Credit: RooM the Agency/Alamy Stock Photo

Cotesia vestalis is a parasitic wasp, which lays its eggs in larvae of the moth Plutella xylostella. With its eggs the wasp releases into the host the symbiotic polydnavirus C. vestalis bracovirus (CvBV) and embryonic cells called teratocytes. Wang et al. now show that the teratocytes and CvBV introduce microRNAs (miRNAs) that can delay larval growth.

The authors took advantage of the recently sequenced genomes of C. vestalis and P. xylostella to identify C. vestalis miRNAs and assess their roles in parasitism. They produced small nuclear RNA libraries from teratocytes and larvae of C. vestalis and identified >150 miRNAs.

Twenty of the teratocyte-expressed miRNAs were encoded by the CvBV provirus (the first polydnavirus miRNAs to be described). Analysis of the expression of several CvBV-encoded miRNAs in P. xylostella larvae at different times after egg deposition showed that they were expressed and indicated that they emerge through both viral infection and expression in teratocytes.

Whereas viral infection could directly deliver miRNAs to host cells, transmission of teratocyte-expressed miRNAs to host cells was suggested to be mediated by extracellular vesicles such as exosomes. Following co-culturing of teratocytes with Pxem_ZJU cells, which is an established P. xylostella cell line, teratocyte-produced exosomes were indeed taken by Pxem_ZJU cells.

Twenty teratocyte-expressed miRNAs — especially miR-375-3p and miR-281-3p, which are also endogenously expressed in P. xylostella — were more abundant in Pxem_ZJU cells co-cultured with teratocytes than in control cells. Moreover, in haemocytes collected from parasitized P. xylostella, most of the miR-281-3p and miR-375-3p miRNAs were of C. vestalis origin. Thus, miRNAs released from teratocytes were taken up by the parasitized larvae.

miRNAs released from teratocytes were taken up by the parasitized larvae

Almost 100 P. xylostella genes were predicted to be targets of the most abundant miRNAs expressed in teratocytes and/or encoded by CvBV. One of the prominent targets was ecdysone receptor (EcR), which was predicted to be targeted by Cve-miR-281-3p (encoded by the C. vestalis genome) and Cve-miR-novel22-5p (encoded by the CvBV genome).

P. xylostella larvae parasitized by C. vestalis exhibit pronounced size reduction and inhibition of metamorphosis. Analysis by dual-luciferase reporter assays supported the targeting of EcR by Cve-miR-281-3p and Cve-miR-novel22-5p, and injection of mimics (agomirs) of either miRNA into larvae inhibited EcR expression. As EcR mediates the response to the moulting hormone ecdysone in insects, its inhibition by Cve-miR-281-3p and Cve-miR-novel22-5p could explain the growth delays of parasitized larvae.

In summary, C. vestalis-derived miRNAs of both CvBV and teratocyte origin — some of which target the same P. xylostella genes — are expressed in host tissues and can contribute to the developmental delays observed in parasitized larvae, at least partly by inhibiting EcR. This could guarantee the availability of host resources for wasp development. As the relationship between the miR-281 family and EcR is conserved in butterflies and moths, other parasites are likely to employ the same strategy.