Prior exposure to B. pertussis shapes the mucosal antibody response to acellular pertussis booster vaccination.

Bordetella pertussis (Bp), the causative agent of pertussis, continues to circulate despite widespread vaccination programs. An important question is whether and how (sub)clinical infections shape immune memory to Bp, particularly in populations primed with acellular pertussis vaccines (aP). Here, we examine the prevalence of mucosal antibodies against non-vaccine antigens in aP-primed children and adolescents of the BERT study (NCT03697798), using antibody binding to a Bp mutant strain lacking aP antigens (Bp_mut). Our study identifies increased levels of mucosal IgG and IgA binding to Bp_mut in older aP-primed individuals, suggesting different Bp exposure between aP-primed birth cohorts, in line with pertussis disease incidence data. To examine whether Bp exposure influences vaccination responses, we measured mucosal antibody responses to aP booster vaccination as a secondary study outcome. Although booster vaccination induces significant increases in mucosal antibodies to Bp in both cohorts, the older age group that had higher baseline antibodies to Bp_ mut shows increased persistence of antibodies after vaccination.

BCG-induced trained immunity enhances acellular pertussis vaccination responses in an explorative randomized clinical trial.

Acellular pertussis (aP) booster vaccines are central to pertussis immunization programs, although their effectiveness varies. The Bacille Calmette-Guérin (BCG) vaccine is a prototype inducer of trained immunity, which enhances immune responses to subsequent infections or vaccinations. While previous clinical studies have demonstrated that trained immunity can protect against heterologous infections, its effect on aP vaccines in humans is unknown. We conducted a clinical study in order to determine the immunological effects of trained immunity on pertussis vaccination. Healthy female volunteers were randomly assigned to either receive BCG followed by a booster dose of tetanus-diphteria-pertussis inactivated polio vaccine (Tdap-IPV) 3 months later (BCG-trained), BCG + Tdap-IPV concurrently, or Tdap-IPV followed by BCG 3 months later. Primary outcomes were pertussis-specific humoral, T- and B-cell responses and were quantified at baseline of Tdap-IPV vaccination and 2 weeks thereafter. As a secondary outcome in the BCG-trained cohort, ex vivo leukocyte responses were measured in response to unrelated stimuli before and after BCG vaccination. BCG vaccination 3 months prior to, but not concurrent with, Tdap-IPV improves pertussis-specific Th1-cell and humoral responses, and also increases total memory B cell responses. These responses were correlated with enhanced IL-6 and IL-1ß production at the baseline of Tdap-IPV vaccination in the BCG-trained cohort. Our study demonstrates that prior BCG vaccination potentiates immune responses to pertussis vaccines and that biomarkers of trained immunity are the most reliable correlates of those responses.

Coffee induces AHR- and Nrf2-mediated transcription in intestinal epithelial cells.

Coffee induces a health-promoting adaptive response of cells in the body. Here, we investigated enterocyte responses to AHR agonists in coffee and measured their transport across a polarized intestinal epithelium. AHR-activating potencies of Turkish, filter, and instant coffee were determined using DR CALUX® bioassay, before and after intestinal metabolization by Caco-2 cells. Furthermore, effects of coffee on induction of AHR- and Nrf2-pathway genes in Caco-2 cells were evaluated by real-time qPCR. Coffee samples showed considerable AHR-activating potencies in DR CALUX® bioassay (up to 79% of positive control activity). After incubation with Caco-2 cells, AHR activity of different coffees was between 35 and 64% of their initial value, suggesting rapid uptake and metabolization by epithelial cells. Expression of AHR-regulated gene CYP1A1 increased up to 41-fold and most Nrf2-pathway genes were up-regulated by coffee. This in vitro study may support the notion that coffee bioactives contribute to antioxidant defense and detoxification processes in vivo.

A Prioritized and Validated Resource of Mitochondrial Proteins in Plasmodium Identifies Unique Biology.

