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The isolation of Streptococcus pneumoniae serotypes in systemic tissues of patients with invasive disease versus the nasopharynx of healthy individuals with asymptomatic carriage varies widely. Some serotypes are hyper-invasive, particularly serotype 1, but the underlying genetics remain poorly understood due to the rarity of carriage isolates, reducing the power of comparison with invasive isolates. Here, we use a well-controlled genome-wide association study to search for genetic variation associated with invasiveness of serotype 1 pneumococci from a serotype 1 endemic setting in Africa. We found no consensus evidence that certain genomic variation is overrepresented among isolates from patients with invasive disease than asymptomatic carriage. Overall, the genomic variation explained negligible phenotypic variability, suggesting a minimal effect on the disease status. Furthermore, changes in lineage distribution were seen with lineages replacing each other over time, highlighting the importance of continued pathogen surveillance. Our findings suggest that the hyper-invasiveness is an intrinsic property of the serotype 1 strains, not specific for a "disease-associated" subpopulation disproportionately harboring unique genomic variation. [ABSTRACT FROM AUTHOR]

Porous silicon (pSi) microparticles, in diverse sizes and shapes, can be functionalized to present pathogen-associated molecular patterns that activate dendritic cells. Intraperitoneal injection of MPL-adsorbed pSi microparticles, in contrast to free MPL, resulted in the induction of local inflammation, reflected in the recruitment of neutrophils, eosinophils and proinflammatory monocytes, and the depletion of resident macrophages and mast cells at the injection site. Injection of microparticle-bound MPL resulted in enhanced secretion of the T helper 1 associated cytokines IFN-γ and TNF-α by peritoneal exudate and lymph node cells in response to secondary stimuli while decreasing the anti-inflammatory cytokine IL-10. MPL-pSi microparticles independently exhibited anti-tumor effects and enhanced tumor suppression by low dose doxorubicin nanoliposomes. Intravascular injection of the MPL-bound microparticles increased serum IL-1β levels, which was blocked by the IL-1 receptor antagonist Anakinra. The microparticles also potentiated tumor infiltration by dendritic cells, cytotoxic T lymphocytes, and F4/80+ macrophages, however, a specific reduction was observed in CD204+ macrophages. [ABSTRACT FROM AUTHOR]

Abstract: Ovalbumin (OVA) is known as a major allergen in egg white. A number of studies have reported the partial T and B cell epitope mapping of OVA using murine models and allergic patients'' sera. Recently, we have reported the IgE-binding regions of the entire OVA molecule using egg allergic patients'' sera. However, the entire epitope mapping of OVA in a murine model has not been completed yet. In the present study, BALB/c mice were administered a solution of OVA using three different entry routes (oral, intraperitoneal and subcutaneous) with their respective adjuvant (cholera toxin, aluminum hydroxide and Freund''s adjuvant). Two nitrocellulose membranes containing 188 overlapping synthetic peptides (with a length of 12 amino acids and an offset of two amino acids) covering the primary sequence of OVA, were probed with the three different BALB/c mice antisera. Antisera obtained from orally challenged mice identified eight IgE epitope regions, i.e. I53D60; V77R84; S103E108; G127T136; E275V280; G301F306; I323A332 and A375S384, while sera raised by intraperitoneal and subcutaneous injections exhibited two (K55D60 and K277L282) and five (K55R58; G127T136; K279L282; T303S308 and I323A332) IgE binding sequences, respectively. The residues critical for the epitope–paratope interactions were finely characterized using the oral immunization serum. Analysis of IgE binding epitopes in mice provides us with potential strategies for design of specific immunotherapy. [Copyright &y& Elsevier]

