Journal of Infection (2010) 61, 516e525

www.elsevierhealth.com/journals/jinf

First session: Chairs & discussants Professor DavidDockrell (Sheffield) & Professor Dietrich Mack(Swansea)

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PRESENCE OF IS1301 IN THE CAPSULEBIOSYNTHESIS LOCUS IN MENINGOCOCCALCARRIAGE AND DISEASE ISOLATES FROM UK

Kugelberg Elisabeth 1, Gollan Bridget 1,Farrance Christopher 1, Bratcher Holly 2,Lucidarme Jay 3, Ibarz-Pavon Ana Belen 2,Maiden Martin 2, Borrow Ray 3, Tang Christoph 1

1 Imperial College London, UK2University of Oxford, UK3Health Protection Agency, Manchester, UK

Introduction

The complement system is critical for immunity against thehuman pathogen Neisseria meningitidis. The current poly-saccharide conjugate vaccine against serogroup C N. menin-gitidis (MenC) has lead to a striking decrease in diseasecaused by this serogroup. We have previously identifiedMenC strains, isolated in Spain from invasive cases, whichavoid killing by bactericidal antibodies in vaccinees’ sera.These isolates were shown to have an insertion of IS1301 inthe intergenic region (IGR) between the operons necessaryfor capsule biosynthesis and export. Insertion of IS1301 leadsto increased amount of polysaccharide capsule and therebyincreased protection against complement-mediated killing.

Methods and results

PCR and sequencing was used to screen >1500 menin-gococcal carriage and disease isolates from UK for thepresence of IS1301 in the IGR. IS1301 was not found in theIGR of MenC vaccine failures but was frequent among se-rogroup B N. meningitidis (MenB) isolates. InterestinglyIS1301 found in UK MenB isolates all differed from IS1301

0163-4453/$36doi:10.1016/j.jinf.2010.09.005

previously found in MenC. IS1301 can be inserted in the op-posite orientation and is always associated with novel poly-morphisms. Human serum assays showed that IS1301 withnovel polymorphisms in the IGR did not mediate enhancedresistance to human serum.

Conclusions

MenC disease in UK vaccine failures is not caused by isolateswith IS1301 in the IGR. There is no significant difference forthe presence of IS1301 in the IGR of MenB disease isolatescompared to carriage isolates. IS1301 with associated novelpolymorphisms in the IGR of UK MenB isolates do not lead tothe resistance phenotype seen for IS1301 in the IGR of MenCisolates.

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THE VACCINE ANTIGEN FACTOR H BINDINGPROTEIN FROM NEISSERIA MENINGITIDIS CAN BEMODIFIED TO REDUCE BINDING TO FACTOR HWITH NO CHANGE IN IMMUNOGENICITY

Tan Lionel 1, Caesar Joe 2, Li Yanwen 1,Exley Rachel 1, Kugelberg Elisabeth 1, Zhang Qian 1,Yan Gabriel 1, Lea Susan 2, Tang Christoph 1

1Centre for Molecular Microbiology & Infection, ImperialCollege London, UK2 Sir William Dunn School of Pathology, University ofOxford, UK

Introduction

Neisseria meningitidis recruits the negative complementregulator, factor H (fH),to its surface via factor H binding

Abstracts 517

protein (fHbp),enabling it to evade the immune system andcause disease. fHbp is a component of two vaccines cur-rently in clinical trials. Binding of fH to fHbp may affectthe immunogenicity and reactogenicity of this novel vac-cine antigen. Cloaking of epitopes within the fH:fHbp bind-ing site may lead to reduced immunogenicity of fHbp andadditionally the fH:fHbp interaction may result in the de-velopment of adverse events. High affinity interactions be-tween fHbp and fH are dependent on two amino acids infHbp, E283 and E304.

Methods and results

We expressed and purified recombinant fHbp with definedamino acid changes (E283A/E304A) at these two criticalresidues. The modified fHbp and wild type fHbp were usedto immunise BALB/c mice. Far Western analysis and surfaceplasmon resonance were used to confirm that there issignificantly reduced fH binding to the modified fHbpcompared to the wild-type protein. However, modifiedfHbp retains immunogenicity, elicits high antibody titresand serum bactericidal activity.

