Monocytes regulate the mechanism of T-cell death by inducing Fas-mediated apoptosis during bacterial infection.

Monocytes and T-cells are critical to the host response to acute bacterial infection but monocytes are primarily viewed as amplifying the inflammatory signal. The mechanisms of cell death regulating T-cell numbers at sites of infection are incompletely characterized. T-cell death in cultures of peripheral blood mononuclear cells (PBMC) showed 'classic' features of apoptosis following exposure to pneumococci. Conversely, purified CD3(+) T-cells cultured with pneumococci demonstrated necrosis with membrane permeabilization. The death of purified CD3(+) T-cells was not inhibited by necrostatin, but required the bacterial toxin pneumolysin. Apoptosis of CD3(+) T-cells in PBMC cultures required 'classical' CD14(+) monocytes, which enhanced T-cell activation. CD3(+) T-cell death was enhanced in HIV-seropositive individuals. Monocyte-mediated CD3(+) T-cell apoptotic death was Fas-dependent both in vitro and in vivo. In the early stages of the T-cell dependent host response to pneumococci reduced Fas ligand mediated T-cell apoptosis was associated with decreased bacterial clearance in the lung and increased bacteremia. In summary monocytes converted pathogen-associated necrosis into Fas-dependent apoptosis and regulated levels of activated T-cells at sites of acute bacterial infection. These changes were associated with enhanced bacterial clearance in the lung and reduced levels of invasive pneumococcal disease

Protective Contributions against Invasive Streptococcus pneumoniae Pneumonia of Antibody and Th17-Cell Responses to Nasopharyngeal Colonisation

The nasopharyngeal commensal bacteria Streptococcus pneumoniae is also a frequent cause of serious infections. Nasopharyngeal colonisation with S. pneumoniae inhibits subsequent re-colonisation by inducing Th17-cell adaptive responses, whereas vaccination prevents invasive infections by inducing antibodies to S. pneumoniae capsular polysaccharides. In contrast, protection against invasive infection after nasopharyngeal colonisation with mutant S. pneumoniae strains was associated with antibody responses to protein antigens. The role of colonisation-induced Th17-cell responses during subsequent invasive infections is unknown. Using mouse models, we show that previous colonisation with S. pneumoniae protects against subsequent lethal pneumonia mainly by preventing bacteraemia with a more modest effect on local control of infection within the lung. Previous colonisation resulted in CD4-dependent increased levels of Th17-cell cytokines during subsequent infectious challenge. However, mice depleted of CD4 cells prior to challenge remained protected against bacteraemia, whereas no protection was seen in antibody deficient mice and similar protection could be achieved through passive transfer of serum. Serum from colonised mice but not antibody deficient mice promoted phagocytosis of S. pneumoniae, and previously colonised mice were able to rapidly clear S. pneumoniae from the blood after intravenous inoculation. Thus, despite priming for a Th17-cell response during subsequent infection, the protective effects of prior colonisation in this model was not dependent on CD4 cells but on rapid clearance of bacteria from the blood by antibody-mediated phagocytosis. These data suggest that whilst nasopharyngeal colonisation induces a range of immune responses, the effective protective responses depend upon the site of subsequent infectio

Beneficial Effects of Prebiotic Saccharomyces cerevisiae

One of the unmet needs for asthma management is a new therapeutic agent with both anti-inflammatory and anti-smooth muscle (ASM) remodeling effects. The mannose receptor (MR) family plays an important role in allergen uptake and processing of major allergens Der p 1 and Fel d 1. We have previously reported that ASM cells express a mannose receptor (ASM-MR) and that mannan derived from Saccharomyces cerevisiae (SC-MN) inhibits mannosyl-rich lysosomal hydrolase-induced bovine ASM cell proliferation. Using a humanized transgenic mouse strain (huASM-MRC2) expressing the human MRC2 receptor in a SM tissue-specific manner, we have demonstrated that ASM hyperplasia/hypertrophy can occur as early as 15 days after allergen challenge in this mouse model and this phenomenon is preventable with SC-MN treatment. This proof-of-concept study would facilitate future development of a potential asthma therapeutic agent with dual function of anti-inflammatory and anti-smooth muscle remodeling effects

A Variable Region within the Genome of Streptococcus pneumoniae Contributes to Strain-Strain Variation in Virulence

