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Recently, it has been reported that eriC and crcB are involved in bacterial fluoride resistance. However, the fluoride-resistance mechanism in oral streptococci remains unclear. BLAST studies showed that two types of eriCs (eriC1 and eriC2) and two types of crcBs (crcB1 and crcB2) are present across 18 oral streptococci, which were identified in ≥ 10% of 166 orally healthy subjects with ≥ 0.01% of the mean relative abundance. They were divided into three groups based on the distribution of these four genes: group I, only eriC1; group II, eriC1 and eriC2; and group III, eriC2, crcB1, and crcB2. Group I consisted of Streptococcus mutans, in which one of the two eriC1s predominantly affected fluoride resistance. Group II consisted of eight species, and eriC1 was responsible for fluoride resistance, but eriC2 was not, in Streptococcus anginosus as a representative species. Group III consisted of nine species, and both crcB1 and crcB2 were crucial for fluoride resistance, but eriC2 was not, in Streptococcus sanguinis as a representative species. Based on these results, either EriC1 or CrcBs play a role in fluoride resistance in oral streptococci. Complementation between S. mutans EriC1 and S. sanguinis CrcB1/CrcB2 was confirmed in both S. mutans and S. sanguinis. However, neither transfer of S. sanguinis CrcB1/CrcB2 into wild-type S. mutans nor S. mutans EriC1 into wild-type S. sanguinis increased the fluoride resistance of the wild-type strain. Co-existence of different F- channels (EriC and CrcB) did not cause the additive effect on fluoride resistance in oral Streptococcus species.

Differentiation between mitis group streptococci (MGS) bacteria in routine laboratory tests has become important for obtaining accurate epidemiological information on the characteristics of MGS and understanding their clinical significance. The most reliable method of MGS species identification is multilocus sequence analysis (MLSA) with seven house-keeping genes; however, because this method is time-consuming, it is deemed unsuitable for use in most clinical laboratories. In this study, we established a scheme for identifying 12 species of MGS ( ) using the MinION nanopore sequencer (Oxford Nanopore Technologies, Oxford, UK) with the taxonomic aligner \"What's in My Pot?\" (WIMP; Oxford Nanopore's cloud-based analysis platform) and Kraken2 pipeline with the custom database adjusted for MGS species identification. The identities of the species in reference genomes ( = 514), clinical isolates ( = 31), and reference strains ( = 4) were confirmed via MLSA. The nanopore simulation reads were generated from reference genomes, and the optimal cut-off values for MGS species identification were determined. For 31 clinical isolates ( = 8, = 17 and = 6) and 4 reference strains ( = 1, = 1, = 1, and = 1), a sequence library was constructed via a Rapid Barcoding Sequencing Kit for multiplex and real-time MinION sequencing. The optimal cut-off values for the identification of MGS species for analysis by WIMP and Kraken2 pipeline were determined. The workflow using Kraken2 pipeline with a custom database identified all 12 species of MGS, and WIMP identified 8 MGS bacteria except , and . The results obtained by MinION with WIMP and Kraken2 pipeline were consistent with the MGS species identified by MLSA analysis. The practical advantage of whole genome analysis using the MinION nanopore sequencer is that it can aid in MGS surveillance. We concluded that MinION sequencing with the taxonomic aligner enables accurate MGS species identification and could contribute to further epidemiological surveys.

The Centers for Disease Control and Prevention's Arctic Investigations Program evaluated whole-genome sequencing (WGS) workflows and bioinformatics pipelines developed by the Centers' Streptococcus Laboratory. We compared WGS-based antimicrobial drug resistance predictions with phenotypic testing for group B (n = 130) and group A (n = 217) Streptococcus and Streptococcus pneumoniae (n = 293). Isolates were collected in Alaska during January 2019-February 2021. We also included a historical phenotypically nonsusceptible subset. Concordances between phenotypic testing and WGS predictions were 99.9% (895/896) for group B Streptococcus, 100% (1,298/1,298) for group A Streptococcus, and 99.98% (3,516/3,517) for S. pneumoniae. Common resistance determinants were ermTR, ermB, and mef for macrolides, tetM for tetracyclines, and gyrA and parC for levofloxacin. S. pneumoniae trimethoprim/sulfamethoxazole nonsusceptibility was associated with folP gene insertions and folA mutations. In 2022, the Arctic Investigations Program transitioned Streptococcus spp. workflows to WGS, enabling more rapid monitoring and prevention of invasive disease.

