An Optimized Small-Scale Rearing System to Support Embryonic Microinjection Protocols for Western Corn Rootworm, Diabrotica virgifera virgifera.

Western corn rootworm (WCR), a major pest of corn, has been reared in laboratories since the 1960s. While established rearing methods are appropriate for maintaining WCR colonies, they are not optimal for performing germline transformation or CRISPR/Cas9-based genome editing. Here we report the development of an optimized rearing system for use in WCR functional genomics research, specifically the development of a system that facilitates the collection of preblastoderm embryos for microinjection as well as gathering large larvae and pupae for downstream phenotypic screening. Further, transgenic-based experiments require stable and well-defined survival rates and the ability to manipulate insects at every life stage. In our system, the WCR life cycle (egg to adult) takes approximately 42 days, with most individuals eclosing between 41 and 45 days post oviposition. Over the course of one year, our overall survival rate was 67%. We used this data to establish a quality control system for more accurately monitoring colony health. Herein, we also offer detailed descriptions for setting up single-pair crosses and conducting phenotypic screens to identify transgenic progeny. This study provides a model for the development of new rearing systems and the establishment of highly controlled processes for specialized purposes.

How Rearing Systems for Various Species of Flies Benefit Humanity.

Flies (Diptera) have played a prominent role in human history, and several fly species are reared at different scales and for different beneficial purposes worldwide. Here, we review the historical importance of fly rearing as a foundation for insect rearing science and technology and synthesize information on the uses and rearing diets of more than 50 fly species in the families Asilidae, Calliphoridae, Coelopidae, Drosophilidae, Ephydridae, Muscidae, Sarcophagidae, Stratiomyidae, Syrphidae, Tachinidae, Tephritidae, and Tipulidae. We report more than 10 uses and applications of reared flies to the well-being and progress of humanity. We focus on the fields of animal feed and human food products, pest control and pollination services, medical wound therapy treatments, criminal investigations, and on the development of several branches of biology using flies as model organisms. We highlight the relevance of laboratory-reared Drosophila melanogaster Meigen as a vehicle of great scientific discoveries that have shaped our understanding of many biological systems, including the genetic basis of heredity and of terrible diseases such as cancer. We point out key areas of fly-rearing research such as nutrition, physiology, anatomy/morphology, genetics, genetic pest management, cryopreservation, and ecology. We conclude that fly rearing is an activity with great benefits for human well-being and should be promoted for future advancement in diverse and innovative methods of improving existing and emerging problems to humanity.

Deliberately added and "cryptic" antioxidants in three artificial diets for insects.

Characteristics of both deliberately added and "cryptic" antioxidants were assayed from hydrophilic and lipophilic extracts from artificial diets for plant bugs, lepidopteran larvae, and green lacewings. Cryptic antioxidants are defined as substances naturally existing in diet ingredients but not deliberately added because of their antioxidant potential. Diets were tested after 1) being freshly produced, 2) stored for 48 h at 4 degrees C, or 3) held for 48 h under rearing room conditions at 27 degrees C. Tests included 1) a general assay of antioxidant capacity known as the ferric-reducing antioxidant power (FRAP) assay. 2) a cation radical-scavenging assay, 3) an ascorbic acid assay, and 4) an assay of inhibition of lipid peroxidation. In all assays, the lepidopteran diet had the highest values for protection against reactive oxygen species (ROS). The lepidopteran diet (with 0.17-0.23-mg equivalents of gallic acid equals total phenolic compounds per gram of diet) had three- to four-fold higher concentrations of phenolic compounds than did either the plant bug diet or the lacewing diet. Unexpectedly, the plant bug and the lacewing diets caused more lipid peroxidation than did the positive controls. This was attributed to the high concentrations of iron in these diets (mainly from chicken eggs), causing an ascorbate-ferric ion-induced lipid peroxidation. Diet storage, measured after 2 d at 27 or 4-6 degrees C, caused no significant declines in overall antioxidant potential. However, storage did lead to decline in ascorbic acid. The FRAP assay offered the best potential as a general, routine test of the potential of various insect diets to resist the destructive effects of ROS. The importance of addressing issues of protection against ROS in insect diets is discussed.