BackgroundThe prevalence of extended-spectrum β-lactamases (ESBLs) have been reported in clinical isolates obtained from various hospitals in Ethiopia. However, there is no data on the prevalence and antibiotic susceptibility patterns of CTX-M type ESBL produced by Gram-negative bacilli. The aim of this study was to determine the frequency and distribution of the blaCTX-M genes and the susceptibility patterns in ESBL producing clinical isolates of Gram-negative bacilli in Jimma University Specialized Hospital (JUSH), southwest Ethiopia.MethodsA total of 224 non-duplicate and pure isolates obtained from clinically apparent infections, were included in the study. Identification of the isolates was performed by MALDI-TOF mass spectrometry. Susceptibility testing and ESBL detection was performed using VITEK® 2, according to EUCAST v4.0 guidelines. Genotypic analysis was performed using Check-MDR CT103 Microarrays.ResultsOf the total 112 (50.0%) isolates screen positive for ESBLs, 63.4% (71/112) tested positive for ESBL encoding genes by Check-MDR array, which corresponds to 91.8% (67/73) of the total Enterobacteriaceae and 10.3% (4/39) of nonfermenting Gram-negative bacilli. Among the total ESBL gene positive isolates, 95.8% (68/71) carried blaCTX-M genes with CTX-M group 1 type15 being predominant (66/68; 97.1% of CTX-M genes). The blaCTX-M carrying Enterobacteriaceae (n = 64) isolates showed no resistance against imipenem and meropenem and a moderate resistance rate against tigecycline (14.1%), fosfomycin (10.9%) and amikacin (1.6%) suggesting the effectiveness of these antibiotics against most isolates. On the other hand, all the blaCTX-M positive Enterobacteriaceae showed a multidrug resistant (MDR) phenotype with remarkable co-resistances (non-susceptibility rates) to aminoglycosides (92.2%), fluoroquinolones (78.1%) and trimethoprim/sulfamethoxazol (92.2%).ConclusionsThis study demonstrates a remarkably high prevalence of blaCTX-M genes among ESBL-producing isolates. The high level of resistance to β-lactam and non-β-lactam antibiotics as well as the trend to a MDR profile associated with the blaCTX-M genes are alarming and emphasize the need for routine diagnostic antimicrobial susceptibility testing for appropriate choice of antimicrobial therapy.Electronic supplementary materialThe online version of this article (10.1186/s12879-018-3436-7) contains supplementary material, which is available to authorized users.