Abstract
Routinely used typing methods including MLST, rep-PCR and whole genome sequencing (WGS) are time-consuming, costly, and often low throughput. Here, we describe a novel mini-MLST scheme for Eschericha coli as an alternative method for rapid genotyping. Using the proposed mini-MLST scheme, 10,946 existing STs were converted into 1,038 Melting Types (MelTs). To validate the new mini-MLST scheme, in silico analysis was performed on 73,704 strains retrieved from EnteroBase resulting in discriminatory power D = 0.9465 (CI 95% 0.9726–0.9736) for mini-MLST and D = 0.9731 (CI 95% 0.9726–0.9736) for MLST. Moreover, validation on clinical isolates was conducted with a significant concordance between MLST, rep-PCR and WGS. To conclude, the great portability, efficient processing, cost-effectiveness, and high throughput of mini-MLST represents immense benefits, even when accompanied with a slightly lower discriminatory power than other typing methods. This study proved mini-MLST is an ideal method to screen and subgroup large sets of isolates and/or quick strain typing during outbreaks. In addition, our results clearly showed its suitability for prospective surveillance monitoring of emergent and high-risk E. coli clones’.
Highlights
Used typing methods including multilocus sequence typing (MLST), rep-PCR and whole genome sequencing (WGS) are time-consuming, costly, and often low throughput
The single nucleotide polymorphisms (SNPs) that do not represent a nucleotide exchange affecting the number of hydrogen bonds (A/T ↔ G/C) were excluded, because these are not commonly detectable by high-resolution melting (HRM)
All SNPs without flanking conserved regions were excluded from future analyses as those are crucial for primer design
Summary
Used typing methods including MLST, rep-PCR and whole genome sequencing (WGS) are time-consuming, costly, and often low throughput. Bacterial genotyping is a powerful tool for strain identification, outbreak investigation and surveillance applications Electrophoretic techniques such as pulse-field gel electrophoresis (PFGE) or repetitive element sequence-based PCR (rep-PCR) have been considered the gold standard methods for bacterial strain tracking for many y ears[2]. Along with the rapid development, WGS technology has become more accessible for routine applications It is still impractical for prospective typing and/or screening typing larger numbers of strains. Data processing and their evaluation are currently WGS’ main disadvantage, demanding both extra computational and human resources Another method that can be used for genotyping is high-resolution melting (HRM). Primer ID adk[322] FW adk[322] RV fumC327 FW fumC327 RV gyrB174 FW gyrB174 RV icd[174] FW icd[174] RV purA263 FW purA263 RV recA91 FW recA91 RV
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