Background: Ascites is a recognized problem in patients with decompensated liver cirrhosis. Spontaneous bacterial peritonitis (SBP) is a massive problem in patients who suffer from cirrhosis with ascites. Early bacterial detection allows great intervention to stop SBP. 16S ribosomal RNA (rRNA) is a universal gene used to detect different bacteria present within a sample. Objectives: This study aimed to evaluate the efficacy of broad range 16S rRNA gene polymerase chain reaction (PCR) in diagnosis of ascitic fluid (AF) infection. Methodology: Fifty cirrhotic ascitic patients were undergone to full history, clinical examination, laboratory tests including, AF specimens analysis for polymorph nuclear leucocytic (PMN) count, culture for bacteria and PCR- for DNA detection of bacteria. Results: Bacteria were separated from 21 (42%) of samples of ascitic fluid, and they were mainly Gram-negative bacteria. The sensitivities of culture for bacteria and PCR in diagnosing AF infection were 53% and 86% respectively, while the accuracies were 62% and 74% respectively relation with PMNL. Conclusions: Bacterial DNA in AF samples may be another method for diagnosis AF infection rather than bacterial culture and PMN count for early detection and treatment of AF infection.