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The article presents abstracts on drug delivery systems which include use of dendriplexes as a novel vaccine-delivery system against anthrax, intravenous infusions of gentamicin in neonate patients, and Overcoming the mucin barrier in the treatment of pulmonary cystic fibrosis.

The article presents abstracts on analytical chemistry which include microemulsion liquid chromatography (MELC) method validation for quantification of terbutaline, assay of ciprofloxacin in powdered tablets by spectroscopy, and the use of dendriplexes as a vaccine-delivery system against anthrax.

Abstract: Samples of human plasma from anthrax vaccine adsorbed (AVA, BioThrax™)-vaccinated individuals were used to demonstrate passive protection of A/J mice from a lethal challenge with the Sterne strain of anthrax bacteria. The maximum concentration of human anti-protective antigen IgG in mouse sera 24h after injection of 260μg of anti-PA IgG was 134μg/ml, declining to 91μg/ml at 72h (half-life=101.7h). Mice showed significant survival (p ≤0.001) after injection of serial dilutions up to 1:4 of the standard plasma and challenged with 100 LD50. Similarly, mice injected with the standard anti-AVA plasma and challenged up to 5 days post-treatment also survived (p ≤0.001). Using a cohort of human plasma to measure passive protection, the best correlation between passive protection and an in vitro assay was found to be with the quantitative toxin neutralization assay (minimum fold increase in odds of survival: 2.71, p =0.0062). These results demonstrate a reliable in vivo neutralization method that correlates with standard in vitro measures of neutralizing antibody levels in plasma from individuals vaccinated with the standard anthrax vaccine. This analytical method may provide additional opportunities to compare the efficacy of improved anthrax vaccines with the licensed vaccine. [Copyright &y& Elsevier]

Bacillus anthracis is a spore-forming bacterium that causes anthrax in humans and in other mammals. The glycoprotein BclA (Bacillus collagen-like protein of anthracis) is a major constituent of the exosporium, the outermost surface of B. anthracis spores. The glycosyl part of BclA is an oligosaccharide composed of 2-O-methyl-4-(3-hydroxy-3-methylbutanamido)-4,6-dideoxy-D-glucose, referred to as anthrose, and three rhamnose residues. A structure similar to anthrose, 4-(3-hydroxy-3-methylbutanamido)-4,6-dideoxy-D-glucose is found in the side chain of the capsular polysaccharide (CPS) of Shewanella spp. MR-4. Under certain growth conditions the bacteria produce a variant CPS lacking one methyl group on the hydroxybutyrate, 4-(3-hydroxybutanamido)-4,6-dideoxy-D-glucose. Contrary to anthrose, neither of the Shewanella CPSs is 2-O methylated. Here, we report that both Shewanella CPS variants react with anti-B. anthracis spore sera. We also found that these antisera reacted with flagellae of Pseudomonas syringae. reported to be glycosylated with a similar terminal saccharide, 4-(3-hydroxy-butanamido)-4,6-dideoxy-2-O-methyl-D-glucose. Sera produced by immunization with Shewanella or P. syringae cells bound to B. anthracis spores but not to Bacillus cereus spores in a fluorescent microscopy assay. These experiments show that methylation of the anthrose at the O-2 of the sugar ring and at the C-3 of 3-hydroxybutyrate are not essential for induction of cross-reactive antibodies. We report the preparation, characterization, and anti-body responses to protein conjugates of the two variants of Shewanella CPS. Both conjugates induced antibodies that bound to both Shewanella CPS variants by ELISA and to B. anthracis spores, as detected by fluorescent microscopy. We propose the use of Shewanella CPS conjugates as a component of an anthrax vaccine. [ABSTRACT FROM AUTHOR]

This research identifies the perceptions of U.S. military service members regarding the Department of Defense Anthrax Vaccine Immunization Program (AVIP). The service members' perceptions were addressed in the dimensions of ethics, effectiveness, and safety, as well as the overall perceptions of the AVIP. The study, conducted in October 2004, randomly selected active duty service members from the uniformed services assigned to a Caribbean military base who participated in the AVIP during the period of 1998 to 2000. Their perceptions were measured with a survey instrument with 14 closed-ended, Likert-scale questions. The research demonstrated that a substantial number of service members disagreed with issues regarding the ethics, safety, and efficacy of the AVIP, We recommend enhanced training and education to increase understanding of the benefits of the AVIP. [ABSTRACT FROM AUTHOR]

