB5.4 and Xenorhabdus WB12.5, which were closely related to X. stockiae, resulted in high mortality of Ae. aegypti (99.99% and 70%, respectively) at 96 hr after exposure. Comparing with control groups, mortality of Ae. aegypti larvae was low (1.11%–6.67%) after exposure for 24–96 hr. Our findings showed the potential of X. stockiae for controlling Ae. aegypti. Further studies are needed to elucidate the mechanisms through which these bacteria kill Ae. aegypti larvae.     

Integration between Steinernema feltiae and some of environmental friendly compounds to control Spodoptera littoralis (Lepidoptera: Noctuidae)

The efficacy of different concentrations of the commercial neem-based insecticide, Nimbicidine^® and the ecdysone agonist compound Methoxyfenozide (RH-2485), was evaluated against larvae of Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae). RH-2485 and Nimbicidine^® significantly (p?<?0.001) reduced adult longevity by 2.7 and 1.9 d at the higher concentrations tested, respectively, but no significant differences were observed at low concentration. The tested compounds strongly affected the reproduction of S. littoralis by producing a high percentage of sterility. Fecundity and fertility were significantly affected by both insecticides. No egg laying has been recorded with the higher concentration 0.5% of Nimbicidine^® and 0.1?ppm of RH-2485, while no significant (p?>?0.05) difference was noticed on total number of eggs laid by the female when lower concentration 0.0001?ppm of RH-2485 was applied in the same stage as compared to the control. In the second part of this study, the invasion rate of Steinernema feltiae was affected by the addition of both Nimbicidine and RH-2485 to the diet of experimental host. At higher concentration of both compounds, the invasion rate was decreased despite the infection rate, while the percentage of invading nematodes increase to 56.7% with the combination treatment indicating a considerable improvement in the efficacy of S. feltiae nematode applied in combination with the lower concentration of Nimbicidine^® over that of nematode alone. On the other hand, the invasion of S. feltiae to the insects that were fed on the diet with the addition of ecdyson compound was strongly decreased with increasing concentration of RH-2485 in host diet.