Lymphoid-Tissue-Resident Commensal Bacteria Promote Members of the IL-10 Cytokine Family to Establish Mutualism.

Physical separation between the mammalian immune system and commensal bacteria is necessary to limit chronic inflammation. However, selective species of commensal bacteria can reside within intestinal lymphoid tissues of healthy mammals. Here, we demonstrate that lymphoid-tissue-resident commensal bacteria (LRC) colonized murine dendritic cells and modulated their cytokine production. In germ-free and antibiotic-treated mice, LRCs colonized intestinal lymphoid tissues and induced multiple members of the IL-10 cytokine family, including dendritic-cell-derived IL-10 and group 3 innate lymphoid cell (ILC3)-derived IL-22. Notably, IL-10 limited the development of pro-inflammatory Th17 cell responses, and IL-22 production enhanced LRC colonization in the steady state. Furthermore, LRC colonization protected mice from lethal intestinal damage in an IL-10-IL-10R-dependent manner. Collectively, our data reveal a unique host-commensal-bacteria dialog whereby selective subsets of commensal bacteria interact with dendritic cells to facilitate tissue-specific responses that are mutually beneficial for both the host and the microbe.

Emergence of Bordetella holmesii as a Causative Agent of Whooping Cough, Barcelona, Spain.

We describe the detection of Bordetella holmesii as a cause of whooping cough in Spain. Prevalence was 3.9% in 2015, doubling to 8.8% in 2016. This emergence raises concern regarding the contribution of B. holmesii to the reemergence of whooping cough and the effectiveness of the pertussis vaccine.

Glycomacropeptide administration attenuates airway inflammation and remodeling associated to allergic asthma in rat.

OBJECTIVE: Glycomacropeptide (GMP) is a bioactive peptide derived from milk that has been reported to exhibit a range of anti-inflammatory and immunomodulatory properties. The aim of this study was to analyze the prophylactic effect of GMP administration on airway inflammation and remodeling in an experimental model of asthmatic rat. METHODS: Animals treated orally with or without GMP (500 mg/kg/day) were ovalbumin-sensitized and -nebulized and several indicators of Th2 response, airway structural changes and inflammatory cells recruitment were evaluated. RESULTS: Treatment with GMP prior and during asthma development resulted in reduction of allergen-specific IgE titers in serum and blood eosinophilia. Also, GMP substantially suppressed the recruitment of inflammatory cells to bronchoalveolar compartment. Histological studies demonstrated that GMP markedly inhibits eosinophils infiltration, goblet cells hyperplasia and collagen deposit in lung tissue. The latter effect was related with an inhibition in transforming growth factor-ß expression. In addition, expression of interleukin-5 and -13 were substantially inhibited in lung while that of interleukin-10 was increased. CONCLUSION: Our results suggest that administration of GMP may prevent the development of an excessive Th2 response in asthma and effectively ameliorates the progression of the disease.

Development of vaccines against pertussis caused by Bordetella holmesii using a mouse intranasal challenge model.

Bordetella holmesii is recognized as the third causative agent of pertussis (whooping cough) in addition to Bordetella pertussis and Bordetella parapertussis. Pertussis caused by B. holmesii is not rare around the world. However, to date, there is no effective vaccine against B. holmesii. We examined the protective potency of pertussis vaccines available in Japan and vaccines prepared from B. holmesii. A murine model of respiratory infection was exploited to evaluate protective potency. No Japanese commercial pertussis vaccines were effective against B. holmesii. In contrast, a wBH vaccine and an aBH vaccine prepared from B. holmesii were both protective. Passive immunization with sera from mice immunized with aBH vaccine established protection against B. holmesii, indicating that B. holmesii-specific serum antibodies might play an important role in protection. Immuno-proteomic analysis with sera from mice immunized with aBH vaccine revealed that the sera recognized a BipA-like protein of B. holmesii. An aBH vaccine prepared from a BipA-like protein-deficient mutant strain did not have a protective effect against B. holmesii. Taken together, our results suggest that the BipA-like protein plays an important role in the protective efficacy of aBH vaccine.

Horizontally acquired divergent O-antigen contributes to escape from cross-immunity in the classical bordetellae.