Summary: Protective antigen (PA) is a central component of anthrax toxin and a major antigen in anthrax vaccines. However, the use of native PA as a vaccine is not optimal. If administered to people who have been freshly exposed to anthrax, PA may actually aid in anthrax toxin formation and thus may pose a serious safety concern for postexposure vaccination applications. A non-functional PA mutant may be a much safer alternative. To identify an improved anthrax vaccine antigen, we examined four non-functional mutants of PA, each being impaired in a critical step of the cellular intoxication pathway of PA. These mutants were Rec− (unable to bind PA-receptors), SSSR (resistant to activation by furin), Oligo− (unable to form oligomers), and DNI (Dominant Negative Inhibitory, unable to form endosomal transmembrane pores). When tested in mice and after three doses of immunization, all four mutants were highly potent in eliciting PA-specific, toxin-neutralizing antibodies, with immunogenicity increasing in the order of PA

Summary: The seroconversion rates and geometric mean concentrations (GMC) of IgG anti-PA for stored sera from U.S. military personnel immunized 3, 4, and 6 times with the U.S. licensed anthrax vaccine adsorbed were studied. Anti-PA IgG concentrations were measured by ELISA. All 246 vaccinees had low but detectable pre-immunization anti-PA IgG (GMC 1.83μg/mL). Three doses elicited a GMC of 59.92μg/mL and a seroconversion rate of 85.3%, four doses elicited a GMC of 157.44μg/mL and 67.9% and the sixth of 276.95μg/mL and 45.5%, respectively. The forth dose elicited 100% seroconversion compared to the pre-immunization level. These results should facilitate comparison between different immunization schedules and new vaccines. [Copyright &y& Elsevier]

A new generation anthrax vaccine is expected to target not only the anthrax protective antigen (PA) protein, but also other virulent factors of Bacillus anthracis. It is also expected to be amenable for rapid mass immunization of a large number of people. This study aimed to address these needs by designing a prototypic triantigen nasal anthrax vaccine candidate that contained a truncated PA (rPA63), the anthrax lethal factor (LF), and the capsular poly-γ-d-glutamic acid (γDPGA) as the antigens and a synthetic double-stranded RNA (dsRNA), polyriboinosinic-polyribocytodylic acid (poly(I:C)) as the adjuvant. This study identified the optimal dose of nasal poly(I:C) in mice, demonstrated that nasal immunization of mice with the LF was capable of inducing functional anti-LF antibodies (Abs), and showed that nasal immunization of mice with the prototypic triantigen vaccine candidate induced strong immune responses against all three antigens. The immune responses protected macrophages against an anthrax lethal toxin challenge in vitro and enabled the immunized mice to survive a lethal dose of anthrax lethal toxin challenge in vivo. The anti-PGA Abs were shown to have complement-mediated bacteriolytic activity. After further optimization, this triantigen nasal vaccine candidate is expected to become one of the newer generation anthrax vaccines. [ABSTRACT FROM AUTHOR]

Abstract: The active component of the licensed human anthrax vaccine (BioThrax™, or AVA) is a Bacillus anthracis toxin known as protective antigen (PA). Second generation anthrax vaccines currently under development are also based on a recombinant form of PA. Since the current and future anthrax vaccines are based on this toxin, it is important that the immunobiology of this protein in vaccinated humans be understood in detail. We have isolated and analyzed the PA-specific antibody repertoire from an AVA-vaccinated individual. When examined at the clonal level, we find an antibody response that is complex in terms of the combinatorial elements and immunoglobulin variable genes employed. All PA-specific antibodies had undergone somatic hypermutation and class switch recombination, both signs of affinity maturation. Although the antigenic epitopes recognized by the response were distributed throughout the PA monomer, the majority of antibodies arising in this individual following vaccination recognize determinants located on the amino-terminal (PA20) sub-domain of the molecule. This latter finding may have implications for the rational design of future PA-based anthrax vaccines. [Copyright &y& Elsevier]

In 2001 the United States experienced several anthrax attacks that attracted significant attention from the medical community, policy analysts and many other professionals, but relatively little attention from economists. This paper builds on the work of Hupert et al. (2002) by using an M/G/1 queuing model to analyze the tension between economic cost minimization and the stringency of medical examinations in the aftermath of an anthrax attack. The theoretical analysis displays ambiguous results. Given this outcome, numerical analysis of the aforementioned tension is conducted with two main results. First, for many values of the model parameters that describe the stringency of the examinations, a tension does exist between economic cost minimization and examination stringency. Second, for many values of the parameter that describes the scale of an anthrax attack a tension does not exist between economic cost minimization and examination stringency. Moreover, when medical examiners are charged with the task of making equitable versus ethical tradeoffs in their provision of medical care, the problem of discerning the lower cost examination regime becomes confounded further. It is the goal of this work to extend the literature regarding bioterrorism and medical responses to include an additional goal of economic cost minimization. [ABSTRACT FROM AUTHOR]

