Pasteurella multocida is a significant pathogen that sporadically infects poultry flocks, often causing high mortality and resulting in considerable economic losses. This study sought to isolate Pasteurella multocida from laying chickens in Karbala province, detection rate, and confirmation the infection via molecular analysis. A total of 90 samples of 30 chickens showing signs of infection (e.g., nasal discharge, anorexia, sudden death) were sampled. Samples included 30 nasal swabs, 30 keel bone, and 30 lungs. Isolation was performed using selective Cystine Glucose Trypticase medium and blood agar under aerobic conditions. Suspected colonies were identified based on their morphology, odour, and biochemical characteristics. Molecular detection of virulence genes was performed using PCR to target specific gene sequences. Chickens exhibited clinical signs including pyrexia, mucopurulent nasal discharge, watery diarrhea, and sudden death. Necropsy revealed severe lung congestion (purulent pneumonia) and generalised internal organ congestion, particularly in the intestines. The isolates formed grey round colonies with a smooth or mucoid, non-hemolytic texture and a characteristic “mousy” odour, 48 hours of aerobic incubation at 37°C. No growth on MacConkey agar was for confirmation with negatively urea test. Culture positivity was highest in nasal swabs (66.67%), followed by keel bones (60%) and lungs (20%). PCR detection was also most successful in keel bones (43.33%), followed by nasal swabs (36.67%) and lungs (10%). Given the aggressive impact of this disease on the poultry industry, there is a need to improve diagnostic monitoring and biosecurity measures to prevent further dissemination in Iraqi poultry farms. The results of the study can be used in veterinary diagnostic practice for the timely detection of Pasteurella multocida and for improving control measures against fowl cholera on poultry farms
pasteurellosis; fowl cholera; culture; ecbJ gene; poultry