| The diamondback moth, Plutella xylostella (Lepidoptera, Plutellidae), is the most destructive pests of economically important Brassicaceae crops. Currently, there are still no effective strategies to control this very prolific insect that can reproduce rapidly in China. To better understand the reproductive regulation of P. xylostella and the consequences on reproductive potential and adaptability, we examined the role of vitellogenin gene Vg in P. xylostella (PxVg), which is a fundamental substance of reproductive regulation in insects. Three experiments will be conducted. First, the morphology of reproductive system in P. xylostella and developmental dynamic of ovaries were surveyed. Subsequently, the molecular characteristics and expression pattern of PxVg, as well as its phylogenetic relationship with other insects were analyzed. Finally, the roles of PxVg in reproductive regulation were studied. This study provided clues for further understanding of the physiological functions of Vg gene, as well as a basis for exploring the reproductive regulatory mechanisms of P. xylostella. The main results were as follows:1. The morphology of female reproductive system and developmental dynamic of ovaries in P. xylostellaThe female reproductive system of P. xylostella consisted of ovaries, lateral oviduct, common oviduct, accessory glands, spermatheca and copulatory pouch. Each ovary was composed of 4 ovarioles. According to the egg morphology and yolk deposition in ovaries at different developmental stages of P. xylostella, the oogenesis was divided into three stages:previtellogenic stage (1 day-old pupa), vitellogenic stage (2-3 day-old pupa), and postvitellogenic stage (newly emerged female).2. The molecular characterization and expression pattern of Vg gene in P. xylostellaPxVg has an open reading frame (ORF) with 5217 base pairs (bp) that encoded for 1738 amino acids with a predicted molecular mass of 196.9 kDa and a proposed isoelectric point of 6.13. The deduced primary structure indicated that the numbers of Alanine and Serine were 150 (up to 8.6%) and 130 (up to 7.5%), which were more than the other amino acids. Sequence alignment analysis indicated that the protein sequence of PxVg contained three conserved domains, as detected in the Vgs of the other insects, including one vitellogenin-N (or lipoprotein aminoterminal) domain near the N-terminus, a von Willebrand factor type D domain (VWD) near the C-terminus, and a DUF1943 functional domain in between, as well as a special domain Zn-finger. There are four putative cleavage sites, including RNER, RARR, RLAR, and RLAR in PxVg. In addition, a large number of conserved motifs, including GL/ICG, DGXR and cysteine residues were identified near the C-terminus of PxVg. An amino acid mutation at GVCG was detected in GL/ICG motif. The polyserine sequence was not found in PxVg. PxVg contained 6 glycosylation sites and 120 phosphorylation sites. The comparative analysis of the deduced amino acid sequences from Vg cDNA fragments of P. xylostella showed high similarity to other Lepidoptera species, such as Antheraea pernyi (40.3%), Actias selene (41.2%) and Samia Cynthia (40.5%). Gene transcript analyses revealed that PxVg could express at different developmental stages with the highest levels at the adult stage. However, with adult aging, the expression level of PxVg gradually declined.3. Effect of Vg gene on fecundity of P. xylostellaInjection of PxVg-siRNA into one day-old female pupa caused severe suppression of Vg mRNA expression within 24 h. Meanwhile, PxVg silencing also had an effect on fecundity of P. xylostella. Preliminary results showed that the number of eggs laid by females within 5 days was significantly decreased due to the PxVg silencing, but no significant difference was observed on the hatching rate of the eggs. |
PDF Full Text Request