Background: Combination vaccines reduce the total number of injections required for each component administered separately and generally provide the same level of disease protection. Yet, physical, chemical, and biological interactions between vaccine components are often detrimental to vaccine safety or efficacy. Methods: As a possible alternative to combination vaccines, we used specially designed microneedles to inject rhesus macaques with four separate recombinant protein vaccines for anthrax, botulism, plague and staphylococcal toxic shock next to each other just below the surface of the skin, thus avoiding potentially incompatible vaccine mixtures. Results: The intradermally-administered vaccines retained potent antibody responses and were well- tolerated by rhesus macaques. Based on tracking of the adjuvant, the vaccines were transported from the dermis to draining lymph nodes by antigen-presenting cells. Vaccinated primates were completely protected from an otherwise lethal aerosol challenge by Bacillus anthracis spores, botulinum neurotoxin A, or staphylococcal enterotoxin B. Conclusion: Our results demonstrated that the physical separation of vaccines both in the syringe and at the site of administration did not adversely affect the biological activity of each component. The vaccination method we describe may be scalable to include a greater number of antigens, while avoiding the physical and chemical incompatibilities encountered by combining multiple vaccines together in one product. [ABSTRACT FROM AUTHOR]

Anthrax spores rank as the leading threat among bioweapons. This article reviews the accumulated evidence for immunization, either active or passive, to counter the malicious release of anthrax spores. The key protective factor in current anthrax vaccines for humans is a protein called protective antigen, which allows ingress of toxins into cells. The US vaccine is licensed to prevent anthrax, regardless of the route of exposure. Its dosing schedule is cumbersome and somewhat painful (shortcomings that may be resolved by ongoing clinical studies). It can be prescribed with the confidence commensurate with dozens of human safety studies and experience in 1.8 million recent vaccinees. For post-exposure prophylaxis, combining antibiotic prophylaxis and active immunization before illness onset may offer the best combination of prompt and sustained protection, especially for people who inhale large doses of spores. To treat anthrax infection, passive immunization using a polyclonal or monoclonal antibody product may offer important clinical benefit, especially if the anthrax bacteria are resistant to multiple antibiotics. [ABSTRACT FROM AUTHOR]

The Centers for Disease Control and Prevention has classified Bacillus anthracis, the causative organism of anthrax, as a category A potential bioterrorism agent. There are critical shortcomings in the US anthrax vaccine program. Rather than depending on the private sector, the government must assume direct production of anthrax vaccine. The development of a capacity capable of preemptive immunization of the public against anthrax should be considered. (Am J Public Health. 2007;97:1945-1951. doi:10.2105/ AJPH.2006.102749) [ABSTRACT FROM AUTHOR]

The article focuses on the assessment of the safety of anthrax immunization in the U.S. Army aircrew members through physical examination. To address questions raised regarding the safety of anthrax vaccine, immunized and un-immunized people have been compared in population-based studies. The study conducted a retrospective evaluation of data from periodic physical examinations collected on anthrax-immunized and un-immunized U.S. Army aircrew members between 1998 and 2005. Detailed information regarding the results of the study is discussed.

The article presents abstracts on pharmacology which include risk mitigation in formulation development strategy, validation in engineering enhanced protein formulation, and understanding the interaction of antigen and adjuvant in an recombinant plasminogen activator (rPA) anthrax vaccine.

Abstract: In this study, we examine the potential of a combinatorial vaccine consisting of the lead-candidate antigens for the next generations of vaccines against anthrax (rPA) and plague (F1-V) with the specific objective of determining synergy or interference between the vaccine components when they are administered separately or together by both traditional parenteral immunization (SC) and mucosal immunization (IN) in the presence of appropriate adjuvants. The most significant findings of the study reported here are that (1) a combinatorial vaccine consisting of equal amounts of F1-V and rPA administered SC is effective at eliciting a robust serum and bronchoalveolar lavage (BAL) antigen-specific IgG and IgG1 response against both antigens in immunized animals, and when administered IN, a robust antigen-specific IgG2a response in the serum and BAL is also induced; (2) there were few instances where either synergy or interference was observed in the combined vaccine administered by either route and those differences occurred soon after the final immunization and were not sustained over time; (3) IN immunization was as effective as SC immunization for induction of antigen-specific serum and BAL antibody responses using the same amount of antigen; (4) the IgG1/IgG2a ratios suggest a strongly biased Type 2 response following SC immunization, while IN immunization produced a more balanced Type 1/Type 2 response; (5) the IgG1/IgG2a ratio was influenced by the route of immunization, the adjuvant employed, and the nature of the antigen. As with previously published studies, there were still detectable levels of circulating anti-F1-V and anti-rPA even 6 months post-primary immunization. These studies provide important insights into the development of new generation biodefense vaccines. [Copyright &y& Elsevier]

