Evaluation of a direct method for detecting extended-spectrum β-lactamase using the Cica-beta test on positive blood culture bottles

The global prevalence of extended-spectrum β-lactamase (ESBL)-producing Enterobacterales is a growing concern, underscoring the need for rapid detection to enable timely, targeted therapy. Conventional methods for ESBL detection, typically involving subculture followed by phenotypic screening and confirmatory tests based on the Clinical and Laboratory Standards Institute guidelines, require extended turnaround times. While rapid molecular diagnostic tests exist, their availability can be limited in some settings due to cost or infrastructure requirements, whereas matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for bacterial identification is more widely adopted. The chromogenic cephalosporin HMRZ-86-based Cica-beta test is a simpler alternative but traditionally requires colonies from subcultures. This study evaluated a direct Cica-beta test using positive blood cultures for rapid extended-spectrum β-lactamase detection. Using bacterial pellets prepared from 109 positive blood cultures of <i>Escherichia coli</i>, <i>Klebsiella pneumoniae</i>, <i>Klebsiella oxytoca</i>, or <i>Proteus mirabilis</i>, the direct Cica-beta test demonstrated high sensitivity (96.3%, 95% confidence interval [CI]: 81.0-99.9) and specificity (100%, 95% CI: 93.5-100), with a positive predictive value of 100% (95% CI: 81.0-100) and a negative predictive value of 98.8% (95% CI: 93.5-100), when compared to conventional methods. This direct method provides a faster alternative to traditional subculture-based testing, enabling prompt antibiotic adjustments. Although challenges remain, such as the potential misclassification of certain β-lactamases, such as AmpC, this approach offers a promising, cost-effective strategy for rapid diagnosis. It has the potential to improve patient outcomes by facilitating timely and appropriate antimicrobial therapy. Further multicenter studies are necessary to validate these findings.IMPORTANCEExtended-spectrum β-lactamase-producing Enterobacterales pose significant challenges in clinical practice due to their resistance to β-lactam antibiotics and association with increased mortality in bloodstream infections. Rapid and accurate identification of these organisms remains crucial for guiding initial antimicrobial therapy and improving patient outcomes. Conventional testing methods are time-consuming, and advanced molecular techniques, such as FilmArray, are costly and not universally available. This study evaluated a novel application of the chromogenic cephalosporin HMRZ-86-based Cica-beta test, a direct method using bacterial pellet from positive blood cultures for extended-spectrum β-lactamase detection. It demonstrated high diagnostic accuracy, with sensitivity and specificity comparable to conventional and FilmArray methods, while significantly reducing result turnaround time. This cost-effective, rapid diagnostic tool offers practical advantages in diverse clinical settings and resource-limited environments. Its implementation has the potential to enhance antimicrobial stewardship programs, support timely clinical decision-making, and aid efforts to combat antibiotic resistance.