Impact of insect-resistant transgenic rice on target insect pests and non-target arthropods in China

Progress on the research and development of insect-resistant transgenic rice, especially expressing insecticidal proteins from Bacillus thuringiensis （Bt）, in China has been rapid in recent years. A number of insect-resistant transgenic rice lines/varieties have passed restricted and enlarged field testing, and several have been approved for productive testing since 2002 in China, although none was approved for commercial use until 2006. Extensive laboratory and field trials have been conducted for evaluation of the efficiency of transgenic rice on target lepidoteran pests and potential ecological risks on non-target arthropods. The efficacy of a number of transgenic rice lines currently tested in China was excellent for control of the major target insect pests, the rice stem borers （Chilo suppressalis, Scirpophaga incertulas, Sesamia inferens） and leaffolder （ Cnaphalocrocis medinalis）, and was better than most insecticides extensively used by millions of farmers at present in China. No significantly negative or unintended effects of transgenic rice on non-target arthropods were found compared with non-transgenic rice. In contrast, most of the current insecticides used for the control of rice stem borers and leaffolders proved harmful to natural enemies, and some insecticides may directly induce resurgence of rice planthoppers. Studies for developing a proactive insect resistance management of transgenic rice in the future are discussed to ensure the sustainable use of transgenic rice.

Parasitism of Pieris rapae （Lepidoptera： Pieridae） by the endoparasitic wasp Pteromalus puparum （Hymenoptera： Pteromalidae）： Effects of parasitism on differential hemocyte counts, micro- and ultra-structures of host hemocytes

Parasitism by the endoparasitic wasp Pteromalus puparum （Hymenoptera： Pteromalidae） by using only its associated venom, can suppress the immunal responses of Pieris rapae （Lepidoptera： Pieridae）. However, up to now, current knowledge of the mech- anisms has been limited. The response of host hemocytes to parasitism was investigated using a combination of light and transmission electron microscopy （TEM）. Five hemocyte types, prohemocytes （PRs）, granulocytes （GRs）, plasmatocytes （PLs）, oenocytoids （OEs） and coagulocytes （COs）, were observed and characterized from both unparasitized and parasitized Pieris rapae pupae. Light microscopy showed that both GRs and PLs became more round and spread abnormally after parasitism, whereas the shape of other types of hemocytes remained unaffected. In addition, the size of PRs and PLs became larger while OEs became smaller. The proportion of PRs significantly increased after parasitism and that of PLs decreased by 43.9%, but there was no significant increase of GRs and OEs. TEM showed that all types of hemocytes except COs were damaged to various degrees after parasitism, especially resulting in electron opaque cytoplasm and nucleus, fewer cell organelles of rough endoplasmic reticulum, mitochondria and vesicles. Our results indicate that parasitism by P. puparum affects differential hemocyte counts and structures of host hemocytes, particularly for GRs and PLs, which may be the main cause of the parasitoid suppressing host cellular immune responses.

Molecular characterization and expression profiles of nicotinic acetylcholine receptors in the rice striped stem borer, Chilo suppressalis （Lepidoptera： Crambidae）

Nicotinic acetylcholine receptors （nAChRs） are members of the cys-loop ligand- gated ion channel （cysLGIC） superfamily, mediating fast synaptic cholinergic transmission in the central nervous system in insects. Insect nAChRs are the molecular targets of economically important insecticides, such as neonicotinoids and spinosad. Identification and characterization of the nAChR gene family in the rice striped stem borer, Chilo suppressa[is, could provide beneficial information about this important receptor gene family and contribute to the investigation of the molecular modes of insecticide action and resistance for current and future chemical control strategies. We searched our C. suppressalis transcriptome database using Bombyx mori nAChR sequences in local BLAST searches and obtained the putative nAChR subunit complementary DNAs （cDNAs） via reverse transcription polymerase chain reaction （RT-PCR） and rapid amplification of cDNA ends methods. Similar to B. mori, C. suppressalis possesses 12 nAChR subunits, including nine c^-type and three/%type subunits. Quantitative RT-PCR analysis revealed the expression profiles of the nAChR subunits in various tissues, including the brain, subesophageal ganglion, thoracic ganglion, abdominal ganglion, hemocytes, fat body, foregut, midgut, hindgut and Malpighian tubules. Developmental expression analyses showed clear differential expression of nAChR subunits throughout the C. suppressalis life cycle. The identification of nAChR subunits in this study will provide a foundation for investigating the diverse roles played by nAChRs in C. suppressalis and for exploring specific target sites for chemicals that control agricultural pests while sparing beneficial species

Cellular and humoral immune interactions between Drosophila and its parasitoids

The immune interactions occurring between parasitoids and their host insects,especially in Drosophila-wasp models,have long been the research focus of insect immunology and parasitology.Parasitoid infestation in Drosophila is counteracted by its multiple natural immune defense systems,which include cellular and humoral immunity,Occurring in the hemocoel,cellular immune responses involve the proliferation,differentiation,migration and spreading of host hemocytes and parasitoid encapsulation by them.Contrastingly,humoral immune responses rely more heavily on melanization and on the Toll,Imd and Jak/Stat immune pathways associated w ith antimicrobial peptides along with stress factors.On the wasps'side,successful development is achieved by introducing various virulence factors to counteract immune responses of Drosophila.Some or all of these factors manipulate the host's immunity for successful parasitism.Here we review current knowledge of the cellular and humoral immune interactions between Drosophila and its parasitoids,focusing on the defense mechanisms used by Drosophila and the strategies evolved by parasitic wasps to outwit it.

