RESUMEN
A mosaic of cross-phylum chemical interactions occurs between all metazoans and their microbiomes. A number of molecular families that are known to be produced by the microbiome have a marked effect on the balance between health and disease1-9. Considering the diversity of the human microbiome (which numbers over 40,000 operational taxonomic units10), the effect of the microbiome on the chemistry of an entire animal remains underexplored. Here we use mass spectrometry informatics and data visualization approaches11-13 to provide an assessment of the effects of the microbiome on the chemistry of an entire mammal by comparing metabolomics data from germ-free and specific-pathogen-free mice. We found that the microbiota affects the chemistry of all organs. This included the amino acid conjugations of host bile acids that were used to produce phenylalanocholic acid, tyrosocholic acid and leucocholic acid, which have not previously been characterized despite extensive research on bile-acid chemistry14. These bile-acid conjugates were also found in humans, and were enriched in patients with inflammatory bowel disease or cystic fibrosis. These compounds agonized the farnesoid X receptor in vitro, and mice gavaged with the compounds showed reduced expression of bile-acid synthesis genes in vivo. Further studies are required to confirm whether these compounds have a physiological role in the host, and whether they contribute to gut diseases that are associated with microbiome dysbiosis.
Asunto(s)
Ácidos y Sales Biliares/biosíntesis , Ácidos y Sales Biliares/química , Metabolómica , Microbiota/fisiología , Animales , Ácidos y Sales Biliares/metabolismo , Ácido Cólico/biosíntesis , Ácido Cólico/química , Ácido Cólico/metabolismo , Fibrosis Quística/genética , Fibrosis Quística/metabolismo , Fibrosis Quística/microbiología , Vida Libre de Gérmenes , Humanos , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/microbiología , Ratones , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismoRESUMEN
Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause of adult life-threatening sepsis and urinary tract infections (UTI). The emergence and spread of multidrug-resistant (MDR) ExPEC strains result in a considerable amount of treatment failure and hospitalization costs, and contribute to the spread of drug resistance amongst the human microbiome. Thus, an effective vaccine against ExPEC would reduce morbidity and mortality and possibly decrease carriage in healthy or diseased populations. A comparative genomic analysis demonstrated a gene encoding an invasin-like protein, termed sinH, annotated as an autotransporter protein, shows high prevalence in various invasive ExPEC phylogroups, especially those associated with systemic bacteremia and UTI. Here, we evaluated the protective efficacy and immunogenicity of a recombinant SinH-based vaccine consisting of either domain-3 or domains-1,2, and 3 of the putative extracellular region of surface-localized SinH. Immunization of a murine host with SinH-based antigens elicited significant protection against various strains of the pandemic ExPEC sequence type 131 (ST131) as well as multiple sequence types in two distinct models of infection (colonization and bacteremia). SinH immunization also provided significant protection against ExPEC colonization in the bladder in an acute UTI model. Immunized cohorts produced significantly higher levels of vaccine-specific serum IgG and urinary IgG and IgA, findings consistent with mucosal protection. Collectively, these results demonstrate that autotransporter antigens such as SinH may constitute promising ExPEC phylogroup-specific and sequence-type effective vaccine targets that reduce E. coli colonization and virulence.
Asunto(s)
Bacteriemia , Infecciones por Escherichia coli , Escherichia coli Patógena Extraintestinal , Infecciones Urinarias , Animales , Humanos , Ratones , Escherichia coli , Sistemas de Secreción Tipo V/genética , Infecciones por Escherichia coli/prevención & control , Escherichia coli Patógena Extraintestinal/genética , Vacunación , Factores de Virulencia/genética , Vacunas Sintéticas , Infecciones Urinarias/prevención & control , Bacteriemia/prevención & control , Inmunoglobulina G/farmacologíaRESUMEN
Extraintestinal pathogenic Escherichia coli (ExPEC) is a leading cause of worldwide morbidity and mortality, the top cause of antimicrobial-resistant (AMR) infections, and the most frequent cause of life-threatening sepsis and urinary tract infections (UTI) in adults. The development of an effective and universal vaccine is complicated by this pathogen's pan-genome, its ability to mix and match virulence factors and AMR genes via horizontal gene transfer, an inability to decipher commensal from pathogens, and its intimate association and co-evolution with mammals. Using a pan virulome analysis of >20,000 sequenced E. coli strains, we identified the secreted cytolysin α-hemolysin (HlyA) as a high priority target for vaccine exploration studies. We demonstrate that a catalytically inactive pure form of HlyA, expressed in an autologous host using its own secretion system, is highly immunogenic in a murine host, protects against several forms of ExPEC infection (including lethal bacteremia), and significantly lowers bacterial burdens in multiple organ systems. Interestingly, the combination of a previously reported autotransporter (SinH) with HlyA was notably effective, inducing near complete protection against lethal challenge, including commonly used infection strains ST73 (CFT073) and ST95 (UTI89), as well as a mixture of 10 of the most highly virulent sequence types and strains from our clinical collection. Both HlyA and HlyA-SinH combinations also afforded some protection against UTI89 colonization in a murine UTI model. These findings suggest recombinant, inactive hemolysin and/or its combination with SinH warrant investigation in the development of an E. coli vaccine against invasive disease.
