Bordetella pertussis isolates in Finland after acellular vaccination: serotype change and biofilm formation.

OBJECTIVES: In Finland, whole cell pertussis vaccine (wP) was introduced in 1952 and was replaced by acellular pertussis vaccine (aP) without fimbrial (FIM) antigen in 2005. We aimed to analyse the changes in serotypes of circulating Bordetella pertussis before and after acellular vaccination and to explore the relationship between biofilm formation and serotype diversity after the introduction of aP vaccine. METHODS: Serotyping of 1399 B. pertussis isolates collected at the Finnish National Reference Laboratory for Pertussis and Diphtheria in Turku, Finland, from 1974 to 2023 was performed by slide agglutination or indirect ELISA. Of 278 isolates collected after 2005, 53 were selected, genotyped for fim3 and fim2 alleles, and tested for biofilm formation. The selection criteria included maintaining a relatively equal distribution of isolates per time interval, ensuring approximately a 50:50 ratio of FIM2 (N = 26) and FIM3 (N = 27) serotypes. The reference strain Tohama I was used as a control. RESULTS: During the wP era, the majority of circulating B. pertussis exhibited the FIM2 serotype. However, FIM3 strains have appeared since 1999 and become prevalent. After the implementation of aP vaccines, the distribution of serotypes has exhibited substantial variability. FIM3 isolates displayed an enhanced biofilm formation compared to FIM2 isolates (Geometric mean value (95% CI): 0.90 (0.79-1.03) vs. 0.75 (0.65-0.85); p < 0.05). Of the 27 FIM3 isolates, 8 harboured fim3-1 and 19 fim3-2 alleles. FIM3 isolates with fim3-2 allele were significantly associated with increased biofilm formation when compared to those with fim3-1 (1.07 (0.96-1.19) vs. 0.61 (0.52-0.72); p < 0.0001). CONCLUSION: Following the implementation of aP vaccines, the distribution of serotypes in Finland has exhibited substantial variability. FIM3 isolates with the fim3-2 allele displayed an enhanced biofilm formation capability compared to FIM2 isolates.

Genome-based prediction of cross-protective, HLA-DR-presented epitopes as putative vaccine antigens for multiple Bordetella species.

IMPORTANCE: Pertussis, caused by Bordetella pertussis, can cause debilitating respiratory symptoms, so whole-cell pertussis vaccines (wPVs) were introduced in the 1940s. However, reactogenicity of wPV necessitated the development of acellular pertussis vaccines (aPVs) that were introduced in the 1990s. Since then, until the COVID-19 pandemic began, reported pertussis incidence was increasing, suggesting that aPVs do not induce long-lasting immunity and may not effectively prevent transmission. Additionally, aPVs do not provide protection against other Bordetella species that are observed during outbreaks. The significance of this work is in determining potential new vaccine antigens for multiple Bordetella species that are predicted to elicit long-term immune responses. Genome-based approaches have aided the development of novel vaccines; here, these methods identified Bordetella vaccine candidates that may be cross-protective and predicted to induce strong memory responses. These targets can lead to an improved vaccine with a strong safety profile while also strengthening the longevity of the immune response.

Repeated Bordetella pertussis Infections Are Required to Reprogram Acellular Pertussis Vaccine-Primed Host Responses in the Baboon Model.

BACKGROUND: The United States has experienced a resurgence of pertussis following the introduction of acellular pertussis (aP) vaccines. This is likely due to the failure of aP vaccines to induce durable immunity and prevent infection, carriage, and transmission. METHODS: To evaluate the impact of aP vaccination on the immune response to infection and test the ability of infection to reprogram aP-imprinted immune responses, we challenged unvaccinated and aP-vaccinated baboons with Bordetella pertussis multiple times and accessed the immune responses and outcomes of infections after each exposure. RESULTS: Multiple infections were required to elicit T-helper 17 responses and protection in aP-vaccinated animals comparable to responses seen in unvaccinated animals after a single challenge. Even after 3 challenges, T-helper 1 responses were not observed in aP-vaccinated animals. Immunoglobulin G responses to vaccine and nonvaccine antigens were not negatively affected in aP-vaccinated animals. CONCLUSIONS: Our results indicate that it is possible to retrain aP-primed immune responses, but it will likely require an optimal booster and multiple doses. Our results in the baboon model suggest that circulation of B. pertussis in aP-vaccinated populations is concentrated in the younger age bands of the population, providing information that can guide improved modeling of B. pertussis epidemiology in aP-vaccinated populations.

