Hemocyte encapsulation reactions of infective juveniles of two Iranian isolates of the entomopathogenic nematodes, Heterorhabditis bacteriophora Poinar (Rhabditina: Heterorhabditidae) and Steinernema feltiae Filipjev (Tylenchina: Steinernematidae), were compared in the economic pest Colorado potato beetle, Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae), and the greater wax moth, Galleria mellonella L. (Lepidoptera: Pyralidae). The former was a more responsive host than the latter and the hemocyte responses occurred sooner and more extensively. Complete encapsulation of some of the nematodes occurred by 4 h post injection for H. bacteriophora in both L. decemlineata and G. mellonella, and by 2 h pi for S. feltiae in L. decemlineata. The percentage of encapsulation from 24 h to 72 h pi in L. decemlineata was 86.2% for S. feltiae and 39% for H. bacteriophora. In G. mellonella there were no encapsulation or melanization responses against S. feltiae, whereas when H. bacteriophora was encapsulated and melanized (16.7%) the encapsulation level was lower than in L. decemlineata. This study may contribute to effectively selecting entomopathogenic nematode species active against significant economic pests based on the latter's cellular immune response.

To determine the LC 50 values of two entomopathogenic nematodes against Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae) prepupae, different concentrations of the nematodes were tested in soil. Because of the different temperature requirements of the two nematode species, bioassay experiments were conducted at 20 ± 1°C and 27 ± 2°C for Steinernema feltiae Filipjev (Rhabditida: Steinernematidae) and Heterorhabditis bacteriophora Poinar (Rhabditida: Heterorhabditidae), respectively. Both the isolates were effective against L. decemlineata. LC 50 values of H. bacteriophora against progeny of field-collected adults and laboratory-reared adults were estimated as 8.5 and 7.6 IJ per prepupa, respectively. For S. feltiae the value was calculated as 51.2 IJ per prepupa against offspring of laboratory-reared adults of L. decemlineata only. Cellular encapsulation of both nematode species was observed. Sublethal nematode concentrations caused wing deformation and delayed metamorphosis which may affect Colorado potato beetle adult fitness.

The Colorado potato beetle (CPB), Leptinotarsa decemlineata Say is the most destructive insect pest of potato in many areas of the world. Little is known about the haemocyte types of the CPB and its plasma phenoloxidase (PO). In this regard, we investigated the haemocyte profile and PO of CPB and its immune response to the entomopathogenic nematode, Steinernema carpocapsae. Five types of haemocytes, the plasmatocytes (~67.4%), granulocytes (~23.5%), oenocytoids (~2.4%), spherulocytes (~0.25%) and prohaemocytes (~6.5%) were identified in fourth instar CPB larvae. Total haemocyte counts (THCs) were significantly reduced in nematode-injected insects compared with control groups (P < 0.05). Nematode cellular encapsulation observed in haemolymph of nematode-injected insects may partially explain decreased THCs. Plasma PO assay showed increased PO activity in nematode-injected insects compared with control groups (P < 0.05). Plasma PO assay on native polyacrylamide gel electrophoresis (PAGE) assay with L-3, 4-dihydroxyphenylalanine as substrate showed five bands (with molecular weights of approximately 200, 118, 68.5, 62.5 and 58.75 kDa).

Lethal effect of the entomopathogenic nematode, Steinernema feltiae, against prepupae of Helicoverpa armigera (Hb) (Lepidoptera: Noctuidae) in soil was evaluated. Plasma phenoloxidase activity of H. armigera prepupae against S. feltiae also was evaluated. In brief, 20-25 infective juveniles (IJs) of S. feltiae in 20 μl Ringer's solution were injected into the hemolymph of H. armigera prepupae in different time intervals. In order to evaluate the effect of the Ringer's solution, it was also injected into the larvae in a separate experiment as positive control. A third group of non-injected insects was used as negative control. Hemolymph of the insects was collected in different time intervals (0 to 24 h post injection). Phenoloxidase (PO) activity of H. armigera hemolymph was determined spectrophotometrically, using L-Dopa as substrate. The LC 20 , LC 50 , and LC 80 values were 4.5, 19, and 76 IJs per insect, respectively. PO activity assay demonstrated higher levels of PO unit in nematode-injected insects compared to control groups. Nematode-injected insects showed the highest plasma PO activity 8 h post-injection.

