Application of Whole-Genome Sequencing for Bacterial Strain Typing in Molecular Epidemiology

Abstract

Nosocomial infections pose a significant threat to patient health; however, the gold standard laboratory method for determining bacterial relatedness (pulsed-field gel electrophoresis [PFGE]) remains essentially unchanged 20 years after its introduction. Here, we explored bacterial whole-genome sequencing (WGS) as an alternative approach for molecular strain typing. We compared WGS to PFGE for investigating presumptive outbreaks involving three important pathogens: vancomycin-resistant Enterococcus faecium (n ‫؍‬ 19), methicillin-resistant Staphylococcus aureus (n ‫؍‬ 17), and Acinetobacter baumannii (n ‫؍‬ 15). WGS was highly reproducible (average < 0.39 differences between technical replicates), which enabled a functional, quantitative definition for determining clonality. Strain relatedness data determined by PFGE and WGS roughly correlated, but the resolution of WGS was superior (P ‫؍‬ 5.6 ؋ 10 ؊8 to 0.016). Several discordant results were noted between the methods. A total of 28.9% of isolates which were indistinguishable by PFGE were nonclonal by WGS. For A. baumannii, a species known to undergo rapid horizontal gene transfer, 16.2% of isolate pairs considered nonidentical by PFGE were clonal by WGS. Sequencing whole bacterial genomes with single-nucleotide resolution demonstrates that PFGE is prone to false-positive and false-negative results and suggests the need for a new gold standard approach for molecular epidemiological strain typing.

Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

“…In this regard, WGS was superior to PFGE due to its enhanced discriminatory power (Table 2) (0.997 versus 0.892), which has been noted in other studies of bacterial outbreaks (31,79,99,100). In particular, our results are similar to those of Salipante and colleagues (79), who used a different approach to identify SNPs and define outbreak strains but arrived at the same conclusions regarding the deficiencies of PFGE in the context of an A. baumannii outbreak. At the molecular level, this reflects the relatively large number of SNPs that can accumulate in genomes before they are detectable by PFGE (79).…”

“…In particular, our results are similar to those of Salipante and colleagues (79), who used a different approach to identify SNPs and define outbreak strains but arrived at the same conclusions regarding the deficiencies of PFGE in the context of an A. baumannii outbreak. At the molecular level, this reflects the relatively large number of SNPs that can accumulate in genomes before they are detectable by PFGE (79). In addition, our study found three variants of the same strain within a single pa-…”

“…Because of the high level of antibiotic resistance associated with some A. baumannii strains, these outbreaks have important consequences for patient care and require the use of considerable resources by infection control personnel. PFGE is routinely used by many hospitals to track nosocomial outbreaks, but several studies indicate that WGS is better suited for this purpose (38,78,79). In 2013, a CRAB outbreak occurred in our hospital and was investigated using PFGE and contact tracing.…”

The whole genome sequencing of Acinetobacter-calcoaceticus-baumannii complex strains involved in suspected outbreak in an Intensive Care Unit of a pediatric hospital

“…[7][8][9] Acinetobacter baumannii is resistant to desiccation and to disinfectants, it is difficult to be treated and to be eradicated and it requires rigorous monitoring programs to prevent and control hospital outbreaks. [10] Guidance documents [11][12][13] on the control of Acinetobacter baumannii nosocomial infections are based on the combined interventions including an active microbiological surveillance, precautions to contain the spread of infections from patient to patient and nursing care protocols for colonized patients. Additional measures regard: antibiotic use restriction, careful hand hygiene, extensive environmental surface decontamination procedures [14] and clinical staff education on how to deal with an outbreak.…”

Genomic Epidemiology of Clostridium botulinum Isolates from Temporally Related Cases of Infant Botulism in New South Wales, Australia

“…Unfortunately, the resolution of standard typing methods and the frequent absence of environmental samples often make it difficult to conclusively link cases or identify environmental reservoirs. The use of whole-genome sequencing (WGS) to compare entire bacterial genomes at the single nucleotide level can, however, greatly improve clustering of bacterial pathogens (39).…”