Mineo Watanabe

Lipopolysaccharides from Bordetella pertussis and Bordetella parapertussis Differently Modulate Human Dendritic Cell Functions Resulting in Divergent Prevalence of Th17-Polarized Responses

Bordetella pertussis and B. parapertussis are the etiological agents of pertussis, yet the former has a higher incidence and is the cause of a more severe disease, in part due to pertussis toxin. To identify other factors contributing to the different pathogenicity of the two species, we analyzed the capacity of structurally different lipooligosaccharide (LOS) from B. pertussis and LPS from B. parapertussis to influence immune functions regulated by dendritic cells. Either B. pertussis LOS and B. parapertussis LPS triggered TLR4 signaling and induced phenotypic maturation and IL-10, IL-12p40, IL-23, IL-6, and IL-1β production in human monocyte-derived dendritic cells (MDDC). B. parapertussis LPS was a stronger inducer of all these activities as compared with B. pertussis LOS, with the notable exception of IL-1β, which was equally produced. Only B. parapertussis LPS was able to induce IL-27 expression. In addition, although MDDC activation induced by B. parapertussis LPS was greatly dependent on soluble CD14, B. pertussis LOS activity was CD14-independent. The analysis of the intracellular pathways showed that B. parapertussis LPS and B. pertussis LOS equally induced IκBα and p38 MAPK phosphorylation, but B. pertussis LOS triggered ERK1/2 phosphorylation more rapidly and at higher levels than B. parapertussis LPS. Furthermore, B. pertussis LOS was unable to induce MyD88-independent gene induction, which was instead activated by B. parapertussis LPS, witnessed by STAT1 phosphorylation and induction of the IFN-dependent genes, IFN regulatory factor-1 and IFN-inducible protein-10. These differences resulted in a divergent regulation of Th cell responses, B. pertussis LOS MDDC driving a predominant Th17 polarization. Overall, the data observed reflect the different structure of the two LPS and the higher Th17 response induced by B. pertussis LOS may contribute to the severity of pertussis in humans.

Whooping cough due to Bordetella parapertussis: an unresolved problem

Bordetella parapertussis is one of the bacteria that causes whooping cough. However, little attention has been paid to this bacterium because it causes a milder illness than Bordetella pertussis and the rate of detection is low, even though research suggests that pertussis vaccines have limited efficacy against B. parapertussis infection. However, recent studies have revealed high rates of detection in patients with whooping cough in some field studies. In this review, the relevant studies of B. parapertussis are summarized and it is demonstrated that it is now necessary to pay greater attention to infections by this bacterium.

Acellular pertussis vaccines in Japan: past, present and future

An antivaccine movement developed in Japan as a consequence of increasing numbers of adverse reactions to whole-cell pertussis vaccines in the mid-1970s. After two infants died within 24 h of the vaccination from 1974 to 1975, the Japanese government temporarily suspended vaccinations. Subsequently, the public and the government witnessed the re-emergence of whooping cough, with 41 deaths in 1979. This series of unfortunate events revealed to the public that the vaccine had, in fact, been beneficial. Furthermore, researchers and the Japanese government proceeded to develop safer pertussis vaccines. Japan now has the most experience worldwide with acellular pertussis vaccines, being the first country to have approved their use. This review describes the major events associated with the Japanese vaccination program. The Japanese experience should be valuable to other countries that are considering the development and use of such vaccines.

Reciprocal protective immunity against Bordetella pertussis and Bordetella parapertussis in a murine model of respiratory infection.

ABSTRACT The protective immunity induced by infection with Bordetella pertussis and with Bordetella parapertussis was examined in a murine model of respiratory infection. Convalescent mice that had been infected by aerosol with B. pertussis or with B. parapertussis exhibited a protective immune response against B. pertussis and also against B. parapertussis. Anti-filamentous hemagglutinin (anti-FHA) serum immunoglobulin G (IgG) and anti-FHA lung IgA antibodies were detected in both mice infected with B. pertussis and those infected with B. parapertussis. Antibodies against pertussis toxin (anti-PT) and against killed B. pertussis cells were detected in mice infected with B. pertussis. Pertactin-specific antibodies and antibodies against killed B. parapertussis cells were detected in mice infected with B. parapertussis. Spleen cells from mice infected with B. pertussis secreted interferon-γ (IFN-γ) in response to stimulation by FHA or PT. Spleen cells from mice infected with B. parapertussis also secreted IFN-γ in response to FHA. Interleukin-4 was not produced in response to any of the antigens tested. The profiles of cytokine secretion in vitro revealed induction of a Th1-biased immune response during convalescence from infection by B. pertussis and byB. parapertussis. It is possible that Th1 and Th2 responses against FHA might be related to the reciprocal protection achieved in our murine model.

Characterization of Serological Responses to Pertussis

ABSTRACT We have compared the use of five nonvaccine antigens to the use of conventional vaccine antigens, pertussis toxin (PT), and filamentous hemagglutinin (FHA) for the serological diagnosis of pertussis by enzyme-linked immunosorbent assay (ELISA). The nonvaccine antigens included the catalytic region of adenylate cyclase toxin (CatACT), the C-terminal region of FHA (C-FHA), lipooligosaccharide (LOS), the peptidoglycan-associated lipoprotein (PAL), and the BrkA protein. The serological responses of individuals with culture-confirmed pertussis were compared to those of adults with no recent history of a coughing disease. An immunoglobulin G (IgG) ELISA for PT was the most sensitive (92.2%) test for the serodiagnosis of pertussis. Of the nonvaccine antigens, ELISA for IgG responses to CatACT (sensitivity, 62.8%), C-FHA (sensitivity, 39.2%), and LOS IgA (sensitivity, 29.4%) were less sensitive but could also distinguish culture-positive individuals from control individuals. The use of a combination of multiple ELISA targets improved the sensitivity of the assay for serological diagnosis. Elevated IgG and IgA antibody titers persisted for more than a year in the individuals with culture-confirmed pertussis.