Vaccination with anthrax vaccine adsorbed (AVA) results in the production of protective antigen (PA) specific antibodies, which play an important protective role against anthrax toxins. Analyzing the specificity of serum antibodies generated in response to AVA vaccination can provide insight into the mechanisms of protective immunity against this important pathogen. The goal of this study was to develop a competitive enzyme linked immunosorbent assay (cELISA) to test human immune serum for antibodies specific for a known lethal toxin neutralizing epitope in PA. PA-specific antibodies in sera from individuals who received the six-dose AVA vaccine series competed for binding to immobilized PA with monoclonal antibody F20G75, which binds to a linear epitope in domain 2 of PA and neutralizes lethal toxin activity in vitro. These results suggest that antibodies in human AVA vaccinee serum recognize the same epitope as F20G75, or one in close proximity to it, and may serve a protective role against anthrax lethal toxin. This assay may be used for serological confirmation of successful immunization against anthrax and for the identification of antibodies in human vaccinee serum that recognize protective epitopes on PA. [ABSTRACT FROM AUTHOR]

Background: In 1998, the United States Department of Defense initiated the Anthrax Vaccine Immunization Program. Concerns about vaccine-related adverse health effects followed, prompting several studies. Although some studies used self-reported vaccination data, the reliability of such data has not been established. The purpose of this study was to compare self-reported anthrax vaccination to electronic vaccine records among a large military cohort and to evaluate the relationship between vaccine history and health outcome data. Methods: Between September 2005 and February 2006 self-reported anthrax vaccination was compared to electronic records for 67,018 participants enrolled in the Millennium Cohort Study between 2001 and 2003 using kappa statistics. Multivariable modeling investigated vaccination concordance as it pertains to subjective health (functional status) and objective health (hospitalization) metrics. Results: Greater than substantial agreement (kappa=0.80) was found between self-report and electronic recording of anthrax vaccination. Of all participants with electronic documentation of anthrax vaccination, 98% self-reported being vaccinated; and of all participants with no electronic record of vaccination, 90% self-reported not receiving a vaccination. There were no differences between vaccinated and unvaccinated participants in overall measures of health. Only the subset of participants who self-reported anthrax vaccination, but had no electronic confirmation, differed from others in the cohort, with consistently lower measures of health as indicated by Medical Outcomes Study 36-Item Short Form Health Survey for Veterans (SF-36V) scores. Conclusions: These results indicate that military members accurately recall their anthrax vaccinations. Results also suggest that anthrax vaccination among Millennium Cohort participants is not associated with self-reported health problems or broad measures of health problems severe enough to require hospitalization. Service members who self-report vaccination with no electronic documentation of vaccination, however, report lower measures of physical and mental health and deserve further research. [Copyright &y& Elsevier]

Abstract: Using a cross-sectional analysis design, we measured serum anti-protective antigen (PA) concentrations in individuals receiving six or fewer US licensed anthrax vaccinations. Samples were collected from 363 individuals with a mean of 29.6±8.42 months after their last vaccination (range 3–57 months). An enzyme-linked immunosorbent assay (ELISA) developed and validated by the Centers for Disease Control and Prevention (CDC) was used to evaluate the range and status of anthrax vaccine-induced serum antibody concentrations. A significant correlation (r =0.73, P ≤0.001) was found to exist between the number of vaccinations received and specific anti-PA immunoglobulin G (IgG) concentrations. We observed two discrete groups comprised of one to three doses (5.9–11.7μg/ml) and four to six doses (26.2–30.2μg/ml). These data indicate that anti-PA IgG is present at low but detectable levels after as few as two vaccinations (5.9±6.43μg/ml). These findings may have significance for anthrax vaccine recipients who are unable to complete the primary or full regimen with this licensed product. [Copyright &y& Elsevier]

Synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs act as immune adjuvants, improving the response elicited by a coadministered vaccine. Combining CpG ODN with anthrax vaccine adsorbed (AVA, the licensed human vaccine) increases the speed, magnitude, and avidity of the resultant antibody response. IgG Abs against anthrax protective antigen (PA) protect mice, guinuea pigs, and rhesus macaques from infection. [ABSTRACT FROM AUTHOR]

Abstract: Specific lots of anthrax vaccine adsorbed administered to members of the U.S. Armed Forces have been alleged to contain squalene, a chemical purported to be associated with illnesses of Gulf War veterans. A method of enhanced sensitivity for determining squalene in anthrax vaccine adsorbed using high-performance liquid chromatography with photodiode array detection has been developed, validated, and applied to 44 bottles of 38 lots of anthrax vaccine. In 43 bottles of 37 lots, no squalene was detected within a detection limit of 1ng/0.5ml dose (2 parts-per-billion). One lot, FAV008, was found to contain trace amounts of squalene at 7, 9, and 1μgl−1, levels considerably below normal human plasma levels (290μgl−1). The overall results of this investigation provide direct evidence for the absence of squalene in nearly all of anthrax vaccine preparations tested. [Copyright &y& Elsevier]

