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Conventional influenza vaccines can prevent infection, but their efficacy depends on the degree of antigenic "match" between the strains used for vaccine preparation and those circulating in the population. A universal influenza vaccine based on invariant regions of the virus, able to provide broadly cross-reactive protection, without requiring continuous manufacturing update, would solve a major medical need. Since the temporal and geographical dominance of the influenza virus type and/or subtype (A/H3, A/H1, or B) cannot yet be predicted, a universal vaccine, like the vaccines currently in use, should include both type A and type B influenza virus components. However, while encouraging preclinical data are available for influenza A virus, no candidate universal vaccine is available for influenza B virus. We show here that a peptide conjugate vaccine, based on the highly conserved maturational cleavage site of the HA 0 precursor of the influenza B virus hemagglutinin, can elicit a protective immune response against lethal challenge with viruses belonging to either one of the representative, non-antigenically cross-reactive influenza B virus lineages. We demonstrate that protection by the HA 0 vaccine is mediated by antibodies, probably through effector mechanisms, and that a major part of the protective response targets the most conserved region of HA 0 , the P1 residue of the scissile bond and the fusion peptide domain. In addition, we present preliminary evidence that the approach can be extended to influenza A virus, although the equivalent HA 0 conjugate is not as efficacious as for influenza B virus.

Background: HCMV gH/gL/pUL128 -131 pentameric complex is a main target for potent neutralizing antibody responses. Results: Soluble gH/gL/pUL128 -131 inhibits viral entry and presents neutralizing epitopes. Conclusion:The biological activities of gH/gL/pUL128 -131 confirm the critical role of the native complex as an immunogen. Significance: The functional differences between gH/gL and gH/gL/pUL128 -131 must be considered when designing vaccines and assessing immune responses.

e Clostridium difficile strains producing binary toxin, in addition to toxin A (TcdA) and toxin B (TcdB), have been associated with more severe disease and increased recurrence of C. difficile infection in recent outbreaks. Binary toxin comprises two subunits (CDTa and CDTb) and catalyzes the ADP-ribosylation of globular actin (G-actin), which leads to the depolymerization of filamentous actin (F-actin) filaments. A robust assay is highly desirable for detecting the cytotoxic effect of the toxin and the presence of neutralizing antibodies in animal and human sera to evaluate vaccine efficacy. We describe here the optimization, using design-of-experiment (DOE) methodology, of a high-throughput assay to measure the toxin potency and neutralizing antibodies (NAb) against binary toxin. Vero cells were chosen from a panel of cells screened for sensitivity and specificity. We have successfully optimized the CDTa-to-CDTb molar ratio, toxin concentration, cell-seeding density, and sera-toxin preincubation time in the NAb assay using DOE methodology. This assay is robust, produces linear results across serial dilutions of hyperimmune serum, and can be used to quantify neutralizing antibodies in sera from hamsters and monkeys immunized with C. difficile binary toxin-containing vaccines. The assay will be useful for C. difficile diagnosis, for epidemiology studies, and for selecting and optimizing vaccine candidates. Clostridium difficile is a Gram-positive bacterium and the primary cause of hospital-acquired diarrhea. C. difficile infection (CDI) occurs when antibiotic treatment disrupts the normal bacterial flora of the intestine, which allows for the colonization of C. difficile bacteria. CDI is characterized by symptoms that range from mild diarrhea to severe and often life-threatening colitis. The incidence of more severe CDI has been increasing in recent years due to the emergence of epidemic hypervirulent strains (1, 2).Two glucosylating toxins, TcdA and TcdB, are considered the main virulence factors of C. difficile. These two toxins catalyze the glucosylation of proteins RhoA, Rac, and Cdc42, which leads to the disruption of actin filaments and collapse of the cell cytoskeleton, disruption of other signaling pathways, and eventually, cell death (3-5). In addition to TcdA and TcdB, hypervirulent strains have been found to express a binary toxin referred to as C. difficile transferase (CDT). Much of what we know of CDT was learned through a comparison to related binary toxins found in other clostridial species, such as Clostridium botulinum C2 toxin and Clostridium perfringens iota toxin (6-8). Binary toxins are characterized as having an enzymatically active "A" component that causes ADP-ribosylation of globular actin (G-actin) and a cell binding and translocation "B" component. For C. difficile, CDTa and CDTb are transcribed monocistronically but translated as two separate proteins. The proposed mechanism of action for the C. difficile binary toxin has been described in the literature (7). Briefly, the precursor C...