Asunto(s)
Infecciones por Escherichia coli , Proteínas de Escherichia coli , Vacunas contra Escherichia coli , Escherichia coli Patógena Extraintestinal , Proteínas Hemolisinas , Animales , Escherichia coli Patógena Extraintestinal/genética , Escherichia coli Patógena Extraintestinal/inmunología , Infecciones por Escherichia coli/prevención & control , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/inmunología , Ratones , Proteínas Hemolisinas/inmunología , Proteínas Hemolisinas/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/inmunología , Vacunas contra Escherichia coli/inmunología , Antígenos Bacterianos/inmunología , Antígenos Bacterianos/genética , Femenino , Factores de Virulencia/genética , Factores de Virulencia/inmunología , Sistemas de Secreción Tipo V/inmunología , Sistemas de Secreción Tipo V/genética , Modelos Animales de Enfermedad , HumanosRESUMEN
Type VIIb secretion systems (T7SSb) in Gram-positive bacteria facilitate physiology, interbacterial competition, and/or virulence via EssC ATPase-driven secretion of small É-helical proteins and toxins. Recently, we characterized T7SSb in group B Streptococcus (GBS), a leading cause of infection in newborns and immunocompromised adults. GBS T7SS comprises four subtypes based on variation in the C-terminus of EssC and the repertoire of downstream effectors; however, the intraspecies diversity of GBS T7SS and impact on GBS-host interactions remains unknown. Bioinformatic analysis indicates that GBS T7SS loci encode subtype-specific putative effectors, which have low interspecies and inter-subtype homology but contain similar domains/motifs and therefore may serve similar functions. We further identify orphaned GBS WXG100 proteins. Functionally, we show that GBS T7SS subtype I and III strains secrete EsxA in vitro and that in subtype I strain CJB111, esxA1 appears to be differentially transcribed from the T7SS operon. Furthermore, we observe subtype-specific effects of GBS T7SS on host colonization, as CJB111 subtype I but not CNCTC 10/84 subtype III T7SS promotes GBS vaginal colonization. Finally, we observe that T7SS subtypes I and II are the predominant subtypes in clinical GBS isolates. This study highlights the potential impact of T7SS heterogeneity on host-GBS interactions.
Asunto(s)
Infecciones Estreptocócicas , Sistemas de Secreción Tipo VII , Recién Nacido , Femenino , Humanos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Sistemas de Secreción Tipo VII/genética , Virulencia , Operón/genética , Genitales Femeninos/metabolismo , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/genética , Streptococcus agalactiae/metabolismo , Vagina/metabolismo , Vagina/microbiologíaRESUMEN
Multidrug-resistant (MDR) Enterococcus faecalis are major causes of hospital-acquired infections. Numerous clinical strains of E. faecalis harbor a large pathogenicity island that encodes enterococcal surface protein (Esp), which is suggested to promote biofilm production and virulence, but this remains controversial. To resolve this issue, we characterized the Esp N-terminal region, the portion implicated in biofilm production. Small angle X-ray scattering indicated that the N-terminal region had a globular head, which consisted of two DEv-Ig domains as visualized by X-ray crystallography, followed by an extended tail. The N-terminal region was not required for biofilm production but instead significantly strengthened biofilms against mechanical or degradative disruption, greatly increasing retention of Enterococcus within biofilms. Biofilm strengthening required low pH, which resulted in Esp unfolding, aggregating, and forming amyloid-like structures. The pH threshold for biofilm strengthening depended on protein stability. A truncated fragment of the first DEv-Ig domain, plausibly generated by a host protease, was the least stable and sufficient to strengthen biofilms at pH ≤ 5.0, while the entire N-terminal region and intact Esp on the enterococcal surface was more stable and required a pH ≤ 4.3. These results suggested a virulence role of Esp in strengthening enterococcal biofilms in acidic abiotic or host environments.