Bordetella pertussis and outer membrane vesicles.

Bordetella pertussis is the causative agent of a respiratory infection called pertussis (whooping cough) that can be fatal in newborns and infants. The pathogen produces a variety of antigenic compounds which alone or simultaneously can damage various host cells. Despite the availability of pertussis vaccines and high vaccination coverage around the world, a resurgence of the disease has been observed in many countries. Reasons for the increase in pertussis cases may include increased awareness, improved diagnostic techniques, low vaccine efficacy, especially acellular vaccines, and waning immunity. Many efforts have been made to develop more effective strategies to fight against B. pertussis and one of the strategies is the use of outer membrane vesicles (OMVs) in vaccine formulations. OMVs are attracting great interest as vaccine platforms since they can carry immunogenic structures such as toxins and LPS. Many studies have been carried out with OMVs from different B. pertussis strains and they revealed promising results in the animal challenge and human preclinical model. However, the composition of OMVs differs in terms of isolation and purification methods, strains, culture, and stress conditions. Although the vesicles from B. pertussis represent an attractive pertussis vaccine candidate, further studies are needed to advance clinical research for next-generation pertussis vaccines. This review summarizes general information about pertussis, the history of vaccines against the disease, and the immune response to these vaccines, with a focus on OMVs. We discuss progress in developing an OMV-based pertussis vaccine platform and highlight successful applications as well as potential challenges and gaps.

[Pertussis worldwide. Vaccinating children and adults]. / La coqueluche dans le monde. Vacciner l'enfant et l'adulte.

Pertussis (whooping cough) is an important cause of morbidity and mortality in infants world-wide, and continues to be a public health concern despite high vaccination coverage. The disease, caused by bacterium Bordetella pertussis, is present in all countries. Before vaccines became widely available in the 1950s, pertussis was one of the most common childhood diseases worldwide. According to WHO, estimation of deaths was 4 millions/year in 1950 and 100 000/year in 2015. But morbidity remains important with a high circulation of the bacterium determining atypical clinical forms after whole cell or acellular vaccines use. This is due mainly to the absence of booster doses in adolescents and adults. Major progress are generalisation of PCR and vaccination of mother during pregnancy. A resurgence of pertussis is observed after generalisation of acellular vaccines use. In China the progression of allele ptxPl was found in all areas following the use of acellular vaccine. This allele, rare before acellullar vaccine, is linked to a macrolide resistance, and reaches more than 30% of strains isolated in hospitalised children.These evolutions must be evaluated in clinical forms and genotyping of all strains, in all areas.

Development of a TLR7/8 agonist adjuvant formulation to overcome early life hyporesponsiveness to DTaP vaccination.

Infection is the most common cause of mortality early in life, yet the broad potential of immunization is not fully realized in this vulnerable population. Most vaccines are administered during infancy and childhood, but in some cases the full benefit of vaccination is not realized in-part. New adjuvants are cardinal to further optimize current immunization approaches for early life. However, only a few classes of adjuvants are presently incorporated in vaccines approved for human use. Recent advances in the discovery and delivery of Toll-like receptor (TLR) agonist adjuvants have provided a new toolbox for vaccinologists. Prominent among these candidate adjuvants are synthetic small molecule TLR7/8 agonists. The development of an effective infant Bordetella pertussis vaccine is urgently required because of the resurgence of pertussis in many countries, contemporaneous to the switch from whole cell to acellular vaccines. In this context, TLR7/8 adjuvant based vaccine formulation strategies may be a promising tool to enhance and accelerate early life immunity by acellular B. pertussis vaccines. In the present study, we optimized (a) the formulation delivery system, (b) structure, and (c) immunologic activity of novel small molecule imidazoquinoline TLR7/8 adjuvants towards human infant leukocytes, including dendritic cells. Upon immunization of neonatal mice, this TLR7/8 adjuvant overcame neonatal hyporesponsiveness to acellular pertussis vaccination by driving a T helper (Th)1/Th17 biased T cell- and IgG2c-skewed humoral response to a licensed acellular vaccine (DTaP). This potent immunization strategy may represent a new paradigm for effective immunization against pertussis and other pathogens in early life.