Abstract Timely surveillance of bacteremia is important for identifying emerging pathogens and to implement effective antimicrobial strategies and public health measures. Organisms in the genera Streptococcus and Enterococcus are often the cause of bacteremia and merit particular attention. The objective of this study was to determine the prevalence of bacteremia caused by several clinically significant Streptococcus species (S. pyogenes, S. agalactiae, S. pneumoniae, S. dysgalactiae/equisimilis, the S. anginosus group (S. anginosus, S. constellatus, S. intermedius), and Enterococcus species (E. faecium and E. faecalis) over a 5 year period (2017-2022). Bacteria isolated from blood culture bottles were identified by MALDI-TOF mass spectrometry (VITEK® MS). Data were obtained from a Laboratory Results Repository in Epic Beaker to include these organisms, isolated in blood cultures (BCs), patient gender, patient age, and the collection date. The data were then consolidated to yield one unique organism per patient per episode. The prevalence of organisms as a percent of the total positive BCs for the overall time period and for each year was determined. Species from the Enterococcus genus were the two most prevalent organisms for each year except for 2017. E. faecalis was the most prevalent overall (57.4%, n = 534) and for each year except for 2020. Combining all analyzed organisms, S. agalactiae was the third most prevalent organism for each year except for 2017 when it was the second and 2020-2021 when it was the fourth. However, within the Streptococcus genus alone, S. agalactiae was the most prevalent (27.5%, n = 281). S. pneumoniae, demonstrating seasonality, was the second most prevalent from 2018-2019, however, it was outranked by S. dysgalactiae/equisimilis from 2020-2022 when it was the fourth and by the S. anginosus group in 2017 and 2020-2022. S. pyogenes was the fourth most prevalent organism in 2017 and the third most in 2018; however, it decreased in subsequent years and was surpassed by all other organisms. Stratified by decade of life, patients in their seventh decade of life (60-69 yr) accounted for the most positive BCs for every organism, except for E. faecalis, followed frequently by patients in their sixth (50-59 yr) and eighth (70-79 yr) decades of life. Patients in their first decade of life (0-9 yr) accounted for a preponderance of positive BCs for E. faecalis, S. agalactiae (highest prevalence <7 days of life), and S. pneumoniae. This study highlighted the relative abundance of bacteremia caused by Enterococcus species. Additionally, it demonstrated the importance of Streptococcus agalactiae bacteremia, especially in the first and seventh decades of life. Finally, this study showed the increasing prevalence of organisms within the Streptococcus anginosus group and Streptococcus dysgalactiae/equisimilis. This analysis demonstrated the dynamics of bacteremia and prevalence among several species within the Enteroccocus and Streptococcus genera.

Streptococcus pneumoniae is one of the most important human pathogens but is closely related to Streptococcus mitis , with which humans live in harmony. The fact that the two species evolved from a common ancestor provides a unique basis for studies of both infection-associated properties and properties important for harmonious coexistence with the host. By detailed comparisons of genomes of the two species and other related streptococci, we identified 224 genes associated with virulence and 25 genes unique to the mutualistic species. The exclusive presence of the virulence factors in S. pneumoniae enhances their potential as vaccine components, as a direct impact on beneficial members of the commensal microbiota can be excluded. Successful adaptation of S. mitis and other commensal streptococci to a harmonious relationship with the host relied on genetic stability and properties facilitating life in biofilms. From a common ancestor, Streptococcus pneumoniae and Streptococcus mitis evolved in parallel into one of the most important pathogens and a mutualistic colonizer of humans, respectively. This evolutionary scenario provides a unique basis for studies of both infection-associated properties and properties important for harmonious coexistence with the host. We performed detailed comparisons of 60 genomes of S. pneumoniae , S. mitis , Streptococcus pseudopneumoniae , the three Streptococcus oralis subspecies oralis , tigurinus , and dentisani , and Streptococcus infantis . Nonfunctional remnants of ancestral genes in both S. pneumoniae and in S. mitis support the evolutionary model and the concept that evolutionary changes on both sides were required to reach their present relationship to the host. Confirmed by screening of >7,500 genomes, we identified 224 genes associated with virulence. The striking difference to commensal streptococci was the diversity of regulatory mechanisms, including regulation of capsule production, a significantly larger arsenal of enzymes involved in carbohydrate hydrolysis, and proteins known to interfere with innate immune factors. The exclusive presence of the virulence factors in S. pneumoniae enhances their potential as vaccine components, as a direct impact on beneficial members of the commensal microbiota can be excluded. In addition to loss of these virulence-associated genes, adaptation of S. mitis to a mutualistic relationship with the host apparently required preservation or acquisition of 25 genes lost or absent from S. pneumoniae . Successful adaptation of S. mitis and other commensal streptococci to a harmonious relationship with the host relied on genetic stability and properties facilitating life in biofilms. IMPORTANCE Streptococcus pneumoniae is one of the most important human pathogens but is closely related to Streptococcus mitis , with which humans live in harmony. The fact that the two species evolved from a common ancestor provides a unique basis for studies of both infection-associated properties and properties important for harmonious coexistence with the host. By detailed comparisons of genomes of the two species and other related streptococci, we identified 224 genes associated with virulence and 25 genes unique to the mutualistic species. The exclusive presence of the virulence factors in S. pneumoniae enhances their potential as vaccine components, as a direct impact on beneficial members of the commensal microbiota can be excluded. Successful adaptation of S. mitis and other commensal streptococci to a harmonious relationship with the host relied on genetic stability and properties facilitating life in biofilms.

