Bordetella Pertussis virulence factors in the continuing evolution of whooping cough vaccines for improved performance.

Despite high vaccine coverage, whooping cough caused by Bordetella pertussis remains one of the most common vaccine-preventable diseases worldwide. Introduction of whole-cell pertussis (wP) vaccines in the 1940s and acellular pertussis (aP) vaccines in 1990s reduced the mortality due to pertussis. Despite induction of both antibody and cell-mediated immune (CMI) responses by aP and wP vaccines, there has been resurgence of pertussis in many countries in recent years. Possible reasons hypothesised for resurgence have ranged from incompliance with the recommended vaccination programmes with the currently used aP vaccine to infection with a resurged clinical isolates characterised by mutations in the virulence factors, resulting in antigenic divergence with vaccine strain, and increased production of pertussis toxin, resulting in dampening of immune responses. While use of these vaccines provide varying degrees of protection against whooping cough, protection against infection and transmission appears to be less effective, warranting continuation of efforts in the development of an improved pertussis vaccine formulations capable of achieving this objective. Major approaches currently under evaluation for the development of an improved pertussis vaccine include identification of novel biofilm-associated antigens for incorporation in current aP vaccine formulations, development of live attenuated vaccines and discovery of novel non-toxic adjuvants capable of inducing both antibody and CMI. In this review, the potential roles of different accredited virulence factors, including novel biofilm-associated antigens, of B. pertussis in the evolution, formulation and delivery of improved pertussis vaccines, with potential to block the transmission of whooping cough in the community, are discussed.

Roads to the development of improved pertussis vaccines paved by immunology.

Current acellular pertussis vaccines have various shortcomings, which may contribute to their suboptimal efficacy and waning immunity in vaccinated populations. This calls for the development of new pertussis vaccines capable of inducing long-lived protective immunity. Immunization with whole cell pertussis vaccines and natural infection with Bordetella pertussis induce distinct and more protective immune responses when compared with immunization with acellular pertussis vaccines. Therefore, the immune responses induced with whole cell vaccine or after infection can be used as a benchmark for the development of third-generation vaccines against pertussis. Here, we review the literature on the immunology of B. pertussis infection and vaccination and discuss the lessons learned that will help in the design of improved pertussis vaccines.

What to do about pertussis vaccines? Linking what we know about pertussis vaccine effectiveness, immunology and disease transmission to create a better vaccine.

Pertussis (whooping cough) is a respiratory disease caused by the bacterium Bordetella pertussis. Despite the implementation of immunization programs and high vaccine coverage in most jurisdictions, pertussis is still one of the most common vaccine-preventable diseases, suggesting that the current vaccines and immunization schedules have not been sufficiently effective. Several factors are thought to contribute to this. The acellular pertussis vaccine that has been used in many jurisdictions since the 1990s is less effective than the previously used whole-cell vaccine, with immunity waning over time. Both whole-cell and acellular pertussis vaccines are effective at reducing disease severity but not transmission, resulting in outbreaks in vaccinated cohorts. In this review, we discuss various limitations of the current approaches to protection from pertussis and outline various options for reducing the burden of pertussis on a population level.

Strategies and new developments to control pertussis, an actual health problem.

The aim of this article is to describe the current epidemiological situation of pertussis, as well as different short-term strategies that have been implemented to alleviate this threat. The state of the art of the development of new vaccines that are expected to provide long-lasting immunity against pertussis was also included.

Characterization of co-purified acellular pertussis vaccines.

Whole-cell pertussis vaccines (WPVs) have been completely replaced by the co-purified acellular vaccines (APVs) in China. To date few laboratory studies were reported for co-purified APVs in terms of their antigenic composition and protective immune responses. To further understand the antigenic composition in co-purified APVs, in the present study 2-dimensional gel electrophoresis-based proteomic technology was used to analyze the composition of co-purified APVs. The results showed that besides the main antigens pertussis toxin (PT) and filamentous hemagglutinin (FHA), co-purified APVs also contained pertactin (PRN), fimbriae (FIM) 2and3 and other minor protein antigens. Of the 9 proteins identified, 3 were differentially presented in products from manufacturer 1 and manufacturer 2. Compared with WPVs and purified APVs, co-purified APVs induced a mixed Th1/Th2 immune response with more toward to a Th1 response than the purified APVs in this study. These results hint that different immune mechanisms might be involved in protection induced by co-purified and purified APVs.

Development of improved pertussis vaccine.

Rates of infection with Bordetella pertussis, the gram-negative bacterium that causes the respiratory disease called whooping cough or pertussis, have not abated and 16 million cases with almost 200,000 deaths are estimated by the WHO to have occurred worldwide in 2008. Despite relatively high vaccination rates, the disease has come back in recent years to afflict people in numbers not seen since the pre-vaccine days. Indeed, pertussis is now recognized as a frequent infection not only in newborn and infants but also in adults. The disease symptoms also can be induced by the non-vaccine-preventable infection with the close species B. parapertussis for which an increasing number of cases have been reported. The epidemiologic situation and current knowledge of the limitations of pertussis vaccine point out the need to design improved vaccines. Several alternative approaches and their challenges are summarized.