An expert panel opinion on protection against pertussis in Mexico: The role of acellular and whole-cell vaccines.

Pertussis is a highly contagious disease caused by Bordetella pertussis, which may be preventable by vaccination. There are two types of vaccines: whole-cell vaccines and acellular vaccines. Since pertussis control worldwide is heterogeneous, re-emergence of whooping cough has been observed in some countries. This re-emergence has been related to several factors: increased susceptibility to infection, better detection of disease, problems in obtaining adequate vaccination coverage, increase in susceptible subjects (mainly under 6 months of age), loss of immunity in adolescents and young adults, and likely genetic and adaptive B. pertussis changes. This paper discusses whole-cell and acellular vaccines' characteristics, advantages, and disadvantages. International recommendations are presented, and the participants' position is offered regarding the influence of the use of acellular vaccines and the potential disadvantages of reintroducing whole-cell vaccines, mainly due to their reactogenicity. Finally, strategies to achieve better control of pertussis in Mexico are discussed.

La tos ferina es una enfermedad causada por Bordetella pertussis. Aunque es altamente contagiosa, puede ser prevenible por vacunación. Existen dos tipos de vacunas: las de células enteras y las acelulares. La tos ferina ha resurgido en algunos países debido a que su control a escala mundial es heterogéneo. Esta reemergencia se ha relacionado con diversos factores: mayor sensibilidad hacia la infección, mejor detección de la enfermedad, problemas para obtener adecuadas coberturas de vacunación, incremento en los sujetos susceptibles (especialmente menores de 6 meses), pérdida de la inmunidad en los adolescentes y adultos jóvenes, y probables cambios genéticos y adaptativos de B. pertussis. En este documento se analizan las características, las ventajas y las desventajas de las vacunas de células enteras y de las vacunas acelulares. Se presentan las recomendaciones internacionales y se ofrece el posicionamiento de los participantes con respecto a la influencia del uso de vacunas acelulares y las desventajas potenciales de volver a utilizar vacunas de células enteras, en especial por su reactogenicidad. Por último, se analizan las estrategias para lograr un mejor control de la tos ferina en México.

Prevalidation of the cAMP-PTx reporter assay for quantitative assessment of pertussis toxin activity.

Testing for inactivation of pertussis toxin and reversion to toxicity in aP vaccines has historically relied on the murine histamine sensitization test, that lacks mechanistic understanding, suffers from standardization problems and is associated with severe animal suffering. Though the regulatory requirements for in vivo testing of acellular pertussis (aP) vaccine products have been waived in Europe, it is still common practice globally. Easy and quantitative in vitro methods are therefore urgently needed. One of the alternatives under development is our reporter cell line - CHO-CRE cells - that carries a cAMP-reporter construct. After exposure to pertussis toxin, cells are stimulated with a low concentration of forskolin to allow detection of pertussis toxin dependent changes in intracellular cAMP levels. Here, the results of two prevalidation studies with purified pertussis toxin and pertussis toxin spiked aP vaccines are described that were performed according to the principles of the ICH Q2(R1) guidelines for a content assay. We confirmed the assay's specificity, accuracy, precision, linearity and range. The cAMP-PTx reporter assay allows for objective, reliable and quantitative assessment of pertussis toxin levels in aP vaccines and can thereby boost broad and global replacement of the histamine sensitization test.

Evolution of Bordetella pertussis in the acellular vaccine era in Norway, 1996 to 2019.