Proteins harboring intrinsically disordered regions (IDRs) lacking stable secondary or tertiary structures are abundant across the three domains of life. These regions have not been systematically studied in prokaryotes. Here, our genome-wide analysis identifies extracytoplasmic serine/threonine-rich IDRs in several biologically important membrane-associated proteins in streptococci. We demonstrate that these IDRs are glycosylated with glucose by glycosyltransferases GtrB and PgtC2 in Streptococcus pyogenes and Streptococcus pneumoniae , and with N-acetylgalactosamine by a Pgf-dependent mechanism in Streptococcus mutans . The absence of glycosylation leads to a defect in biofilm formation under ethanol-stressed conditions in S. mutans . We link this phenotype to the C-terminal IDR of the post-translocation chaperone PrsA. Our data reveal that O -linked glycosylation protects the IDR-containing proteins from proteolytic degradation and is critical for the biological function of PrsA in biofilm formation. Here, the authors identify mechanisms of glycosylation of intrinsically disordered regions present in streptococci membrane proteins, uncovering a functional role for glycosylation in Streptococcus mutans chaperone PrsA.

Purpose Streptococcal bloodstream infections (BSIs) are common, yet prognostic factors are poorly investigated. We aimed to investigate the mortality according to streptococcal species and seasonal variation. Methods Patients with streptococcal BSIs from 2008 to 2017 in the Capital Region of Denmark were investigated, and data were crosslinked with nationwide registers for the identification of comorbidities. A multivariable logistic regression analysis was performed to assess mortality according to streptococcal species and season of infection. Results Among 6095 patients with a streptococcal BSI (mean age 68.1 years), the 30-day mortality was 16.1% and the one-year mortality was 31.5%. With S. pneumoniae as a reference, S. vestibularis was associated with a higher adjusted mortality both within 30 days (odds ratio (OR) 2.89 [95% confidence interval (CI) 1.20–6.95]) and one year (OR 4.09 [95% CI 1.70–9.48]). One-year mortality was also higher in S. thermophilus , S. constellatus , S. parasanguinis , S. salivarius , S. anginosus , and S. mitis/oralis . However, S. mutans was associated with a lower one-year mortality OR 0.44 [95% CI 0.20–0.97], while S. gallolyticus was associated with both a lower 30-day (OR 0.42 [95% CI 0.26–0.67]) and one-year mortality (OR 0.66 [95% CI 0.48–0.93]). Furthermore, with infection in the summer as a reference, patients infected in the winter and autumn had a higher association with 30-day mortality. Conclusions The mortality in patients with streptococcal BSI was associated with streptococcal species. Further, patients with streptococcal BSIs infected in the autumn and winter had a higher risk of death within 30 days, compared with patients infected in the summer.

