Many vaccines induce protective immunity via antibodies. Recent studies have used systems biological approaches to determine signatures that predict vaccine immunity in humans, but whether there is a ‘universal signature’ that can predict antibody responses to any vaccine, is unknown. Here we performed systems analyses of immune responses to the meningococcal polysaccharide and conjugate vaccines in healthy adults, in the broader context of our previous studies with the yellow fever and two influenza vaccines. To achieve this, we performed a large-scale network integration of public human blood transcriptomes, and systems-scale databases in specific biological contexts, and deduced a set of blood transcription modules. These modules revealed distinct transcriptional signatures of antibody responses to different classes of vaccines providing key insights into primary viral, protein recall and anti-polysaccharide responses. These results illuminate the early transcriptional programs orchestrating vaccine immunity in humans, and demonstrate the power of integrative network modeling.

The pneumococcal polysaccharide vaccine is recommended as a means of preventing invasive disease in the elderly. We compared responses to the 23-valent polysaccharide vaccine in 46 previously unvaccinated, healthy, institutionalized elderly persons (mean age, 85.5 years) with those in 12 healthy younger adults (mean age, 37 years) by measuring prevaccination and postvaccination serum IgG antibody concentrations (by ELISA), functional antibody activity (by opsonophagocytosis), IgG antibody avidity, and passive protection in mice. Postvaccination IgG antibody concentrations for two serotypes (6B and 19F) of the five studied (4, 6B, 14, 19F, and 23F) were significantly lower in elderly than in younger adults; however, opsonophagocytic activity was significantly reduced for all serotypes in the elderly. Sera with reduced opsonophagocytic activity (titer, <64) correlated with low IgG antibody avidity and protected mice poorly against pneumococcal challenge. In elderly persons receiving polysaccharide vaccination, there was a significant reduction in the functionality of postvaccination antibodies, and this appeared to increase with advanced age.

No abstract

The bioterrorism-associated human anthrax epidemic in the fall of 2001 highlighted the need for a sensitive, reproducible, and specific laboratory test for the confirmatory diagnosis of human anthrax. The Centers for Disease Control and Prevention developed, optimized, and rapidly qualified an enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G (IgG) antibodies to Bacillus anthracis protective antigen (PA) in human serum. The qualified ELISA had a minimum detection limit of 0.06 µg/mL, a reliable lower limit of detection of 0.09 µg/mL, and a lower limit of quantification in undiluted serum specimens of 3.0 µg/mL anti-PA IgG. The diagnostic sensitivity of the assay was 97.8%, and the diagnostic specificity was 94.2%. A competitive inhibition anti-PA IgG ELISA was also developed to enhance diagnostic specificity to 100%. The anti-PA ELISAs proved valuable for the confirmation of cases of cutaneous and inhalational anthrax and evaluation of patients in whom the diagnosis of anthrax was being considered.

Host protection against pneumococcal disease is primarily mediated by phagocytosis. We developed and standardized an opsonophagocytic assay using HL-60 cells (human promyelocytic leukemia cells). Fifty-five serum samples were analyzed for the presence of functional antibody against seven pneumococcal serogroups or serotypes (4, 6B, 9V, 14, 18C, 19F, and 23F) by using differentiated HL-60 cells (granulocytes) and peripheral blood leukocytes (PBLs). Six of the 55 serum samples were from unvaccinated adult volunteers, 31 serum samples were from adults who received one dose of the 14-valent or the 23-valent polysaccharide vaccine, and 18 serum samples were from 16-month-old infants who received four doses of an investigational 7-valent polysaccharide-protein conjugate vaccine. The results of an opsonophagocytic assay with HL-60 cells correlated highly with those of an assay with PBLs as effector cells (median r for seven serotypes ‫؍‬ 0.87; P < 0.01). Opsonophagocytic titers were compared with the immunoglobulin G antibody concentrations determined by enzyme-linked immunosorbent assay (ELISA). The r values for serogroups or serotypes 4