We described the population structure of Bordetella pertussis (B. pertussis) in Norway from 1996 to 2019 and determined if there were evolutionary shifts and whether these correlated with changes in the childhood immunization program. We selected 180 B. pertussis isolates, 22 from the whole cell vaccine (WCV) era (1996-1997) and 158 from the acellular vaccine (ACV) era (1998-2019). We conducted whole genome sequencing and determined the distribution and frequency of allelic variants and temporal changes of ACV genes. Norwegian B. pertussis isolates were evenly distributed across a phylogenetic tree that included global strains. We identified seven different allelic profiles of ACV genes (A-F), in which profiles A1, A2, and B dominated (89%), all having pertussis toxin (ptxA) allele 1, pertussis toxin promoter (ptxP) allele 3, and pertactin (prn) allele 2 present. Isolates with ptxP1 and prn1 were not detected after 2007, whereas the prn2 allele likely emerged prior to 1972, and ptxP3 before the early 1980s. Allele conversions of ACV genes all occurred prior to the introduction of ACV. Sixteen percent of our isolates showed mutations within the prn gene. ACV and its booster doses (implemented for children in 2007 and adolescents in 2013) might have contributed to evolvement of a more uniform B. pertussis population, with recent circulating strains having ptxA1, ptxP3, and prn2 present, and an increasing number of prn mutations. These strains clearly deviate from ACV strains (ptxA1, ptxP1, prn1), and this could have implications for vaccine efficiency and, therefore, prevention and control of pertussis.

The nearest neighbor nuclei method to objectify analysis of pertussis toxin-induced clustering.

The in vivo histamine sensitization test (HIST) has historically been performed to guarantee the safety of acellular per­tussis vaccine batches. Non-compliance of batches is primarily associated with the presence of low levels of pertussis toxin (PTx). Because of ethical, standardization and scientific reasons, a variety of alternative in vitro approaches have been studied to replace the lethal HIST. A broadly applied and partially accepted method is the CHO cell clustering test, which is based on the clustered growth pattern of CHO cells when exposed to minute amounts of PTx. One of the major hurdles for global application of the CHO clustering test is the manual assessment of the clusters, which is associated with suboptimal reproducibility of test outcomes and is time-consuming. Here, various parameters of CHO cell nuclei were evaluated in search for a reliable, objective read-out parameter. We demonstrate that the distance between each nucleus and its nearest neighbor (3N method) is the most suitable parameter to assess clustered cell growth. This method detects 2.8 mIU PTx/mL and thereby complies with the requirement set for the sensitivity of the CHO clustering test based on visual reading. In commercial acellular pertussis vaccines spiked with PTx, the method detects 45 mIU/mL PTx, which is substantially lower than the 181-725 mIU/mL PTx detected by visual interpretation. The 3N method thus allows objective and sensitive assessment of CHO clustering and thereby encourages broad and global implementation of the in vitro test as an alternative to the HIST.

Comparing the pertussis antibody levels of healthy children immunized with four doses of DTap-IPV/Hib (Pentaxim) combination vaccine and DTaP vaccine in Quzhou, China.

Despite the high coverage of pertussis vaccines in high-income countries, pertussis resurgence has been reported in recent years, and has stimulated interest in the effects of vaccines and vaccination strategies. Immunoglobulin G (IgG) antibodies against pertussis toxoid (PT), filamentous hemagglutinin (FHA), and pertactin (PRN) after immunization with four doses of co-purified or component vaccines were determined by enzyme-linked immunosorbent assay (ELISA). Serological data of PT-IgG geometric mean concentrations (GMCs) over time since vaccination were used to fit the mathematical models. A total of 953 children were included in this study; 590 participants received four doses of the component acellular vaccine and 363 participants received four doses of the co-purified acellular vaccine. The GMCs and the seropositivity rate of pertussis IgG were significantly influenced by the production methods, and the immunogenicity of the component acellular vaccine was superior to that of the co-purified acellular vaccine. The fitted mathematical models for the component acellular vaccine and the co-purified acellular vaccine were Y=91.20e-0.039x and Y=37.71x-0.493, respectively. The initial GMCs of the component acellular vaccine was higher than that of the co-purified acellular vaccine, but both were similar at 72 months after immunization. Pertussis IgG levels waned over time after four doses of acellular pertussis vaccine, regardless of whether component or co-purified vaccine was used. The development and promotion of component acellular pertussis vaccines should be accelerated in China, and booster doses of pertussis vaccine in adolescents, adults, and pregnant women should be employed.

