Prediction of Antimicrobial Resistance in Clinical Enterococcus faecium Isolates Using a Rules-Based Analysis of Whole-Genome Sequences.

Enterococcus faecium is a major cause of clinical infections, often due to multidrug-resistant (MDR) strains. Whole-genome sequencing (WGS) is a powerful tool to study MDR bacteria and their antimicrobial resistance (AMR) mechanisms. In this study, we used WGS to characterize E. faecium clinical isolates and test the feasibility of rules-based genotypic prediction of AMR. Clinical isolates were divided into derivation and validation sets. Phenotypic susceptibility testing for ampicillin, vancomycin, high-level gentamicin, ciprofloxacin, levofloxacin, doxycycline, tetracycline, and linezolid was performed using the Vitek 2 automated system, with confirmation and discrepancy resolution by broth microdilution, disk diffusion, or gradient diffusion when needed. WGS was performed to identify isolate lineage and AMR genotype. AMR prediction rules were derived by analyzing the genotypic-phenotypic relationship in the derivation set. Phylogenetic analysis demonstrated that 88% of isolates in the collection belonged to hospital-associated clonal complex 17. Additionally, 12% of isolates had novel sequence types. When applied to the validation set, the derived prediction rules demonstrated an overall positive predictive value of 98% and negative predictive value of 99% compared to standard phenotypic methods. Most errors were falsely resistant predictions for tetracycline and doxycycline. Further analysis of genotypic-phenotypic discrepancies revealed potentially novel pbp5 and tet(M) alleles that provide insight into ampicillin and tetracycline class resistance mechanisms. The prediction rules demonstrated generalizability when tested on an external data set. In conclusion, known AMR genes and mutations can predict E. faecium phenotypic susceptibility with high accuracy for most routinely tested antibiotics, providing opportunities for advancing molecular diagnostics.

The effect of transport temperature and time on the recovery of antimicrobial-resistant Enterobacterales in stool.

Surveillance for antibiotic-resistant (AR) bacteria is challenging. We evaluated AR Enterobacterales survival in stool over various transport conditions. Stool in Cary-Blair medium was spiked with AR Enterobacterales, held at 3 °C, 20 °C, or 37 °C, and cultured on days 3, 8, and 15. Stool from US international travelers sent through the US mail was also screened. We compared recovery rates using Fisher's exact tests and linear regression models. AR Enterobacterales recovery reduced with time (86% versus 75% versus 61% at days 3, 8, and 15; Beta for linear trend=-0.02, r2=0.99, P=0.02) and colder temperatures [56% (3 °C) versus 89% (20 °C) versus 86% (37 °C); P=0.003]. Traveler sample recovery also reduced with transport time (Beta for linear trend=-0.03, r2=0.70, P=0.01) but not with season [20% (cold) versus 22% (warm), P=0.7]. AR Enterobacterales are found over variable transport conditions, providing rationale for expanding surveillance sample processing timelines.

Evaluation of a Screening Method for the Detection of Colistin-Resistant Enterobacteriaceae in Stool.

Emergence of mobile colistin resistance (mcr)-containing Enterobacteriaceae is a public health threat, prompting enhanced surveillance through the Centers for Disease Control and Prevention. We evaluated a selective culture medium for the isolation of Enterobacteriaceae with non-wild-type colistin minimum inhibitory concentrations, including those with mcr-1 genes, in spiked stool samples.