Asunto(s)
Infecciones por Bacterias Grampositivas , Proteínas de la Membrana , Proteínas Bacterianas/metabolismo , Biopelículas , Enterococcus/genética , Enterococcus/metabolismo , Enterococcus faecalis , Humanos , Proteínas de la Membrana/metabolismo , Péptido Hidrolasas/metabolismoRESUMEN
BACKGROUND: Maternal rectovaginal colonization by group B Streptococcus (GBS) increases the risk of perinatal GBS disease that can lead to death or long-term neurological impairment. Factors that increase the risk of rectovaginal GBS carriage are incompletely understood resulting in missed opportunities for detecting GBS in risk-based clinical approaches. There is a lacking consensus on whether gestational diabetes mellitus (GDM) is a risk factor for rectovaginal GBS. This systematic review and meta-analysis aims to address current conflicting findings and determine whether GDM should be clinically considered as a risk factor for maternal GBS colonization. METHODS: Peer-reviewed studies that provided GDM prevalence and documented GBS vaginal and/or rectal colonization in women with and without GDM were included in this analysis. From study inception to October 30, 2023, we identified 6,275 relevant studies from EMBASE and PUBMED of which 19 were eligible for inclusion. Eligible studies were analyzed and thoroughly assessed for risk of bias with a modified Newcastle-Ottawa Scale that interrogated representativeness and comparability of cohorts, quality of reporting for GDM and GBS status, and potential bias from other metabolic diseases. Results were synthesized using STATA 18 and analyzed using random-effects meta-analyses. RESULTS: Studies encompassed 266,706 women from 10 different countries, with study periods spanning from 1981 to 2020. Meta-analysis revealed that gestational diabetes is associated with a 16% increased risk of rectovaginal GBS carriage (OR 1.16, CI 1.07-1.26, P = 0.003). We also performed subgroup analyses to assess independent effects of pregestational vs. gestational diabetes on risk of maternal GBS carriage. Pregestational diabetes (Type 1 or Type 2 diabetes mellitus) was also associated with an increased risk of 76% (pooled OR 1.76, CI 1.27-2.45, P = 0.0008). CONCLUSIONS: This study achieved a consensus among previously discrepant observations and demonstrated that gestational diabetes and pregestational diabetes are significant risk factors for maternal rectovaginal carriage of GBS. Recognition of GDM as a risk factor during clinical decisions about GBS screening and intrapartum antibiotic prophylaxis may decrease the global burden of GBS on maternal-perinatal health.
Asunto(s)
Diabetes Gestacional , Complicaciones Infecciosas del Embarazo , Recto , Infecciones Estreptocócicas , Streptococcus agalactiae , Vagina , Humanos , Diabetes Gestacional/epidemiología , Femenino , Embarazo , Factores de Riesgo , Infecciones Estreptocócicas/epidemiología , Vagina/microbiología , Complicaciones Infecciosas del Embarazo/epidemiología , Complicaciones Infecciosas del Embarazo/microbiología , Recto/microbiologíaRESUMEN
Group B Streptococcus (GBS) is a pervasive neonatal pathogen accounting for a combined half a million deaths and stillbirths annually. The most common source of fetal or neonatal GBS exposure is the maternal microbiota. GBS asymptomatically colonizes the gastrointestinal and vaginal mucosa of 1 in 5 individuals globally, although its precise role in these niches is not well understood. To prevent vertical transmission, broad-spectrum antibiotics are administered to GBS-positive mothers during labor in many countries. Although antibiotics have significantly reduced GBS early-onset neonatal disease, there are several unintended consequences, including an altered neonatal microbiota and increased risk for other microbial infections. Additionally, the incidence of late-onset GBS neonatal disease remains unaffected and has sparked an emerging hypothesis that GBS-microbe interactions in developing neonatal gut microbiota may be directly involved in this disease process. This review summarizes our current understanding of GBS interactions with other resident microbes at the mucosal surface from multiple angles, including clinical association studies, agriculture and aquaculture observations, and experimental animal model systems. We also include a comprehensive review of in vitro findings of GBS interactions with other bacterial and fungal microbes, both commensal and pathogenic, along with newly established animal models of GBS vaginal colonization and in utero or neonatal infection. Finally, we provide a perspective on emerging areas of research and current strategies to design microbe-targeting prebiotic or probiotic therapeutic intervention strategies to prevent GBS disease in vulnerable populations.
