Search bioRxiv⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2022.03.13.484133

Direct colorimetry of imipenem decomposition as a novel cost effective method for detecting carabapenamase producing bacteria

Kotsakis, S. D.·Lambropoulou, A.·Miliotis, G.·Tzelepi, E.·Miriagou, V.·Tzouvelekis, L. S.
View original source ↗Export metadata ↓

Abstract

In the absence of a molecule that would collectively inhibit both metallo-{beta}-lactamases and serine reactive carbapenemases, containment of their genes spreading is the main weapon currently available for confronting carbepenem resistance in hospitals. Cost effective methodologies rapidly detecting carbapenemase producing enterobacteria (CPE) would facilitate such measures. Herein a low cost CPE detection method was developed that was based on the direct colorimetry of the yellow shift caused by the accumulation of diketopiperazines - products of the acid catalyzed imipenem oligomerization - induced by carbapenemase action on dense solutions of imipenem/cilastatin. The reactions were studied by spectrophotometry in the visible spectrum using preparations of {beta}-lactamases from the four molecular classes. The effects of various buffers on reactions containing the potent carbapenemases NDM-1 and NMC-A were monitored at 405 nm. Optimal conditions were used for the analysis of cell suspensions and the assay was evaluated using 38 selected enterobacteria including 29 CPE as well as nine carbapenemase-negative strains overexpressing other {beta}-lactamases. The development of the yellow color was specific for carbapenemase containing enzyme preparations and the maximum intensity was achieved in acidic or un-buffered conditions in the presence of zinc. When applied on bacterial cell suspensions the assay could detect CPE with 96.7 % sensitivity and 100 % specificity with results being comparable to those obtained with the CARBA NP technique. Direct colorimetry of carbapenemase-induced imipenem decomposition required minimum reagents while exhibited high accuracy in detecting CPE. Therefore it should be considered for screening purposes after further clinical evaluation. ImportanceCurrently, spread of multi-drug resistant (MDR) carbapenemase-producing enterobacteria (CPE), mostly in the clinical setting, is among the most pressing public health problems worldwide. In order to effectively control CPE, use of reliable and affordable methods detecting carbapenemase genes or the respective {beta}-lactamases is of vital importance. Herein we developed a novel method, based on a previously undescribed phenomenon, which can detect CPE with few reagents by direct colorimetry of bacterial suspensions and imipenem/cilastatin mixtures.

Source connections

Explore related subjects

microbiologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Kotsakis, S. D., Lambropoulou, A., Miliotis, G., Tzelepi, E., Miriagou, V., Tzouvelekis, L. S.. 2022-03-13. Direct colorimetry of imipenem decomposition as a novel cost effective method for detecting carabapenamase producing bacteria. https://doi.org/10.1101/2022.03.13.484133

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

A population-scale landscape of the subgingival microbiome reveals divergent routes to periodontal dysbiosis

Periodontitis is an archetypical mucosal inflammatory disease in which microbiome dysbiosis at the tooth-epithelial interface interacts with host genetic and behavioral risk factors to drive immune-mediated tissue destruction. Although subgingival microbiome compositional shifts are thought to parallel disease severity, microbiome variation at the population-level and its relationship to periodontal clinical phenotypes and disease-modifying factors remain poorly defined. Here, we use unsupervised manifold learning to map the compositional landscape of the subgingival microbiome in 1,355 adults spanning periodontal health to severe periodontitis. We identified eight latent microbiome states organized along a branching continuum from eubiosis to dysbiosis. An intermediate microbial configuration marked ecological destabilization and bifurcation into two distinct periodontitis-associated dysbiotic trajectories, distinguished by links to gingival inflammation and smoking. Although the microbiome trajectories broadly tracked periodontal destruction, a minority of individuals showed discordant microbiome-clinical phenotypes, with some individuals with periodontitis retaining otherwise eubiotic microbiomes enriched for low-abundance pathobionts, while some cases of health or mild disease had highly dysbiotic communities, suggesting distinct host susceptibility. Together, these findings define a population-scale ecological landscape of the subgingival microbiome, reveal divergent trajectories to periodontal dysbiosis, and highlight heterogeneity in the relationship between microbial community structure and clinical disease expression.

microbiology↗
bioRxiv2026

Beta-lactam enhancement against methicillin-resistant Staphylococcus aureus by cell wall blockade is autolysis-dependent: a butyrolactone derivative as case in point

Methicillin-resistant Staphylococcus aureus (MRSA) is non-susceptible to beta-lactams. Blockade of cell wall biosynthesis is a potential target for beta-lactam enhancement but requires further investigation. A butyrolactone derivative enhanced beta-lactams against MRSA strains by reducing the availability of D-Ala-D-Ala. Unlike D-cycloserine, it did not inhibit D-Ala-D-Ala ligase (Ddl). Nor did it show an additive or synergistic effect when combined with cycloserine, indicating a unique mechanism for blocking cell wall precursor production that does not involve the traditional Lipid II pathway. Notably, beta-lactam potentiation by our chemical or D-cycloserine was highly dependent on the intrinsic autolytic ability of the tested MRSA strains. Strains that resisted lysis upon Triton X-100 exposure showed a minimal increase in beta-lactam susceptibility, whereas highly autolytic strains showed significant changes in their beta-lactam MICs. We have thus identified autolytic ability as the Achilles Heel in the strategy of targeting cell wall biosynthesis for beta-lactam potentiation.

microbiology↗
bioRxiv2026

Rapid and largely reversible shifts in the canine fecal metabolome during dietary change

Diet can rapidly change the fecal metabolome, but less is known about recovery after the original diet is restored. We used untargeted UPLC-MS metabolomics to analyze 72 fecal samples from nine Pumi dogs during an owner-managed switch from dry food to raw food and back to dry food. Diet phase accounted for a large proportion of variation in both ionization modes. More than 13,000 LC-MS features changed at the first sampling point after the switch to raw food, with a similarly large response after return to dry food. Among features significant in both comparisons, more than 99% changed in opposite directions. At the final sampling point, no positive-mode (ESI+) features and only 13 negative-mode (ESI-) features differed from the second dry-food baseline under the same threshold. BARF-associated patterns persisted in analyses excluding individual dogs and in pedigree-adjusted candidate models, although individual feature effects depended on normalization. Putative metabolites from several biochemical classes differed in their response and recovery. The fecal metabolome therefore changed rapidly and returned largely toward baseline, with differences among dogs.

microbiology↗