Multiple-Locus Variable Number Tandem Repeat Analysis (MLVA) and Tandem Repeat Sequence Typing (TRST), helpful tools for subtyping Staphylococcus lugdunensis

Abstract

Staphylococcus lugdunensis is an emergent virulent coagulase-negative Staphylococcus that is increasingly responsible for severe infections. In an attempt to generate informative sequence data for subtyping S. lugdunensis, we selected and sequenced seven polymorphic variable number of tandem repeats (VNTRs) to develop two new methods: a classic length-based multiple-locus VNTR analysis (MLVA) method and a tandem repeat sequence typing (TRST) method. We assessed their performances compared to two existing methods, multilocus sequence typing (MLST) and multivirulence-locus sequence typing (MVLST) for 128 isolates from diverse clinical settings and geographical origins. The clustering achieved by the four methods was highly congruent, with MLVA discriminating within clonal complexes as defined by MLST. Indeed, MLVA was highly discriminant compared to MLST and MVLST in terms of number of genotypes as well as diversity indexes. Sequencing of the seven VNTRs showed that they were stable, and analysis of sequence polymorphisms provided superior discriminatory power. The typeability, reproducibility, and epidemiological concordance of these new methods were excellent. Of note, no link between clustering and clinical settings was identified. This study demonstrates that MLVA and TRST provide valuable information for molecular epidemiological study of S. lugdunensis, and represent promising tools to distinguish between strains of homogenous lineages in this clonal species.

fbl-Typing of Staphylococcus lugdunensis: A Frontline Tool for Epidemiological Studies, but Not Predictive of Fibrinogen Binding Ability

“…fbl -typing results were compared with those of MLST and TRST for 128 isolates representative of the genetic diversity of S. lugdunensis (Dahyot et al, 2018). Twenty-five STs belonging to 7 CCs, 69 TRTs, and 60 fbl -types were identified for these isolates (Table 2).…”

“…Staphylococcus lugdunensis is a significant human pathogen with distinct clinical and microbiological characteristics compared to other CoNS (Frank et al, 2008; Becker et al, 2014). To better understand the genetic background and population structure of S. lugdunensis , several sequence-based methods have been developed, such as MLST, MVLST and more recently TRST (Chassain et al, 2012; Didi et al, 2014; Dahyot et al, 2018). Phylogenetic analysis has shown the clonal population structure and the mutational evolution of this species.…”

“…Among them, ten pairs of isolates were collected from ten patients at time intervals ranging from 0 to 17 days. A first panel, the “TRST panel,” included 128 S. lugdunensis isolates previously characterized by MLST and TRST and collected from five regions of France and from Sweden (Dahyot et al, 2018). A further set of 106 clinical isolates recovered from 101 patients was collected at Rouen University Hospital in France from January 2016 to December 2016.…”

“…These markers can be used to build allelic profiles in MLVA systems, in order to discriminate between clonal bacterial populations (Van Belkum et al, 2007). Recently, we developed the first two VNTR-based schemes for S. lugdunensis typing: a classic length-based MLVA method and a sequence-based MLVA method known as the TRST method (Dahyot et al, 2018). These typing methods were more discriminating than MLST and MVLST, and represent promising tools for molecular epidemiological studies of S. lugdunensis .…”

Multi-locus Variable-number Tandem-repeat Analysis of the Fish-pathogenic Bacterium <em>Yersinia ruckeri</em> by Multiplex PCR and Capillary Electrophoresis

“…The technique is based on repeat number variation in selected variable-number tandem-repeat (VNTR) loci, resulting in output data that is highly transportable, making comparison of profiled isolates towards online databases and across labs straightforward. Although MLST remains the gold standard for epidemiological typing of many bacterial pathogens, an increasing number of studies identify a significantly higher discriminatory power of MLVA 8,9,10 . Several protocols have also been published targeting fish-pathogenic bacteria, such as Francisella noatunensis, Edwardsiella piscicida and Renibacterium salmoninarum 11,12,13 .…”