Live attenuated vaccines against pertussis.

The intensive use of pertussis vaccines has dramatically reduced the incidence of whooping cough during the 20th century. However, recent outbreaks in countries with high vaccination coverage illustrate the shortcomings of current vaccination regimens, and immunity induced by the most recent, acellular vaccines wanes much faster than anticipated. As an alternative, live attenuated vaccine candidates have recently been developed in order to mimic natural infection, which induces long-lasting immunity. One of them has successfully completed a Phase I trial in humans and is now undergoing further product and clinical developments. This article describes the development of such vaccines, discusses their advantages over existing vaccines and their interesting bystander properties as powerful anti-inflammatory agents, which widens their potential use far beyond that for protection against whooping cough.

Pertussis: pertussis control strategies and the options for improving current vaccines.

Pertussis is resurgent in many countries, perhaps owing in part to waning immunity after acellular pertussis vaccination. We consider the options for improving current vaccines as well as other strategies to control pertussis.

Pathogenesis and histopathology of pertussis: implications for immunization.

Pertussis is a unique infectious disease in that it can be severe and fatal but occurs without fever and other evidence of an inflammatory illness. The authors with others have studied the histopathology of fatal pertussis and also the unique characteristics of severe pertussis in young infants. Histopathologic observations from approximately 100 years ago, and from recent evaluation, indicate that the histopathologic changes of the upper respiratory tract of patients with fatal pertussis are often relatively normal unless there is a secondary bacterial infection. Bordetella pertussis contains many protein antigens and perhaps a polysaccharide capsule which contribute to the infectious process. However, only two of these antigens contribute to clinical illness. These antigens are pertussis toxin and the yet to be identified 'cough toxin'. The authors speculate as to the nature of the 'cough toxin' and discuss the implications of their observations and concepts for the future control of pertussis.

Bordetella pertussis pathogenesis: current and future challenges.

Pertussis, also known as whooping cough, has recently re-emerged as a major public health threat despite high levels of vaccination against the aetiological agent Bordetella pertussis. In this Review, we describe the pathogenesis of this disease, with a focus on recent mechanistic insights into B. pertussis virulence-factor function. We also discuss the changing epidemiology of pertussis and the challenges facing vaccine development. Despite decades of research, many aspects of B. pertussis physiology and pathogenesis remain poorly understood. We highlight knowledge gaps that must be addressed to develop improved vaccines and therapeutic strategies.

Possible options for new pertussis vaccines.

Increasing evidence that the currently available acellular pertussis vaccines are not providing optimal control of pertussis in the United States and many other countries has stimulated interest in improvements of the current vaccines and in the development of new vaccines. A better understanding of the limitations of the current vaccines and the basis for the pertussis resurgence is needed to design improved vaccines. This article outlines several alternate approaches and summarizes the challenges related to the development of new or modified vaccines.

Outer membrane vesicles derived from Bordetella parapertussis as an acellular vaccine against Bordetella parapertussis and Bordetella pertussis infection.

Bordetella parapertussis, a close related species of B. pertussis, can also cause the disease named pertussis or whooping cough. The number of cases caused by this related pathogen has risen sustained in the last years. The widely used cellular (wP) or acellular (aP) pertussis vaccines have little or no efficacy against B. parapertussis. In an effort to devise an effective acellular vaccine against B. parapertussis infection, outer membrane vesicles (OMVs) were obtained from B. parapertussis. Proteomic analysis of the resulting OMVs, designated OMVsBpp, evidenced the presence of several surface immunogens including pertactin. The characterized OMVsBpp were used in murine B. parapertussis intranasal challenge model to examine their protective capacity when administered by systemic route. Immunized BALB/c mice were challenged with sublethal doses of B. parapertussis. Significant differences between immunized animals and the negative control group were observed (p<0.001). OMVsBpp protected against B. parapertussis infection, whereas current commercial aP vaccine showed little protection against such pathogen. More interestingly, protection induced by OMVsBpp against B. pertussis was comparable to our previously designed vaccine consisting in OMVs derived from B. pertussis (OMVsBp). For these experiments we used as a positive control the current commercial aP vaccine in high dose. As expected aP offered protection against B. pertussis in mice. Altogether the results presented here showed that the OMVs from B. parapertussis are an attractive vaccine candidate to protect against whooping cough induced by B. parapertussis but also by B. pertussis.

Bordetella pertussis vaccine strains and circulating isolates in Serbia.