Duration of protection against Bordetella pertussis infection elicited by whole-cell and acellular vaccine priming in Polish children and adolescents.

BACKGROUND: In the context of reported resurgence of pertussis in the last decade, researchers hypothesized that acellular (aP) pertussis vaccines elicit a shorter-lived protection compared to whole-cell (wP) pertussis vaccines. However, in the studies seeking to demonstrate this hypothesis, exposure to each vaccine type was not concurrent, and contradictory epidemiologic modeling questioned its validity. The context of pertussis vaccination history in Poland, with both vaccine types used concurrently in comparable proportions, provided an opportunity to investigate this hypothesis. We sought to compare waning of protection by primary series vaccine type by measuring anti-pertussis toxin antibody concentrations as proxy for recent infection. MATERIALS AND METHODS: Serological samples from 2,745 children and adolescents aged ≥5 years and <16 years and with completed 5-dose pertussis vaccination series were tested by ELISA for pertussis toxin (PT) antibodies. Participants were stratified by type of priming vaccine (wP or aP). Vaccination timeliness and priming-specific trends in anti-PT antibody levels by time since last vaccine dose were analyzed. RESULTS: A total of 1,161 (42.5%) children received wP vaccines, and 1,314 (48.1%) received aP vaccines for their primary series and toddler booster. Overall, 53.57% of the subjects received doses 2-4 in a timely manner, while only 41.52% received all 5 doses at the recommended intervals. Using GMCs or seropositivity measures, both priming groups showed a re-increase in anti-PT antibody levels signing infection in recent years from 8 years after the school-entry booster onward. Comparisons did not show any significant differences between the two groups in the timing or intensity of this re-increase. CONCLUSION: Our results clearly confirm that vaccine-elicited immunity against pertussis wanes among adolescents even after a complete infant, toddler and school-entry vaccination series. The timing and intensity of the waning of protection appear similar with whole-cell as with acellular pertussis vaccines.

A system-view of Bordetella pertussis booster vaccine responses in adults primed with whole-cell versus acellular vaccine in infancy.

The increased incidence of whooping cough worldwide suggests that current vaccination against Bordetella pertussis infection has limitations in quality and duration of protection. The resurgence of infection has been linked to the introduction of acellular vaccines (aP), which have an improved safety profile compared with the previously used whole-cell (wP) vaccines. To determine immunological differences between aP and wP priming in infancy, we performed a systems approach of the immune response to booster vaccination. Transcriptomic, proteomic, cytometric, and serologic profiling revealed multiple shared immune responses with different kinetics across cohorts, including an increase of blood monocyte frequencies and strong antigen-specific IgG responses. Additionally, we found a prominent subset of aP-primed individuals (30%) with a strong differential signature, including higher levels of expression for CCL3, NFKBIA, and ICAM1. Contrary to the wP individuals, this subset displayed increased PT-specific IgE responses after boost and higher antigen-specific IgG4 and IgG3 antibodies against FHA and FIM2/3 at baseline and after boost. Overall, the results show that, while broad immune response patterns to Tdap boost overlap between aP- and wP-primed individuals, a subset of aP-primed individuals present a divergent response. These findings provide candidate targets to study the causes and correlates of waning immunity after aP vaccination.

Lack of evidence supporting a role of IFN-ß and TGF-ß in differential polarization of Bordetella pertussis specific-T cell responses.

