BACKGROUND: Beta-lactamase enzymes are considered the most important factor of resistance against b-lactam antibiotics among gram-negative bacteria. In recent years, the production of extended-spectrum b-lactamases has been prevailed among bacteria, especially bacteria of animal origin, and this is important in terms of public health. OBJECTIVE: The prupose of this study is to evaluate the presence of extended-spectrum b-lactamase (ESBL)-genes blaCTX-M, blaTEM and blaSHV in E. coli isolates recovered from fecal samples of apparently healthy water buffaloes (Bubalus bubalis) using polymerase chain reaction. METHODS: In this study, 105 isolates of E. coli, which were obtained from 135 fecal samples of water buffaloes from different areas of West Azerbaijan province (33 isolates from Urmia, 33 isolates from Khoy, 24 isolates from Piranshahr and 15 isolates from Miandoab), were identified using biochemical characteristics as well as 23S rRNA gene amplification. Then, the presence of CTX-M, TEM, and SHV groups of ESBL genes were evaluated among the studied E. coli isolates by the PCR method. RESULTS: In the studied isolates, 47 out of 105 E. coli isolates (44.8%) contained blaCTX-M gene and 37 isolates (35.2%) harbored blaTEM gene. Also, 17 isolates (16.2%) contained both blaCTX-M and blaTEM genes simultaneously. According to the results, blaSHV gene was not detected among the studied isolates. Also, no significant difference was seen in distribution of ESBL genes among the studied regions. CONCLUSIONS: The results of this study indicate that water Buffalo gastrointestinal E. coli is reservoir for ESBLs, especially CTX-M and TEM types, and this should be considered in terms of public health and the transfer of resistance genes to pathogenic bacteria.