Asunto(s)
Enfermedades del Recién Nacido , Complicaciones Infecciosas del Embarazo , Infecciones Estreptocócicas , Femenino , Animales , Recién Nacido , Humanos , Embarazo , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae , Antibacterianos , Red Social , Complicaciones Infecciosas del Embarazo/microbiologíaRESUMEN
Background: Streptococcus agalactiae (group B Streptococcus [GBS]) asymptomatically colonizes approximately 20% of adults; however, GBS causes severe disease in susceptible populations, including newborns, pregnant women, and elderly individuals. In shifting between commensal and pathogenic states, GBS reveals multiple mechanisms of virulence factor control. Here we describe a GBS protein that we named "biofilm regulatory protein A" (BrpA) on the basis of its homology with BrpA from Streptococcus mutans. Methods: We coupled phenotypic assays, RNA sequencing, human neutrophil and whole-blood killing assays, and murine infection models to investigate the contribution of BrpA to GBS physiology and virulence. Results: Sequence analysis identified BrpA as a LytR-CpsA-Psr enzyme. Targeted mutagenesis yielded a GBS mutant (ΔbrpA) with normal ultrastructural morphology but a 6-fold increase in chain length, a biofilm defect, and decreased acid tolerance. GBS ΔbrpA stimulated increased neutrophil reactive oxygen species and proved more susceptible to human and murine blood and neutrophil killing. Notably, the wild-type parent outcompeted ΔbrpA GBS in murine sepsis and vaginal colonization models. RNA sequencing of ΔbrpA uncovered multiple differences from the wild-type parent, including pathways of cell wall synthesis and cellular metabolism. Conclusions: We propose that BrpA is an important virulence regulator and potential target for design of novel antibacterial therapeutics against GBS.
Asunto(s)
Proteínas Bacterianas/fisiología , Inmunidad Innata/inmunología , Streptococcus agalactiae/inmunología , Streptococcus agalactiae/patogenicidad , Animales , Biopelículas , Línea Celular , Femenino , Interacciones Huésped-Patógeno/inmunología , Interacciones Huésped-Patógeno/fisiología , Humanos , Ratones , Neutrófilos/inmunología , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/química , Streptococcus agalactiae/fisiologíaRESUMEN
Urinary tract infections (UTIs) caused by the human fungal pathogen Candida albicans and related species are prevalent in hospitalized patients, especially those on antibiotic therapy, with indwelling catheters, or with predisposing conditions such as diabetes or immunodeficiency. Understanding of key host defenses against Candida UTI is critical for developing effective treatment strategies. Tamm-Horsfall glycoprotein (THP) is the most abundant urine protein, with multiple roles in renal physiology and bladder protection. THP protects against bacterial UTI by blocking bacterial adherence to the bladder epithelium, but its role in defense against fungal pathogens is not yet described. Here we demonstrate that THP restricts colonization of the urinary tract by C. albicans THP binds to C. albicans hyphae, but not the yeast form, in a manner dependent on fungal expression of the Als3 adhesion glycoprotein. THP directly blocks C. albicans adherence to bladder epithelial cells in vitro, and THP-deficient mice display increased fungal burden in a C. albicans UTI model. This work outlines a previously unknown role for THP as an essential component for host immune defense against fungal urinary tract infection.
Asunto(s)
Candida albicans/patogenicidad , Candidiasis/inmunología , Infecciones Urinarias/inmunología , Sistema Urinario/microbiología , Uromodulina/inmunología , Animales , Candidiasis/orina , Línea Celular , Femenino , Proteínas Fúngicas/genética , Humanos , Hifa/patogenicidad , Ratones , Ratones Noqueados , Unión Proteica , Infecciones Urinarias/microbiología , Uromodulina/farmacología , Urotelio/microbiologíaRESUMEN
BACKGROUND: Composition of the vaginal microbiota has significant influence on female urogenital health and control of infectious disease. Murine models are widely utilized to characterize host-pathogen interactions within the vaginal tract, however, the composition of endogenous vaginal flora remains largely undefined with modern microbiome analyses. Here, we employ 16S rRNA amplicon sequencing to establish the native microbial composition of the vaginal tract in adult C57Bl/6 J mice. We further interrogate the impact of estrous cycle and introduction of the human vaginal pathobiont, group B Streptococcus (GBS) on community state type and stability, and conversely, the impact of the vaginal microbiota on GBS persistence. RESULTS: Sequencing analysis revealed five distinctive community states of the vaginal microbiota dominated largely by Staphylococcus and/or Enterococcus, Lactobacillus, or a mixed population. Stage of estrus did not impact microbial composition. Introduction of GBS decreased community stability at early timepoints; and in some mice, GBS became the dominant bacterium by day 21. Endogenous Staphylococcus abundance correlated with GBS ascension into the uterus, and increased community stability in GBS-challenged mice. CONCLUSIONS: The murine vaginal flora is diverse and fluctuates independently of the estrous cycle. Endogenous flora may impact pathogen colonization and dissemination and should be considered in urogenital infection models.