Background: Pneumococcus kills over one million children annually and over 90 % of these deaths occur in low-income countries especially in Sub-Saharan Africa (SSA) where HIV exacerbates the disease burden. In SSA, serotype 1 pneumococci particularly the endemic ST217 clone, causes majority of the pneumococcal disease burden. To understand the evolution of the virulent ST217 clone, we analysed ST217 whole genomes from isolates sampled from African and Asian countries. Methods: We analysed 226 whole genome sequences from the ST217 lineage sampled from 9 African and 4 Asian countries. We constructed a whole genome alignment and used it for phylogenetic and coalescent analyses. We also screened the genomes to determine presence of antibiotic resistance conferring genes. Results: Population structure analysis grouped the ST217 isolates into five sequence clusters (SCs), which were highly associated with different geographical regions and showed limited intracontinental and intercontinental spread. The SCs showed lower than expected genomic sequence, which suggested strong purifying selection and small population sizes caused by bottlenecks. Recombination rates varied between the SCs but were lower than in other successful clones such as PMEN1. African isolates showed higher prevalence of antibiotic resistance genes than Asian isolates. Interestingly, certain West African isolates harbored a defective chloramphenicol and tetracycline resistance-conferring element (Tn5253) with a deletion in the loci encoding the chloramphenicol resistance gene (cat pC194), which caused lower chloramphenicol than tetracycline resistance. Furthermore, certain genes that promote colonisation were absent in the isolates, which may contribute to serotype 1’s rarity in carriage and consequently its lower recombination rates. Conclusions: The high phylogeographic diversity of the ST217 clone shows that this clone has been in circulation globally for a long time, which allowed its diversification and adaptation in different geographical regions. Such geographic adaptation reflects local variations in selection pressures in different locales. Further studies will be required to fully understand the biological mechanisms which makes the ST217 clone highly invasive but unable to successfully colonise the human nasopharynx for long durations which results in lower recombination rates. Electronic supplementary material The online version of this article (doi:10.1186/s12879-016-1987-z) contains supplementary material, which is available to authorized users.

Streptococcus pneumoniae causes a high burden of invasive pneumococcal disease (IPD) globally, especially in children from resource-poor settings. Like many bacteria, the pneumococcus can import DNA from other strains or even species by transformation and homologous recombination, which has allowed the pneumococcus to evade clinical interventions such as antibiotics and pneumococcal conjugate vaccines (PCVs). Pneumococci are enclosed in a complex polysaccharide capsule that determines the serotype; the capsule varies in size and is associated with properties including carriage prevalence and virulence. We determined and quantified the association between capsule and recombination events using genomic data from a diverse collection of serotypes sampled in Malawi. We determined both the amount of variation introduced by recombination relative to mutation (the relative rate) and how many individual recombination events occur per isolate (the frequency). Using univariate analyses, we found an association between both recombination measures and multiple factors associated with the capsule, including duration and prevalence of carriage. Because many capsular factors are correlated, we used multivariate analysis to correct for collinearity. Capsule size and carriage duration remained positively associated with recombination, although with a reduced P value, and this effect may be mediated through some unassayed additional property associated with larger capsules. This work describes an important impact of serotype on recombination that has been previously overlooked. While the details of how this effect is achieved remain to be determined, it may have important consequences for the serotype-specific response to vaccines and other interventions.

Mother’s milk is widely accepted as nutritious and protective to the newborn mammals by providing not only macronutrients but also immune-defensive factors. However, the mechanisms accounting for these benefits are not fully understood. Here we show that maternal very-low-density-lipoprotein receptor (VLDLR) deletion in mice causes the production of defective milk containing diminished level of platelet-activating factor acetylhydrolase (PAFAH). As a consequence, the nursing neonates suffer from alopecia, anemia and growth retardation owing to elevated levels of pro-inflammatory platelet-activating factors (PAFs). VLDLR deletion significantly impairs the expression of phospholipase A2 group 7 (Pla2g7) in macrophages, which decreases PAFAH secretion. Exogenous oral supplementation of neonates with PAFAH effectively rescues the toxicity. These findings not only reveal a novel role of VLDLR in suppressing inflammation by maintaining macrophage PAFAH secretion, but also identify the maternal VLDLR as a key genetic program that ensures milk quality and protects the newborns., Chemistry and Chemical Biology

