Carrion beetles of the subfamily Nicrophorinae search and bury a carcass that they utilize for reproduction. After burial, the carcass is coated with oral secretions that delay its decomposition. Previously, oral secretions of Nicrophorus marginatus were found to show antimicrobial activity, whereas secretion of N. carolinus lacked significant activity. Here, we tested the effects of temperature, sex of the beetle, and food type on the antimicrobial properties of oral secretions of both species. Unlike previous findings, we found that oral secretions of N. carolinus had antimicrobial activity. Temperature had significant effects on the amount of secretion protein. When protein concentrations were standardized to 1 µ/ml, N. marginatus secretions had higher antimicrobial activity at cooler temperatures, while N. carolinus had higher activity at warmer ones. The sex of the beetle did not affect antimicrobial activity for either species. Beetles of both species that were fed whole rats contained more protein in their oral secretions than beetles fed with equally sized pieces of raw ground beef. After standardizing the resulting protein concentrations to 1 µg/ml, antimicrobial activity of oral secretion increased for N. carolinus after rat feeding, but not for N. marginatus. Our results highlight key ecological differences between these closely related species. In addition, they demonstrate the importance of experiments being conducted under varying environmental conditions when evaluating species for potential antimicrobial compounds.