Plasmodium species have a single mitochondrion that is essential for their survival and has been successfully targeted by antimalarial drugs. Most mitochondrial proteins are imported into this organelle, and our picture of the Plasmodium mitochondrial proteome remains incomplete. Many data sources contain information about mitochondrial localization, including proteome and gene expression profiles, orthology to mitochondrial proteins from other species, coevolutionary relationships, and amino acid sequences, each with different coverage and reliability. To obtain a comprehensive, prioritized list of Plasmodium falciparum mitochondrial proteins, we rigorously analyzed and integrated eight data sets using Bayesian statistics into a predictive score per protein for mitochondrial localization. At a corrected false discovery rate of 25%, we identified 445 proteins with a sensitivity of 87% and a specificity of 97%. They include proteins that have not been identified as mitochondrial in other eukaryotes but have characterized homologs in bacteria that are involved in metabolism or translation. Mitochondrial localization of seven Plasmodium berghei orthologs was confirmed by epitope labeling and colocalization with a mitochondrial marker protein. One of these belongs to a newly identified apicomplexan mitochondrial protein family that in P. falciparum has four members. With the experimentally validated mitochondrial proteins and the complete ranked P. falciparum proteome, which we have named PlasmoMitoCarta, we present a resource to study unique proteins of Plasmodium mitochondria. IMPORTANCE The unique biology and medical relevance of the mitochondrion of the malaria parasite Plasmodium falciparum have made it the subject of many studies. However, we actually do not have a comprehensive assessment of which proteins reside in this organelle. Many omics data are available that are predictive of mitochondrial localization, such as proteomics data and expression data. Individual data sets are, however, rarely complete and can provide conflicting evidence. We integrated a wide variety of available omics data in a manner that exploits the relative strengths of the data sets. Our analysis gave a predictive score for the mitochondrial localization to each nuclear encoded P. falciparum protein and identified 445 likely mitochondrial proteins. We experimentally validated the mitochondrial localization of seven of the new mitochondrial proteins, confirming the quality of the complete list. These include proteins that have not been observed mitochondria before, adding unique mitochondrial functions to P. falciparum.

Uncovering Distinct Primary Vaccination-Dependent Profiles in Human Bordetella Pertussis Specific CD4+ T-Cell Responses Using a Novel Whole Blood Assay.

To advance research and development of improved pertussis vaccines, new immunoassays are needed to qualify the outcome of Bordetella pertussis (Bp) specific CD4+ T-cell differentiation. Here, we applied a recently developed whole blood assay to evaluate Bp specific CD4+ T-cell responses. The assay is based on intracellular cytokine detection after overnight in vitro Bp antigen stimulation of diluted whole blood. We show for the first time that CD4+ T-cell memory of Th1, Th2, and Th17 lineages can be identified simultaneously in whole blood. Participants ranging from 7 to 70 years of age with different priming backgrounds of whole-cell pertussis (wP) and acellular pertussis (aP) vaccination were analyzed around an acellular booster vaccination. The assay allowed detection of low frequent antigen-specific CD4+ T-cells and revealed significantly elevated numbers of activated and cytokine-producing CD4+ T-cells, with a significant tendency to segregate recall responses based on primary vaccination background. A stronger Th2 response hallmarked an aP primed cohort compared to a wP primed cohort. In conclusion, analysis of Bp specific CD4+ T-cell responses in whole blood showed separation based on vaccination background and provides a promising tool to assess the quantity and quality of CD4+ T-cell responses induced by vaccine candidates.

Use of Microarray Datasets to generate Caco-2-dedicated Networks and to identify Reporter Genes of Specific Pathway Activity.

Intestinal epithelial cells, like Caco-2, are commonly used to study the interaction between food, other luminal factors and the host, often supported by microarray analysis to study the changes in gene expression as a result of the exposure. However, no compiled dataset for Caco-2 has ever been initiated and Caco-2-dedicated gene expression networks are barely available. Here, 341 Caco-2-specific microarray samples were collected from public databases and from in-house experiments pertaining to Caco-2 cells exposed to pathogens, probiotics and several food compounds. Using these datasets, a gene functional association network specific for Caco-2 was generated containing 8937 nodes 129711 edges. Two in silico methods, a modified version of biclustering and the new Differential Expression Correlation Analysis, were developed to identify Caco-2-specific gene targets within a pathway of interest. These methods were subsequently applied to the AhR and Nrf2 signalling pathways and altered expression of the predicted target genes was validated by qPCR in Caco-2 cells exposed to coffee extracts, known to activate both AhR and Nrf2 pathways. The datasets and in silico method(s) to identify and predict responsive target genes can be used to more efficiently design experiments to study Caco-2/intestinal epithelial-relevant biological processes.