The present study was conducted on 145 clinically diseased fish samples, 55 Nile tilapia; Oreochromus niloticus (O.niloticus); 65 African catfish (C. gariepinus), and 25 Grey mullet, Mugilcephalus (M. cephalus) of various sizes. Fishes were collected from different fish markets at Qalubiya Governorate, Egypt, during the period from September 2021 to May 2022 for isolation of Pseudomonas (Ps) aeruginosa with molecular studying of its virulence factors. The results cleared that 43 Ps. aeruginosa   isolates (13.8%) were recovered as follow;14/55 from O. niloticus (25.4%), 19/65 from C. gariepinus (29.2%) and 10/25 from M. cephalus (40.0%). The isolated Ps. aeruginosa were highly resistant for ampicillin; penicillin-G followed by oxacillin; amoxicillin; cefotaxime and tetracycline. In contrast, they were sensitive to gentamicin followed by norfloxacin, ciprofloxacin and doxycycline. PCR results showed that, exotoxin A (toxA); outer membrane lipoprotein L (oprL) and exo polysaccharide synthesis locus (pslA) virulence gene were detected in three out of six studied Ps. aeruginosa isolates and exotoxin S (exoS) virulence gene was detected in five out of six studied ones. It is concluded that Ps. Aeruginosa has multiple antibiotic resistances and virulence genes, therefore it is an important pathogen causes Pseudomonas septicemia in fish and could have a public health concern for the consumers