Hemagglutinin Of Pandemic Influenza Research Articles

Antigenic Site Variation in the Hemagglutinin of Pandemic Influenza A(H1N1)pdm09 Viruses between 2009-2017 in Ukraine.

The hemagglutinin (HA) is a major influenza virus antigen, which, once recognized by antibodies and substitutions in HA genes, helps virus in escaping the human immune response. It is therefore critical to perform genetic and phylogenetic analysis of HA in circulating influenza viruses. We performed phylogenetic and genetic analysis of isolates from Ukraine, the vaccine strain and reference strains were used to phylogenetically identify trends in mutation locations and substitutions. Ukrainian isolates were collected between 2009–2017 and clustered in the influenza genetic groups 2, 6, 7, and 8. Genetic changes were observed in each of the antigenic sites: Sa – S162T, K163Q, K163I; Sb – S185T, A186T, S190G, S190R; Ca1 – S203T, R205K, E235V, E235D, S236P; Ca2 – P137H, H138R, A141T, D222G, D222N; Cb – A73S, S74R, S74N. In spite of detected mutations in antigenic sites, Ukrainian isolates retained similarity to the vaccine strain A/California/07/09 circulated during 2009–2017. However, WHO recommended a new vaccine strain A/Michigan/45/2015 for the Southern Hemisphere after the emergence of the new genetic groups 6B.1 and 6B.2. Our study demonstrated genetic variability of HA protein of A(H1N1)pdm09 viruses isolated in 2009–2017 in Ukraine. Influenza surveillance is very important for understanding epidemiological situations.

Immune Correlates of Protection Induced by Virus-Like Particles Containing 2009 H1N1 Pandemic Influenza HA, NA or M1 Proteins

ABSTRACTInfluenza virus-like particle (VLPs) vaccines are a promising alternative to conventional egg-based vaccines. Evaluation of vaccine efficacy induced by HA-M1 VLPs, NA-M1 VLPs or M1 VLPs against virus challenge infection would provide important insight into vaccine design strategy. In this study, we generated VLPs containing hemagglutinin (HA), neuraminidase (NA) or M1 proteins derived from the A/California/04/09. Mice were immunized intramuscularly with HA-M1, NA-M1 or M1 VLPs and protective immunity was evaluated by assessing lung virus loads against low (5LD50) or high (100LD50) lethal dose of homologous virus challenges. High levels of virus-specific serum IgG antibody responses were induced in mice after HA-M1 VLPs immunization, whereas low or no IgG antibody responses were detected from immunization with NA-M1 VLPs or M1 VLPs, independently. Mice that were immunized with HA-M1 VLPs showed below the limit of detection on lung virus loads against low dose (5LD50) of challenge and significant reduction against high dose (100LD50) of challenge infection. Mice that were immunized with NA-M1 or M1 VLPs also displayed reduced lung viral loads compared to naïve control. In vitro cultures of cells from mouse spleen and bone marrow revealed that HA-M1 VLPs and NA-M1 VLPs induced higher levels of antibody-secreting cell (ASC) responses compared to naïve control, whereas M1 VLPs showed no ASC responses. HA-M1, NA-M1 or M1 VLPs immunization demonstrated varying degree of protection with respect to body weight changes and survival rates, which are consistent with the levels of antibody responses in sera and ASC responses from spleen and bone marrow.

Influence of copy number on the expression levels of pandemic influenza hemagglutinin recombinant protein in methylotrophic yeast Pichia pastoris

The hemagglutinin (HA) gene of novel Swine Origin Influenza A/California/04/2009 (H1N1) was engineered for expression in Pichia pastoris as a soluble secreted protein. The full length HA-synthetic gene having α secretory tag under the control of AOX1 promoter was integrated into P. pastoris genome through homologous recombination. The resultant Pichia clones having single and multiple copy integrants of the expression cassettes were screened for the expression of full length HA protein in the culture supernatant. In order to completely exploit the expression potential of the P. pastoris expression system, a systematic investigation on the influence of gene copy number on the expression of the recombinant protein was made. A panel of Pichia clones carrying increasing copies of the heterologous gene was selected based on Geneticin resistance and SYBR green-based quantitative real-time PCR approach. Using these strategies, recombinant Pichia transformants carrying up to a maximum of four to six copies of the transgene were identified. After optimising the expression conditions for shaker flask culture, the resultant clones demonstrated that the increase in copy number results in a proportional elevation in the expression level of H1N1HA recombinant protein. Our findings clearly suggest that the gene dosage effect play a vital role in high level expression of the pandemic Influenza HA protein in yeast system.

Quasispecies of the D225G Substitution in the Hemagglutinin of Pandemic Influenza A(H1N1) 2009 Virus from Patients with Severe Disease in Hong Kong, China

The D225G (aspartic acid to glycine) substitution in the hemagglutinin of H1N1 influenza virus may alter its receptor-binding specificity. Direct analysis of polymorphisms in 126 amino acids spanning the receptor-binding site in the hemagglutinin of pandemic H1N1 2009 virus from 117 clinical specimens in Hong Kong found the D225G substitution for 7 (12.5%) of 57 patients with severe disease and for 0 (0%) of 60 patients with mild disease. D225G quasispecies were identified mainly in endotracheal aspirate samples and were identified less frequently in nasopharyngeal aspirate samples from patients with severe disease. Continuous monitoring of the prevalence and tissue tropism of this variant during its circulation among humans is important.