Streptococcus anginosus promotes gastric inflammation, atrophy, and tumorigenesis in mice.

Streptococcus anginosus (S. anginosus) was enriched in the gastric mucosa of patients with gastric cancer (GC). Here, we show that S. anginosus colonized the mouse stomach and induced acute gastritis. S. anginosus infection spontaneously induced progressive chronic gastritis, parietal cell atrophy, mucinous metaplasia, and dysplasia in conventional mice, and the findings were confirmed in germ-free mice. In addition, S. anginosus accelerated GC progression in carcinogen-induced gastric tumorigenesis and YTN16 GC cell allografts. Consistently, S. anginosus disrupted gastric barrier function, promoted cell proliferation, and inhibited apoptosis. Mechanistically, we identified an S. anginosus surface protein, TMPC, that interacts with Annexin A2 (ANXA2) receptor on gastric epithelial cells. Interaction of TMPC with ANXA2 mediated attachment and colonization of S. anginosus and induced mitogen-activated protein kinase (MAPK) activation. ANXA2 knockout abrogated the induction of MAPK by S. anginosus. Thus, this study reveals S. anginosus as a pathogen that promotes gastric tumorigenesis via direct interactions with gastric epithelial cells in the TMPC-ANXA2-MAPK axis.

Neurons Are the Inflammatory Problem.

The discovery that reflex neuronal circuits control immunity provided a new mechanistic understanding of infection and injury. Pinho-Ribeiro et al. (2018) report that lethal tissue destruction from streptococcal necrotizing fasciitis ("flesh-eating disease") occurs because sensory neurons reflexively inhibit frontline immune responses.

Blocking Neuronal Signaling to Immune Cells Treats Streptococcal Invasive Infection.

The nervous system, the immune system, and microbial pathogens interact closely at barrier tissues. Here, we find that a bacterial pathogen, Streptococcus pyogenes, hijacks pain and neuronal regulation of the immune response to promote bacterial survival. Necrotizing fasciitis is a life-threatening soft tissue infection in which "pain is out of proportion" to early physical manifestations. We find that S. pyogenes, the leading cause of necrotizing fasciitis, secretes streptolysin S (SLS) to directly activate nociceptor neurons and produce pain during infection. Nociceptors, in turn, release the neuropeptide calcitonin gene-related peptide (CGRP) into infected tissues, which inhibits the recruitment of neutrophils and opsonophagocytic killing of S. pyogenes. Botulinum neurotoxin A and CGRP antagonism block neuron-mediated suppression of host defense, thereby preventing and treating S. pyogenes necrotizing infection. We conclude that targeting the peripheral nervous system and blocking neuro-immune communication is a promising strategy to treat highly invasive bacterial infections. VIDEO ABSTRACT.

Antibacterial Nucleoside-Analog Inhibitor of Bacterial RNA Polymerase.

Drug-resistant bacterial pathogens pose an urgent public-health crisis. Here, we report the discovery, from microbial-extract screening, of a nucleoside-analog inhibitor that inhibits bacterial RNA polymerase (RNAP) and exhibits antibacterial activity against drug-resistant bacterial pathogens: pseudouridimycin (PUM). PUM is a natural product comprising a formamidinylated, N-hydroxylated Gly-Gln dipeptide conjugated to 6'-amino-pseudouridine. PUM potently and selectively inhibits bacterial RNAP in vitro, inhibits bacterial growth in culture, and clears infection in a mouse model of Streptococcus pyogenes peritonitis. PUM inhibits RNAP through a binding site on RNAP (the NTP addition site) and mechanism (competition with UTP for occupancy of the NTP addition site) that differ from those of the RNAP inhibitor and current antibacterial drug rifampin (Rif). PUM exhibits additive antibacterial activity when co-administered with Rif, exhibits no cross-resistance with Rif, and exhibits a spontaneous resistance rate an order-of-magnitude lower than that of Rif. PUM is a highly promising lead for antibacterial therapy.

An extracellular bacterial pathogen modulates host metabolism to regulate its own sensing and proliferation.

