The Pseudomonas aeruginosa population has a nonclonal epidemic structure. It is generally composed of a limited number of widespread clones selected from a background of many rare and unrelated genotypes recombining at high frequency. Due to the increasing prevalence of nosocomial infections caused by multidrug-resistant/extensively drug-resistant (MDR/XDR) strains, it is advisable to implement infection control measures. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) are considered the gold standard methods in bacterial typing, despite being limited by cost, staff, and instrumental demands. Here, we present a novel mini-MLST scheme for P. aeruginosa rapid genotyping based on high-resolution melting analysis. Using the proposed mini-MLST scheme, 3,955 existing sequence types (STs) were converted into 701 melting types (MelTs), resulting in a discriminatory power of D = 0.993 (95% confidence interval [CI], 0.992 to 0.994). Whole-genome sequencing of 18 clinical isolates was performed to support the newly designed mini-MLST scheme. The clonal analysis of STs belonging to MelTs associated with international high-risk clones (HRCs) performed by goeBURST software revealed that a high proportion of the included STs are highly related to HRCs and have also been witnessed as responsible for serious infections. Therefore, mini-MLST provides a clear warning for the potential spread of P. aeruginosa clones recognized as MDR/XDR strains with possible serious outcomes. IMPORTANCE In this study, we designed a novel mini-MLST typing scheme for Pseudomonas aeruginosa. Its great discriminatory power, together with ease of performance and short processing time, makes this approach attractive for prospective typing of large isolate sets. Integrating the novel P. aeruginosa molecular typing scheme enables the development and spread of MDR/XDR high-risk clones to be investigated.

• Klíčová slova
• Pseudomonas aeruginosa, high-resolution melting, mini-MLST, molecular epidemiology, strain typing,
• MeSH
• buněčné klony MeSH
• genotyp MeSH
• lidé MeSH
• molekulární epidemiologie metody   MeSH
• multilokusová sekvenční typizace MeSH
• prospektivní studie MeSH
• pseudomonádové infekce * epidemiologie   mikrobiologie   MeSH
• Pseudomonas aeruginosa * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Staphylococcus aureus is a major bacterial human pathogen that causes a wide variety of clinical manifestations. The main aim of the presented study was to determine and optimize a novel sequencing independent approach that enables molecular typing of S. aureus isolates and elucidates the transmission of emergent clones between patients. In total, 987 S. aureus isolates including both methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) isolates were used to evaluate the novel typing approach combining high-resolution melting (HRM) analysis of multilocus sequence typing (MLST) genes (mini-MLST) and spa gene (spa-HRM). The novel approach's discriminatory ability was evaluated by whole-genome sequencing (WGS). The clonal relatedness of tested isolates was set by the BURP and BURST approach using spa and MLST data, respectively. Mini-MLST classified the S. aureus isolates into 38 clusters, followed by spa-HRM classifying the isolates into 101 clusters. The WGS proved HRM-based methods to effectively differentiate between related S. aureus isolates. Visualizing evolutionary relationships among different spa-types provided by the BURP algorithm showed comparable results to MLST/mini-MLST clonal clusters. We proved that the combination of mini-MLST and spa-HRM is rapid, reproducible, and cost-efficient. In addition to high discriminatory ability, the correlation between spa evolutionary relationships and mini-MLST clustering allows the variability in population structure to be monitored. IMPORTANCE Rapid and cost-effective molecular typing tools for Staphylococcus aureus epidemiological applications such as transmission tracking, source attribution and outbreak investigations are highly desirable. High-resolution melting based methods are effective alternative to those based on sequencing. Their good reproducibility and easy performance allow prospective typing of large set of isolates while reaching great discriminatory power. In this study, we established a new epidemiological approach to S. aureus typing. This scheme has the potential to greatly improve epidemiological investigations of S. aureus.

• Klíčová slova
• MLST, MRSA, MSSA, high-resolution melting, mini-MLST, spa-typing, whole-genome sequencing,
• MeSH
• kontrola infekce * MeSH
• lidé MeSH
• methicilin rezistentní Staphylococcus aureus genetika   izolace a purifikace   MeSH
• molekulární typizace metody   MeSH
• multilokusová sekvenční typizace MeSH
• prospektivní studie MeSH
• reprodukovatelnost výsledků MeSH
• sekvenování celého genomu MeSH
• stafylokokové infekce diagnóza   mikrobiologie   MeSH
• Staphylococcus aureus klasifikace   genetika   izolace a purifikace   MeSH
• techniky typizace bakterií metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH