Cytokine biomarkers of exacerbations in sputum from Chronic Obstructive Pulmonary Disease patients: a prospective cohort study.

BACKGROUND: We determined the relationships between cytokine expression in sputum and clinical data to characterise and understand Chronic Obstructive Pulmonary Disease (COPD) exacerbations in COPD patients. METHODS: We measured 30 cytokines in 936 sputum samples, collected at stable state (ST) and exacerbation (EX) visits from 99 participants in the Acute Exacerbation and Respiratory InfectionS in COPD (AERIS) study (NCT01360398, www.clinicaltrials.gov). We determined their longitudinal expression and examined differential expression based on disease status or exacerbation type. RESULTS: Of the cytokines, 17 were suitable for analysis. As for disease states, in EX sputum samples, IL-17A, TNF-α, IL-1ß, and IL-10 were significantly increased compared to ST sputum samples, but a logistic mixed model could not predict disease state. As for exacerbation types, bacteria-associated exacerbations showed higher expression of IL-17A, TNF-α, IL-1ß, and IL-1α. IL-1α, IL-1ß, and TNF-α were identified as suitable biomarkers for bacteria-associated exacerbation. Bacteria-associated exacerbations also formed a cluster separate from other exacerbation types in principal component analysis. CONCLUSIONS: Measurement of cytokines in sputum from COPD patients could help identify bacteria-associated exacerbations based on increased concentrations of IL-1α, IL-1ß, or TNF-α. This finding may provide a point-of-care assessment to distinguish a bacterial exacerbation of COPD from other exacerbation types.

Neisseria meningitidis is structured in clades associated with restriction modification systems that modulate homologous recombination.

Molecular data on a limited number of chromosomal loci have shown that the population of Neisseria meningitidis (Nm), a deadly human pathogen, is structured in distinct lineages. Given that the Nm population undergoes substantial recombination, the mechanisms resulting in the evolution of these lineages, their persistence in time, and the implications for the pathogenicity of the bacterium are not yet completely understood. Based on whole-genome sequencing, we show that Nm is structured in phylogenetic clades. Through acquisition of specific genes and through insertions and rearrangements, each clade has acquired and remodeled specific genomic tracts, with the potential to impact on the commensal and virulence behavior of Nm. Despite this clear evidence of a structured population, we confirm high rates of detectable recombination throughout the whole Nm chromosome. However, gene conversion events were found to be longer within clades than between clades, suggesting a DNA cleavage mechanism associated with the phylogeny of the species. We identify 22 restriction modification systems, probably acquired by horizontal gene transfer from outside of the species/genus, whose distribution in the different strains coincides with the phylogenetic clade structure. We provide evidence that these clade-associated restriction modification systems generate a differential barrier to DNA exchange consistent with the observed population structure. These findings have general implications for the emergence of lineage structure and virulence in recombining bacterial populations, and they could provide an evolutionary framework for the population biology of a number of other bacterial species that show contradictory population structure and dynamics.

Computational modeling of microfluidic data provides high-throughput affinity estimates for monoclonal antibodies.

Affinity measurement is a fundamental step in the discovery of monoclonal antibodies (mAbs) and of antigens suitable for vaccine development. Innovative affinity assays are needed due to the low throughput and/or limited dynamic range of available technologies. We combined microfluidic technology with quantum-mechanical scattering theory, in order to develop a high-throughput, broad-range methodology to measure affinity. Fluorescence intensity profiles were generated for out-of-equilibrium solutions of labelled mAbs and their antigen-binding fragments migrating along micro-columns with immobilized cognate antigen. Affinity quantification was performed by computational data analysis based on the Landau probability distribution. Experiments using a wide array of human or murine antibodies against bacterial or viral, protein or polysaccharide antigens, showed that all the antibody-antigen capture profiles (n = 841) generated at different concentrations were accurately described by the Landau distribution. A scale parameter W, proportional to the full-width-at-half-maximum of the capture profile, was shown to be independent of the antibody concentration. The W parameter correlated significantly (Pearson's r [p-value]: 0.89 [3 × 10-8]) with the equilibrium dissociation constant KD, a gold-standard affinity measure. Our method showed good intermediate precision (median coefficient of variation: 5%) and a dynamic range corresponding to KD values spanning from ~10-7 to ~10-11 Molar. Relative to assays relying on antibody-antigen equilibrium in solution, even when they are microfluidic-based, the method's turnaround times were decreased from 2 days to 2 h. The described computational modelling of antibody capture profiles represents a fast, reproducible, high-throughput methodology to accurately measure a broad range of antibody affinities in very low volumes of solution.

Dissecting the Human Response to Staphylococcus aureus Systemic Infections.

Staphylococcus aureus is a common human commensal and the leading cause of diverse infections. To identify distinctive parameters associated with infection and colonization, we compared the immune and inflammatory responses of patients with a diagnosis of invasive S. aureus disease to healthy donors. We analyzed the inflammatory responses founding a pattern of distinctive cytokines significantly higher in the patients with invasive disease. The measure of antibody levels revealed a wide antibody responsiveness from all subjects to most of the antigens, with significantly higher response for some antigens in the invasive patients compared to control. Moreover, functional antibodies against toxins distinctively associated with the invasive disease. Finally, we examined the genomic variability of isolates, showing no major differences in genetic distribution compared to a panel of representative strains. Overall, our study shows specific signatures of cytokines and functional antibodies in patients with different primary invasive diseases caused by S. aureus. These data provide insight into human responses towards invasive staphylococcal infections and are important for guiding the identification of novel preventive and therapeutic interventions against S. aureus.

Antibody avidity, persistence, and response to antigen recall: comparison of vaccine adjuvants.

Differences in innate immune 'imprinting' between vaccine adjuvants may mediate dissimilar effects on the quantity/quality of persisting adaptive responses. We compared antibody avidity maturation, antibody/memory B cell/CD4+ T cell response durability, and recall responses to non-adjuvanted fractional-dose antigen administered 1-year post-immunization (Day [D]360), between hepatitis B vaccines containing Adjuvant System (AS)01B, AS01E, AS03, AS04, or Alum (NCT00805389). Both the antibody and B cell levels ranked similarly (AS01B/E/AS03 > AS04 > Alum) at peak response, at D360, and following their increases post-antigen recall (D390). Proportions of high-avidity antibodies increased post-dose 2 across all groups and persisted at D360, but avidity maturation appeared to be more strongly promoted by AS vs. Alum. Post-antigen recall, frequencies of subjects with high-avidity antibodies increased only markedly in the AS groups. Among the AS, total antibody responses were lowest for AS04. However, proportions of high-avidity antibodies were similar between groups, suggesting that MPL in AS04 contributes to avidity maturation. Specific combinations of immunoenhancers in the AS, regardless of their individual nature, increase antibody persistence and avidity maturation.

4CMenB Immunization Induces Serum Bactericidal Antibodies Against Non-Serogroup B Meningococcal Strains in Adolescents.

INTRODUCTION: Invasive meningococcal disease (IMD) is an important public health concern. In developed countries, most IMD is caused by meningococcal serogroup B (MenB) and two protein-based MenB vaccines are currently available: the four-component vaccine 4CMenB (Bexsero, GSK) and the bivalent vaccine MenB-FHbp (Trumenba, Pfizer). Genes encoding the 4CMenB vaccine antigens are also present in strains belonging to other meningococcal serogroups. METHODS: To evaluate the potential of 4CMenB vaccination to protect adolescents against non-MenB IMD, we tested the bactericidal activity of sera from immunized adolescents on 147 (127 European and 20 Brazilian) non-MenB IMD isolates, with a serum bactericidal antibody assay using human complement (hSBA). Serum pools were prepared using samples from randomly selected participants in various clinical trials, pre- and post-vaccination: 12 adolescents who received two doses of 4CMenB 2 months apart, and 10 adolescents who received a single dose of a MenACWY conjugate vaccine (as positive control). RESULTS: 4CMenB pre-immune sera killed 7.5% of the 147 non-MenB isolates at hSBA titers ≥ 1:4. In total, 91 (61.9%) tested isolates were killed by post-dose 2 pooled sera at hSBA titers ≥ 1:4, corresponding to 44/80 (55.0%) MenC, 26/35 (74.3%) MenW, and 21/32 (65.6%) MenY isolates killed. CONCLUSION: 4CMenB vaccination in adolescents induces bactericidal killing of non-MenB isolates, suggesting that mass vaccination could impact IMD due to serogroups other than MenB.

Multicomponent meningococcal serogroup B vaccination elicits cross-reactive immunity in infants against genetically diverse serogroup C, W and Y invasive disease isolates.