Lysosomal processing of lethal toxin (LTX) is a key event in the pathogenesis of anthrax. This study investigated the ability of chloroquine (CQ) to interfere with this processing and thereby to reduce suppression of T lymphocytes. T lymphocytes isolated from blood were activated, by cross-linking of CD3, in both the absence and presence of LTX and CQ and then were assayed by flow cytometry and immunoblotting. LTX was found to disrupt intracellular signaling, and it down-regulated T lymphocyte function. CQ significantly reduced the harmful effects of LTX and protected the activation and cytokine production of T lymphocytes. This effect may indicate a promising strategy in the treatment of anthrax. [ABSTRACT FROM AUTHOR]

Summary: Background: We report the results of a phase I dose escalation, safety and immunogenicity trial of a new recombinant protective antigen (rPA102) anthrax vaccine. Methods: Hundred healthy volunteers were randomized in a 4:1 ratio to receive intramuscular doses of rPA102 in the following formulations: 5, 25, 50, or 75μg of rPA102 in 82.5μg aluminum hydroxide adjuvant at 0, 4, and 8 weeks; or the US licensed Anthrax Vaccine Adsorbed (AVA) at weeks 0 and 4. Findings: Local reactogenicity (mostly pain) was more common with AVA than with rPA102 following the first (94.7% versus 44.4%; p <0.001) and the second (84.2% versus 35.4%; p <0.001) vaccinations. Systemic reactogenicity (mostly headache) was more common among rPA102 vaccinees, but only following the first vaccination (49.4% versus 15.8%; p =0.025). A dose-response relationship for anti-PA antibodies was present after the 2nd and 3rd vaccinations. Two weeks following the 2nd vaccination, the geometric mean titers (GMT) for lethal toxin neutralization activity (TNA), for the 5, 25, 50 and 75μg rPA102 and AVA groups were 38.6, 75.4, 373.9, 515.3, and 855.2, respectively. The geometric mean concentrations (GMC) measured by anti-PA IgG ELISA were 3.7, 11.5, 25.9, 44.1, and 171.6, respectively. Two weeks following the 3rd vaccination, TNA GMTs for the four rPA102 groups, were: 134.7, 719.7, 2116.6, 2422.4; and ELISA GMCs were: 22.9, 104.7, 196.4, and 262.6, respectively. Interpretation: No clinically serious or dose-related toxicity or reactogenicity was observed. The TNA response after two injections of the 75μg dose of rPA102 was similar to the response after two injections of AVA. The third rPA102 vaccination substantially increased the antibody response. [Copyright &y& Elsevier]

Prevention of inhalational anthrax after Bacillus anthracis spore exposure requires a prolonged course of antibiotic prophylaxis. In response to the 2001 anthrax attack in the United States, ≈10,000 people were offered 60 days of antibiotic prophylaxis to prevent inhalational anthrax, but adherence to this regimen was poor. We sought to determine whether a short course of antibiotic prophylaxis after exposure could protect non-human primates from a high-dose spore challenge if vaccination was combined with antibiotics. Two groups of 10 rhesus macaques were exposed to ≈1,600 LD50 of spores by aerosol. Both groups were given ciprofloxacin by orogastric tube twice daily for 14 days, beginning 1-2 h after exposure. One group also received three doses of the licensed human anthrax vaccine (anthrax vaccine adsorbed) after exposure. In the ciprofloxacin-only group, four of nine monkeys (44%) survived the challenge. In contrast, all 10 monkeys that received 14 days of antibiotic plus anthrax vaccine adsorbed survived (P = 0.011). Thus postexposure vaccination enhanced the protection afforded by 14 days of antibiotic prophylaxis alone and completely protected animals against inhalational anthrax. These data provide evidence that postexposure vaccination can shorten the duration of antibiotic prophylaxis required to protect against inhalational anthrax and may impact public health management of a bioterrorism event. [ABSTRACT FROM AUTHOR]

The objectives were to illustrate the ease with which one might attribute concomitant or subsequent illness to an exposure such as the anthrax vaccine and to demonstrate an approach that keeps the significance of such exposures in appropriate perspective. A 43-year-old, active duty, Army officer presents with a variety of nonspecific common symptoms and raises concerns about the relationship of his symptoms to receipt of the anthrax vaccine. He is admitted for an evaluation that includes a series of diagnostic tests and consultations. The course of his illness and the corresponding evaluation are reviewed using a series of questions and accompanying discussions to highlight key points regarding diagnostic considerations, the anthrax vaccine, and the ultimate identification of the correct diagnosis. [ABSTRACT FROM AUTHOR]