BACKGROUND: Horizontal gene transfer (HGT) allows for rapid spread of genetic material between species, increasing genetic and phenotypic diversity. Although HGT contributes to adaptation and is widespread in many bacteria, others show little HGT. This study builds on previous work to analyze the evolutionary mechanisms contributing to variation within the locus encoding a prominent antigen of the classical bordetellae. RESULTS: We observed amongst classical bordetellae discrete regions of the lipopolysaccharide O-antigen locus with higher sequence diversity than the genome average. Regions of this locus had less than 50% sequence similarity, low dN/dS ratios and lower GC content compared to the genome average. Additionally, phylogenetic tree topologies based on genome-wide SNPs were incongruent with those based on genes within these variable regions, suggesting portions of the O-antigen locus may have been horizontally transferred. Furthermore, several predicted recombination breakpoints correspond with the ends of these variable regions. To examine the evolutionary forces that might have selected for this rare example of HGT in bordetellae, we compared in vitro and in vivo phenotypes associated with different O-antigen types. Antibodies against O1- and O2-serotypes were poorly cross-reactive, and did not efficiently kill or mediate clearance of alternative O-type bacteria, while a distinct and poorly immunogenic O-antigen offered no protection against colonization. CONCLUSIONS: This study suggests that O-antigen variation was introduced to the classical bordetellae via HGT through recombination. Additionally, genetic variation may be maintained within the O-antigen locus because it can provide escape from immunity to different O-antigen types, potentially allowing for the circulation of different Bordetella strains within the same host population.

Epidemiologic and laboratory features of a large outbreak of pertussis-like illnesses associated with cocirculating Bordetella holmesii and Bordetella pertussis--Ohio, 2010-2011.

BACKGROUND: During 9 May 2010-7 May 2011, an outbreak of pertussis-like illness (incidence, 80 cases per 100 000 persons) occurred in Franklin County, Ohio. The majority of cases were identified by IS481-directed polymerase chain reaction (PCR), which does not differentiate among Bordetella species. We sought to determine outbreak etiology and epidemiologic characteristics. METHODS: We obtained demographic, clinical, and vaccination-related data from the Ohio Disease Reporting System and Impact Statewide Immunization Information System. We tested sera from 14 patients for anti-pertussis toxin (PT) antibodies and used species-specific PCR on 298 nasopharyngeal specimens. RESULTS: Reported cases totaled 918. IS481 results were available for 10 serologically tested patients; 5 of 10 had discordant anti-PT antibody and IS481 results, suggestive of Bordetella holmesii, which lacks PT and harbors IS481. We identified specific Bordetella species in 164 of 298 specimens tested with multitarget PCR; B. holmesii and Bordetella pertussis were exclusively detected among 48 (29%) and 112 (68%), respectively; both were detected in 4 (2%). Among 48 patients with B. holmesii infections, 63% were aged 11-18 years, compared with 35% of 112 patients with B. pertussis infections (P = .001). Symptoms were similar among B. holmesii- and B. pertussis-infected patients. Adolescent pertussis ("Tdap") booster vaccinations were more effective against B. pertussis than B. holmesii (effectiveness: 67% and 36%, respectively; 95% confidence intervals, 38%-82% and -33% to 69%, respectively). CONCLUSIONS: We report the first documented mixed outbreak of B. pertussis and B. holmesii infections. Bordetella holmesii particularly affected adolescents. Although laboratory capacity limitations might inhibit routine use of multitarget PCR for clinical diagnosis, focused testing and enhanced surveillance might improve understanding the burden of B. holmesii infection.

Lack of cross-protection against Bordetella holmesii after pertussis vaccination.

Bordetella holmesii, a species closely related to B. pertussis, has been reported sporadically as a cause of whooping cough-like symptoms. To investigate whether B. pertussis-induced immunity is protective against infection with B. holmesii, we conducted an analysis using 11 human respiratory B. holmesii isolates collected during 2005-2009 from a highly B. pertussis-vaccinated population in Massachusetts. Neither whole-cell (wP) nor acellular (aP) B. pertussis vaccination conferred protection against these B. holmesii isolates in mice. Although T-cell responses induced by wP or aP cross-reacted with B. holmesii, vaccine-induced antibodies failed to efficiently bind B. holmesii. B. holmesii-specific antibodies provided in addition to wP were sufficient to rapidly reduce B. holmesii numbers in mouse lungs. Our findings suggest the established presence of B. holmesii in Massachusetts and that failure to induce cross-reactive antibodies may explain poor vaccine-induced cross-protection.

Dissecting human T cell responses against Bordetella species.