The bacterial factors responsible for the variation in invasive potential between different clones and serotypes of Streptococcus pneumoniae are largely unknown. Therefore, the isolation of rare serotype 1 carriage strains in Indigenous Australian communities provided a unique opportunity to compare the genomes of non-invasive and invasive isolates of the same serotype in order to identify such factors. The human virulence status of non-invasive, intermediately virulent and highly virulent serotype 1 isolates was reflected in mice and showed that whilst both human non-invasive and highly virulent isolates were able to colonize the murine nasopharynx equally, only the human highly virulent isolates were able to invade and survive in the murine lungs and blood. Genomic sequencing comparisons between these isolates identified 8 regions >1 kb in size that were specific to only the highly virulent isolates, and included a version of the pneumococcal pathogenicity island 1 variable region (PPI-1v), phage-associated adherence factors, transporters and metabolic enzymes. In particular, a phage-associated endolysin, a putative iron/lead permease and an operon within PPI-1v exhibited niche-specific changes in expression that suggest important roles for these genes in the lungs and blood. Moreover, in vivo competition between pneumococci carrying PPI-1v derivatives representing the two identified versions of the region showed that the version of PPI-1v in the highly virulent isolates was more competitive than the version from the less virulent isolates in the nasopharyngeal tissue, blood and lungs. This study is the first to perform genomic comparisons between serotype 1 isolates with distinct virulence profiles that correlate between mice and humans, and has highlighted the important role that hypervariable genomic loci, such as PPI-1v, play in pneumococcal disease. The findings of this study have important implications for understanding the processes that drive progression from colonization to invasive disease and will help direct the development of novel therapeutic strategies

Contribution of a genomic accessory region encoding a putative cellobiose phosphotransferase system to virulence of Streptococcus Pneumoniae

Streptococcus pneumoniae (the pneumococcus) is a formidable human pathogen, responsible for massive global morbidity and mortality. The ability to utilize carbohydrates in a variety of host niches appears to be integral to pneumococcal pathogenesis. In this study we investigated a genomic island, which includes a ROK family protein, a putative cellobiose phosphotransferase system (PTS) and a putative sulfatase. This accessory region is widespread in the pneumococcus in strains of various serotypes and levels of virulence. We have performed simple bioinformatic analysis of the region and investigated its role in vivo in 2 strains with markedly different virulence profiles (WCH206 of serotype 3, ST180; Menzies5 of serotype 11A, ST662). Deleting and replacing the entire island with an antibiotic resistance cassette caused the virulent serotype 3 strain to become attenuated in a murine pneumonia/sepsis model. Further mutants were constructed and used to show that various components of the island contribute significantly to the fitness of WCH206 in a variety of niches of this model, including the nasopharynx, ears and blood, but especially in the lungs. In addition, the island conferred a competitive advantage in nasopharyngeal colonization for the serotype 11A strain, which was essentially avirulent in the pneumonia/sepsis model. The contribution of this island to both pathogenesis and colonization may explain why this accessory region is widespread in the pneumococcus.Lauren J. McAllister, Abiodun D. Ogunniyi, Uwe H. Stroeher and James C. Pato

Extracellular matrix formation enhances the ability of streptococcus pneumoniae to cause invasive disease

Extent: 17p.During infection, pneumococci exist mainly in sessile biofilms rather than in planktonic form, except during sepsis. However, relatively little is known about how biofilms contribute to pneumococcal pathogenesis. Here, we carried out a biofilm assay on opaque and transparent variants of a clinical serotype 19F strain WCH159. After 4 days incubation, scanning electron microscopy revealed that opaque biofilm bacteria produced an extracellular matrix, whereas the transparent variant did not. The opaque biofilm-derived bacteria translocated from the nasopharynx to the lungs and brain of mice, and showed 100- fold greater in vitro adherence to A549 cells than transparent bacteria. Microarray analysis of planktonic and sessile bacteria from transparent and opaque variants showed differential gene expression in two operons: the lic operon, which is involved in choline uptake, and in the two-component system, ciaRH. Mutants of these genes did not form an extracellular matrix, could not translocate from the nasopharynx to the lungs or the brain, and adhered poorly to A549 cells. We conclude that only the opaque phenotype is able to form extracellular matrix, and that the lic operon and ciaRH contribute to this process. We propose that during infection, extracellular matrix formation enhances the ability of pneumococci to cause invasive disease.Claudia Trappetti, Abiodun D. Ogunniyi, Marco R. Oggioni and James C. Pato

Pneumococcal Virulence Gene Expression and Host Cytokine Profiles during Pathogenesis of Invasive Disease▿ †