Asunto(s)
Bacterias/aislamiento & purificación , Ratones/microbiología , Microbiota , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/crecimiento & desarrollo , Vagina/microbiología , Animales , Bacterias/clasificación , Bacterias/genética , Bacterias/crecimiento & desarrollo , Modelos Animales de Enfermedad , Femenino , Humanos , Ratones Endogámicos C57BL , ARN Ribosómico 16S/genética , Streptococcus agalactiae/genética , Streptococcus agalactiae/aislamiento & purificaciónRESUMEN
Urinary tract infections are a major problem in human medicine for which better understanding of native immune defenses may reveal new pathways for therapeutic intervention. Tamm-Horsfall glycoprotein (THP), the most abundant urinary protein, interacts with bacteria including uropathogenic Escherichia coli (UPEC) as well host immune cells. In addition to its well-studied functions to antagonize bacterial colonization, we hypothesize that THP serves a critical host defense function through innate immune modulation. Using isolated human neutrophils, we found that THP binds neutrophils and that this interaction reduces reactive oxygen species generation, chemotaxis and killing of UPEC. We discovered that THP engages the inhibitory neutrophil receptor sialic acid-binding Ig-like lectin-9 (Siglec-9), and mouse functional ortholog Siglec-E, in a manner dependent on sialic acid on its N-glycan moieties. THP-null mice have significantly more neutrophils present in the urine compared with wild-type mice, both with and without the presence of inflammatory stimuli. These data support THP as an important negative regulator of neutrophil activation in the urinary tract, with dual functions to counteract bacterial colonization and suppress excessive inflammation within the urinary tract.
Asunto(s)
Antígenos CD/metabolismo , Antígenos de Diferenciación de Linfocitos B/metabolismo , Infecciones por Escherichia coli/inmunología , Escherichia coli/inmunología , Neutrófilos/inmunología , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico/metabolismo , Infecciones Urinarias/inmunología , Sistema Urinario/metabolismo , Uromodulina/metabolismo , Animales , Bacteriólisis , Células Cultivadas , Quimiotaxis , Humanos , Inmunidad Innata , Inmunomodulación , Ratones , Ratones Noqueados , Ácido N-Acetilneuramínico/metabolismo , Activación Neutrófila , Unión Proteica , Especies Reactivas de Oxígeno/metabolismo , Sistema Urinario/inmunología , Uromodulina/genéticaRESUMEN
Streptococcus agalactiae (group B streptococcus [GBS]) colonizes the rectovaginal tract in 20% to 30% of women and during pregnancy can be transmitted to the newborn, causing severe invasive disease. Current routine screening and antibiotic prophylaxis have fallen short of complete prevention of GBS transmission, and GBS remains a leading cause of neonatal infection. We have investigated the ability of Streptococcus salivarius, a predominant member of the native human oral microbiota, to control GBS colonization. Comparison of the antibacterial activities of multiple S. salivarius strains by use of a deferred-antagonism test showed that S. salivarius strain K12 exhibited the broadest spectrum of activity against GBS. K12 effectively inhibited all GBS strains tested, including disease-implicated isolates from newborns and colonizing isolates from the vaginal tract of pregnant women. Inhibition was dependent on the presence of megaplasmid pSsal-K12, which encodes the bacteriocins salivaricin A and salivaricin B; however, in coculture experiments, GBS growth was impeded by K12 independently of the megaplasmid. We also demonstrated that K12 adheres to and invades human vaginal epithelial cells at levels comparable to GBS. Inhibitory activity of K12 was examined in vivo using a mouse model of GBS vaginal colonization. Mice colonized with GBS were treated vaginally with K12. K12 administration significantly reduced GBS vaginal colonization in comparison to nontreated controls, and this effect was partially dependent on the K12 megaplasmid. Our results suggest that K12 may have potential as a preventative therapy to control GBS vaginal colonization and thereby prevent its transmission to the neonate during pregnancy.
Asunto(s)
Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/transmisión , Streptococcus agalactiae/crecimiento & desarrollo , Streptococcus/fisiología , Vagina/microbiología , Animales , Línea Celular , Técnicas de Cocultivo , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , RatonesRESUMEN
Group B streptococcus (GBS) can cause severe disease in susceptible hosts, including newborns, pregnant women, and the elderly. GBS serine-rich repeat (Srr) surface glycoproteins are important adhesins/invasins in multiple host tissues, including the vagina. However, exact molecular mechanisms contributing to their importance in colonization are unknown. We have recently determined that Srr proteins contain a fibrinogen-binding region (BR) and hypothesize that Srr-mediated fibrinogen binding may contribute to GBS cervicovaginal colonization. In this study, we observed that fibrinogen enhanced wild-type GBS attachment to cervical and vaginal epithelium, and that this was dependent on Srr1. Moreover, purified Srr1-BR peptide bound directly to host cells, and peptide administration in vivo reduced GBS recovery from the vaginal tract. Furthermore, a GBS mutant strain lacking only the Srr1 "latching" domain exhibited decreased adherence in vitro and decreased persistence in a mouse model of GBS vaginal colonization, suggesting the importance of Srr-fibrinogen interactions in the female reproductive tract.