The entry routes and translocation mechanisms of microorganisms or particulate materials into the central nervous system remain obscure We report here that Streptococcus pneumoniae (pneumococcus), or polystyrene microspheres of similar size, appear in the meninges of the dorsal cortex of mice within minutes of inhaled delivery. Recovery of viable bacteria from dissected tissue and fluorescence microscopy show that up to at least 72 h, pneumococci and microspheres were predominantly found in the outer of the two meninges: the pachymeninx. No pneumococci were found in blood or cerebrospinal fluid. Intravital imaging through the skull, aligned with flow cytometry showed recruitment and activation of LysM + cells in the dorsal pachymeninx at 5 and 10 hours following intranasal infection. Imaging of the cribriform plate suggested that both pneumococci and microspheres entered through the foramina via an inward flow of fluid connecting the nose to the pachymeninx. Our findings bring new insight into the varied mechanisms of pneumococcal invasion of the central nervous system, but they are also pertinent to the delivery of drugs to the brain and the entry of airborne particulate matter into the cranium. IMPORTANCE Using two-photon imaging, we show that pneumococci translocate from the nasopharynx to the dorsal meninges of a mouse in the absence of any bacteria found in blood or cerebrospinal fluid. Strikingly, this takes place within minutes of inhaled delivery of pneumococci, suggesting the existence of an inward flow of fluid connecting the nasopharynx to the meninges, rather than a receptor-mediated mechanism. We also show that this process is size dependent, as microspheres of the same size as pneumococci can translocate along the same pathway, while larger size microspheres cannot. Furthermore, we describe the host response to invasion of the outer meninges. Our study provides a completely new insight into the key initial events that occur during the translocation of pneumococci directly from the nasal cavity to the meninges, with relevance to the development of intranasal drug delivery systems and the investigations of brain damage caused by inhaled air pollutants.

Streptococcus pneumoniae is a frequent colonizer of the human nasopharynx and a major cause of life-threating invasive infections such as pneumonia, meningitis and sepsis. Over 1 million people die every year due to invasive pneumococcal disease (IPD), mainly in developing countries. Serotype 1 is a common cause of IPD; however, unlike other serotypes, it is rarely found in the carrier state in the nasopharynx, which is often considered a prerequisite for disease. The aim of this study was to understand this dichotomy. We used murine models of carriage and IPD to characterize the pathogenesis of African serotype 1 (sequence type 217) pneumococcal strains obtained from the Queen Elizabeth Central Hospital in Blantyre, Malawi. We found that ST217 pneumococcal strains were highly virulent in a mouse model of invasive pneumonia, but in contrast to the generally accepted assumption, can also successfully establish nasopharyngeal carriage. Interestingly, we found that cocolonizing serotypes may proliferate in the presence of serotype 1, suggesting that acquisition of serotype 1 carriage could increase the risk of developing IPD by other serotypes. RNA sequencing analysis confirmed that key virulence genes associated with inflammation and tissue invasiveness were upregulated in serotype 1. These data reveal important new insights into serotype 1 pathogenesis, with implications for carriage potential and risk of invasive disease through interactions with other cocolonizing serotypes, an often-overlooked factor in transmission and disease progression. IMPORTANCE The pneumococcus causes serious diseases such as pneumonia, sepsis, and meningitis and is a major cause of morbidity and mortality worldwide. Serotype 1 accounts for the majority of invasive pneumococcal disease cases in sub-Saharan Africa but is rarely found during nasopharyngeal carriage. Understanding the mechanisms leading to nasopharyngeal carriage and invasive disease by this serotype can help reduce its burden on health care systems worldwide. In this study, we also uncovered the potential impact of serotype 1 on disease progression of other coinfecting serotypes, which can have important implications for vaccine efficacy. Understanding the interactions between different serotypes during nasopharyngeal carriage may lead to improved intervention methods and therapies to reduce pneumococcal invasive disease levels., (Copyright © 2020 Bricio-Moreno et al.)