Successful infection depends on the ability of the pathogen to gain nutrients from the host. The extracellular pathogenic bacterium group A Streptococcus (GAS) causes a vast array of human diseases. By using the quorum-sensing sil system as a reporter, we found that, during adherence to host cells, GAS delivers streptolysin toxins, creating endoplasmic reticulum stress. This, in turn, increases asparagine (ASN) synthetase expression and the production of ASN. The released ASN is sensed by the bacteria, altering the expression of ∼17% of GAS genes of which about one-third are dependent on the two-component system TrxSR. The expression of the streptolysin toxins is strongly upregulated, whereas genes linked to proliferation are downregulated in ASN absence. Asparaginase, a widely used chemotherapeutic agent, arrests GAS growth in human blood and blocks GAS proliferation in a mouse model of human bacteremia. These results delineate a pathogenic pathway and propose a therapeutic strategy against GAS infections.

Potential for Maternally Administered Vaccine for Infant Group B Streptococcus.

BACKGROUND: Natural history studies have correlated serotype-specific anti-capsular polysaccharide (CPS) IgG in newborns with a reduced risk of group B streptococcal disease. A hexavalent CPS-cross-reactive material 197 glycoconjugate vaccine (GBS6) is being developed as a maternal vaccine to prevent invasive group B streptococcus in young infants. METHODS: In an ongoing phase 2, placebo-controlled trial involving pregnant women, we assessed the safety and immunogenicity of a single dose of various GBS6 formulations and analyzed maternally transferred anti-CPS antibodies. In a parallel seroepidemiologic study that was conducted in the same population, we assessed serotype-specific anti-CPS IgG concentrations that were associated with a reduced risk of invasive disease among newborns through 89 days of age to define putative protective thresholds. RESULTS: Naturally acquired anti-CPS IgG concentrations were associated with a reduced risk of disease among infants in the seroepidemiologic study. IgG thresholds that were determined to be associated with 75 to 95% reductions in the risk of disease were 0.184 to 0.827 µg per milliliter. No GBS6-associated safety signals were observed among the mothers or infants. The incidence of adverse events and of serious adverse events were similar across the trial groups for both mothers and infants; more local reactions were observed in the groups that received GBS6 containing aluminum phosphate. Among the infants, the most common serious adverse events were minor congenital anomalies (umbilical hernia and congenital dermal melanocytosis). GBS6 induced maternal antibody responses to all serotypes, with maternal-to-infant antibody ratios of approximately 0.4 to 1.3, depending on the dose. The percentage of infants with anti-CPS IgG concentrations above 0.184 µg per milliliter varied according to serotype and formulation, with 57 to 97% of the infants having a seroresponse to the most immunogenic formulation. CONCLUSIONS: GBS6 elicited anti-CPS antibodies against group B streptococcus in pregnant women that were transferred to infants at levels associated with a reduced risk of invasive group B streptococcal disease. (Funded by Pfizer and the Bill and Melinda Gates Foundation; C1091002 ClinicalTrials.gov number, NCT03765073.).

Unmutated but T cell dependent IgM antibodies targeting Streptococcus suis play an essential role in bacterial clearance.

Streptococcus suis serotype 2 is an important encapsulated bacterial swine pathogen and zoonotic agent for which no effective vaccine exists. The interaction with B cells and the humoral response against S. suis are poorly understood despite their likely relevance for a potential vaccine. We evaluated germinal center (GC) B cell kinetics, as well as the production and role of S. suis-specific antibodies following infections in a mouse model. We found that mice infected with S. suis developed GC that peaked 13-21 days post-infection. GC further increased and persisted upon periodic reinfection that mimics real life conditions in swine farms. Anti-S. suis IgM and several IgG subclasses were produced, but antibodies against the S. suis capsular polysaccharide (CPS) were largely IgM. Interestingly, depletion of total IgG from the wild-type mice sera had no effect on bacterial killing by opsonophagocytosis in vitro. Somatic hypermutation and isotype switching were dispensable for controlling the infection or anti-CPS IgM production. However, T cell-deficient (Tcrb-/-) mice were unable to control bacteremia, produce optimal anti-CPS IgM titers, or elicit antibodies with opsonophagocytic activity. SAP deficiency, which prevents GC formation but not extrafollicular B cell responses, ablated anti S. suis-IgG production but maintained IgM production and eliminated the infection. In contrast, B cell deficient mice were unable to control bacteremia. Collectively, our results indicate that the antibody response plays a large role in immunity against S. suis, with GC-independent but T cell-dependent germline IgM being the major effective antibody specificities. Our results further highlight the importance IgM, and potentially anti-CPS antibodies, in clearing S. suis infections and provide insight for future development of S. suis vaccines.

