Phase I safety and immunogenicity study of a candidate meningococcal disease vaccine based on Neisseria lactamica outer membrane vesicles.

Natural immunity to meningococcal disease in young children is associated epidemiologically with carriage of commensal Neisseria species, including Neisseria lactamica. We have previously demonstrated that outer membrane vesicles (OMVs) from N. lactamica provide protection against lethal challenge in a mouse model of meningococcal septicemia. We evaluated the safety and immunogenicity of an N. lactamica OMV vaccine in a phase I placebo-controlled, double-blinded clinical trial. Ninety-seven healthy young adult male volunteers were randomized to receive three doses of either an OMV vaccine or an Alhydrogel control. Subsequently, some subjects who had received the OMV vaccine also received a fourth dose of OMV vaccine, 6 months after the third dose. Injection site reactions were more frequent in the OMV-receiving group, but all reactions were mild or moderate in intensity. The OMV vaccine was immunogenic, eliciting rises in titers of immunoglobulin G (IgG) against the vaccine OMVs, together with a significant booster response, as determined by an enzyme-linked immunosorbent assay. Additionally, the vaccine induced modest cross-reactive immunity to six diverse strains of serogroup B Neisseria meningitidis, including IgG against meningococcal OMVs, serum bactericidal antibodies, and opsonophagocytic activity. The percentages of subjects showing > or =4-fold rises in bactericidal antibody titer obtained were similar to those previously reported for the Norwegian meningococcal OMV vaccine against the same heterologous meningococcal strain panel. In conclusion, this N. lactamica OMV vaccine is safe and induces a weak but broad humoral immune response to N. meningitidis.

A Neisseria meningitidis NMB1966 mutant is impaired for invasion of respiratory epithelial cells, survival in human blood and for virulence in vivo.

We sought to determine whether NMB1966, encoding a putative ABC transporter, has a role in pathogenesis. Compared to its isogenic wild-type parent strain Neisseria meningitidis MC58, the NMB1966 knockout mutant was less adhesive and invasive for human bronchial epithelial cells, had reduced survival in human blood and was attenuated in a systemic mouse model of infection. The transcriptome of the wild-type and the NMB1966 mutant was compared. The data are consistent with a previous functional assignment of NMB1966 being the ABC transporter component of a glutamate transporter operon. Forty-seven percent of all the differentially regulated genes encoded known outer membrane proteins or pathways generating complex surface structures such as adhesins, peptidoglycan and capsule. The data show that NMB1966 has a role in virulence and that remodelling of the outer membrane and surface/structures is associated with attenuation of the NMB1966 mutant.

Characterization of the key antigenic components and pre-clinical immune responses to a meningococcal disease vaccine based on Neisseria lactamica outer membrane vesicles.

Serogroup B strains are now responsible for over 80% of meningococcal disease in the UK and no suitable vaccine is available that confers universal protection against all serogroup B strains. Neisseria lactamica shares many antigens with the meningococcus, except capsule and the surface protein PorA. Many of these antigens are thought to be responsible for providing cross-protective immunity to meningococcal disease. We have developed an N. lactamica vaccine using methods developed for meningococcal outer membrane vesicle (OMV) vaccines. The major antigenic components were identified by excision of 11 major protein bands from an SDS-PAGE gel, followed by mass spectrometric identification. These bands contained at least 22 proteins identified from an unassembled N. lactamica genome, 15 of which having orthologues in published pathogenic Neisseria genomes. Western blotting revealed that most of these bands were immunogenic, and antibodies to these proteins generally cross-reacted with N. meningitidis proteins. Sera from mice and rabbits immunized with either N. lactamica or N. meningitidis OMVs produced comparable cross-reactive ELISA titres against OMVs prepared from a panel of diverse meningococcal strains. Mice immunized with either N. meningitidis or N. lactamica OMVs showed no detectable serum bactericidal activity against the panel of target strains except N. meningitidis OMV sera against the homologous strain. Similarly, rabbit antisera to N. lactamica OMVs elicited little or no bactericidal antibodies against the panel of serogroup B meningococcal strains. However, such antisera did mediate opsonophagocytosis, suggestingthat this may did mediate opsonophagocytosis, suggesting that this may be a mechanism by which this vaccine protects in a mouse model of meningococcal bacteraemia.

