Invasive Streptococcus oralis Expressing Serotype 3 Pneumococcal Capsule, Japan.

We report 2 adult cases of invasive disease in Japan caused by Streptococcus oralis that expressed the serotype 3 pneumococcal capsule and formed mucoid colonies. Whole-genome sequencing revealed that the identical serotype 3 pneumococcal capsule locus and hyl fragment were recombined into the genomes of 2 distinct S. oralis strains.

Identification and genetic engineering of pneumococcal capsule-like polysaccharides in commensal oral streptococci.

Capsular polysaccharides (CPS) in Streptococcus pneumoniae are pivotal for bacterial virulence and present extensive diversity. While oral streptococci show pronounced antigenicity toward pneumococcal capsule-specific sera, insights into evolution of capsule diversity remain limited. This study reports a pneumococcal CPS-like genetic locus in Streptococcus parasanguinis, a predominant oral Streptococcus. The discovered locus comprises 15 genes, mirroring high similarity to those from the Wzy-dependent CPS locus of S. pneumoniae. Notably, S. parasanguinis elicited a reaction with pneumococcal 19B antiserum. Through nuclear magnetic resonance analysis, we ascertained that its CPS structure matches the chemical composition of the pneumococcal 19B capsule. By introducing the glucosyltransferase gene cps19cS from a pneumococcal serotype 19C, we successfully transformed S. parasanguinis antigenicity from 19B to 19C. Furthermore, substituting serotype-specific genes, cpsI and cpsJ, with their counterparts from pneumococcal serotype 19A and 19F enabled S. parasanguinis to generate 19A- and 19F-specific CPS, respectively. These findings underscore that S. parasanguinis harbors a versatile 19B-like CPS adaptable to other serotypes. Remarkably, after deleting the locus's initial gene, cpsE, responsible for sugar transfer, we noted halted CPS production, elongated bacterial chains, and diminished biofilm formation. A similar phenotype emerged with the removal of the distinct gene cpsZ, which encodes a putative autolysin. These data highlight the importance of S. parasanguinis CPS for biofilm formation and propose a potential shared ancestry of its CPS locus with S. pneumoniae. IMPORTANCE: Diverse capsules from Streptococcus pneumoniae are vital for bacterial virulence and pathogenesis. Oral streptococci show strong responses to a wide range of pneumococcal capsule-specific sera. Yet, the evolution of this capsule diversity in relation to microbe-host interactions remains underexplored. Our research delves into the connection between commensal oral streptococcal and pneumococcal capsules, highlighting the potential for gene transfer and evolution of various capsule types. Understanding the genetic and evolutionary factors that drive capsule diversity in S. pneumoniae and its related oral species is essential for the development of effective pneumococcal vaccines. The present findings provide fresh perspectives on the cross-reactivity between commensal streptococci and S. pneumoniae, its influence on bacteria-host interactions, and the development of new strategies to manage and prevent pneumococcal illnesses by targeting and modulating commensal streptococci.

Electron Microscopy to Study the Fine Structure of the Pneumococcal Cell.

Electron microscopy allows for studying bacterial ultrastructure at high resolutions. Two types of electron microscopes are used for this purpose. The transmission electron microscope allows for access to inner bacterial ultrastructure when imaging ultrathin sections as well as cell wall-attached structures by negative staining, whereas scanning electron microscopy allows for the detection of structures on the bacterial cell surface alone or to study the interplay between pneumococci and their host cells. This chapter deals with recommendations for well-adapted methodologies to examine pneumococcal ultrastructure in detail. Especially, we focus on the preservation of the pneumococcal capsular polysaccharide, which represents an important virulence factor of pneumococci. Since capsules are highly hydrated structures, the introduction of a new fixation protocol involving lysine acetate, ruthenium red, and osmium (LRR fixation) results in a very well-preserved capsular structure in such a way that the amount of capsular material bound on the bacterial surface can be compared within different serotypes. In our method, capsular ultrastructure is preserved without the need for serotype-specific antibodies, which have been used in other studies to preserve the pneumococcal capsule. In addition, the new LRR fixation allows for studying the presence or absence of capsular material during adhesion and invasion of pneumococci on epithelial or endothelial host cells in cell culture experiments.

A novel genetic variant of Streptococcus pneumoniae serotype 11A discovered in Fiji.

OBJECTIVES: As part of annual cross-sectional Streptococcus pneumoniae carriage surveys in Fiji (2012-2015), we detected pneumococci in over 100 nasopharyngeal swabs that serotyped as '11F-like' by microarray. We examined the genetic basis of this divergence in the 11F-like capsular polysaccharide (cps) locus compared to the reference 11F cps sequence. The impact of this diversity on capsule phenotype, and serotype results using genetic and serologic methods were determined. METHODS: Genomic DNA from representative 11F-like S. pneumoniae isolates obtained from the nasopharynx of Fijian children was extracted and subject to whole genome sequencing. Genetic and phylogenetic analyses were used to identify genetic changes in the cps locus. Capsular phenotypes were evaluated using the Quellung reaction and latex agglutination. RESULTS: Compared to published 11F sequences, the wcwC and wcrL genes of the 11F-like cps locus are phylogenetically divergent, and the gct gene contains a single nucleotide insertion within a homopolymeric region. These changes within the DNA sequence of the 11F-like cps locus have modified the antigenic properties of the capsule, such that 11F-like isolates serotype as 11A by Quellung reaction and latex agglutination. CONCLUSIONS: This study demonstrates the ability of molecular serotyping by microarray to identify genetic variants of S. pneumoniae and highlights the potential for discrepant results between phenotypic and genotypic serotyping methods. We propose that 11F-like isolates are not a new serotype but rather are a novel genetic variant of serotype 11A. These findings have implications for invasive pneumococcal disease surveillance as well as studies investigating vaccine impact.

Discovery of Streptococcus pneumoniae serogroup 35 variants in Australian patients.

OBJECTIVES: Streptococcus pneumoniae isolates from Australian invasive pneumococcal disease cases displaying an atypical 35B phenotype. Whole genome sequencing was used to analyse these strains and identify changes to the capsule gene regions. METHODS: Four atypical serogroup 35 isolates from Australian reference laboratories were unable to be assigned to one of the four known group 35 serotypes by the Quellung serotyping method. Genetic characterization of the capsule locus was performed by bioinformatic analysis of whole genome sequencing data for all isolates. RESULTS: Genetic analysis identified four independent disruptions to the wciG gene, which encodes an O-acetyltransferase responsible for the O-acetylation of the 6Galß1 residue in the capsular polysaccharide repeat unit of serotype 35B. CONCLUSIONS: This is the first published report on the incidence and capsular gene characteristics of a S. pneumoniae 35B variant.