Abstract: Whereas Pasteur patented the biotechnological processes that he invented between 1857 and 1873 in the agro-food domain, he did not file any patents on the artificial vaccine preparation processes that he subsequently developed. This absence of patents can probably be explained by the 1844 patent law in France that established the non-patentable status of pharmaceutical preparations and remedies, including those for use in veterinary medicine. Despite the absence of patents, the commercial exploitation of the anthrax vaccine in the 1880s and 1890s led to a technical and commercial monopoly by Pasteur’s laboratory as well as the founding of a commercial company to diffuse the vaccine abroad. Pasteur repeatedly refused to transfer his know-how and anthrax vaccine production methods to foreign laboratories, on the grounds that he wished to control the quality of the vaccines produced. Indeed, it was relatively difficult to transfer a method that was not yet perfectly stabilized in the early 1880s. Pasteur also wanted to maintain the monopoly of his commercial company and to increase the profits from vaccine sales so that the Institut Pasteur could be financially independent. The ‘Pasteur anthrax vaccine’ operating licences are described and analysed in detail in this article. [Copyright &y& Elsevier]

Abstract: The UK anthrax vaccine uses the culture supernatant of toxigenic non-encapsulated Bacillus anthracis as a crude source for protective antigen (PA). The precise amount of PA is not known. We developed a single radial immuno-diffusion (SRD) assay and an indirect ELISA to measure PA in desorbed anthrax vaccines. Based on 23 batches, the PA contents varied from 19.1 to 88.8μgml−1, with an average of 39.6μgml−1. Analysis of four batches by ELISA revealed considerably lower levels of PA. This discrepancy can be explained by competition of other proteins for binding sites, which results in an artificially low amount of bound PA per well. We conclude that the SRD assay is a reproducible method for the measurement of PA and this assay will contribute to quality control and improve the specifications of current anthrax vaccines. [Copyright &y& Elsevier]

Current human anthrax vaccines available in the United States and Europe consist of alum-precipitated supernatant material from cultures of a toxigenic, nonencapsulated strain of Bacillus anthracis. The major component of human anthrax vaccine that confers protection is protective antigen (PA). A second-generation human vaccine using the recombinant PA (rPA) is being developed. In this study, to prevent the toxicity and the degradation of the native rPA by proteases, we constructed two PA variants, delPA (163-168) and delPA (313-314), that lack trypsin (S163-R164-K165-K166-R167-S168) or chymotrypsin cleavage sequence (F313-F314), respectively. These proteins were expressed in Bacillus brevis 47-5Q. The delPAs were fractionated from the culture supernatant of B. brevis by ammonium sulfate at 70% saturation, followed by anion exchange chromatography on a Hitrap Q, Hiload 16/60 superdex 200 gel filtration column and phenyl sepharose hydrophobic interaction column. In accordance with previous reports, both delPA proteins combined with lethal factor protein did not show any cytotoxicity on J774A.1 cells. The delPA (163- 168) and delPA (313-314) formulated either in Rehydragel HPA or MPL-TDM-CWS (Ribi-Trimix), elicited a comparable amount of anti-PA and neutralizing antibodies to those of native rPA in guinea pigs, and confers full protection of guinea pigs from 50 · LD50 of fully virulent B. anthracis spore challenges. Ribi-Trimix was significantly more effective in inducing anti-PA and neutralizing antibodies than Rehydragel HPA. These results indicate the possibility of delPA (163-168) and delPA (313-314) proteins being developed into nontoxic, effective and stable recombinant vaccine candidates. [ABSTRACT FROM AUTHOR]

Comments on the unintentional exposure of workers to Bacillus anthracis at the Children's Hospital Oakland Research Institute in California. Overview of research being conducted at the laboratory; Importance of the implementation of BSL-2 practices among laboratory personnel who are exposed to infectious cultures; Recommendation on routine anthrax vaccination of persons who work with production quantities or concentrations of B. anthracis cultures.

Anthrax vaccination has been used in an effort to prevent infection should anthrax be used as a biological weapon, and widespread use has been considered in the event of another anthrax attack on American soil, but the long-term impact of anthrax vaccination on reproductive outcome is unknown. We found that exposure to the anthrax vaccine by males who were undergoing assisted reproduction did not negatively impact semen parameters, fertilization rate, embryo quality, or clinical pregnancy rates. [Copyright &y& Elsevier]

Rashes are frequent and potentially serious adverse consequences of smallpox vaccination. Life-threatening rashes must be differentiated from benign, self-limiting ones. Generalized vaccinia, erythema multiforme, and folliculitis are distinct self-limiting entities but may be difficult to differentiate from one another. Two cases of folliculitis after smallpox vaccination are described. Both patients received anthrax vaccination within 2 weeks before smallpox vaccination. Both presented with a papulopustular rash 9 days after smallpox vaccination. Although the rashes were initially diagnosed as erythema multiforme. the clinical features were more consistent with folliculitis. Self-limiting rashes after smallpox vaccination are common and may be difficult to distinguish from each other. These rashes are clinically distinct, with characteristic features. Improvement in diagnosis may help classify the frequency and risk of rashes after smallpox vaccination. The association of vaccine-associated folliculitis, anthrax vaccine, and other potential antigenic triggers should be further explored. [ABSTRACT FROM AUTHOR]