Effects of low doses of lactitol on faecal microflora, pH, short chain fatty acids and gastrointestinal symptomology.

BACKGROUND: Lactitol (4-beta-D: -galactopyranosyl-D: -glucitol) is a sugar alcohol used as a sweetener. Previous studies have shown that it has a beneficial effect on intestinal microflora. AIMS OF THE STUDY: To determine whether low doses of lactitol had beneficial effects without eliciting adverse gastrointestinal symptoms. METHODS: Faecal bacterial populations (total anaerobes, total aerobes, enterobacteria, bifidobacteria and lactobacilli), faecal pH and faecal short chain fatty acids (SCFA) were studied in a randomized longitudinal study of 75 non-adapted healthy adults before and after consumption of low doses of lactitol. Subjects consumed 25 g tablets of milk chocolate containing 10 g sweetener as sucrose:lactitol in ratios of 10:0, 5:5 or 0:10 daily for 7 d. RESULTS: No significant changes in faecal bacterial counts occurred in the 10:0 or 5:5 sucrose:lactitol groups. There were no significant changes in faecal anaerobes, aerobes, Enterobacteriaceae or lactobacilli during the study period in subjects consuming 0:10 sucrose:lactitol but there was a significant increase (P = 0.017) in bifidobacteria. There were no significant changes in faecal pH and SCFA for the 10:0 or 5:5 sucrose:lactitol groups but a significant decrease (P = 0.02) in faecal pH and significant increases (P = 0.001) in concentrations of propionic and butyric acids were observed in the 0:10 sucrose:lactitol group. There were few adverse symptoms of gastrointestinal intolerance to the daily consumption of 10 g lactitol. CONCLUSIONS: The results show that low doses of lactitol can beneficially affect the faecal flora without eliciting gross symptoms of intolerance and that lactitol can be classified as a prebiotic.

The development of a meningococcal disease vaccine based on Neisseria lactamica outer membrane vesicles.

Serogroup B meningococcal disease remains a serious problem in many countries and no effective vaccine is currently available. Immunological and epidemiological evidence suggests that carriage of commensal Neisseria species is involved in the development of natural immunity against meningococcal disease. Neisseria lactamica has many surface structures in common with Neisseria meningitidis and may be the most important of these species. We have produced extensive pre-clinical data, which indicate that N. lactamica outer membrane vesicles (OMVs) may provide a vaccine effective against diverse disease-causing meningococcal strains. Immunisation with N. lactamica OMVs protected against lethal challenge with diverse meningococcal isolates in a mouse intraperitoneal challenge model of meningococcal disease and we are developing this vaccine for use in a phase I safety and immunogenicity study in adult volunteers. We have shown that OMVs produced from bacteria grown under iron-limited or iron-rich conditions provide equivalent protection in the mouse infection model and thus OMVs produced from iron-rich will be used. Sterile filtration of N. lactamica OMVs has proved difficult but this has been improved by resuspending the vesicles in a buffer, which increases their surface zeta potential. The vaccine is currently being manufactured and validated ELISA protocols have been developed for the analysis of serological responses.

Antimicrobial effects of a novel Triclosan/zinc citrate dentifrice against mixed culture oral biofilms.

AIM: To compare the antimicrobial efficacy in vitro of a new fluoride system containing 0.3% Triclosan and 2.0% zinc citrate with a standard fluoride toothpaste against bacteria commonly isolated from dental plaque. METHOD: Mature biofilms of fixed depth containing ten oral bacterial species were developed in a model system and then exposed to the formulations both in situ and ex situ. RESULTS: The standard fluoride toothpaste had limited activity against the Gram positive species following 1 and 5 minutes ex situ exposure (p = 0.004 and p = 0.02, respectively) but none against the Gram negative species; also, no overall effect following repeated pulsing in situ was observed. In contrast, viable counts of Gram positive and Gram negative species were markedly reduced following ex situ exposure for 1 and 5 minutes to the new formulation (Gram positive: p = 0.0007 and p = 0.04; Gram negative: p = 0.04 and p = 0.00001, respectively). Counts of Gram positive bacteria were reduced by 99.9% over a 4-day in situ pulsing period, while no Gram negative species were recoverable from the biofilm by day 4. CONCLUSION: The new toothpaste formulation had a broad spectrum of antimicrobial activity against biofilm consortia containing bacteria associated with both dental caries and periodontal diseases.