To quantify heat tolerance in insects, two manual observation measures are typically implemented: the time to physiological collapse at a static noxious temperature (time to knockdown; TKD) or the temperature at which collapse occurs as temperature increases (critical thermal maximum; CTmax). Both assay modalities focus on physiological collapse, neglecting the prior behavioral processes. In this study, the locomotion response of Drosophila melanogaster to relatively high temperature (39 and 40.5 °C) was quantified using the TriKinetics Drosophila Activity Monitor (DAM2 system). The absence of locomotion was defined as the state of physiological collapse resulting from extended exposure to high temperature. An easy-to-use executable application that allows the user to automatically extract individual TKD from the activity data was developed. For validation, manual TKD assays were performed in parallel to automated assays across multiple factors, including sex, hardening, recovery time after hardening, and assay temperature, which gave similar results. In terms of behavioral aspects, heat hardening consistently led to reduced activity during a subsequent heat stress, irrespective of assay temperature, sex, or recovery time after hardening. Our automated heat tolerance assay utilizing the DAM2 system is one way to expand the scope of the heat tolerance phenotype to include a behavioral component in conjunction with the traditional TKD measure.
Thermotolerance , Animals , Drosophila melanogaster/genetics , Hot Temperature , Phenotype , Drosophila
Drosophila melanogaster Nora virus (DmNV) is a novel picorna-like virus first characterized in 2006. Since then, Nora virus has been detected in several non-Drosophila species, including insects in the Orders Hymenoptera, Lepidoptera, Coleoptera, and Orthoptera. The objective of this study was to determine if DmNV could infect individuals of other species of invertebrates besides D. melanogaster. The presence of DmNV in native invertebrates and commercially available stocks was determined. Laboratory-reared D. yakuba, D. mercatorum, Gryllodes sigillatus, Tenebrio molitor, Galleria mellonella, and Musca domestica were intentionally infected with DmNV. In addition, native invertebrates were collected and D. melanogaster stocks were purchased and screened for DmNV presence using reverse transcription-polymerase chain reaction (RT-PCR) before being intentionally infected for study. All Drosophila species and other invertebrates, except M. domestica, that were intentionally infected with DmNV ended up scoring positive for the virus via RT-PCR. DmNV infection was also detected in three native invertebrates (Spilosoma virginica, Diplopoda, and Odontotaenius disjunctus) and all commercially available stocks tested. These findings suggest that DmNV readily infects individuals of other species of invertebrates, while also appearing to be an endemic virus in both wild and laboratory D. melanogaster populations. The detection of DmNV in commercially available stocks presents a cautionary message for scientists using these stocks in studies of virology and immunology.
