Treatment strategies for acute uncomplicated appendicitis have evolved and now conservative antibacterial treatment is recommended over surgical treatment, especially for paediatric patients. The aim of this study was to evaluate microbiota in paediatric patients with acute uncomplicated and complicated appendicitis, and antibacterial susceptibility of the causative microorganisms. Bacteriological identification was conducted using the VITEK2 analyser. Antibacterial susceptibility tests were performed and the results were evaluated in accordance with the recommendations of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) “Clinical breakpoints and dosing of antibiotics” (Version 7.0, January 2019). Serodiagnosis of Yersinia enterocolitica was performed using indirect haemagglutination. The results revealed differences in microbiota in cases of acute complicated and acute uncomplicated appendicitis. Pseudomonas aeruginosa was identified more frequently in cases of acute complicated appendicitis. Mixed culture was prevalent in cases of both acute complicated and acute uncomplicated appendicitis. Very few positive extended spectrum beta-lactamase (ESBL) Escherichia coli cultures were identified. Most of strains of Pseudomonas aeruginosa were resistant to amoxicillin with clavulanic acid, ertapenem, ampicillin and cefotaxime. Some of E. coli isolates were resistant to ampicillin and to amoxicillin with clavulanic acid.