Streptolysin S is required for Streptococcus pyogenes nasopharyngeal and skin infection in HLA-transgenic mice.

Streptococcus pyogenes is a human-specific pathogen that commonly colonizes the upper respiratory tract and skin, causing a wide variety of diseases ranging from pharyngitis to necrotizing fasciitis and toxic shock syndrome. S. pyogenes has a repertoire of secreted virulence factors that promote infection and evasion of the host immune system including the cytolysins streptolysin O (SLO) and streptolysin S (SLS). S. pyogenes does not naturally infect the upper respiratory tract of mice although mice transgenic for MHC class II human leukocyte antigens (HLA) become highly susceptible. Here we used HLA-transgenic mice to assess the role of both SLO and SLS during both nasopharyngeal and skin infection. Using S. pyogenes MGAS8232 as a model strain, we found that an SLS-deficient strain exhibited a 100-fold reduction in bacterial recovery from the nasopharynx and a 10-fold reduction in bacterial burden in the skin, whereas an SLO-deficient strain did not exhibit any infection defects in these models. Furthermore, depletion of neutrophils significantly restored the bacterial burden of the SLS-deficient bacteria in skin, but not in the nasopharynx. In mice nasally infected with the wildtype S. pyogenes, there was a marked change in localization of the tight junction protein ZO-1 at the site of infection, demonstrating damage to the nasal epithelia that was absent in mice infected with the SLS-deficient strain. Overall, we conclude that SLS is required for the establishment of nasopharyngeal infection and skin infection in HLA-transgenic mice by S. pyogenes MGAS8232 and provide evidence that SLS contributes to nasopharyngeal infection through the localized destruction of nasal epithelia.

Type II and IV toxin-antitoxin systems coordinately stabilize the integrative and conjugative element of the ICESa2603 family conferring multiple drug resistance in Streptococcus suis.

Integrative and conjugative elements (ICEs) play a vital role in bacterial evolution by carrying essential genes that confer adaptive functions to the host. Despite their importance, the mechanism underlying the stable inheritance of ICEs, which is necessary for the acquisition of new traits in bacteria, remains poorly understood. Here, we identified SezAT, a type II toxin-antitoxin (TA) system, and AbiE, a type IV TA system encoded within the ICESsuHN105, coordinately promote ICE stabilization and mediate multidrug resistance in Streptococcus suis. Deletion of SezAT or AbiE did not affect the strain's antibiotic susceptibility, but their duple deletion increased susceptibility, mainly mediated by the antitoxins SezA and AbiEi. Further studies have revealed that SezA and AbiEi affect the genetic stability of ICESsuHN105 by moderating the excision and extrachromosomal copy number, consequently affecting the antibiotic resistance conferred by ICE. The DNA-binding proteins AbiEi and SezA, which bind palindromic sequences in the promoter, coordinately modulate ICE excision and extracellular copy number by binding to sequences in the origin-of-transfer (oriT) and the attL sites, respectively. Furthermore, AbiEi negatively regulates the transcription of SezAT by binding directly to its promoter, optimizing the coordinate network of SezAT and AbiE in maintaining ICESsuHN105 stability. Importantly, SezAT and AbiE are widespread and conserved in ICEs harbouring diverse drug-resistance genes, and their coordinated effects in promoting ICE stability and mediating drug resistance may be broadly applicable to other ICEs. Altogether, our study uncovers the TA system's role in maintaining the genetic stability of ICE and offers potential targets for overcoming the dissemination and evolution of drug resistance.

The emergence and diversification of a zoonotic pathogen from within the microbiota of intensively farmed pigs.

