Circadian Activity and Clock Genes in Pachycrepoideus vindemmiae: Implications for Field Applications and Circadian Clock Mechanisms of Parasitoid Wasps

Abstract

Despite the importance of circadian rhythms in insect behavior, our understanding of circadian activity and the molecular oscillatory mechanism in parasitoid wasp circadian clocks is limited. In this study, behavioral activities expected to be under the control of the endogenous circadian system were characterized in an ectoparasitoid wasp, Pachycrepoideus vindemmiae. Most adults exhibited emergence between late night and early morning, while mating only occurred during the daytime, with a peak at midday. Oviposition had three peaks in the early morning, late day, or early night and late night. Additionally, we identified eight putative clock genes from P. vindemmiae. The quantitative PCR (qPCR) results indicate that most clock genes showed significant rhythmic expressions. Our comparative analysis of clock genes in P. vindemmiae and 43 other parasitoid wasps revealed that none of the wasps possessed the timeless and cry1 genes commonly found in some other insect species, suggesting that the circadian clock system in parasitoid wasps is distinct from that in other non-Hymenoptera insects such as Drosophila. Thus, this study attempted to build the first hypothetical circadian clock model for a parasitoid wasp, thus generating hypotheses and providing a platform for the future functional characterization of P. vindemmiae clock genes as well as those of other parasitoid wasps. Finally, these findings on P. vindemmiae circadian activity will aid the development of effective field release programs for biological control, which can be tested under field conditions.

Keywords: Pachycrepoideus vindemmiae; circadian rhythms; clock genes; field application; parasitoid wasp.

Figure 1

Pachycrepoideus vindemmiae males and females emergence(A), mating (B) and oviposition (C) patterns under a photoperiod of 12: 12 h (light/dark). Data are presented as the means ± standard deviation. The different letters are significantly different based on one-way analysis of variance (ANOVA) and Tukey’s test with differences considered significant at p < 0.05.

Figure 2

Phylogenetic relationships of TIMEOUT proteins from parasitoid wasps including Pachycrepoideus vindemmiae and TIMELESS proteins from Aedes aegypti (Aaeg), Anopheles gambiae (Agam), Antheraea pernyi (Aper), Bombyx mori (Bmor), Drosophila melanogaster (Dmel), Danaus plexippus (Dple), Gryllus bimaculatus (Gbim), Periplaneta americana (Pame) and Tribolium castaneum (Tcas). GenBank numbers of TIMELESS proteins are listed after the abbreviations of species names. For TIMEOUT proteins, tip labels show the protein names in InsectBase 2.0 and the species information is listed in Table S1. The phylogenetic tree is constructed using the maximum likelihood method. The best models is JTT+G4. Red dots at the nodes denote bootstrap values greater than 500 from 1000 trials.

Figure 3

Phylogenetic relationships of DNA photolyases and CRYs. The phylogenetic tree of amino acid sequences was constructed using the maximum likelihood method. The best model is LG+I+G4. Bootstrap values below 70% were removed from the phylogenetic tree. The functional character is mapped onto this phylogenetic tree based on [1]. Orange letters indicate the proteins that can repress CLK: CYC (BMAL)-mediated transcription in cell culture. Blue letters indicate the proteins that lack this ability in cell culture. Black and red letters indicate the proteins whose transcriptional repressive activity remain unknown. The tip labels of parasitoid wasps show the protein names in InsectBase 2.0. Species information for parasitoid wasps is listed in Table S1 and other species are selected based on [1], whose tip labels consist of protein names, species names and Genbank numbers.

Figure 4

qPCR results showing the mRNA abundance levels of the eight clock genes in females (solid red lines) and males (solid blue lines) (A–H). Data are presented as the means ± standard deviation. The different letters are significantly different based on one-way analysis of variance (ANOVA) and Tukey’s test with differences considered significant at p < 0.05.

Figure 5

A hypothetical circadian clock model for Pachycrepoideus vindemmiae.