Bordetella Pertussis (BP) vaccine-induced immunity is waning worldwide despite excellent vaccine coverage. Replacement of the whole-cell inactivated vaccine (wP) by an acellular subunit vaccine (aP) is thought to play a major role and to be associated with the recurrence of whooping cough. Previously, we detected that the polarization towards a Th2 and Th1/Th17 response in aP and wP vaccinees, respectively, persists upon aP boosting in adolescents and adults. Additionally, IL-9 and TGF-ß were found to be up-regulated in aP-primed donors and network analysis further identified IFN-ß as a potential upstream regulator of IL-17 and IL-9. Based on these findings, we hypothesized that IFN-ß produced following aP vaccination may lead to increased IL-9 and decreased IL-17 production. Also, due to the well characterized role of TGF-ß in both Th17 and Th9 differentiation, we put forth that TGF-ß addition to BP-stimulated CD4 + T cells might modulate IL-17 and IL-9 production. To test this hypothesis, we stimulated in vitro cultures of PBMC or isolated naive CD4 + T cells from aP vs wP donors with a pool of BP epitopes and assessed the effect of IFN-ß or TGF-ß in proliferative responses as well as in the cytokine secretion of IL-4, IL-9, IL-17, and IFN-γ. IFN-ß reduced BP-specific proliferation in PBMC as well as cytokine production but increased IL-9, IL-4, and IFN-γ cytokines in naïve CD4 + T cells. These effects were independent of the childhood vaccination received by the donors. Similarly, TGF-ß reduced BP-specific proliferation in PBMC but induced proliferation in naïve CD4 + T cells. However, stimulation was associated with a generalized inhibition of cytokine production regardless of the original aP or wP vaccination received by the donors. Our study suggests that key T cell functions such as cytokine secretion are under the control of antigen stimulation and environmental cues but molecular pathways different than the ones investigated here might underlie the long-lasting differential cytokine production associated with aP- vs wP-priming in childhood vaccination.

The epidemic of erythromycin-resistant Bordetella pertussis with limited genome variation associated with pertussis resurgence in China.

BACKGROUND: The resurgence of Bordetella pertussis infections leading to whooping cough is a concern in many parts of the world. The number of pertussis cases in China has increased significantly since 2013. RESEARCH DESIGN AND METHODS: In this study, whole-genome sequencing analysis was performed for 388 B. pertussis strains isolated in China from the 1970s to 2018, combining 594 published strains from around the world. RESULTS: This study revealed that lineage V diverged about 50 years ago in China, while lineage IV is dominant in the other countries. It also revealed that the erythromycin-resistant sub-lineages Va, Vb, and Vc with limited genomic variation emerged 11 ~ 12 years ago. These three sub-lineages were identified after the co-purified acellular vaccines (cp-ACVs) completely replaced the previous whole cell vaccines (WCVs) after the national immunization program of 2012. It suggests that the cp-ACVs cannot induce immunity that is potent enough to restrict the spread of the lineage V, antibiotic abuse further favors the spread of this lineage in China. CONCLUSIONS: These findings demand a reassessment of the immunization strategy and development of new vaccines in China to stop the resurgence and drug resistance of B. pertussis.

Overcoming Waning Immunity in Pertussis Vaccines: Workshop of the National Institute of Allergy and Infectious Diseases.

Despite high vaccine coverage in many parts of the world, pertussis is resurging in a number of areas in which acellular vaccines are the primary vaccine administered to infants and young children. This is attributed in part to the suboptimal and short-lived immunity elicited by acellular pertussis vaccines and to their inability to prevent nasal colonization and transmission of the etiologic agent Bordetella pertussis In response to this escalating public health concern, the National Institute of Allergy and Infectious Diseases held the workshop "Overcoming Waning Immunity in Pertussis Vaccines" in September 2019 to identify issues and possible solutions for the defects in immunity stimulated by acellular pertussis vaccines. Discussions covered aspects of the current problem, gaps in knowledge and possible paths forward. This review summarizes presentations and discussions of some of the key points that were raised by the workshop.

Comparison of whole-cell versus acellular pertussis vaccine effectiveness in school clusters of pertussis, France, 2013.

OBJECTIVES AND METHOD: We conducted a prospective study in 2013 to compare the whole-cell versus acellular pertussis vaccines effectiveness and duration of protection, following the occurrence of pertussis clusters. RESULTS: During seven school outbreaks, we identified 102 clinical pertussis cases, including 10 cases biologically confirmed by Bordetella pertussis specific PCR, among a cohort of 305 children in 2nd to 6th grade. The risk of pertussis when vaccinated with an acellular vaccine alone was 1.6 (RR=1.6; 95% CI=1.1-2.5) times higher than when vaccinated with a whole-cell vaccine or using a combined schedule. CONCLUSIONS: The limited duration of protection conferred by the acellular vaccine reinforces the 2013 introduction of the pertussis booster at six years old.