The ability of particulate materials to provoke inflammatory immune responses has been well documented. In the case of endogenous and environmental particulates, these effects can often lead to pathological disorders. In contrast, particulate adjuvants incorporated into vaccines promote immune responses, which in turn provide efficient protection against infectious diseases. In recent years, studies have revealed that the NLRP3 inflammasome plays a key role in particulate-driven inflammation and its associated cytotoxicity. Hence, this chapter covers protocols useful to (1) assess NLRP3 inflammasome activation triggered by particulate adjuvants or materials in mouse bone marrow-derived dendritic cell (BMDCs) differentiated cultures, and (2) measure particle-induced cytotoxicity. More specifically, protocols are described for the preparation and differentiation of BMDCs, their priming and stimulation using particulate NLRP3 agonists such as monosodium urate monohydrate (MSU) and the vaccine adjuvant alum. We then detail protocols to assess particulate-driven cytotoxicity via flow cytometry using annexin V-propidium iodide (PI) and novel dye LIVE/DEAD(®) aqua stain. General considerations are provided that warn against the use of endotoxin-contaminated particles and emphasize the use of experimental controls. Suggestions are also outlined for further assessment of the immunomodulatory effects of particulate materials in vivo using the mouse peritonitis model.

For all newborn mammals, mother's milk is the perfect nourishment, crucial for their postnatal development. Here we report that, unexpectedly, maternal western diet consumption in mice causes the production of toxic milk that contains excessive long chain and saturated fatty acids, which triggers ceramide accumulation and inflammation in the nursing neonates, manifested as alopecia. This neonatal toxicity requires Toll-like-receptors (TLR), but not gut microbiota, because TLR2/4 deletion or TLR4 inhibition confers resistance, whereas germ-free mice remain sensitive. These findings unravel maternal western diet-induced inflammatory milk secretion as a novel aspect of the metabolic syndrome at the maternal offspring interface.

Egg has ben documented as a rich source for the supply of biologically active peptides. This study characterizes the immunomodulatory effects of an egg white enzymatic hydrolysate (EWH) using a BALB/c mouse model of egg allergy. Mice were orally sensitized to egg white and subsequently gavaged with EWH. ELISA results indicated significant reductions of both serum histamine and specific IgE titers in EWH-fed mice, accompanied by a repression of both IL-4 and IFN-gamma production in spleen cell cultures. Similarly, real-time RT-PCR analyses highlighted decreased mRNA expression of IFN-gamma and IL-12 (Th1-biased), as well as lower ratios of IL-4 and IL-13 mRNA (Th2-biased). On the other hand, increased intestinal expressions of TGF-beta and FOXp3 mRNA were determined in EWH-fed mice, suggesting induction of local regulatory mechanisms. The presence of immunodominant epitopes was proposed to be responsible for the immunomodulatory effects observed.

The emergence of egg allergy has had both industrial and clinical implications. In industrialized countries, egg allergy accounts for one of the most prevalent food hypersensitivities, especially in children. Atopic dermatitis represents the most common clinical manifestation in infancy; however, the range of clinical signs is broad and encompasses life-threatening anaphylaxis. The dominant egg allergens are proteins and are mainly present in the egg white, for example, ovalbumin, ovomucoid, ovotransferrin, and lysozyme. However, egg yolk also displays low-level allergenicity, for example, alpha-livetin. Strict avoidance of the offending food remains the most common recommendation for egg-allergic individuals. Nevertheless, the omnipresence of egg-derived components in prepackaged or prepared foods makes it difficult. Therefore, more efficient preventive approaches are investigated to protect consumers from inadvertent exposure and ensuing adverse reactions. On the one hand, commercial kits have become readily available that allow for the detection of egg contaminants at trace levels. On the other hand, attempts to produce hypoallergenic egg-containing products through food-processing techniques have met with promising results, but the approach is limited due to its potentially undesirable effects on the unique functional and sensory attributes of egg proteins. Therefore, the development of preventive or curative strategies for egg allergy remains strongly warranted. Pilot studies have suggested that oral immunotherapy (IT) with raw or cooked preparations of egg may represent a safe alternative, immediately available to allergic subjects, but remains applicable to only nonanaphylactic patients. Due to the limitations of conventional IT, novel forms of immunotherapy are sought based on information obtained from the molecular characterization of major egg allergens. In the past decade, promising approaches to the treatment and prevention of egg allergy have been explored and include, among others, the production of hypoallergenic recombinant egg proteins, the development of customized peptides, and bacterial-mediated immunotherapy. Nonspecific approaches have also been evaluated, and preliminary trials with the use of probiotic bacteria have yielded encouraging results. The current understanding of egg allergens offers novel approaches toward the making of food products safe for human consumption and the development of efficient immunotherapeutic strategies.