BACKGROUND: The multicomponent meningococcal serogroup B vaccine (4CMenB) is currently indicated for active immunization against invasive meningococcal disease caused by Neisseria meningitidis serogroup B (MenB). However, genes encoding the 4CMenB antigens are also variably present and expressed in strains belonging to other meningococcal serogroups. In this study, we evaluated the ability of antibodies raised by 4CMenB immunisation to induce complement-mediated bactericidal killing of non-MenB strains. METHODS: A total of 227 invasive non-MenB disease isolates were collected between 1 July 2007 and 30 June 2008 from England and Wales, France, and Germany; 41 isolates were collected during 2012 from Brazil. The isolates were subjected to genotypic analyses. A subset of 147 isolates (MenC, MenW and MenY) representative of the meningococcal genetic diversity of the total sample were tested in the human complement serum bactericidal antibody assay (hSBA) using sera from infants immunised with 4CMenB. RESULTS: Serogroup and clonal complex repertoires of non-MenB isolates were different for each country. For the European panel, MenC, MenW and MenY isolates belonged mainly to ST-11, ST-22 and ST-23 complexes, respectively. For the Brazilian panel, most MenC and MenW isolates belonged to the ST-103 and ST-11 complexes, respectively, and most MenY isolates were not assigned to clonal complexes. Of the 147 non-MenB isolates, 109 were killed in hSBA, resulting in an overall coverage of 74%. CONCLUSION: This is the first study in which 147 non-MenB serogroup isolates have been analysed in hSBA to evaluate the potential of a MenB vaccine to cover strains belonging to other serogroups. These data demonstrate that antibodies raised by 4CMenB are able to induce bactericidal killing of 109 non-MenB isolates, representative of non-MenB genetic and geographic diversity. These findings support previous evidence that 4CMenB immunisation can provide cross-protection against non-MenB strains in infants, which represents an added benefit of 4CMenB vaccination.

Pooled-sera hSBA titres predict individual seroprotection in infants and toddlers vaccinated with 4CMenB.

UNLABELLED: The Serum Bactericidal Antibody assay with human complement (hSBA) using individual immune sera is a surrogate of protection for meningococcal vaccines. Strain coverage of 4CMenB, a licensed vaccine against serogroup B meningococcal (MenB) disease, has been extensively assessed in hSBA using pooled sera, directly or through the Meningococcal Antigen Typing System (MATS). The extent to which pooled-sera hSBA titres reflect individual protection is not yet fully understood. We analysed more than 17000 individual hSBA titres from infants and toddlers vaccinated with 4CMenB, pooled-serum hSBA titres from subsets therein and MATS data from a 40 strain panel representative of invasive MenB disease in England and Wales. Individual hSBA titres segregated in two normal distributions, respectively from responding and non-responding subjects (fit_model-data: r=0.996, p-values <0.05). No individual subject showed abnormally high titres compared to the distributions. Also, when sera from the same subjects were tested individually and in pool, pooled-sera titre and average of individual titres from the same group were substantially indistinguishable (r=0.97, p-value <<0.001). We identified a robust mathematical relationship between the mean of individual hSBA titres and the proportion of subjects achieving a protective titre (seroprotection rate, r=0.95, p-value <<0.001). Using this relation, the seroprotection rate in 15 groups of vaccinees tested against 11 diverse meningococcal isolates was accurately predicted by the hSBA titre of the respective pooled sera (average prediction error 9%). Finally, strains defined covered by MATS had on average 77% predicted seroprotection rate (interquartile range, IQR: 66-100%) and 39% for non-covered strains (IQR: 19-46%). We conclude that seroprotection rates in infants and toddlers vaccinated with 4CMenB can be accurately predicted by pooled-serum hSBA, and that strain coverage defined by MATS is associated with high seroprotection rates. SUMMARY: The Serum Bactericidal Antibody assay (SBA) from individual sera is a surrogate of protection for meningococcal vaccines. We show that SBA performed on pooled sera predicts individual protection.

Building an insurance against modern pandemics.

The response to the first influenza pandemic of the 21st century benefited from the extensive preparation for an avian influenza pandemic and the mild nature of the 2009 A/H1N1 swine influenza virus. However, the pandemic demonstrated the limited ability to predict influenza pandemics, to anticipate levels of cross-protection, and to deliver vaccines in a timely manner, particularly to low income countries. The lessons learned from the 2009 H1N1 pandemic are of paramount importance to develop more effective preparations against future pandemics.