Conclusions

The results indicate that fHbp can be successfully modifiedto reduce fH binding while not affecting the immunogenic-ity of this antigen. Modified fHbps are thus potentialantigens for inclusion in future vaccines against N. meningi-tidis.

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MEMORY B CELL RESPONSES FOLLOWINGIMMUNISATION WITH POLYSACCHARIDE ANDCONJUGATE QUADRIVALENT MENINGOCOCCALACYW135 VACCINES IN HEALTHYADULT VOLUNTEERS

Ramasamy Maheshi, Clutterbuck Elizabeth,Bowman Jaclyn, Snape MatthewD, Mpelembue Mushiya, Haworth Kathryn,Pollard Andrew JUniversity of Oxford, UK

Introduction

Neisseria meningitidis is a leading cause of meningitis andsepticaemia. The development of quadrivalent protein-poly-saccharide conjugate vaccines against serogroups A, C,W135&Yoffers the possibility of broader protection against the or-ganism across all age groups. We present results of a plannedinterim analysis of an ongoing open-label randomised clinicaltrial in healthy adult volunteers aged 18-70 comparing 2doses of a conjugate quadrivalent ACWY vaccine onemonth apart (Group 1) with one dose of a polysaccharide

quadrivalent ACWY vaccine (ACWYVax�) followed by onedose of a conjugate quadrivalent ACWY vaccine one monthlater (Group 2).

Methods and results

Sixty participants were immunised with two doses of vaccineat days 0 and 28. Meningococcal polysaccharide-specificmemory B cells were enumerated in peripheral blood atdays 0, 28 and 56 by cultured EliSpot assay. Median memorycell numbers were similar at baseline in the two groups(medians of 0.5-2.5 cells per million cultured lymphocytes).By both 28 and 56 days median memory cell numbers werehigher in Group 1 than in Group 2 (5.3, 8.5 2.5 and 2.8 permillion cultured lymphocytes for serogroups A, C, W135 & Yrespectively vs. 2.0, 2.5, 1.0 and 0.75 at day 56).

Conclusions

Two doses of conjugate vaccine generate larger memoryresponses than conjugate vaccine preceded by polysac-charide vaccine. This probably represents polysaccharidedriven hypo-responsiveness, but might indicate differ-ences in the magnitude or phenotype of cells respondingto the two different vaccines (follicular B cells to con-jugate vaccine in Group 1 mediating a T dependentresponse, compared to T independent responses to poly-saccharide antigens in Group 2, mediated by B1 or marginalzone B cells.) Notably, the serogroup A component of thevaccines behaves in the same way as other polysaccharidesdespite data indicating that it may act as a T-dependentantigen.

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MONOCYTE DEATH PROCESSES ARE SUBVERTEDBY MENINGOCOCCI

Webster SJ, Daigneault M, Marriott HM, Bewley MA,Read RC, Dockrell DHUniversity of Sheffield Medical School, UK

Monocytes, unlike differentiated macrophages, demon-strate rapid ingestion of bacteria but lack sustained anti-bacterial responses. We challenged highly pure primaryhuman monocytes with bacteria (Streptococcus pneumo-niae (Spn), Escherichia coli (Eco), Klebsiella pneumoniae(Kpn), Neisseria lactamica (Nla) and Neisseria meningitidis(Nme) and determined the kinetics and mechanisms ofcell death. Apoptosis was determined by flow cytometry orfluorescence microscopy, alternative death processesby electron microscopy or fluorescence microscopy andintracellular ATP estimated by chemiluminescence.All bacteria were rapidly phagocytosed and with theexception of Nme induced rapid loss of viability over4-12h. Spn and Nla induced classical apoptosis as did Ecoand Kpn when antibiotics limited bacterial numbers. In theabsence of antibiotics Eco and Kpn produced caspase-1activation, membrane permeabilisation and extracellular