Pneumococcal disease continues to account for significant morbidity and mortality worldwide. For the development of novel prophylactic and therapeutic strategies against the disease spectrum, a complete understanding of pneumococcal behavior in vivo is necessary. We evaluated the expression patterns of the proven and putative virulence factor genes adcR, cbpA, cbpD, cbpG, cpsA, nanA, pcpA, piaA, ply, psaA, pspA, and spxB after intranasal infection of CD1 mice with serotype 2, 4, and 6A pneumococci by real-time reverse transcription-PCR. Simultaneous gene expression patterns of selected host immunomodulatory molecules, CCL2, CCL5, CD54, CXCL2, interleukin-6, and tomor necrosis factor alpha, were also investigated. We show that pneumococcal virulence genes are differentially expressed in vivo, with some genes demonstrating niche- and serotype-specific differential expression. The in vivo expression patterns could not be attributed to in vitro differences in expression of the genes in transparent and opaque variants of the three strains. The host molecules were significantly upregulated, especially in the lungs, blood, and brains of mice. The pneumococcal-gene expression patterns support their ascribed roles in pathogenesis, providing insight into which protein combinations might be more appropriate as vaccine antigens against invasive disease. This is the first simultaneous comparison of bacterial- and host gene expression in the same animal during pathogenesis. The strategy provides a platform for prospective evaluation of interaction kinetics between invading pneumococci and human patients in culture-positive cases and should be feasible in other infection models

Understanding Engagement with a Physical Health Service: A Qualitative Study of Patients with Severe Mental Illness

Objective: Individuals with severe mental illness (SMI) are disproportionally affected by medical comorbidities, resulting in poor physical health and premature death. Despite this, care for chronic medical conditions is suboptimal, and there is limited research that explores this phenomenon from the patient’s perspective. The aim of this study was to identify barriers and facilitators of engagement with a physical health service experienced by individuals with SMI. Methods: Adults with SMI were recruited from a large psychiatric hospital and offered referral to a physical health service focused on the prevention and treatment of obesity and diabetes. Interviews were conducted at referral, 3, and 6 months. Data from 56 interviews of 24 participants were analyzed using the framework method to identify factors influencing engagement. Results: Barriers to engagement were identified at individual, medical program, and health system levels. Factors influencing the individual experience included difficulty in care coordination, affective symptomatology, and ability to bond with providers. Factors at the program level included difficulty adjusting to the clinic environment and the inability to achieve treatment goals. Factors at the system level included challenges in attending multiple appointments in a fragmented health system, lack of social support, and financial constraints. Conclusions: This qualitative study suggests that traditional models of medical care for chronic conditions pose challenges for many individuals with SMI and contribute to health disparities. Adaptation of medical care to populations with SMI and close collaboration between medical and mental health services are necessary to improve medical care and, subsequently, health outcomes. </jats:sec

Beneficial Effects of Prebiotic Saccharomyces cerevisiae Mannan on Allergic Asthma Mouse Models

One of the unmet needs for asthma management is a new therapeutic agent with both anti-inflammatory and anti-smooth muscle (ASM) remodeling effects. The mannose receptor (MR) family plays an important role in allergen uptake and processing of major allergens Der p 1 and Fel d 1. We have previously reported that ASM cells express a mannose receptor (ASM-MR) and that mannan derived from Saccharomyces cerevisiae (SC-MN) inhibits mannosyl-rich lysosomal hydrolase-induced bovine ASM cell proliferation. Using a humanized transgenic mouse strain (huASM-MRC2) expressing the human MRC2 receptor in a SM tissue-specific manner, we have demonstrated that ASM hyperplasia/hypertrophy can occur as early as 15 days after allergen challenge in this mouse model and this phenomenon is preventable with SC-MN treatment. This proof-of-concept study would facilitate future development of a potential asthma therapeutic agent with dual function of anti-inflammatory and anti-smooth muscle remodeling effects

Murine Models of Eosinophil Function in Fungal and Viral Infections

Eosinophils are granulocytes that were historically considered to be terminally differentiated at the time of bone marrow egress. However, more recent evidence provides a new outlook on these cells as complex immunomodulators that are involved in host defense and homeostasis. Our work established a role for eosinophils as mediators of antiviral immune responses during influenza in hosts that were sensitized and challenged with fungal allergens. Herein, we describe methods for working with murine eosinophils in the context of influenza A virus