The successful use of Bacillus anthracis as a lethal biological weapon has prompted renewed research interest in the development of more effective vaccines against anthrax. The disease consists of three critical components: spore, bacillus, and toxin, elimination of any of which confers at least partial protection against anthrax. Current remedies rely on postexposure antibiotics to eliminate bacilli and pre- and postexposure vaccination to target primarily toxins. Vaccines effective against toxin have been licensed for human use, but need improvement. Vaccines against bacilli have recently been developed by us and others. Whether effective vaccines will be developed against spores is still an open question. An ideal vaccine would confer simultaneous protection against spores, bacilli, and toxins. One step towards this goal is our dually active vaccine, designed to destroy both bacilli and toxin. Existing and potential strategies towards potent and effective anthrax vaccines are discussed in this review. [ABSTRACT FROM AUTHOR]

It has been more than 100 years since the realization that microbes are capable of causing disease. In that time, we have learned a great deal as to how each organism has adapted to the immune system so as to avoid elimination. As well, we have also learned an immense amount since Louis Pasteur first proposed that the solution to infectious diseases was to culture the microbes and attenuate their virulence, so as to use them as vaccines. From the optimism and promise of the 19th century and immunization as the ultimate answer to the invasion by the microbial world, to the scientific realities of the 21st century, it is of interest to retrace the steps of the earliest microbiologists cum immunologists, to realize how far we've come, as well as how far we yet have to go. This editorial focuses on the history of anthrax as a microbial disease, and the earliest efforts at producing a vaccine for its prevention. [ABSTRACT FROM AUTHOR]

Abstract: Background:: During February 2003 a number of Australian sailors were returned home from their deployment to the Persian Gulf after refusing anthrax vaccination. This paper examines the media coverage of this episode as a case study in how controversies about vaccine safety escalate. Methods:: Frame analysis of articles from major Australian newspapers (n = 83) and transcripts of radio and television news and current affairs programs (n = 22) to identify the main supportive and oppositional themes used in reportage and media debate. Findings:: Initially, the major news frames were supportive of the vaccine refusing soldiers, and conveyed a sense of distrust of the government’s actions. These initial themes were rapidly re-framed and new dominant discourses appeared. First, sailors went from brave whistleblowers to being portrayed as deserters and cowards. Second, proponents shifted from their portrayal as faceless regulators to personal risk takers embodied in a well-respected Major General having the vaccine. Third, the voluntary nature of the vaccine was emphasised, thus dousing the flames of implied coercion. Conclusion:: Marked shifts in the representation of vaccine opponents and proponents possibly contributed to the rapid diminishment of media interest in the story. [Copyright &y& Elsevier]

The current UK anthrax vaccine is an alum precipitate prepared from static culture filtrate of the avirulent, unencapsulated Sterne strain of Bacillus anthracis. Protective antigen (PA) is regarded as the major immunogen in the vaccine and production conditions are intended to maximize the PA content. However, the precise composition of the vaccine is unknown and there are concerns that the observed side effects of vaccination may be caused by residual enzymatically active toxin components. Two-dimensional gel electrophoresis (2DGE) was used to define the protein components of the current UK anthrax vaccine. Consistency of composition was assessed by examining batches spanning 14 years of vaccine production. The reproducibility of the 2DGE technique was assessed by repeated analysis of selected vaccine batches. For two recently produced batches, between 86.7 and 88.8% of the spots could be matched. However, for one older batch, reproducibility of the spot pattern was considerably less, with a mean similarity of 53.4%. This difference may be explained by a change in production or because of decay during storage. Variation between the recently produced batches ranged from 72.9 to 84.3%, whereas the similarity between these and old batches was comparatively low at between 30 and 59%.Our results demonstrate that, as expected, the major antigen present in the vaccine is PA. The 83 and 63kDa species are dominant but there are numerous lower molecular weight fragments resulting from proteolytic cleavage. In addition, we have established the presence of the toxin components, oedema factor and lethal factor, and S-layer proteins, EA1 and SAP. Mass spectrometry has also enabled us to identify several bacterial cell-derived proteins present in the vaccine, including PA, enolase, fructose-bisphosphate aldolase, nucleoside diphosphate kinase and a 60kDa heat shock protein. The use of proteomics can provide useful information on the antigenic make up of this vaccine and the consistency of vaccine production. [Copyright &y& Elsevier]

This study was conducted to examine whether U.S. Army personnel receiving ≥ 1 dose of anthrax vaccine adsorbed (AVA ) between March 1998 and February 2002 were at higher risk of disability than unvaccinated personnel. We studied a historical cohort study of 716, 833 active-duty soldiers (154, 456 vaccinated) followed for 4.25 years to determine rates of evaluation for disability discharge. Cox proportional hazards models compared estimated risk of evaluation for disability, accounting for occupation and sociodemographics. Adjusted hazard ratio (HR) and 95 % confidence interval (CI) was 0.96 (CI = 0.92–0.99). Separate adjusted HRs for men, women, permanent and temporary disability, musculoskeletal and neurologic conditions were similar, ranging from O. 90 to 1.04. Latency assumptions did not affect results. Anthrax vaccination does not increase risk of disability. This finding may be partially the result of factors influencing selection for vaccination or vaccine tolerance. [ABSTRACT FROM AUTHOR]

