Evaluation of the NG-Test® CTX-M MULTI lateral flow immunoassay and genomic discovery of a blaCTX-M-27 variant with a premature stop codon

Abstract

Background

Timely identification of extended-spectrum beta-lactamase (ESBL)-producing bacteria supports early antimicrobial optimization, especially for patients with invasive infections. NG-Test® CTX-M MULTI (CTX-M LFA) is a rapid immunochromatographic assay for detection of Cefotaximase-Munich (CTX-M)-type ESBL. We evaluated its performance against existing molecular platforms.

Methods

A sampling of archived Gram-negative bacteria (including Enterobacterales and non-fermenters) was included in the study. All isolates underwent testing for CTX-M ESBL by CTX-M LFA. CTX-M LFA results were compared to results from molecular methods, laboratory-developed (LD)-PCR (2013-2019) or BCID2 (2022-2023). Discordances were resolved with a combination of ESBL phenotypic testing and an alternative PCR platform, or whole-genome sequencing.

Results

Seventy-eight isolates were included in the study, all of which underwent CTX-M LFA and molecular testing (n = 47 by LD-PCR, n = 31 by BCID2). Post-discordance analysis showed positive percent agreement of 89.7 % (95 %CI 71.5-97.2 %), negative percent agreement 100 % (95 %CI 90.9-100 %) and overall categorical agreement 96.2 %, κ=0.92 (95 %CI 88.4-99.0 %). One E. coli was negative by CTX-M LFA but positive for bla_CTX-M by BCID2. Whole-genome sequencing identified a non-functional bla_CTX-M-27 gene with adenine insertion at position 70 in codon 24, causing a frameshift mutation resulting in a premature stop codon (codon 58).

Conclusion

CTX-M LFA improved performance in comparison with molecular methods by accurately identifying an isolate with a prematurely truncated protein as a true-negative for CTX-M. This highlights potential limitations of molecular-based resistance detection. CTX-M LFA was simple to perform and yielded results within 15 minutes.