Bovine mastitis caused by P. aeruginosa has received more attention in the recent years as it causes significant economic losses to dairy farms and to dairy industry as a whole. P. aeruginosa mastitis is a multifactorial disease needs interaction of pathogen, host and environment. The emergence and spread of antimicrobial resistance in P. aeruginosa is one of the most important health issues worldwide. This study aimed to isolate P. aeruginosa from mastitic milk samples, study its associated virulence markers and studying their AMR either phenotypically or genotypicaly by detection of ARG in addition to studying immunological parameters in milk and serum samples. 200 milk samples were collected from mastitic cows and buffaloes and examined for existence of P. aeruginosa, the recovered isolates were identified by conventional bacteriological procedure then confirmed by MALDI-TOF revealing a recovery rate (7%) in the examined samples. Antimicrobial susceptibility test showed that isolates exhibited high resistance rates to ampicillin, amoxicillin erythromycin and SXT (100%), followed by ceftriaxone and cefidizime, (92.8%), moderate resistance to chloramphenicol (64.2%) and doxycycline (57.4%). Meanwhile isolates showed high susceptibilities to ciprofloxacin, gentamycin, amakacin, impenim and colistin. PCR results showed amplification of oprL, toxA and pslA virulence genes in all examined isolates. ARG detected by PCR were ampC, blaTEM, blaCTX, sul1 and ermB. Result showed significant increase in gene expression of Bhb1, IL6 and in lysozyme activity in serum samples, meanwhile milk samples showed increase in Bhb1 and decrease in IL10 with no change in IL6 gene expression when compared with the control.