Objective: The objective of this study was to determine whether anthrax vaccine resulted in adverse health effects in Canadian Forces members 8 months after vaccination. Methods: A quasi-experimental, retrospective chart review was undertaken for two groups within the Canadian Forces, one group that received anthrax vaccination and another that did not. Information on symptoms, diagnoses, and injuries for 848 persons for which there were approximately 35.000 chart entries was abstracted from charts over a 4.5-year period and was coded using the International Statistical Classification of Diseases and Related Health Problems. 10th edition. Results; The chart retrieval rate was 84%. The mean number of chart entries per person was higher in the comparison group (43.4) than in the vaccine group (38.2). No statistically significant differences were seen in the percent change before and after vaccination in the number of chart entries for specific diagnoses and symptoms for the vaccine group compared with the comparison group. Visual inspection of the time trend in rates showed no unexplained increases in the rate of diagnosis and symptoms in the vaccine group after vaccination. Conclusion: This study found no evidence that the anthrax vaccination resulted in an increase in adverse health effects in the 8-month period after vaccination. [ABSTRACT FROM AUTHOR]

The potency test for the anthrax vaccine currently licensed for human use in the United States (Anthrax Vaccine Adsorbed) involves the protection of actively immunized guinea pigs from a lethal challenge with a virulent strain of Bacillus anthracis. Lethal challenge tests entail the use of specialized containment facilities for the safe and secure handling of the challenge strain. This potential difficulty, plus humane considerations, have prompted us to investigate non-lethal, alternative immunogenicity assays that could be considered as potency tests not only for the current vaccine, but also for vaccines under development. Immunogenicity tests will require suitable measurement of an antibody response to relevant antigens, by methods such as enzyme linked immunosorbent assay (ELISA) or a toxin neutralization assay. Any assay chosen for this purpose should be adequately validated and reproducible by other laboratories. Validation of an analytical procedure requires the demonstration that the assay is suitable for its intended purpose. The objective of this work was to study the performance of an anti-PA-ELISA designed to assess the antibody response to anthrax vaccines in mice. Validation studies were performed according to the guidelines of the International Conference of Harmonization (ICH), and we have established the working range of the assay (37–1159EU/mL) on the bases of the following parameters: linearity (20–1159EU/mL; r2=0.99; p-value=0.21), accuracy (91–118% recovery), precision (≤20%CV, repeatability; ≤9 and ≤21%CV, intermediate precision per day and per analyst, respectively), detection limit (5EU/mL), and quantification limit (37EU/mL). We believe that assay specificity and the above characteristics are adequate to allow this ELISA to be considered for use in a mouse immunogenicity (potency) test of anthrax vaccines, and for the standardization of reagents. [Copyright &y& Elsevier]

The article discusses Stevens-Johnson Syndrome (SJS) after immunization with smallpox, anthrax, and tetanus vaccines. Smallpox, a highly contagious illness, was declared eradicated in 1980, but recent bioterrorism concerns have renewed interest in smallpox prevention and treatment. Immunization with smallpox vaccine is effective but can produce numerous adverse reactions, ranging in seventy from minor to life threatening. Researchers describe a patient in whom SJS developed after immunization with smallpox, anthrax, and tetanus vaccines.

A recombinant protective antigen (rPA)-based enzyme-linked immunosorbent assay (ELISA) was developed to measure the serological response of female A/J mice after inoculation with the new rPA-based anthrax vaccine. Several fundamental parameters of the ELISA were evaluated: specificity, precision, accuracy, linearity, and stability. Experimental results suggested that the quantitative anti-rPA IgG ELISA could be used to measure antibody levels in female A/J mice and may be useful as a potency assay to monitor consistency of manufacture of a rPA-based vaccine for planned clinical trials. [Copyright &y& Elsevier]

The use of the concept of ‘direct’ versus ‘indirect’ causal effects is common, not only in statistics but also in many areas of social and economic sciences. The related terms of ‘biomarkers’ and ‘surrogates’ are common in pharmacological and biomedical sciences. Sometimes this concept is represented by graphical displays of various kinds. The view here is that there is a great deal of imprecise discussion surrounding this topic and, moreover, that the most straightforward way to clarify the situation is by using potential outcomes to define causal effects. In particular, I suggest that the use of principal stratification is key to understanding the meaning of direct and indirect causal effects. A current study of anthrax vaccine will be used to illustrate ideas. [ABSTRACT FROM AUTHOR]