Proteome changes in the plasma of Pieris rapae parasitized by the endoparasitoid wasp Pteromalus puparum

Parasitism by the endoparasitoid wasp Pteromalus puparum causes alterations in the plasma proteins of Pieris rapae. Analysis of plasma proteins using a proteomic approach showed that seven proteins were differentially expressed in the host pupae after 24-h parasitism. They were masquerade-like serine proteinase homolog (MSPH), enolase (Eno), bilin-binding protein (BBP), imaginal disc growth factor (IDGF), ornithine decarboxylase (ODC), cellular retinoic acid binding protein (CRABP), and one unknown function protein. The full length cDNA sequences of MSPH, Eno, and BBP were successfully cloned using rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR). Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis indicated that the transcript levels of MSPH and BBP in the fat bodies of host pupae were inducible in response to the parasitism and their variations were consistent with translational changes of these genes after parasitism, while the transcript levels of Eno and IDGF were not affected by parasitism. This study will contribute to the better understanding of the molecular bases of parasitoid-induced host alterations associated with innate immune responses, detoxification, and energy metabolism.

Effects of starvation on the vitellogenesis, ovarian development and fecundity in the ectoparasitoid, Nasonia vitripennis （Hymenoptera： Pteromalidae）

A female-specific protein, vitellogenin （Vg）, and its corresponding egg vitellin （Vt） are identified in the ectoparasitic wasp Nasonia vitripennis. Both native Vt and Vg have a molecular mass of about 350 kDa, which is composed of two subunits of approximately 190 kDa and 165 kDa under reducing and denaturing conditions （sodium dodecyl sulfate-polyacrylamide gel electrophoresis）. An indirect sandwich enzyme-linked immunosorbent assay developed with both monoclonal and polyclonal antibodies against N. vitripennis Vt. Vg was first detected in the hemolymph on the 10th day after parasitism, and was first observed in oocytes on the 12th day. In adults deprived of food, the highest hemolymph Vg level occurred at the time of adult eclosion and the highest level of Vt in ovaries was found at 30 h after eclosion. In contrast, feeding adults with 20% sucrose resulted in the reduction of Vt uptake by ovaries and the extension of life span, but had little effect on Vg production. Deprived of hosts, starvation of female wasps had no significant effects on ovariole growth and oocyte maturation before the wasps died. However, starvation of female wasps supplied with hosts accelerated the wasps laying progeny into hosts, but resulted in a decrease of total progeny production by comparison with wasps fed with 20% sucrose.

Heat shock protein genes （hsp20, hsp75 and hsp90） from Pieris rapae： Molecular cloning and transcription in response to parasitization by Pteromalus puparum

Most molecular work on the roles of heat shock proteins （hsps） in host-parasite interaction has focused on vertebrates, rather than invertebrates. Here the full length complementary DNA （cDNA） sequences of three hsp genes （hsp20, hsp75 and hsp90） were amplified from Pieris rapae, and their transcriptional responsiveness to parasitization by the endoparasitic wasp Pteromalus puparum were investigated. The cDNA sequence analysis of hsp20, hsp75 and hsp90 revealed open reading frames of 531, 2328 and 2 157 bp in length, which encode proteins with calculated molecular weights of 19.5, 75.48 and 82.7 kDa, respectively. The comparison of amino acid sequences showed that P rapae hsp20 shared highly divergent homology to that of other insects, while hsp75 and hsp90 showed high homology to their counterparts of other species. The expression analysis indicated that these three genes were influenced in response to parasitization by P. puparum. The hsp20 transcripts in parasitized pupae were higher compared to non- parasitized pupae. The expression of hsp75 and hsp90 were down-regulated following parasitization. The results indicate that hsps are involved in host-parasitoid interactions.

An endoparasitoid uses its egg surface proteins to regulate its host immune response

With proteomic analysis,we identified 379 egg surface proteins from an endoparasitoid,Cotesia chilonis.Proteins containing conserved enzymatic domains constitute a large proportion of egg surface components.Some proteins,such as superoxidase dismutase,homolog of C.rubecula 32-kDa protein,and immunoevasive protein-2A,are classical parasitism factors that have known functions in host immunity regulation.Melanization assays revealed that a novel egg surface protein,C.chilonis egg surface serpin domain-containing protein had the same function as a C.chilonis venom serpin,as both suppressed host melanization in a dose-dependent manner.C.chilonis egg surface serpin domain-containing protein is mainly transcribed in C.chilonis oocytes with follicular cells,and it is located on both the anterior and posterior sides of the mature egg surface.Additionally,we used LC-MS/MS to identify 586 binding proteins sourced from C.suppressalis plasma located on the eggshell surface of C.chilonis,which included some immunity-related proteins.These results not only indicate that C.chilonis uses its egg surface proteins to reduce the immune response of its host but also imply that endoparasitoid egg surface proteins might be a new parasitism factor involved in host immune regulation.