The mustard leaf beetle, Phaedon cochleariae, as a screening model for exogenous RNAi-based control of coleopteran pests.

RNA interference (RNAi) is a promising, selective pest control technology based on the silencing of targeted genes mediated by the degradation of mRNA after the ingestion of double-stranded (ds) RNA. However, the identification of the best target genes remains a challenge, because large scale screening is only feasible in lab model systems and it remains unclear, to what degree such data can be transferred to pest species. Here, we report on our efforts to transfer target genes found in a lab model to the mustard leaf beetle, Phaedon cochleariae. The mustard leaf beetle can be reared easily and resource-efficient in large quantities all year round and is an established chrysomelid pest for higher throughput screening approaches in the crop protection industry. Mustard leaf beetle transcriptome sequencing and assembly revealed genes orthologous to those previously described as highly efficient RNAi targets in the model beetle Tribolium castaneum. First, we observed mortality after injection of dsRNA targeting the respective orthologous genes in 2nd instar mustard beetle larvae. Next, we adopted a robust, automated multi-well plate foliar RNAi screening procedure with 2nd instar larvae of the mustard leaf beetle to assess those genes. Indeed, foliar application and oral uptake of dsRNA targeting the same genes resulted in larval mortality as well. The most effective target genes with a strong (lethal) phenotype - at dsRNA doses as low as 300 ng/leaf disc (equal to 9.6 g/ha) - were srp54k, rop, αSNAP, rpn7 and rpt3. Rather limited effects were observed after application of dsRNA targeting cactus, shibire and PP-α, though they had previously been shown to be highly lethal in red flour beetle. Importantly, our experiments demonstrated that the overall efficacy pattern obtained after oral dsRNA application was well correlated with the results obtained after dsRNA injection. RT-qPCR confirmed significant target gene knock-down after normalization by employing three reference genes shown to be stably expressed across life stages. In summary, several RNAi targeted genes elicited a strong lethal phenotype and significant target gene knock-down after feeding, suggesting P. cochleariae as a potential coleopteran screening model for foliarly applied exogenous RNAi.

A Dynamic Energy Budget Approach for the Prediction of Development Times and Variability in Spodoptera frugiperda Rearing.

A major challenge in insect rearing is the need to provide certain life cycle stages at a given time for the initiation of experimental trials. The timing of delivery, organism quality, and variability directly affect the outcome of such trials. Development times and intraspecific variability are directly linked to the availability of food and to the ambient temperature. Varying temperature regimes is an approach to adapt development times to fulfill experimental needs without impairment of larval quality. However, current practices of temperature setting may lead to increased variability in terms of development times and the frequency of particular life stages at a given point in time. In this study, we analyzed how resource availability and ambient temperature may affect the larval development of the economically important noctuid species Spodoptera frugiperda by means of dynamic energy budget modeling. More specifically, we analyzed how rearing practices such as raising of temperatures may affect the variability in larval development. Overall, the presented modeling approach provides a support system for decisions that must be made for the timely delivery of larvae and reduction of variability.