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Infectious serositisis a considerable economic problem in duck industry caused by Riemerella anatipestifer. The current study was conducted to investigate the circulating R. anatipestifer in ducks in Assiut Province and assessing their antimicrobial susceptibility. One-hundred and twenty diseased or freshly dead ducks aging 1-18 weeks were examined. Naturally infected birds showed respiratory, nervous, and locomotor disturbances, and low body weight. R. anatipestifer was detected in 16.6% (20) of birds. Among the bacteriologically positive 20 birds, only 10 could be identified by PCR as R. anatipestifer with a prevalence rate of 8.33%. The sensitivity biogram revealed that all the obtained isolates were sensitive to amoxicillin, doxycycline, and flumequine while resistance to streptomycin, chloramphenicol, ampicillin, erythromycin, spectinomycin, and cephradine was observed. On the basis of MIC, all isolates had 90- 100% sensitivity to doxycycline and amoxicillin, respectively. Experimentally, the isolated R.anatipestifer strains showed pathogenicity to 14-days-old ducklings.