Parasitoid wasps inject venom containing complex bioactive compounds to regulate the immune response and development of host arthropods and sometime paralyze host arthropods. Although extensive studies have been conducted on the identification of venom proteins in larval parasitoids, relatively few studies have examined the pupal parasitoids. In our current study, a combination of transcriptomic and proteomic methods was used to identify 64 putative venom proteins from Pachycrepoideus vindemmiae, an ectoparasitoid of Drosophila. Expression analysis revealed that 20 tested venom proteins have 419-fold higher mean expression in the venom apparatus than in other wasp tissues, indicating their specialization to venom. Comparisons of venom proteins from P. vindemmiae and other five species spanning three parasitoid families detected a core set of “ancient” orthologs in Pteromalidae. Thirty-five venom proteins of P. vindemmiae were assigned to the orthologous groups by reciprocal best matches with venoms of other pteromalids, while the remaining 29 were not. Of the 35 categories, twenty-seven have orthologous relationships with Nasonia vitripennis venom proteins and 25 with venoms of Pteromalus puparum. More distant relationships detected that five and two venom proteins of P. vindemmiae are orthologous with venoms of two Figitidae parasitoids and a Braconidae representative, respectively. Moreover, twenty-two venoms unique to P. vindemmiae were also detected, indicating considerable interspecific variation of venom proteins in parasitoids. Phylogenetic reconstruction based on a set of single-copy genes clustered P. vindemmiae with P. puparum, N. vitripennis, and other members of the family Pteromalidae. These findings provide strong evidence that P. vindemmiae venom proteins are well positioned for future functional and evolutionary studies.

The immunological interaction between Drosophila melanogaster and its larval parasitoids has been thoroughly investigated, however, little is known about the interaction between the host and its pupal parasitoids. Pachycrepoideus vindemmiae, a pupal ectoparasitoid of D. melanogaster, injects venom into its host while laying eggs on the puparium, which regulates host immunity and interrupts host development. To resist the invasion of parasitic wasps, various immune defense strategies have been developed in their hosts as a consequence of co-evolution. In this study, we mainly focused on the host immunomodulation by P. vindemmiae and thoroughly investigated cellular and humoral immune response, including cell adherence, cell viability, hemolymph melanization and the Toll, Imd, and JAK/STAT immune pathways. Our results indicated that venom had a significant inhibitory effect on lamellocyte adherence and induced plasmatocyte cell death. Venom injection and in vitro incubation strongly inhibited hemolymph melanization. More in-depth investigation revealed that the Toll and Imd immune pathways were immediately activated upon parasitization, followed by the JAK/STAT pathway, which was activated within the first 24 h post-parasitism. These regulatory effects were further validated by qPCR. Our present study manifested that P. vindemmiae regulated the cellular and humoral immune system of host D. melanogaster in many aspects. These findings lay the groundwork for studying the immunological interaction between D. melanogaster and its pupal parasitoid.

As a major invasive pest in China, Spodoptera frugiperda (Smith) (Lepidoptera: Noctuidae) has caused great damage to crops. Hymenopteran parasitoids, especially the braconid wasps, play crucial roles in depressing pest populations. However, there was little information about the ideal storage of parasitoids to achieve their mass-rearing. Here, we identified a dominant parasitoid of S. frugiperda, Microplitis manilae (Ashmead) (Hymenoptera: Braconidae), in the Hainan province of China with a field parasitism rate of 5.66–19.10%. The investigation of biological parameters revealed that the parasitism rate of M. manilae significantly decreased with an increase in both wasp adult longevity and host age, and the wasp of 1–3 d post eclosion performed best on the first instar of host larvae, showing the highest parasitism rate. We also discovered that the decreased temperature from 30 to 20 °C greatly extended the longevity of wasp adults, and a similar result was observed after feeding on 10% sucrose water compared with sterile water. Then, the effects of different pupal cold storage temperatures (4 and 10 °C), storage period (prepupa, middle-aged pupa, late-aged pupa) and storage time (5, 10 or 20 d) on the emergence rate, parasitism rate, female proportion and longevity of M. manilae were investigated. The results demonstrated that the middle-aged wasp pupae stored at 10 °C for 5–10 d possessed a stronger parasitic ability and longer longevity. These findings may promote the flexibility and efficacy of large-scale production of M. manilae, thus contributing to its biological field control against S. frugiperda.

Chalcone synthase (CHS) is a key rate-limiting enzyme in the flavonoid synthesis pathway. Flavonoids are crucial secondary metabolites that play significant roles in plant growth, development, and stress resistance. The CHS gene (BrCHS) family in Chinese cabbage has not yet been studied. We identified 10 BrCHS genes distributed across 7 chromosomes in the Chinese cabbage genome. Their encoded proteins all contain the Chal_Sti_Synt_C (PF02797) and Chal_Sti_Synt_N (PF00195) domains and can be classified into two groups based on systematic evolution analysis. These BrCHS genes contain 2–4 exons and numerous cis-acting elements responsive to light, hormones, stress, growth and development in the BrCHS gene promoters. We also revealed that the expression of BrCHS2 and BrCHS8 increased under treatment with methyl jasmonate, salt, or drought stress. Virus-induced gene silencing (VIGS) of BrCHS4 inhibited the expression of BrCHS4 and reduced the flavonoid and anthocyanin contents in leaves. Ten BrCHS family genes are present in the genome of Chinese cabbage. These BrCHS genes seemingly maintained similar characteristics and functionalities during evolution. Our results demonstrated that BrCHS4 is involved in flavonoid and anthocyanin accumulation in Chinese cabbage and identified candidate genes for purple Chinese cabbage breeding.