To identify the minimal structures that may be important for the creation of a synthetic and/or recombinant vaccine against whooping cough, human T cell clones were obtained against Bordetella antigens. Cloned peripheral blood T lymphocytes from an immune donor were grown in IL-2 and tested for proliferation in response to inactivated Bordetella species (B. pertussis, B. parapertussis, and B. bronchiseptica) and mutants deficient for the expression of virulence-associated antigens. All the T cell clones obtained were CD4+8- and recognized specifically the Bordetella antigens when presented by autologous B cells. On the basis of the responsiveness to the whole inactivated bacteria, it was possible to cluster the 12 clones obtained into four groups with the following specificity: (1) filamentous hemagglutinin (FHA); (2) B. pertussis-specific antigens; (3) virulence-associated Bordetella-specific antigens; and (4) nonvirulence-associated Bordetella-specific antigens. Using two new B. pertussis deletion mutants, clone 6 (representative of cluster 1) was found to recognize the COOH terminus of FHA. Furthermore, three out of four clones of cluster 3 were specifically stimulated by the soluble 69-kD protein from the outer membrane of B. pertussis. Surprisingly, none of the twelve clones obtained by stimulation in vitro with whole inactivated bacteria recognized pertussis toxin (PT), which is believed to be the most important protein to be included in an acellular vaccine. However, when a new generation of clones was obtained using soluble PT as the in vitro stimulus, it was observed that 11 clones of this group recognized this antigen. Thus, PT does not seem to be the most representative antigen on the whole inactivated bacteria, although T cell memory against PT exists in a donor who had the disease several years ago.

Human T cell clones define S1 subunit as the most immunogenic moiety of pertussis toxin and determine its epitope map.

Human T lymphocyte clones specific for pertussis toxin (PT) were used to analyze the fine specificity of the response to PT, the basic component of new acellular vaccines against whooping cough. The majority (83%) of the clones specific for PT recognized S1, the subunit that in animal models has been shown to be highly immunogenic. To map T cell epitopes on S1, 18 S1-specific clones were tested for recognition of recombinant fragments representing NH2-terminal and COOH-terminal deletions of S1 and two recombinant S1 subunits containing amino acid substitutions. This approach led to the identification of three regions of the protein as the sequences containing T cell antigenic sites: 1-42, 181-211, and 212-235. Synthetic peptides were eventually used for a finer localization of the T cell epitopes. Two peptides, one of 13 residues (27-39) at the NH2 terminus and one of 24 residues (171-194) at the COOH terminus, stimulated proliferation of three and four clones, respectively. Both peptides are recognized in association with HLA DR1 molecules. These results stress the role of S1 in the immune response to PT and provide data useful for the development of a recombinant or synthetic antipertussis vaccine containing T cell epitopes from S1.

Immunomodulation as a Novel Strategy for Prevention and Treatment of Bordetella spp. Infections.

Well-adapted pathogens have evolved to survive the many challenges of a robust immune response. Defending against all host antimicrobials simultaneously would be exceedingly difficult, if not impossible, so many co-evolved organisms utilize immunomodulatory tools to subvert, distract, and/or evade the host immune response. Bordetella spp. present many examples of the diversity of immunomodulators and an exceptional experimental system in which to study them. Recent advances in this experimental system suggest strategies for interventions that tweak immunity to disrupt bacterial immunomodulation, engaging more effective host immunity to better prevent and treat infections. Here we review advances in the understanding of respiratory pathogens, with special focus on Bordetella spp., and prospects for the use of immune-stimulatory interventions in the prevention and treatment of infection.

Enhancement of immune response against Bordetella spp. by disrupting immunomodulation.

Well-adapted pathogens must evade clearance by the host immune system and the study of how they do this has revealed myriad complex strategies and mechanisms. Classical bordetellae are very closely related subspecies that are known to modulate adaptive immunity in a variety of ways, permitting them to either persist for life or repeatedly infect the same host. Exploring the hypothesis that exposure to immune cells would cause bordetellae to induce expression of important immunomodulatory mechanisms, we identified a putative regulator of an immunomodulatory pathway. The deletion of btrS in B. bronchiseptica did not affect colonization or initial growth in the respiratory tract of mice, its natural host, but did increase activation of the inflammasome pathway, and recruitment of inflammatory cells. The mutant lacking btrS recruited many more B and T cells into the lungs, where they rapidly formed highly organized and distinctive Bronchial Associated Lymphoid Tissue (BALT) not induced by any wild type Bordetella species, and a much more rapid and strong antibody response than observed with any of these species. Immunity induced by the mutant was measurably more robust in all respiratory organs, providing completely sterilizing immunity that protected against challenge infections for many months. Moreover, the mutant induced sterilizing immunity against infection with other classical bordetellae, including B. pertussis and B. parapertussis, something the current vaccines do not provide. These findings reveal profound immunomodulation by bordetellae and demonstrate that by disrupting it much more robust protective immunity can be generated, providing a pathway to greatly improve vaccines and preventive treatments against these important pathogens.