The expansion and intensification of livestock production is predicted to promote the emergence of pathogens. As pathogens sometimes jump between species, this can affect the health of humans as well as livestock. Here, we investigate how livestock microbiota can act as a source of these emerging pathogens through analysis of Streptococcus suis, a ubiquitous component of the respiratory microbiota of pigs that is also a major cause of disease on pig farms and an important zoonotic pathogen. Combining molecular dating, phylogeography, and comparative genomic analyses of a large collection of isolates, we find that several pathogenic lineages of S. suis emerged in the 19th and 20th centuries, during an early period of growth in pig farming. These lineages have since spread between countries and continents, mirroring trade in live pigs. They are distinguished by the presence of three genomic islands with putative roles in metabolism and cell adhesion, and an ongoing reduction in genome size, which may reflect their recent shift to a more pathogenic ecology. Reconstructions of the evolutionary histories of these islands reveal constraints on pathogen emergence that could inform control strategies, with pathogenic lineages consistently emerging from one subpopulation of S. suis and acquiring genes through horizontal transfer from other pathogenic lineages. These results shed light on the capacity of the microbiota to rapidly evolve to exploit changes in their host population and suggest that the impact of changes in farming on the pathogenicity and zoonotic potential of S. suis is yet to be fully realized.

Membrane vesicle delivery of a streptococcal M protein disrupts the blood-brain barrier by inducing autophagic endothelial cell death.

M family proteins are critical virulence determinants of Streptococci. Streptococcus equi subsp. zooepidemicus (SEZ) are Group C streptococci that cause meningitis in animals and humans. SzM, the M protein of SEZ, has been linked to SEZ brain invasion. Here, we demonstrate that SzM is important in SEZ disruption of the blood-brain barrier (BBB). SEZ release SzM-bound membrane vesicles (MVs), and endocytosis of these vesicles by human brain endothelial microvascular cells (hBMECs) results in SzM-dependent cytotoxicity. Furthermore, administration of SzM-bound MVs disrupted the murine BBB. A CRISPR screen revealed that SzM cytotoxicity in hBMECs depends on PTEN-related activation of autophagic cell death. Pharmacologic inhibition of PTEN activity prevented SEZ disruption of the murine BBB and delayed mortality. Our data show that MV delivery of SzM to host cells plays a key role in SEZ pathogenicity and suggests that MV delivery of streptococcal M family proteins is likely a common streptococcal virulence mechanism.

Phosphorylation of GntR reduces Streptococcus suis oxidative stress resistance and virulence by inhibiting NADH oxidase transcription.

GntR transcription factor of Streptococcus suis serotype 2 (SS2) is a potential substrate protein of STK, but the regulation mechanisms of GntR phosphorylation are still unclear. This study confirmed that STK phosphorylated GntR in vivo, and in vitro phosphorylation experiments showed that STK phosphorylated GntR at Ser-41. The phosphomimetic strain (GntR-S41E) had significantly reduced lethality in mice and reduced bacterial load in the blood, lung, liver, spleen, and brain of infected mice compared to wild-type (WT) SS2. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) experiments demonstrated that the promoter of nox was bound by GntR. The phosphomimetic protein GntR-S41E cannot bind to the promoter of nox, and the nox transcription levels were significantly reduced in the GntR-S41E mutant compared to WT SS2. The virulence in mice and the ability to resist oxidative stress of the GntR-S41E strain were restored by complementing transcript levels of nox. NOX is an NADH oxidase that catalyzes the oxidation of NADH to NAD+ with the reduction of oxygen to water. We found that NADH is likely accumulated under oxidative stress in the GntR-S41E strain, and higher NADH levels resulted in increased amplified ROS killing. In total, we report GntR phosphorylation could inhibit the transcription of nox, which impaired the ability of SS2 to resist oxidative stress and virulence.

CD1 and iNKT cells mediate immune responses against the GBS hemolytic lipid toxin induced by a non-toxic analog.

