Predicting antimicrobial susceptibilities for Escherichia coli and Klebsiella pneumoniae isolates using whole genomic sequence data

Abstract

ObjectivesWhole-genome sequencing potentially represents a single, rapid and cost-effective approach to defining resistance mechanisms and predicting phenotype, and strain type, for both clinical and epidemiological purposes. This retrospective study aimed to determine the efficacy of whole genome-based antimicrobial resistance prediction in clinical isolates of Escherichia coli and Klebsiella pneumoniae.MethodsSeventy-four E. coli and 69 K. pneumoniae bacteraemia isolates from Oxfordshire, UK, were sequenced (Illumina HiSeq 2000). Resistance phenotypes were predicted from genomic sequences using BLASTn-based comparisons of de novo-assembled contigs with a study database of >100 known resistance-associated loci, including plasmid-associated and chromosomal genes. Predictions were made for seven commonly used antimicrobials: amoxicillin, co-amoxiclav, ceftriaxone, ceftazidime, ciprofloxacin, gentamicin and meropenem. Comparisons were made with phenotypic results obtained in duplicate by broth dilution (BD Phoenix). Discrepancies, either between duplicate BD Phoenix results or between genotype and phenotype, were resolved with gradient diffusion analyses.ResultsA wide variety of antimicrobial resistance genes were identified, including blaCTX-M, blaLEN, blaOKP, blaOXA, blaSHV, blaTEM, aac(3′)-Ia, aac-(3′)-IId, aac-(3′)-IIe, aac(6′)-Ib-cr, aadA1a, aadA4, aadA5, aadA16, aph(6′)-Id, aph(3′)-Ia, qnrB and qnrS, as well as resistance-associated mutations in chromosomal gyrA and parC genes. The sensitivity of genome-based resistance prediction across all antibiotics for both species was 0.96 (95% CI: 0.94–0.98) and the specificity was 0.97 (95% CI: 0.95–0.98). Very major and major error rates were 1.2% and 2.1%, respectively.ConclusionsOur method was as sensitive and specific as routinely deployed phenotypic methods. Validation against larger datasets and formal assessments of cost and turnaround time in a routine laboratory setting are warranted.

Whole-Genome Sequencing for Detecting Antimicrobial Resistance in Nontyphoidal Salmonella

“…By providing definitive genotype information, WGS offers the highest practical resolution for characterizing an individual microbe. This includes the full complement of resistance determinants (7,8), including resistance to compounds not routinely tested phenotypically. Bacteria that have identical resistance patterns caused by different mechanisms can also be differentiated by WGS.…”

Whole-Genome Sequencing Analysis Accurately Predicts Antimicrobial Resistance Phenotypes in Campylobacter spp

“…Recent studies have shown that WGS analysis can potentially be a single, rapid, and cost-effective approach to define resistance genotypes and predict resistance phenotypes of bacteria with great sensitivity and specificity (1)(2)(3)(4).…”

Establishing Genotypic Cutoff Values To Measure Antimicrobial Resistance in Salmonella

“…For most bacteria, the evidence for using WGS to infer antimicrobial susceptibility data is sparse, although some studies have shown a high correlation between resistance phenotypes and the presence of resistance genes or resistance-associated mutations. This agreement has been best demonstrated with enteric bacteria, including Escherichia coli, Campylobacter spp., and nontyphoidal Salmonella (8)(9)(10)(11)(12). In addition, other DNAbased methods have been used to detect resistance determinants and even guide therapy for infections caused by Mycobacterium tuberculosis, Staphylococcus aureus, HIV, and other pathogens (13)(14)(15).…”