Integrated Signaling Pathways Mediate Bordetella Immunomodulation, Persistence, and Transmission.

The mammalian immune system includes a sophisticated array of antimicrobial mechanisms. However, successful pathogens have developed subversive strategies to detect, modulate, and/or evade immune control and clearance. Independent disciplines study host immunology and bacterial pathogenesis, but interkingdom signaling between bacteria and host during natural infection remains poorly understood. An efficient natural host infection system has revealed complex communication between Bordetella spp. and mice, identified novel regulatory mechanisms, and demonstrated that bordetellae can respond to microenvironment and inflammatory status cues. Understanding these bacterial signaling pathways and their complex network that allows precisely timed expression of numerous immunomodulatory factors will serve as a paradigm for other organisms lacking such a powerful experimental infection system. VIDEO ABSTRACT.

Modeling systems-level regulation of host immune responses.

Many pathogens are able to manipulate the signaling pathways responsible for the generation of host immune responses. Here we examine and model a respiratory infection system in which disruption of host immune functions or of bacterial factors changes the dynamics of the infection. We synthesize the network of interactions between host immune components and two closely related bacteria in the genus Bordetellae. We incorporate existing experimental information on the timing of immune regulatory events into a discrete dynamic model, and verify the model by comparing the effects of simulated disruptions to the experimental outcome of knockout mutations. Our model indicates that the infection time course of both Bordetellae can be separated into three distinct phases based on the most active immune processes. We compare and discuss the effect of the species-specific virulence factors on disrupting the immune response during their infection of naive, antibody-treated, diseased, or convalescent hosts. Our model offers predictions regarding cytokine regulation, key immune components, and clearance of secondary infections; we experimentally validate two of these predictions. This type of modeling provides new insights into the virulence, pathogenesis, and host adaptation of disease-causing microorganisms and allows systems-level analysis that is not always possible using traditional methods.

Bordetella adenylate cyclase toxin: a swift saboteur of host defense.

Bordetella that infect mammals produce a multifunctional repeat in toxin (RTX) adenylate cyclase toxin known as CyaA, an excellent example of bacterial sophistication in subverting host defense. Recent reports show that interaction of CyaA with tracheal epithelial cells aids adhesion of Bordetella to ciliated mucosa and induces production of the pro-inflammatory cytokine interleukin, IL-6. Myeloid phagocytes, attracted to the site of infection are the target of freshly secreted CyaA that binds to the alpha(M)beta2 integrin (CD11b/CD18), penetrates cells and promptly suppresses their bactericidal functions by converting cellular ATP to cAMP. Such uncontrolled cAMP signaling can also drive CD11b-expressing immature dendritic cells into a semi-mature state, possibly hijacking them to shape the local adaptive immune response towards tolerance of the pathogen.

[Whooping cough from infants to adults]. / La coqueluche de l'enfant a l'adulte.

Neither natural nor vaccine-induced immunity to pertussis lasts for life. Before intensive vaccination of toddlers, pertussis was essentially a pediatric disease. Mortality and morbidity have fallen drastically in parts of the world where toddlers have been systematically protected with efficacious whole-cell vaccines. The infection is now more common in adults with waning specific immunity, who can go on to infect newborns too young to be vaccinated, with potentially dramatic consequences. For these reasons, the vaccine strategy was recently modified in several countries, with the introduction of vaccine boosters for children, adolescents and young adults, as well as for healthcare workers in contact with newborns. In France, vaccine boosters were introduced for adolescents in 1998 and for adults in contact with newborns in 2004, leading to a decrease in morbidity in adolescents and very young adults. It is important to inform the medical community, and the families with neonates, of these changes in vaccine strategy. Efficient surveillance and notification systems are needed in all countries. Surveillance must include the use of standardized diagnostic tests and reagents. Bacterial isolates should be thoroughly characterized at the genomic, transcriptomic and proteomic levels.

Bacterial Metabolism in the Host Environment: Pathogen Growth and Nutrient Assimilation in the Mammalian Upper Respiratory Tract.

