RNA-seq analysis of gene expression changes during pupariation in Bactrocera dorsalis (Hendel) (Diptera: Tephritidae).

BACKGROUND: The oriental fruit fly, Bactrocera dorsalis (Hendel) has been considered to be one of the most important agricultural pest around the world. As a holometabolous insect, larvae must go through a metamorphosis process with dramatic morphological and structural changes to complete their development. To better understand the molecular mechanisms of these changes, RNA-seq of B. dorsalis from wandering stage (WS), late wandering stage (LWS) and white puparium stage (WPS) were performed. RESULTS: In total, 11,721 transcripts were obtained, out of which 1914 genes (578 up-regulated and 1336 down-regulated) and 2047 genes (655 up-regulated and 1392 down-regulated) were found to be differentially expressed between WS and LWS, as well as between WS and WPS, respectively. Of these DEGs, 1862 and 1996 genes were successfully annotated in various databases. The analysis of RNA-seq data together with qRT-PCR validation indicated that during this transition, the genes in the oxidative phosphorylation pathway, and genes encoding P450s, serine protease inhibitor, and cuticular proteins were down-regulated, while the serine protease genes were up-regulated. Moreover, we found some 20-hydroxyecdysone (20E) biosynthesis and signaling pathway genes had a higher expression in the WS, while the genes responsible for juvenile hormone (JH) synthesis, degradation, signaling and transporter pathways were down-regulated, suggesting these genes might be involved in the process of larval pupariation in B. dorsalis. For the chitinolytic enzymes, the genes encoding chitinases (chitinase 2, chitinase 5, chitinase 8, and chitinase 10) and chitin deacetylase might play the crucial role in the degradation of insect chitin with their expressions significantly increased during the transition. Here, we also found that chitin synthase 1A might be involved in the chitin synthesis of cuticles during the metamorphosis in B. dorsalis. CONCLUSIONS: Significant changes at transcriptional level were identified during the larval pupariation of B. dorsalis. Importantly, we also obtained a vast quantity of RNA-seq data and identified metamorphosis associated genes, which would all help us to better understand the molecular mechanism of metamorphosis process in B. dorsalis.

Genome-wide annotation of cuticular proteins in the oriental fruit fly (Bactrocera dorsalis), changes during pupariation and expression analysis of CPAP3 protein genes in response to environmental stresses.

Cuticular proteins (CPs) are essential components of the insect cuticle as they create a structural and protective shield and may have a role in insect development. In this paper, we studied the CPs in the oriental fruit fly (Bactrocera dorsalis), one of the most economically important pests in the Tephritidae family around the world. The availability of a complete genome sequence (NCBI Assembly: ASM78921v2) allowed the identification of 164 CP genes in B. dorsalis. Comparative analysis of the CPs in B. dorsalis with those in the model insect Drosophila melanogaster and the closely related Ceratitis capitata, and CPs from mosquitoes, Lepidoptera, Hymenoptera and Coleoptera identified Diptera-specific genes and cuticle development patterns. Analysis of their evolutionary relationship revealed that some CP families had evolved according to the phylogeny of the different insect species, while others shared a closer relationship based on domain architecture. Subsequently, transcriptome analysis showed that while most of the CPs (60-100% of the family members) are expressed in the epidermis, some were also present in internal organs such as the fat body and the reproductive organs. Furthermore, the study of the expression profiles throughout development revealed a profound change in the expression of CPs during the formation of the puparium (pupariation). Further analysis of the expression profiles of the CPAP3 genes under various environmental stresses revealed them to be involved in the response to pesticides and arid and extreme temperatures conditions. In conclusion, the data provide a particular overview of CPs and their evolutionary and transcriptional dynamics, and in turn they lay a molecular foundation to explore their roles in the unique developmental process of insect metamorphosis and stress responses.

Corazonin Signaling Is Required in the Male for Sperm Transfer in the Oriental Fruit Fly Bactrocera dorsalis.

Corazonin (Crz) is a widely distributed neuropeptide (or neurohormone) in insects with diverse physiological functions. The present study aimed to reveal the functions of Crz and its receptor (CrzR) in the regulation of sexual behavior and fertility in male Bactrocera dorsalis. Tissue-specific expression analyses showed that the BdCrz transcript was most abundant in the central nervous system (CNS), and the BdCrzR transcript was most abundant in both the fat body and CNS. Immunochemical localization confirmed that three pairs of Crz-immunoreactive neurons are located in the dorsolateral protocerebrum region of male adult brain. Importantly, RNAi-mediated Crz knockdown lengthened mating duration in males, and knockdown of Crz or CrzR strongly decreased male fertility in the following 3 days, while the courtship behavior and mating efficiency were not affected. The reduced number of sperm in the reproductive organs of mated females indicated that Crz knockdown in males reduced sperm transfer. The findings of this study indicate that Crz contributes to the reproductive physiology of the oriental fruit fly B. dorsalis by regulating sperm transfer in male adults.