Asunto(s)
Adhesinas Bacterianas/fisiología , Fibrinógeno/metabolismo , Glicoproteínas de Membrana/fisiología , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/patogenicidad , Vagina/microbiología , Animales , Adhesión Bacteriana , Línea Celular , Modelos Animales de Enfermedad , Epitelio/microbiología , Femenino , Ratones , Serina , Streptococcus agalactiae/fisiología , Enfermedades Vaginales/microbiologíaRESUMEN
Streptococcus agalactiae (Group Bâ Streptococcus, GBS) is a frequent commensal organism of the vaginal tract of healthy women. However, GBS can transition to a pathogen in susceptible hosts, but host and microbial factors that contribute to this conversion are not well understood. GBS CovR/S (CsrR/S) is a two component regulatory system that regulates key virulence elements including adherence and toxin production. We performed global transcription profiling of human vaginal epithelial cells exposed to WT, CovR deficient, and toxin deficient strains, and observed that insufficient regulation by CovR and subsequent increased toxin production results in a drastic increase in host inflammatory responses, particularly in cytokine signalling pathways promoted by IL-8 and CXCL2. Additionally, we observed that CovR regulation impacts epithelial cell attachment and intracellular invasion. In our mouse model of GBS vaginal colonization, we further demonstrated that CovR regulation promotes vaginal persistence, as infection with a CovR deficient strainresulted in a heightened host immune response as measured by cytokine production and neutrophil activation. Using CXCr2 KO mice, we determined that this immune alteration occurs, at least in part, via signalling through the CXCL2 receptor. Taken together, we conclude that CovR is an important regulator of GBS vaginal colonization and loss of this regulatory function may contribute to the inflammatory havoc seen during the course of infection.
Asunto(s)
Regulación de la Expresión Génica , Proteínas Represoras/metabolismo , Transducción de Señal , Streptococcus agalactiae/inmunología , Streptococcus agalactiae/patogenicidad , Vagina/inmunología , Vagina/microbiología , Animales , Proteínas Bacterianas/metabolismo , Línea Celular , Modelos Animales de Enfermedad , Células Epiteliales/inmunología , Células Epiteliales/microbiología , Femenino , Perfilación de la Expresión Génica , Interacciones Huésped-Patógeno , Humanos , Ratones , Ratones Noqueados , Receptores de Interleucina-8B/genética , Receptores de Interleucina-8B/inmunología , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiologíaRESUMEN
The composition of the vaginal microbiota is linked to numerous reproductive health problems, including increased susceptibility to infection, pregnancy complications, and impaired vaginal tissue repair; however, the mechanisms contributing to these adverse outcomes are not yet fully defined. In this review, we highlight recent clinical advancements associating vaginal microbiome composition and function with health outcomes. Subsequently, we provide a summary of emerging models employed to identify microbe-microbe interactions contributing to vaginal health, including metagenomic sequencing, multi-omics approaches, and advances in vaginal microbiota cultivation. Last, we review new in vitro, ex vivo, and in vivo models, such as organoids and humanized microbiota murine models, used to define and mechanistically explore host-microbe interactions at the vaginal mucosa.
Asunto(s)
Medicina Clínica , Microbiota , Embarazo , Femenino , Humanos , Animales , Ratones , Metagenoma , Interacciones Microbiota-Huesped , VaginaRESUMEN
Urinary neutrophils are a hallmark of urinary tract infection (UTI), yet the mechanisms governing their activation, function, and efficacy in controlling infection remain incompletely understood. Tamm-Horsfall glycoprotein (THP), the most abundant protein in urine, uses terminal sialic acids to bind an inhibitory receptor and dampen neutrophil inflammatory responses. We hypothesized that neutrophil modulation is an integral part of THP-mediated host protection. In a UTI model, THP-deficient mice showed elevated urinary tract bacterial burdens, increased neutrophil recruitment, and more severe tissue histopathological changes compared to WT mice. Furthermore, THP-deficient mice displayed impaired urinary NETosis during UTI. To investigate the impact of THP on NETosis, we coupled in vitro fluorescence-based NET assays, proteomic analyses, and standard and imaging flow cytometry with peripheral human neutrophils. We found that THP increases proteins involved in respiratory chain, neutrophil granules, and chromatin remodeling pathways, enhances NETosis in an ROS-dependent manner, and drives NET-associated morphologic features including nuclear decondensation. These effects were observed only in the presence of a NETosis stimulus and could not be solely replicated with equivalent levels of sialic acid alone. We conclude that THP is a critical regulator of NETosis in the urinary tract, playing a key role in host defense against UTI.