For evaluating lethal effects of abamectin on T. urticae leaf dipping method was used. Then T. brassicae adults were treated with LC 10 and LC 50 values obtained from probit analysis of the mite bioassay results using toxicity bioassay with fresh residue method. For evaluating residual effects of abamectin, bean plants were sprayed with twentieth of recommended field dose of abamectin (equal to 25 ppm of trade product). Leaf pieces were prepared from treated plants in different time intervals and the adult mites were transferred on them. Mortality was recorded after 24 h. Probit analysis of lethal experiment against T. urticae revealed 0.05, 0.417 and 3.26 ppm from trade product as LC 10 , LC 50 and LC 90 values, respectively. Results of residual effects experiment showed that mite mortality was 100% in 1 day after plant spraying which decreased to 55.62% in 21 days after spraying. LC 10 and LC 50 values of abamectin tended to 53.87 and 72.57% mortality of T. brassicae, respectively.

During a survey of entomopathogenic nematodes (EPNs) in Moghan region of Iran in 2015, a steinernematid species was isolated, using the Galleria-baiting method. Based on its morphological and phylogenetic analysis of molecular data, the isolate was identified as Steinernema carpocapsae. The ITS rDNA sequence was deposited in NCBI (National Center for Biotechnology Information) with accession number of MF187616.1. Nucleotide row data was edited, using MEGA 6.0 software, and homologous sequences were involved in analysis, using Blast software. Sequences were aligned using Clustal W. Bootstrap analysis. The phylogenetic tree was constructed by the maximum likelihood method, using MEGA 6.0 software, and Steinernema feltiae was used as out-group. The turnip moth, Agrotis segetum Denis and Schiffermuller (Lepidoptera: Noctuidae), is one of the most important and destructive cutworm pests in Moghan. Therefore, the lethal effect of S. carpocapsae isolate Moghan (IRMoghan) was evaluated in a soil assay against the last instar larvae of A. segetum under laboratory conditions. The bioassay results showed high susceptibility of the larvae to S. carpocapsae. The LC 10 , LC 50 , and LC 90 values were 9.9, 54.13, and 246.2 IJs (infective juveniles) per larva of the pest, respectively (χ 2 = 7.36; df = 3, P value = 0.061). Reproduction of the EPNs within the dissected cadavers was observed. The bioassay results indicated that the new isolate is a promising biocontrol agent against A. segetum larvae with success recycling through them.

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The potato tuber moth PTM, Phthorimaea operculella, is one of the most economically important potato pests worldwide. In the present study, the potential of Steinernema feltiae and Steinernema carpocapsae for controlling PTM in potato tubers was evaluated compared to alpha-cypermethrin. Steinernema carpocapsae in both concentrations (12.6×10 6 IJs and 6.2×10 6 IJs) showed a lower number of emerged insects than alpha-cypermethrin (10 mg ai l -1 ).Alpha-cypermethrin (20 mg ai l -1 ) showed the highest efficacy against PTM (81.17%), S.carpocapsae (12.6×10 6 IJs), and alpha-cypermethrin (10 mg ai l -1 ) showed similar efficacy (72.53%) while S. feltiae (6.2×10 6 IJs) showed the lowest efficacy (39.04%). The results showed that S.carpocapsae in both concentrations and S. feltiae (12.6×10 6 IJs) were efficient the same as alpha-cypermethrin (10 mg ai l -1 ) having no environmental and health adverse impacts issued in the chemical insecticides usage. Both concentrations of alpha-cypermethrin and S. carpocapsae showed the least tuber damage with no significant differences, while it was as high as the control (59.26%) in both concentrations of S. feltiae. This promising finding introduces EPNs as a part of the potato tuber protection program in storage. Accordingly, EPNs can be considered as an appropriate alternative to synthetic chemicals for PTM control without any residue and health problems.