CRISPR Typing Increases the Discriminatory Power of Streptococcus agalactiae Typing Methods.

Clémence Beauruelle,Claire Poyart,Thierry Cochard,Ludovic Treluyer,Philippe Lanotte,Laurent Mereghetti,Clément Lier,Philippe Glaser,Adeline Pastuszka

doi:10.3389/fmicb.2021.675597

Abstract

We explored the relevance of a Clustered regularly interspaced short palindromic repeats (CRISPR)-based genotyping tool for Streptococcus agalactiae typing and we compared this method to current molecular methods [multi locus sequence typing (MLST) and capsular typing]. To this effect, we developed two CRISPR marker schemes (using 94 or 25 markers, respectively). Among the 255 S. agalactiae isolates tested, 229 CRISPR profiles were obtained. The 94 and 25 markers made it possible to efficiently separate isolates with a high diversity index (0.9947 and 0.9267, respectively), highlighting a high discriminatory power, superior to that of both capsular typing and MLST (diversity index of 0.9017 for MLST). This method has the advantage of being correlated with MLST [through analysis of the terminal direct repeat (TDR) and ancestral spacers] and to possess a high discriminatory power (through analysis of the leader-end spacers recently acquired, which are the witnesses of genetic mobile elements encountered by the bacteria). Furthermore, this “one-shot” approach presents the benefit of much-reduced time and cost in comparison with MLST. On the basis of these data, we propose that this method could become a reference method for group B Streptococcus (GBS) typing.

Highlights

Streptococcus agalactiae or group B Streptococcus (GBS) is the leading cause of neonatal infections and an emerging pathogen in adults, in elderly and immunocompromised patients (Phares et al, 2008; Skoff et al, 2009; Slotved and Hoffmann, 2020; Vuillemin et al, 2021)
We explored the relevance of the CRISPR1-based genotyping tool, and we compared this method to current molecular standards
We developed two schemes of markers selection (94 or 25 selected markers corresponding to spacers and terminal direct repeat (TDR)), and we simultaneously characterized these isolates by multi locus sequence typing (MLST) and capsular typing

Summary

Introduction

Streptococcus agalactiae or group B Streptococcus (GBS) is the leading cause of neonatal infections and an emerging pathogen in adults, in elderly and immunocompromised patients (Phares et al, 2008; Skoff et al, 2009; Slotved and Hoffmann, 2020; Vuillemin et al, 2021). MLST, a sequence-based method, is widely used to investigate the population structure and genetic lineage of GBS and is currently the reference method for GBS typing This approach is based on the combination of alleles for seven housekeeping genes (Jones et al, 2003). ST can be clustered in clonal complexes (CCs) when six of the seven alleles are in common (Feil et al, 2004) These methods have highlighted the involvement of serotype III and ST17, in causing more invasive neonate diseases (Manning et al, 2009). Despite the clonality observed within GBS populations causing invasive disease, increasing diversity has appeared, the need to investigate the population structure Between these two typing methods, while serotyping is relatively easy to perform in the laboratory, it is insufficiently discriminating to compare isolates. As the sequences targeted evolve slowly, MLST is not highly discriminant for epidemiological studies and local surveillance, and isolates are not easy to distinguish at the ST level (Radtke et al, 2010; Haguenoer et al, 2011; Sabat et al, 2013)

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