RESUMEN
The mammalian Siglec receptor sialoadhesin (Siglec1, CD169) confers innate immunity against the encapsulated pathogen group B Streptococcus (GBS). Newborn lung macrophages have lower expression levels of sialoadhesin at birth compared with the postnatal period, increasing their susceptibility to GBS infection. In this study, we investigate the mechanisms regulating sialoadhesin expression in the newborn mouse lung. In both neonatal and adult mice, GBS lung infection reduced Siglec1 expression, potentially delaying acquisition of immunity in neonates. Suppression of Siglec1 expression required interactions between sialic acid on the GBS capsule and the inhibitory host receptor Siglec-E. The Siglec1 gene contains multiple STAT binding motifs, which could regulate expression of sialoadhesin downstream of innate immune signals. Although GBS infection reduced STAT1 expression in the lungs of wild-type newborn mice, we observed increased numbers of STAT1+ cells in Siglece-/- lungs. To test if innate immune activation could increase sialoadhesin at birth, we first demonstrated that treatment of neonatal lung macrophages ex vivo with inflammatory activators increased sialoadhesin expression. However, overcoming the low sialoadhesin expression at birth using in vivo prenatal exposures or treatments with inflammatory stimuli were not successful. The suppression of sialoadhesin expression by GBS-Siglec-E engagement may therefore contribute to disease pathogenesis in newborns and represent a challenging but potentially appealing therapeutic opportunity to augment immunity at birth.
Asunto(s)
Animales Recién Nacidos , Ratones Noqueados , Ácido N-Acetilneuramínico , Factor de Transcripción STAT1 , Lectina 1 Similar a Ig de Unión al Ácido Siálico , Infecciones Estreptocócicas , Streptococcus agalactiae , Animales , Ratones , Streptococcus agalactiae/inmunología , Ácido N-Acetilneuramínico/metabolismo , Lectina 1 Similar a Ig de Unión al Ácido Siálico/metabolismo , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiología , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT1/genética , Inmunidad Innata , Ratones Endogámicos C57BL , Pulmón/inmunología , Pulmón/microbiología , Pulmón/metabolismo , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/metabolismo , Femenino , Macrófagos/inmunología , Macrófagos/metabolismo , Lectinas/metabolismo , Lectinas/genética , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico/metabolismo , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico/genética , Antígenos CD/metabolismo , Antígenos CD/genética , Antígenos de Diferenciación de Linfocitos BRESUMEN
Group B Streptococcus (GBS) is a pervasive perinatal pathogen, yet factors driving GBS dissemination in utero are poorly defined. Gestational diabetes mellitus (GDM), a complication marked by dysregulated immunity and maternal microbial dysbiosis, increases risk for GBS perinatal disease. Using a murine GDM model of GBS colonization and perinatal transmission, we find that GDM mice display greater GBS in utero dissemination and subsequently worse neonatal outcomes. Dual-RNA sequencing reveals differential GBS adaptation to the GDM reproductive tract, including a putative glycosyltransferase (yfhO), and altered host responses. GDM immune disruptions include reduced uterine natural killer cell activation, impaired recruitment to placentae, and altered maternofetal cytokines. Lastly, we observe distinct vaginal microbial taxa associated with GDM status and GBS invasive disease status. Here, we show a model of GBS dissemination in GDM hosts that recapitulates several clinical aspects and identifies multiple host and bacterial drivers of GBS perinatal disease.