Although hemolytic lipids have been discovered from many human pathogens including Group B Streptococcus (GBS), strategies that neutralize their function are lacking. GBS is a leading cause of pregnancy-associated neonatal infections, and adult GBS infections are on the rise. The GBS hemolytic lipid toxin or granadaene, is cytotoxic to many immune cells including T and B cells. We previously showed that mice immunized with a synthetic nontoxic analog of granadaene known as R-P4 had reduced bacterial dissemination during systemic infection. However, mechanisms important for R-P4 mediated immune protection was not understood. Here, we show that immune serum from R-P4-immunized mice facilitate GBS opsonophagocytic killing and protect naïve mice from GBS infection. Further, CD4+ T cells isolated from R-P4-immunized mice proliferated in response to R-P4 stimulation in a CD1d- and iNKT cell-dependent manner. Consistent with these observations, R-P4 immunized mice lacking CD1d or CD1d-restricted iNKT cells exhibit elevated bacterial burden. Additionally, adoptive transfer of iNKT cells from R-P4 vaccinated mice significantly reduced GBS dissemination compared to adjuvant controls. Finally, maternal R-P4 vaccination provided protection against ascending GBS infection during pregnancy. These findings are relevant in the development of therapeutic strategies targeting lipid cytotoxins.

Type 2 Interleukin-4 Receptor Signaling in Neutrophils Antagonizes Their Expansion and Migration during Infection and Inflammation.

Neutrophils are the first immune cells recruited to sites of inflammation and infection. However, patients with allergic disorders such as atopic dermatitis show a paucity of skin neutrophils and are prone to bacterial skin infections, suggesting that allergic inflammation curtails neutrophil responses. Here we have shown that the type 2 cell signature cytokine interleukin-4 (IL-4) hampers neutrophil expansion and migration by antagonizing granulocyte colony-stimulating factor (G-CSF) and chemokine receptor-mediated signals. Cutaneous bacterial infection in mice was exacerbated by IL-4 signaling and improved with IL-4 inhibition, each outcome inversely correlating with neutrophil migration to skin. Likewise, systemic bacterial infection was worsened by heightened IL-4 activity, with IL-4 restricting G-CSF-induced neutrophil expansion and migration to tissues by affecting CXCR2-CXCR4 chemokine signaling in neutrophils. These effects were dependent on IL-4 acting through type 2 IL-4 receptors on neutrophils. Thus, targeting IL-4 might be beneficial in neutropenic conditions with increased susceptibility to bacterial infections.

Enolase of Streptococcus suis serotype 2 promotes biomolecular condensation of ribosomal protein SA for HBMECs apoptosis.

BACKGROUND: Ribosomal protein SA (RPSA) of human brain microvascular endothelial cells (HBMECs) can transfer from the cytosol to the cell surface and act as a receptor for some pathogens, including Streptococcus suis serotype 2 (SS2), a zoonotic pathogen causing meningitis in pigs and humans. We previously reported that SS2 virulence factor enolase (ENO) binds to RPSA on the cell surface of HBMECs and induces apoptosis. However, the mechanism that activates RPSA translocation to the cell surface and induces ENO-mediated HBMEC apoptosis is unclear. RESULTS: Here, we show that RPSA localization and condensation on the host cell surface depend on its internally disordered region (IDR). ENO binds to the IDR of RPSA and promotes its interaction with RPSA and vimentin (VIM), which is significantly suppressed after 1,6-Hexanediol (1,6-Hex, a widely used tool to disrupt phase separation) treatment, indicating that ENO incorporation and thus the concentration of RPSA/VIM complexes via co-condensation. Furthermore, increasing intracellular calcium ions (Ca2+) in response to SS2 infection further facilitates the liquid-like condensation of RPSA and aggravates ENO-induced HBMEC cell apoptosis. CONCLUSIONS: Together, our study provides a previously underappreciated molecular mechanism illuminating that ENO-induced RPSA condensation activates the migration of RPSA to the bacterial cell surface and stimulates SS2-infected HBMEC death and, potentially, disease progression. This study offers a fresh avenue for investigation into the mechanism by which other harmful bacteria infect hosts via cell surfaces' RPSA.

Streptococcus suis Serotype 2 Type IV Secretion Effector SspA-1 Induces Proinflammatory Cytokine Production via TLR2 Endosomal and Type I Interferon Signaling.