BopB is a type III secreted protein in Bordetella bronchiseptica and is required for cytotoxicity against cultured mammalian cells

The cytotoxicity of Bordetella bronchiseptica to infected cells is known to be dependent on a B. bronchiseptica type III secretion system. Although the precise mechanism of the type III secretion system is unknown, BopN, BopD and Bsp22 have been identified as type III secreted proteins. In order to identify other proteins secreted via the type III secretion machinery in Bordetella, a type III mutant was generated, and its secretion profile was compared with that of the wild‐type strain. The results showed that the wild‐type strain, but not the type III mutant, secreted a 40‐kDa protein into the culture supernatant. This protein was identified as BopB by the analysis of its N‐terminal amino acid sequence. Severe cytotoxicity such as necrosis was induced in L2 cells by infection with the wild‐type B. bronchiseptica. In contrast, this effect was not observed by the BopB mutant infection. The haemolytic activity of the BopB mutant was greatly impaired compared with that of the wild‐type strain. The results of a digitonin assay strongly suggested that BopB was translocated into HeLa cells infected with the wild‐type strain. Taken together, our results demonstrate that Bordetella secretes BopB via a type III secretion system during infection. BopB may play a role in the formation of pores in the host plasma membrane which serve as a conduit for the translocation of effector proteins into host cells.

Synergic Effect of Genotype Changes in Pertussis Toxin and Pertactin on Adaptation to an Acellular Pertussis Vaccine in the Murine Intranasal Challenge Model

ABSTRACT The Bordetella pertussis pertussis toxin and pertactin (Prn) are protective antigens and are contained in acellular pertussis vaccines. Polymorphisms in the A subunit of pertussis toxin (PtxA) and pertactin have been proposed to mediate vaccine resistance and contribute to pertussis reemergence. To test this hypothesis, previous studies compared clinical isolates expressing different alleles for the proteins. However, other virulence factors or virulence factor expression levels also may vary, confounding the analysis. To overcome these limitations, we constructed isogenic mutants of B. pertussis Tohama expressing the alleles ptxA1 or ptxA2 and prn1 or prn2 and compared the efficacies of an acellular pertussis vaccine against the mutants in a mouse model. While the vaccine was effective against all of the B. pertussis strains regardless of the allele expression pattern, the strain expressing ptxA1 and prn2 displayed a survival advantage over the other strains. These results suggest that an allele shift to the ptxA1 prn2 genotype may play a role in the emergence of pertussis in vaccinated populations.

Role of Systemic and Mucosal Immune Responses in Reciprocal Protection against Bordetella pertussis and Bordetella parapertussis in a Murine Model of Respiratory Infection

ABSTRACT The roles of systemic humoral immunity, cell-mediated immunity, and mucosal immunity in reciprocal protective immunity against Bordetella pertussis and Bordetella parapertussis were examined by using a murine model of respiratory infection. Passive immunization with serum from mice infected with B. pertussis established protective immunity against B. pertussis but not against B. parapertussis. Protection against B. parapertussis was induced in mice that had been injected with serum from mice infected with B. parapertussis but not from mice infected with B. pertussis. Adoptive transfer of spleen cells from mice infected with B. pertussis or B. parapertussis also failed to confer reciprocal protection. To examine the role of mucosal immunity in reciprocal protection, mice were infected with preparations of either B. pertussis or B. parapertussis, each of which had been incubated with the bronchoalveolar wash of mice that were convalescing after infection with B. pertussis or B. parapertussis. Such incubation conferred reciprocal protection against B. pertussis and B. parapertussis on infected mice. The data suggest that mucosal immunity including secreted immunoglobulin A in the lungs might play an important role in reciprocal protective immunity in this murine model of respiratory infection.

Efficacy of chemically cross-linked antigens for acellular pertussis vaccine.

The efficacy of chemically cross-linked antigens for acellular pertussis vaccine was investigated by a murine respiratory infection model. Filamentous hemagglutinin (FHA), pertussis toxin (PT) and pertactin, prepared from a supernatant fluid of Bordetella pertussis culture or the bacterial cells, were chemically cross-linked by the addition of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide (NHSS). The biological activities of FHA and PT were inactivated by the cross-linking reaction. The cross-linked antigens induced potent protective immunity against B. pertussis more than the vaccine prepared by formalin treatment (0.2 M). These data suggest a possibility that the vaccine prepared by using EDC and NHSS is more effective than the formalin-treated vaccine.

Efficacy of pertussis vaccines consisted of antigens detoxified with tea-leaf catechins

The ability of tea-leaf catechins to detoxify agents was examined. Filamentous hemagglutinin (FHA) and pertussis toxin (PT) were detoxified by the catechins at an extraordinarily lower concentration compared with that of formalin. The sera from the mice immunized by the catechin-treated antigens recognized, not only catechin-treated, but also untreated antigens. Furthermore, catechin-treated PT induced the antibody to neutralize PT activity in the sera of the immunized mice. Pertussis vaccines were prepared including antigens detoxified by the treatment of catechins and intraperitoneally injected into mice. Protection against Bordetella pertussis infection was shown in mice immunized with the vaccines prepared by treatment with catechins. These data suggest that catechins are effective toxoiding agents for preparing a pertussis vaccine.