MUNICIPAL sewage, septic tanks, fertilizers, animal feedlots and irrigation wastewater are the common sources of groundwater pollution in rural areas. Sedimentation pools of wastewater treatment plants are possibly suggested to be another source of pollution. Two wastewater treatment plants located at Qalyubia governorate named Qalyub (QWTP) and Shubra Al-khayma (SHWTP) were selected to carry out the present study; two samples from influent and effluent were collected seasonally from both plants. Furthermore, 6 groundwater samples were collected from nearby rural houses of both plants. The physicochemical and microbiological properties were evaluated for all samples. Moreover, Box-PCR for 12 strains isolated from wastewater and groundwater at Qalyub district was carried out. Water quality assessment studies proven that the effectiveness of the two wastewater treatment plants under investigation is questionable, especially QWTP. The percentages of removing of TSS, COD, BOD, ammonia, total coliform and fecal coliform counts were 87, 74, 88, 66, 94 and 89%, respectively, for QWTP, and 85, 89, 93, 86, 94 and 83%, respectively, for SHWTP. The groundwater at Qalyub district was contaminated with Mn, Fe, ammonia, BOD, coliform bacteria, fecal streptococci, E. coli, Aerumonas hydrophilla and Pseudomonas aeruginosea, while the groundwater at Shubra Al-khayma district was contaminated with Mn, coliform bacteria, fecal streptococci and P. aeruginosea. Microbial and chemical evaluation of groundwater, as well as, Box-PCR results proven that, the contamination of the tested groundwater might be of other source than water treatment plants.