In Serbia, whole cell pertussis vaccine was introduced in 1957. Current composition of the vaccine has been used since 1985 and contains four autochthonous strains of Bordetella pertussis isolated from 1957 to 1984. To monitor changes in bacterial population, 70 isolates collected from 1953 to 2000 were studied together with the vaccine strains. The methods included serotyping of fimbriae (Fim), genotyping of pertactin (prn) and pertussis toxin S1 subunit (ptxA), and pulsed-field gel electrophoresis analysis. Shift from ptxA2 to ptxA1 has been observed in isolates since the late of 1960s. All isolates from 1980 to 1984 harbored ptxA1. Re-appearance of the ptxA2 allele followed an addition of the two strains harboring ptxA1 in the vaccine in 1985. The allele prn1 was predominant among the Serbian isolates, though prn3 and prn11 have been detected since 1981 and 1984. The allele prn2 was found only in two strains isolated in 2000. Serotype Fim2.3 disappeared before 1980 and serotype Fim2 became predominant since then. The Serbian vaccine strains showed differences in ptxA and prn. The results of this present study indicate that the B. pertussis population in Serbia is different from other vaccinated populations and that this difference may be related to the vaccine used.

Scaling-up vaccine production: implementation aspects of a biomass growth observer and controller.

This study considers two aspects of the implementation of a biomass growth observer and specific growth rate controller in scale-up from small- to pilot-scale bioreactors towards a feasible bulk production process for whole-cell vaccine against whooping cough. The first is the calculation of the oxygen uptake rate, the starting point for online monitoring and control of biomass growth, taking into account the dynamics in the gas-phase. Mixing effects and delays are caused by amongst others the headspace and tubing to the analyzer. These gas phase dynamics are modelled using knowledge of the system in order to reconstruct oxygen consumption. The second aspect is to evaluate performance of the monitoring and control system with the required modifications of the oxygen consumption calculation on pilot-scale. In pilot-scale fed-batch cultivation good monitoring and control performance is obtained enabling a doubled concentration of bulk vaccine compared to standard batch production.

[Effect of solution environment on the purification of pertussis toxin].

The low recovery of pertussis toxin (PT) and the low resolving efficiency of chromatography, due to the instability of PT in low salt condition, are the main challenges for its purification. We aplied 2 mol/L urea to prevent the aggregation and disassociation of PT during the purification by ion-exchange chromatography (IEC) and gel filtration chromatography (GFC). The effect of urea on the purification of PT was studied by ELISA assay and non-reduced SDS-PAGE. The activity recoveries of PT and filamentous hemagglutinin (FHA) in IEC and GFC, the resolution efficiency in GFC and the purities of PT and FHA were improved obviously by adding 2 mol/L urea in the mobile phase. The results highlight the potential application of urea in the acellular pertussis vaccine (APV) manufacture procedure.

Storage at -3 degrees C for 24 h alters the immunogenicity of pertussis vaccines.

The immunogenicity of pertussis antigens in an acellular and a whole-cell triple antigen vaccine used for childhood immunisation was assessed in murine models after storage of vaccines below 0 degree C. Swiss outbred and Balb/c mice received DTPa or DTPw vaccine or placebo. Vaccines were stored at 2-8 degrees C (ideal), or at -3 degrees C for 24 h. Pre and post immunisation IgG responses to pertussis toxin (PT), filamentous haemagglutinin (FHA) and pertactin (PRN) were measured using enzyme immunoassays (EIA). In Balb/c mice, responses to pertactin after receiving adversely stored DTPa were significantly reduced (P = 0.005, difference in GMCs 145.9% [24.6-385.4%]). A reduction in GMC to pertactin was also seen in response to adversely stored DTPw (P = 0.190,224.1% [83.8-599.2%]). Outbred mice receiving adversely stored DTPa had lower IgG antibody responses to FHA than those receiving correctly stored vaccine (P = 0.002,522.2% [26.1-2155.6%]). Outbred mice also had a significantly lower response to FHA after administration of DTPw (P = 0.009,14.0% [3.8-51.9%]). Responses to DTPa in both strains generally were greater than those to DTPw. Storage of pertussis vaccines below 0 degree C appears to alter the immunogenicity of PRN and FHA. Further study is required to determine the effects of such storage on vaccine protective efficacy.

Acellular pertussis vaccine in children with perinatal human immunodeficiency virus-type 1 infection.

The immunogenicity of an acellular pertussis vaccine containing genetically detoxified pertussis toxin, filamentous haemoagglutinin and pertactin was studied in 12 children [median age: 45 (6-107) months] with perinatal human immunodeficiency virus-type 1 (HIV-1) infection. Antibody response to all antigens was observed in six cases and another children 3 reacted to two or one antigen(s), but titres were lower than those from healthy controls. Antibody titre fold-rise correlated with preimmunization CD4-positive cell counts. Significant titres were still detectable 4 months after the third dose. The acellular vaccine is immunogenic in a portion of children with perinatal HIV-1 infection but early vaccination might be more effective, taking advantage of still adequate CD4-positive cell numbers.