No abstract

Lot 89SF has been the reference standard serum pool used in pneumococcal enzyme-linked immunosorbent assays (ELISAs) since 1990. In 2005, it was estimated that there remained between 2 and 5 years' supply of lot 89SF. Since lot 89SF was the reference standard used in the evaluation of the seven-valent pneumococcal conjugate vaccine Prevnar (PCV7), the link to clinical efficacy would be severed if stocks became completely depleted. Furthermore, demonstration of immune responses comparable to those elicited by PCV7 is a licensure approach used for new pneumococcal conjugate vaccines, so a replacement reference standard was required. A total of 278 volunteers were immunized with the 23-valent unconjugated polysaccharide vaccine Pneumovax II, and a unit of blood was obtained twice within 120 days following immunization. Plasma was prepared, pooled, and confirmed to be free from hepatitis B virus (HBV), hepatitis C virus (HCV), and HIV. The pooled serum was poured at 6 ml per vial into 15,333 vials and lyophilized. Immunological bridging of 007sp to 89SF was used to establish equivalent reference values for 13 pneumococcal capsular serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F and 23F) by five independent laboratories. Antibody concentrations in 007sp were established relative to the lot 89SF reference preparation using the WHO reference ELISA. Subsequently, 12 existing WHO calibration sera had concentrations reassigned for 13 pneumococcal serotypes using new serum 007sp as the reference, and these were compared to concentrations relative to the original reference serum. Agreement was excellent for the 12 WHO calibration sera. The 007sp preparation has replaced 89SF as the pneumococcal reference standard. Sufficient quantity of this new preparation is available such that, with judicious use, it should be available for at least 25 years.Streptococcus pneumoniae human reference serum lot 89SF has been used as a standard serum in enzyme-linked immunosorbent assay (ELISA) designed to measure IgGspecific antibody for individual pneumococcal capsular polysaccharides since 1990. This standard serum greatly facilitated the standardization of ELISA methodologies during a critical period when the first polysaccharide-conjugate vaccines were being evaluated for licensure. Lot 89SF consists of pooled, defibrinated plasma collected in the late 1980s from 17 individuals vaccinated with a 23-valent pneumococcal polysaccharide vaccine (Pnu-imune; Lederle), a 4-valent meningococcal polysaccharide vaccine (Menomune; Connaught), and Haemophilus influenzae type b conjugate vaccine (ProHibit; Connaught). The final volume of this collection was approximately 10 liters (3 ml/vial). Serotypespecific weight-based values for IgG, IgA, and IgM were derived for serotypes 1,3,4,5, 6B, 7F, 9V, 14, 18C, 19F, and 23F for lot 89SF by Quateart et al. (5). Assignments for the additional serotypes in the 23-valent pneumococcal polysaccharide vaccine were subsequently bridged from the assignments for the original 11 serotypes (6). The lot 89...

The pca branch of the beta-ketoadipate pathway in Pseudomonas putida is responsible for the complete degradation of p-hydroxybenzoate through ortho cleavage of the initial pathway metabolite, protocatechuate. The pcaR regulatory locus has been found to be required for both induction of all of the genes within the pca regulon (pcaBDC, pcaIJ, and pcaF) and the chemotactic response of the bacteria to aromatic compounds. Insertional inactivation mutagenesis, using Tn5 and mini-Tn5 transposons, was used to locate, clone, and sequence this pcaR regulatory gene. The pcaR gene product, when overexpressed in Escherichia coli, possessed a specific affinity for the pcaIJ promoter region and demonstrated that the entire PcaR protein was required for this function. The deduced amino acid sequence of the PcaR regulatory peptide bears little resemblance to its counterpart in the other branch of the pathway, CatR, but exhibits significant homology to its regulatory antecedent, PobR, which regulates the initial breakdown of p-hydroxybenzoate into protocatechuate. Comparisons of the pcaIJ and pcaR promoter regions revealed conservation of a 15-bp sequence centered around the -10 region in both sequences. This, together with previously defined deletional studies with the pcaIJ promoter region, suggests that PcaR exerts its regulatory effect through protein-DNA interactions within this region, which would be unusually close to the transcriptional start site of pcaIJ for a positive regulator.