A total of 100 random samples of raw milk were collected from different markets in Port- Said city aseptically for isolation and identification of Pseudomonas species. The results revealed that the Pseudomonas species could be detected in a percentage of 6% from raw milk samples. Six strains were identified biochemical as Pseudomonas aeruginosa. This study confirms the rapidity and sensitivity of 16SrDNA analysis in identifying strains which contribute in early monitoring, accurate analysis and control of microbial risks in food products. The described methodology has special relevance in raw milk quality control and safety. Molecular characterizations of P. aeruginosa were confirmed using 16SrDNA of Pseudomonas spp. and 16SrDNA of P. aeruginosa by conventional PCR. Which were developed with specific primers for the detection of different virulence genes (oprL, toxA, lasI, rhlR, ExoU, ExoS and ecfX) of P. aeruginosa, which may be considered a significant in food safety threat. P. aeruginosa possesses a variety of virulence genes that may contribute to its pathogenicity. Our results showed that oprL and oxA genes were detected in all 6 tested strains of P. aeruginosa, while lasI gene was detected in strains 1,2,3,4. But ExoS gene was detected only in 1, 2, and 3 strains. On the other hand Rhl1 gene was detected in 1, 3, and 4 tested strains. ExoU, rhlR, and ecfX genes not detect in any tested strains. PCR method is rapid than other diagnostic methods for the identification of P. aeruginosa strains, without using additional biochemical tests and It is a useful technique for detection its virulence genes. The public health hazard of this microorganism, as well as recommended measures to improve quality status of raw milk was discussed.