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Porphyromonas gingivalis is a key periodontal pathogen that has been implicated in the aetiology of chronic adult periodontitis. The aim of this study was to characterize two potential vaccine candidates (PG32 and PG33) identified from a previous genomic sequence analysis. Gene knockout studies suggested that these proteins play an important role in bacterial growth and are transcriptionally linked. Analysis of 14 laboratory and clinical isolates of P. gingivalis found that in all strains, both genes were present with a high level of conservation and that the two proteins were also expressed in vitro. Truncated recombinant PG32 and PG33 proteins were produced in Escherichia coli in an attempt to increase the solubility of the proteins while retaining their native conformation. While most of the truncated proteins remained insoluble, two truncated proteins showed good solubility and high levels of protection in the P. gingivalis murine lesion model and may be considered as potential vaccine candidates for further testing in models of human periodontal disease.

Influenza is a highly contagious, acute, febrile respiratory infection that can have fatal consequences particularly in individuals with chronic illnesses. Sporadic reports suggest that intravenous immunoglobulin (IVIg) may be efficacious in the influenza setting. We investigated the potential of human IVIg to ameliorate influenza infection in ferrets exposed to either the pandemic H1N1/09 virus (pH1N1) or highly pathogenic avian influenza (H5N1). IVIg administered at the time of influenza virus exposure led to a significant reduction in lung viral load following pH1N1 challenge. In the lethal H5N1 model, the majority of animals given IVIg survived challenge in a dose dependent manner. Protection was also afforded by purified F(ab′)2 but not Fc fragments derived from IVIg, supporting a specific antibody-mediated mechanism of protection. We conclude that pre-pandemic IVIg can modulate serious influenza infection-associated mortality and morbidity. IVIg could be useful prophylactically in the event of a pandemic to protect vulnerable population groups and in the critical care setting as a first stage intervention.

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Follicular helper T (TFH) cells control antibody responses by supporting antibody affinity maturation and memory formation. Inadequate TFH function has been found in individuals with ineffective responses to vaccines, but the mechanism underlying TFH regulation in vaccination is not understood. Here, we report that lower serum levels of the metabolic hormone leptin associate with reduced vaccine responses to influenza or hepatitis B virus vaccines in healthy populations. Leptin promotes mouse and human TFH differentiation and IL-21 production via STAT3 and mTOR pathways. Leptin receptor deficiency impairs TFH generation and antibody responses in immunisation and infection. Similarly, leptin deficiency induced by fasting reduces influenza vaccination-mediated protection for the subsequent infection challenge, which is mostly rescued by leptin replacement. Our results identify leptin as a regulator of TFH cell differentiation and function and indicate low levels of leptin as a risk factor for vaccine failure.

Please cite this paper as: Verity et al. (2011) Rapid generation of pandemic influenza virus vaccine candidate strains using synthetic DNA. Influenza and Other Respiratory Viruses DOI:10.1111/j.1750‐2659.2011.00273.x. Background  Vaccination is considered the most effective means of reducing influenza burden. The emergence of H5N1 and pandemic spread of novel H1N1/2009 viruses reinforces the need to have strategies in place to rapidly develop seed viruses for vaccine manufacture. Methods  Candidate pandemic vaccine strains consisting of the circulating strain haemagglutinin (HA) and neuraminidase (NA) in an A/PR/8/34 backbone were generated using alternative synthetic DNA approaches, including site‐directed mutagenesis of DNA encoding related virus strains, and rapid generation of virus using synthetic DNA cloned into plasmid vectors. Results  Firstly, synthetic A/Bar Headed Goose/Qinghai/1A/2005 (H5N1) virus was generated from an A/Vietnam/1194/2004 template using site‐directed mutagenesis. Secondly, A/Whooper Swan/Mongolia/244/2005 (H5N1) and A/California/04/09 (H1N1) viruses were generated using synthetic DNA encoding the viral HA and NA genes. Replication and antigenicity of the synthetic viruses were comparable to that of the corresponding non‐synthetic viruses. Conclusions  In the event of an influenza pandemic, the use of these approaches may significantly reduce the time required to generate and distribute the vaccine seed virus and vaccine manufacture. These approaches also offer the advantage of not needing to handle wild‐type virus, potentially diminishing biocontainment requirements.

In Australia, during the 2010 Southern Hemisphere (SH) influenza season, there was an unexpected increase in post-marketing adverse event reports of febrile seizures (FS) in children under 5 years of age shortly after vaccination with the CSL 2010 SH trivalent influenza vaccine (CSL 2010 SH TIV) compared to previous CSL TIVs and other licensed 2010 SH TIVs. In an accompanying study, we described the contribution to these adverse events of the 2010 SH influenza strains as expressed in the CSL 2010 SH TIV using in vitro cytokine/chemokine secretion from whole blood cells and induction of NF-κB activation in HEK293 reporter cells. The aim of the present study was to identify the root cause components that elicited the elevated cytokine/chemokine and NF-κB signature. Our studies demonstrated that the pyrogenic signal was associated with a heat-labile, viral-derived component(s) in the CSL 2010 SH TIV. Further, it was found that viral lipid-mediated delivery of short, fragmented viral RNA was the key trigger for the increased cytokine/chemokine secretion and NF-κB activation. It is likely that the FS reported in children <5 years were due to a combination of the new influenza strains included in the 2010 SH TIV and the CSL standard method of manufacture preserving strain-specific viral components of the new influenza strains (particularly B/Brisbane/60/2008 and to a lesser extent H1N1 A/California/07/2009). These combined to heighten immune activation of innate immune cells, which in a small proportion of children <5 years of age is associated with the occurrence of FS. The data also demonstrates that CSL TIVs formulated with increased levels of splitting agent (TDOC) for the B/Brisbane/60/2008 strain can attenuate the pro-inflammatory signals in vitro, identifying a potential path forward for generating a CSL TIV indicated for use in children <5 years.