Recycling treated wastewater is considered one of the most critical sectors in recycling. Its benefit fell beyond the disposability of sewage flow that extends to the possibility of reusing the water. Wastewater treatment in any system depends exclusively on maximizing the activity of the microbial community. Wastewater samples were collected from two systems under the same administration in one of the largest wastewater treatment plants (WWTPs) in the Middle East (Al-Jabal Al-Asfar). Five major collection points for the daily assessment of the plant were used for collecting samples. In this study, real-time quantification polymerase chain reaction (qRT-PCR) was used to quantify the microbial community to evaluate the two treatment systems in the plant. Based on the results, the microbial community in each of the five major points from both systems was quantified based on the linear model produced from qRT-PCR. One of the two systems was active and efficient, in contrast to the other, which lacked profound control over the treatment. The current study suggests the routine usage of qRT-PCR as a crucial molecular genetics technique to evaluate the microbial community within the wastewater to ensure reliable and efficient treatment of the raw sewage. The study recommends the full assessment of the inactive system within the plant, encouraging the usage of compatible microbes to enrich and activate the returned activated sludge (RAS) for better performance.