Asunto(s)
Diabetes Gestacional , Microbiota , Infecciones Estreptocócicas , Embarazo , Femenino , Humanos , Animales , Ratones , Transmisión Vertical de Enfermedad Infecciosa , Citocinas , Vagina/microbiología , Streptococcus , Streptococcus agalactiae , Infecciones Estreptocócicas/microbiologíaRESUMEN
Streptococcus agalactiae (group B Streptococcus [GBS]) is a Gram-positive bacterium that colonizes the cervicovaginal tract in approximately 25% of healthy women. Although colonization is asymptomatic, GBS can be vertically transmitted to newborns peripartum, causing severe disease such as pneumonia and meningitis. Current prophylaxis, consisting of late gestation screening and intrapartum antibiotics, has failed to completely prevent transmission, and GBS remains a leading cause of neonatal sepsis and meningitis in the United States. Lack of an effective vaccine and emerging antibiotic resistance necessitate exploring novel therapeutic strategies. We have employed a host-directed immunomodulatory therapy using a novel peptide, known as EP67, derived from the C-terminal region of human complement component C5a. Previously, we have demonstrated in vivo that EP67 engagement of the C5a receptor (CD88) effectively limits staphylococcal infection by promoting cytokine release and neutrophil infiltration. Here, using our established mouse model of GBS vaginal colonization, we observed that EP67 treatment results in rapid clearance of GBS from the murine vagina. However, this was not dependent on functional neutrophil recruitment or CD88 signaling, as EP67 treatment reduced the vaginal bacterial load in mice lacking CD88 or the major neutrophil receptor CXCr2. Interestingly, we found that EP67 inhibits GBS growth in vitro and in vivo and that antibacterial activity was specific to Streptococcus species. Our work establishes that EP67-mediated clearance of GBS is likely due to direct bacterial killing rather than to enhanced immune stimulation. We conclude that EP67 may have potential as a therapeutic to control GBS vaginal colonization.
Asunto(s)
Antibacterianos/farmacología , Complemento C5a/química , Péptidos/farmacología , Infecciones del Sistema Genital/tratamiento farmacológico , Infecciones Estreptocócicas/tratamiento farmacológico , Streptococcus agalactiae/efectos de los fármacos , Secuencia de Aminoácidos , Animales , Antibacterianos/síntesis química , Recuento de Colonia Microbiana , Femenino , Expresión Génica , Humanos , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Neutrófilos/inmunología , Neutrófilos/metabolismo , Péptidos/síntesis química , Receptor de Anafilatoxina C5a/genética , Receptor de Anafilatoxina C5a/inmunología , Receptores de Interleucina-8B/genética , Receptores de Interleucina-8B/inmunología , Infecciones del Sistema Genital/inmunología , Infecciones del Sistema Genital/microbiología , Especificidad de la Especie , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/crecimiento & desarrollo , Vagina/efectos de los fármacos , Vagina/microbiologíaRESUMEN
Vaginal microbiota composition is associated with differential risk of urogenital infection. Although vaginal Lactobacillus spp. are thought to confer protection through acidification, bacteriocin production, and immunomodulation, lack of an in vivo model system that closely resembles the human vaginal microbiota remains a prominent barrier to mechanistic discovery. We performed 16S rRNA amplicon sequencing of wildtype C57BL/6J mice, commonly used to study pathogen colonization, and found that the vaginal microbiome composition varies highly both within and between colonies from three distinct vivaria. Because of the strong influence of environmental exposure on vaginal microbiome composition, we assessed whether a humanized microbiota mouse ( HMb mice) would model a more human-like vaginal microbiota. Similar to humans and conventional mice, HMb mice vaginal microbiota clustered into five community state types ( h mCST). Uniquely, HMb mice vaginal communities were frequently dominated by Lactobacilli or Enterobacteriaceae . Compared to genetically-matched conventional mice, HMb mice were less susceptible to uterine ascension by urogenital pathobionts group B Streptococcus (GBS) and Prevotella bivia , but no differences were observed with uropathogenic E. coli . Specifically, vaginal Enterobacteriaceae and Lactobacillus were associated with the absence of uterine GBS. Anti-GBS activity of HMb mice vaginal E. coli and L. murinus isolates, representing Enterobacteriaceae and Lactobacillus respectively, were characterized in vitro and in vivo . Although L. murinus reduced GBS growth in vitro , vaginal pre-inoculation with HMb mouse-derived E. coli , but not L. murinus , conferred protection against vaginal GBS burden. Overall, the HMb mice are an improved model to elucidate the role of endogenous microbes in conferring protection against urogenital pathogens. IMPORTANCE: An altered vaginal microbiota, typically with little to no levels of Lactobacillus , is associated with increased susceptibility to urogenital infections, although mechanisms driving this vulnerability are not fully understood. Despite known inhibitory properties of Lactobacillus against urogenital pathogens, clinical studies with Lactobacillus probiotics have shown mixed success. In this study, we characterize the impact of the vaginal microbiota on urogenital pathogen colonization using a humanized microbiota mouse model that more closely mimics the human vaginal microbiota. We found several vaginal bacterial taxa that correlated with reduced pathogen levels but showed discordant effects in pathogen inhibition between in vitro and in vivo assays. We propose that this humanized microbiota mouse platform is an improved model to describe the role of the vaginal microbiota in protection against urogenital pathogens. Furthermore, this model will be useful in testing efficacy of new probiotic strategies in the complex vaginal environment.