Global climate change is allowing the invasion of insect pests into new areas without natural competitors and/or predators. The dipteran Drosophila suzukii has invaded both the Americas and Europe, becoming a serious problem for fruit crops. Control methods for this pest are still based on the use of pesticides, but less invasive and more sustainable methods, such as biocontrol, are needed. Variations in environmental conditions can affect the efficacy of bioinsecticides influencing their behavior and physiology besides that of the target insects. In this work, we developed a system that simulates the daily temperature fluctuations (DTFs) detected in the environment, with the aim of studying the influence of temperature on biocontrol processes. We investigated the effects of DTFs on the efficacy of four bioinsecticides. Results showed that DTFs modify the efficacy of some entomopathogens while they are ineffective on others. Specifically, the bacterium Bacillus thuringiensis is the most effective bioinsecticide under all conditions tested, i.e., low DTF (11–22 °C) and high DTF (17–33 °C) compared to constant temperature (25 °C). In contrast, nematodes are more sensitive to changes in temperature: Steinernema carpocapsae loses efficacy at low DTF, while Steinernema feltiae and Heterorhabditis bacteriophora are not effective in controlling the target dipteran. This work provides a basis for reviewing biological control methods against invasive species in the current context of climate change.