For many years, the University of Chicago administered sulfamethoxazole-trimethoprim sulfate (SMZ-TMP) oral suspension to select immunocompromised mouse colonies via the drinking water. In 2014, SMZ-TMP oral suspension was placed on back-order and medicated diet with a different sulfonamide, sulfadiazine-trimethoprim (SDZ-TMP) was used as a replacement. Months after this transition, sentinel mice from the same room as one of the remaining immunocompromised colonies on this diet were found dead or appeared sick. Necropsies revealed cardiomegaly, and histology confirmed myocardial fibrosis in the first 4 sentinel mice examined, consistent with cardiomyopathy. Subsequent sequential monitoring of 2 sentinel mice via echocardiography showed their progression toward decreased cardiac function. Investigation of the housing room revealed that the sentinel mice had been accidently placed on SDZ-TMP diet upon entering the colony housing room. This case report describes cardiomyopathy in 6 ICR mice after long term consumption of SDZ-TMP medicated feed.