Non-phenotypic tests to detect and characterize antibiotic resistance mechanisms in Enterobacteriaceae

Abstract

In the past two decades, we have observed a rapid increase of infections due to multidrug-resistant Enterobacteriaceae. Regrettably, these isolates possess genes encoding for extended-spectrum β-lactamases (e.g., blaCTX-M, blaTEM, blaSHV) or plasmid-mediated AmpCs (e.g., blaCMY) that confer resistance to last-generation cephalosporins. Furthermore, other resistance traits against quinolones (e.g., mutations in gyrA and parC, qnr elements) and aminoglycosides (e.g., aminoglycosides modifying enzymes and 16S rRNA methylases) are also frequently co-associated. Even more concerning is the rapid increase of Enterobacteriaceae carrying genes conferring resistance to carbapenems (e.g., blaKPC, blaNDM). Therefore, the spread of these pathogens puts in peril our antibiotic options. Unfortunately, standard microbiological procedures require several days to isolate the responsible pathogen and to provide correct antimicrobial susceptibility test results. This delay impacts the rapid implementation of adequate antimicrobial treatment and infection control countermeasures. Thus, there is emerging interest in the early and more sensitive detection of resistance mechanisms. Modern non-phenotypic tests are promising in this respect, and hence, can influence both clinical outcome and healthcare costs. In this review, we present a summary of the most advanced methods (e.g., next-generation DNA sequencing, multiplex PCRs, real-time PCRs, microarrays, MALDITOF MS, and PCR/ESI MS) presently available for the rapid detection of antibiotic resistance genes in Enterobacteriaceae. Taking into account speed, manageability, accuracy, versatility, and costs, the possible settings of application (research, clinic, and epidemiology) of these methods and their superiority against standard phenotypic methods are discussed.

Intestinal Carriage of Carbapenemase-Producing Organisms: Current Status of Surveillance Methods

“…There are several NAAT-based methodologies that may be employed to detect carbapenemase genes in bacterial isolates (169). Theoretically, all of them can be used to screen stool specimens.…”

“…Next-generation sequencing (NGS) may be another option, although it is not readily available in most clinical laboratories at this time (169). NGS remains expensive due to high equipment acquisition costs and the need for significant computer processing power and data storage (170).…”

“…The target gene of the pathogen is then labeled and hybridized to the immobilized probe. This reaction is then measured with a scanner (169). Microarrays are difficult to standardize (169), and reports describing the use of microarrays to directly screen for ␤-lactamase genes in stool are not available.…”

“…This reaction is then measured with a scanner (169). Microarrays are difficult to standardize (169), and reports describing the use of microarrays to directly screen for ␤-lactamase genes in stool are not available. Most assays, however, can be used to confirm and characterize the ␤-lactamase gene from suspicious colonies of a screening culture.…”

Carbapenem-Resistant Non-Glucose-Fermenting Gram-Negative Bacilli: the Missing Piece to the Puzzle

“…Many nonphenotypic-based methods have been developed for the detection of CROs/CPOs, including nucleic amplification methods, microarrays, matrix-assisted laser desorption ionizationtime of flight mass spectrometry (MALDI-TOF MS), UV spectrometry, and next-generation sequencing (Table 3) (63).…”