The subtilisin-like protease-1 (SspA-1) plays an important role in the pathogenesis of a highly virulent strain of Streptococcus suis 2. However, the mechanism of SspA-1-triggered excessive inflammatory response is still unknown. In this study, we demonstrated that activation of type I IFN signaling is required for SspA-1-induced excessive proinflammatory cytokine production. Further experiments showed that the TLR2 endosomal pathway mediates SspA-1-induced type I IFN signaling and the inflammatory response. Finally, we mapped the major signaling components of the related pathway and found that the TIR adaptor proteins Mal, TRAM, and MyD88 and the downstream activation of IRF1 and IRF7 were involved in this pathway. These results explain the molecular mechanism by which SspA-1 triggers an excessive inflammatory response and reveal a novel effect of type I IFN in S. suis 2 infection, possibly providing further insights into the pathogenesis of this highly virulent S. suis 2 strain.

Group B Streptococcus transcriptome when interacting with brain endothelial cells.

Bacterial meningitis is a life-threatening infection of the central nervous system (CNS) that occurs when bacteria are able to cross the blood-brain barrier (BBB) or the meningeal-cerebrospinal fluid barrier (mBCSFB). The BBB and mBCSFB comprise highly specialized brain endothelial cells (BECs) that typically restrict pathogen entry. Group B Streptococcus (GBS or Streptococcus agalactiae) is the leading cause of neonatal meningitis. Until recently, identification of GBS virulence factors has relied on genetic screening approaches. Instead, we here conducted RNA-seq analysis on GBS when interacting with induced pluripotent stem cell-derived BECs (iBECs) to pinpoint virulence-associated genes. Of the 2,068 annotated protein-coding genes of GBS, 430 transcripts displayed significant changes in expression after interacting with BECs. Notably, we found that the majority of differentially expressed GBS transcripts were downregulated (360 genes) during infection of iBECs. Interestingly, codY, encoding a pleiotropic transcriptional repressor in low-G + C Gram-positive bacteria, was identified as being highly downregulated. We conducted qPCR to confirm the codY downregulation observed via RNA-seq during the GBS-iBEC interaction and obtained codY mutants in three different GBS background parental strains. As anticipated from the RNA-seq results, the [Formula: see text]codY strains were more adherent and invasive in two in vitro BEC models. Together, this demonstrates the utility of RNA-seq during the BEC interaction to identify GBS virulence modulators. IMPORTANCE: Group B Streptococcus (GBS) meningitis remains the leading cause of neonatal meningitis. Research work has identified surface factors and two-component systems that contribute to GBS disruption of the blood-brain barrier (BBB). These discoveries often relied on genetic screening approaches. Here, we provide transcriptomic data describing how GBS changes its transcriptome when interacting with brain endothelial cells. Additionally, we have phenotypically validated these data by obtaining mutants of a select regulator that is highly down-regulated during infection and testing on our BBB model. This work provides the research field with a validated data set that can provide an insight into potential pathways that GBS requires to interact with the BBB and open the door to new discoveries.

Dpr-mediated H2O2 resistance contributes to streptococcus survival in a cystic fibrosis airway model system.

The cystic fibrosis (CF) lung environment is conducive to the colonization of bacteria as polymicrobial biofilms, which are associated with poor clinical outcomes for persons with CF (pwCF). Streptococcus spp. are highly prevalent in the CF airway, but its role in the CF lung microbiome is poorly understood. Some studies have shown Streptococcus spp. to be associated with better clinical outcomes for pwCF, while others show that high abundance of Streptococcus spp. is correlated with exacerbations. Our lab previously reported a polymicrobial culture system consisting of four CF-relevant pathogens that can be used to study microbial behavior in a more clinically relevant setting. Here, we use this model system to identify genetic pathways that are important for Streptococcus sanguinis survival in the context of the polymicrobial community. We identified genes related to reactive oxygen species as differentially expressed in S. sanguinis monoculture versus growth of this microbe in the mixed community. Genetic studies identified Dpr as important for S. sanguinis survival in the community. We show that Dpr, a DNA-binding ferritin-like protein, and PerR, a peroxide-responsive transcriptional regulator of Dpr, are important for protecting S. sanguinis from phenazine-mediated toxicity in co-culture with Pseudomonas aeruginosa and when exposed to hydrogen peroxide, both of which mimic the CF lung environment. Characterizing such interactions in a clinically relevant model system contributes to our understanding of microbial behavior in the context of polymicrobial biofilm infections. IMPORTANCE: Streptococcus spp. are recognized as a highly prevalent pathogen in cystic fibrosis (CF) airway infections. However, the role of this microbe in clinical outcomes for persons with CF is poorly understood. Here, we leverage a polymicrobial community system previously developed by our group to model CF airway infections as a tool to investigate a Pseudomonas-Streptococcus interaction involving reactive oxygen species (ROS). We show that protection against ROS is required for Streptococcus sanguinis survival in a clinically relevant polymicrobial system. Using this model system to study interspecies interactions contributes to our broader understanding of the complex role of Streptococcus spp. in the CF lung.