Background: Streptococcus suis is a zoonotic pathogen that infects pigs and can occasionally cause serious infections in humans. S. suis infections occur sporadically in human Europe and North America, but a recent major outbreak has been described in China with high levels of mortality. The mechanisms of S. suis pathogenesis in humans and pigs are poorly understood. Methodology/Principal Findings: The sequencing of whole genomes of S. suis isolates provides opportunities to investigate the genetic basis of infection. Here we describe whole genome sequences of three S. suis strains from the same lineage: one from European pigs, and two from human cases from China and Vietnam. Comparative genomic analysis was used to investigate the variability of these strains. S. suis is phylogenetically distinct from other Streptococcus species for which genome sequences are currently available. Accordingly, approximately 40% of the approximately 2 Mb genome is unique in comparison to other Streptococcus species. Finer genomic comparisons within the species showed a high level of sequence conservation; virtually all of the genome is common to the S. suis strains. The only exceptions are three approximately 90 kb regions, present in the two isolates from humans, composed of integrative conjugative elements and transposons. Carried in these regions are coding sequences associated with drug resistance. In addition, small-scale sequence variation has generated pseudogenes in putative virulence and colonization factors. Conclusions/Significance: The genomic inventories of genetically related S. suis strains, isolated from distinct hosts and diseases, exhibit high levels of conservation. However, the genomes provide evidence that horizontal gene transfer has contributed to the evolution of drug resistance.

Streptococcus infantarius subsp. infantarius (Sii) belongs to the Streptococcus bovis/Streptococcus equinus complex associated with several human and animal infections. Sii is a predominant bacterium in spontaneously fermented milk products in Africa. The genome sequence of Sii strain CJ18 was compared with that of other Streptococcus species to identify dairy adaptations including genome decay such as in Streptococcus thermophilus, traits for its competitiveness in spontaneous milk fermentation and to assess potential health risks for consumers. The genome of Sii CJ18 harbors several unique regions in comparison to Sii ATCC BAA-102T, among others an enlarged exo- and capsular polysaccharide operon; Streptococcus thermophilus-associated genes; a region containing metabolic and hypothetical genes mostly unique to CJ18 and the dairy isolate Streptococcus gallolyticus subsp. macedonicus; and a second oligopeptide transport operon. Dairy adaptations in CJ18 are reflected by a high percentage of pseudogenes (4.9%) representing genome decay which includes the inactivation of the lactose phosphotransferase system (lacIIABC) by multiple transposases integration. The presence of lacS and lacZ genes is the major dairy adaptation affecting lactose metabolism pathways also due to the disruption of lacIIABC.We constructed mutant strains of lacS, lacZ and lacIIABC and analyzed the resulting strains of CJ18 to confirm the redirection of lactose metabolism via LacS and LacZ.Natural competence genes are conserved in both Sii strains, but CJ18 contains a lower number of CRISPR spacers which indicates a reduced defense capability against alien DNA. No classical streptococcal virulence factors were detected in both Sii strains apart from those involved in adhesion which should be considered niche factors. Sii-specific virulence factors are not described. Several Sii-specific regions encoding uncharacterized proteins provide new leads for virulence analyses and investigation of the unclear association of dairy and clinical Sii with human diseases. The genome of the African dairy isolate Sii CJ18 clearly differs from the human isolate ATCC BAA-102T. CJ18 possesses a high natural competence predisposition likely explaining the enlarged genome. Metabolic adaptations to the dairy environment are evident and especially lactose uptake corresponds to S. thermophilus. Genome decay is not as advanced as in S. thermophilus (10-19%) possibly due to a shorter history in dairy fermentations.

Streptococcus pneumoniae is a major cause of morbidity and mortality worldwide. Virulence-associated proteins common and conserved among all capsular types now represent the best strategy to combat pneumococcal infections. Our aim was to identify conserved targets in pneumococci that showed positive prediction for lipoprotein and extracellular subcellular location using bioinformatics programs and verify the distribution and the degree of conservation of these targets in pneumococci. These targets can be considered potential vaccine candidate to be evaluated in the future. A set of 13 targets were analyzed and confirmed the presence in all pneumococci tested. These 13 genes were highly conserved showing around >96 % of amino acid and nucleotide identity, but they were also present and show high identity in the closely related species Streptococcus mitis, Streptococcus oralis, and Streptococcus pseudopneumoniae. S. oralis clusters away from S. pneumoniae, while S. pseudopneumoniae and S. mitis cluster closer. The divergence between the selected targets was too small to be observed consistently in phylogenetic groups between the analyzed genomes of S. pneumoniae. The proteins analyzed fulfill two of the initial criteria of a vaccine candidate: targets are present in a variety of different pneumococci strains including different serotypes and are conserved among the samples evaluated.