Pathogens evolve in specific host niches and microenvironments that provide the physical and nutritional requirements conducive to their growth. In addition to using the host as a source of food, bacterial pathogens must avoid the immune response to their presence. The mammalian upper respiratory tract is a site that is exposed to the external environment, and is readily colonized by bacteria that live as resident flora or as pathogens. These bacteria can remain localized, descend to the lower respiratory tract, or traverse the epithelium to disseminate throughout the body. By virtue of their successful colonization of the respiratory epithelium, these bacteria obtain the nutrients needed for growth, either directly from host resources or from other microbes. This chapter describes the upper respiratory tract environment, including its tissue and mucosal structure, prokaryotic biota, and biochemical composition that would support microbial life. Neisseria meningitidis and the Bordetella species are discussed as examples of bacteria that have no known external reservoirs but have evolved to obligately colonize the mammalian upper respiratory tract.

Structural variability and originality of the Bordetella endotoxins.

Structural studies of Bordetella endotoxins (LPSs) have revealed remarkable differences: (i) between their LPSs and those of other bacterial pathogens; (ii) among the LPSs of the seven identified Bordetella species; and (iii) among the LPSs of some Bordetella strains. The lipid As have the "classical" bisphosphorylated diglucosamine backbone but tend to have fewer and species-specific fatty acid components compared to those of other genera. Nevertheless, three strains of B. bronchiseptica have at least three different fatty acid distributions; however, the recently identified B. hinzii and B. trematum LPSs had identical lipid A structures. The B. pertussis core is a dodecasaccharide multi-branched structure bearing amino and carboxylic groups. Another unusual feature is the presence of free amino sugars in the central core region and a complex distal trisaccharide unit containing five amino groups of which four are acetylated and one is methylated. The B. pertussis LPS does not have O-chains and that of B. trematum had only a single O-unit, unlike the LPSs of all the other species of the smooth-type. The O-chain-free cores of non-B. pertussis LPSs were always built on the B. pertussis core model but most were species-specifically incomplete. The LPS structures of three B. bronchiseptica strains were found to be different from each other. The O-chains of B. bronchiseptica and B. parapertussis were almost identical and had some features in common with B. hinzii O-chain. Serological analyses are consistent with the determined LPS structures.

Pertussis.

Pertussis as an infectious disease exhibits a number of unusual features, both with respect to its epidemiology and to the interaction of the bacteia with the infected host. Evidence that a virus may in some cases be the etiology of the syndrome has been reviewed and seems to be have been established in certain instances. Pertussis caused by a virus would not be difficult to accept insofar as the respiratory manifestations of the disease are concerned. It is difficult, however, to reconcile the hematologic changes of pertussis with what is presently known about the usual virus infection of humans. The clinical syndrome itself is suficiently unique as to be recognized in most cases. The use of antibiotics has considerably reduced the mortality of the disease by allowing treatment of complications. The other serious complication of disease, pertussis encephalopathy, remains a problem. The possible occurrence of hypoglycemia during pertussis has been noted and deserves further documentation especially in that it may contribute to the encephalopathy...

Polymerase chain reaction for the identification of Bordetella pertussis and Bordetella parapertussis.

The polymerase chain reaction (PCR) for the detection of Bordetella pertussis and Bordetella parapertussis DNA in clinical samples was well documented by recent studies. Different regions in Bordetella pertussis DNA have been successfully used as targets for this method by various authors. In this work we report the usefulness of the PCR assay also for speciating Bordetellae isolates in those cases where the biochemical and serological tests gave inconclusive results.

Characterization of the common antigenic lipopolysaccharide O-chains produced by Bordetella bronchiseptica and Bordetella parapertussis.

Representative strains of Bordetella bronchiseptica and B. parapertussis were found to produce smooth lipopolysaccharides (LPS) having identical antigenic O-polysaccharide components composed of linear unbranched polymers of 1,4-linked 2,3-diacetamido-2,3-dideoxy-alpha-L-galacto-pyranosyluronic acid residues. These LPSs differed from the LPS of B. pertussis which produces only rough-type LPS, devoid of O-polysaccharide. While B. bronchiseptica and B. parapertussis had chemically and immunologically identical O-polysaccharide structures, their core oligosaccharide components differed. The core oligosaccharide of B. parapertussis was chemically distinct from the core of B. bronchiseptica which appeared to be structurally and immunologically similar to a core oligosaccharide of B. pertussis.