Intestinal parasites continue to pose a significant threat to human health worldwide, particularly among children. Contaminated water and soil serve as major transmission vehicles for these parasites and intestinal protists are among the most prevalent parasites in both developed and developing nations. Traditionally, parasites have been studied using human or animal fecal samples, while studying them in environmental samples has been challenging due to technical limitations. However, advancements in Next-Generation Sequencing (NGS) and bioinformatic approaches now enable the detection of parasite DNA in environmental samples. In this study, we applied a metataxonomic and phylogenetic strategy to detect and classify DNA of protists present in sewage sludge from two major cities in Colombia: Medellin and Cali. We successfully detected several human pathogenic parasites including Giardia intestinalis, Entamoeba histolytica, and Blastocystis sp., among other protists, in all sludge samples examined. We also investigated the entry and exit of parasite DNA from the San Fernando wastewater treatment plant (WWTP). We observed a higher number of parasite DNA sequences in the plant's influent wastewater, but we also detected the discharge of DNA from pathogenic parasites in both effluent waters and biosolids.