Antibiotic resistance has become a global threat in which the anthropogenically influenced aquatic environment represents not only a reservoir for the spread of antibiotic resistant bacteria (ARB) among humans and animals but also an environment where resistance genes are introduced into natural microbial ecosystems. Wastewater is one of the sources of antibiotic resistance. The aim of this research was the evaluation of wastewater impact on the spread of antibiotic resistance in the water environment. In this study, qPCR was used to detect antibiotic resistance genes (ARGs)—blaCTX-M-15, blaCTX-M-32, ampC, blaTEM, sul1, tetM and mcr-1 and an integron detection primer (intl1). Detection of antibiotic resistant Escherichia coli was used as a complement to the observed qPCR results. Our results show that the process of wastewater treatment significantly reduces the abundances of ARGs and ARB. Nevertheless, treated wastewater affects the ARGs and ARB number in the receiving river.