Specific interaction between Group B Streptococcus CC17 hypervirulent clone and phagocytes.

Streptococcus agalactiae also named Group B Streptococcus (GBS) is the most significant pathogen causing invasive infections, such as bacteremia and meningitis, in neonates. Worldwide epidemiological studies have shown that a particular clonal complex (CC) of capsular serotype III, the CC17, is strongly associated with meningitis in neonates and is therefore, designated as the hypervirulent clone. Macrophages are a permissive niche for intracellular bacteria of all GBS clones. In this study, we deciphered the specific interaction of GBS CC17 strains with macrophages. Our study revealed that CC17 strains are phagocytosed at a higher rate than GBS non-CC17 strains by human monocytes and macrophages both in cellular models and in primary cells. CC17-enhanced phagocytosis is due to an initial enhanced-attachment step to macrophages mediated by the CC17-specific surface protein HvgA and the PI-2b pilus (Spb1). We showed that two different inhibitors of scavenger receptors (fucoidan and poly(I)) specifically inhibited CC17 adhesion and phagocytosis while not affecting those of non-CC17 strains. Once phagocytosed, both CC17 and non-CC17 strains remained in a LAMP-1 positive vacuole that ultimately fuses with lysosomes where they can survive at similar rates. Finally, both strains displayed a basal egress which occurs independently from actin and microtubule networks. Our findings provide new insights into the interplay between the hypervirulent GBS CC17 and major players of the host's innate immune response. This enhanced adhesion, leading to increased phagocytosis, could reflect a peculiar capacity of the CC17 lineage to subvert the host immune defenses, establish a niche for persistence or disseminate.

Manganese uptake by MtsABC contributes to the pathogenesis of human pathogen group A streptococcus by resisting host nutritional immune defenses.

The interplay between host nutritional immune mechanisms and bacterial nutrient uptake systems has a major impact on the disease outcome. The host immune factor calprotectin (CP) limits the availability of essential transition metals, such as manganese (Mn) and zinc (Zn), to control the growth of invading pathogens. We previously demonstrated that the competition between CP and the human pathogen group A streptococcus (GAS) for Zn impacts GAS pathogenesis. However, the contribution of Mn sequestration by CP in GAS infection control and the role of GAS Mn acquisition systems in overcoming host-imposed Mn limitation remain unknown. Using a combination of in vitro and in vivo studies, we show that GAS-encoded mtsABC is a Mn uptake system that aids bacterial evasion of CP-imposed Mn scarcity and promotes GAS virulence. Mn deficiency caused by either the inactivation of mtsC or CP also impaired the protective function of GAS-encoded Mn-dependent superoxide dismutase. Our ex vivo studies using human saliva show that saliva is a Mn-scant body fluid, and Mn acquisition by MtsABC is critical for GAS survival in human saliva. Finally, animal infection studies using wild-type (WT) and CP-/- mice showed that MtsABC is critical for GAS virulence in WT mice but dispensable in mice lacking CP, indicating the direct interplay between MtsABC and CP in vivo. Together, our studies elucidate the role of the Mn import system in GAS evasion of host-imposed metal sequestration and underscore the translational potential of MtsABC as a therapeutic or prophylactic target.