Increasing human population size and income growth are causing an increasing demand for food and feed. Insects are a more sustainable alternative to conventional animal source proteins, as they can convert waste and by-products from the agricultural industry into biomass for commercial feed for livestock and, potentially, serve as a food source for human consumption. Moreover, insects together with their microorganisms have been shown to play a pivotal role in the development of insects and in the breakdown of complex growth substrates, and are, therefore, closely tied to insect production. This study aims to determine if the removal of egg- and substrate-associated microorganisms impacts larval performance (growth, final biomass, and the survival rate) of Linnaeus. Four treatments are tested: disinfected eggs and non-autoclaved substrate, non-disinfected eggs and autoclaved substrate, disinfected eggs and autoclaved substrate, and a control without any removal of microbiota. The results show a significant decrease in the final biomass of larvae subjected to the treatments with only disinfected eggs, only autoclaved substrate, and both compared to the control, and a significant decrease in survival rate for non-disinfected eggs and autoclaved substrate and disinfected eggs and autoclaved substrate compared to the control group. Moreover, larval growth shows a significant difference across days within all treatments. Together, this suggests that the microorganisms of housefly eggs and the growth substrate play an important role in biomass, which is critical in commercial insect production. Together this suggest, that more studies are needed to examine these parameters with respect to more commercially relevant substrates.