Modified vaccinia virus Ankara expressing the hemagglutinin of pandemic (H1N1) 2009 virus induces cross-protective immunity against Eurasian 'avian-like' H1N1 swine viruses in mice

Abstract

Objectives: To examine cross-reactivity between hemagglutinin (HA) derived from A/California/7/09 (CA/09) virus and that derived from representative Eurasian "avian-like" (EA) H1N1 swine viruses isolated in Italy between 1999 and 2008 during virological surveillance in pigs.

Design: Modified vaccinia virus Ankara (MVA) expressing the HA gene of CA/09 virus (MVA-HA-CA/09) was used as a vaccine to investigate cross-protective immunity against H1N1 swine viruses in mice.

Sample: Two classical swine H1N1 (CS) viruses and four representative EA-like H1N1 swine viruses previously isolated during outbreaks of respiratory disease in pigs on farms in Northern Italy were used in this study.

Setting: Female C57BL/6 mice were vaccinated with MVA/HA/CA/09 and then challenged intranasally with H1N1 swine viruses.

Main outcome measures: Cross-reactive antibody responses were determined by hemagglutination- inhibition (HI) and virus microneutralizing (MN) assays of sera from MVA-vaccinated mice. The extent of protective immunity against infection with H1N1 swine viruses was determined by measuring lung viral load on days 2 and 4 post-challenge.

Results and conclusions: Systemic immunization of mice with CA/09-derived HA, vectored by MVA, elicited cross-protective immunity against recent EA-like swine viruses. This immune protection was related to the levels of cross-reactive HI antibodies in the sera of the immunized mice and was dependent on the similarity of the antigenic site Sa of H1 HAs. Our findings suggest that the herd immunity elicited in humans by the pandemic (H1N1) 2009 virus could limit the transmission of recent EA-like swine HA genes into the influenza A virus gene pool in humans.

Keywords: Avian-like; cross-protection; hemagglutinin; influenza; pandemic (H1N1) 2009; transmission.

Figure 1

Phylogenetic tree of the HA1 region (nucleotides 88–981) of the HA gene of the H1N1 swine influenza viruses used in this study (indicated in bold and underlined) as well as that of the swine and human influenza viruses in the database. The unrooted tree was generated with the mega4 program using the neighbor-joining method. Bootstrap values were calculated on 1000 replicates, and only values higher 70% are shown.

Figure 2

Effect of MVA-HA-CA/09 vaccination on the replication of H1N1 viruses in mouse lung. Groups of mice were vaccinated with two doses, 3 weeks apart, of 10^7·3 or 10^6·3 pfu of MVA-HA-CA/09. Control mice received PBS only on the same schedule. Four weeks later, the mice were challenged intranasally with 10⁵ TCID₅₀ of the indicated H1N1 viruses. Virus titers in the lungs of four mice per group sacrificed on days 2 and 4 post-infection are expressed as the mean ±SE log₁₀TCID₅₀/g of tissue. The lower detection limit is denoted by the dashed horizontal line. One of two similar experiments is shown. For virus abbreviations, see Table 1. *P < 0·05, **P < 0·001 compared with control mice by unpaired Student's t test.

Figure 3

Comparison of the HA antigenic sites. Amino acids sequences (H1 open reading frame numbering, including the signal peptide) of the CS viruses and the Italian swine viruses used in this study were aligned to the CA/09 HA sequence. Major antigenic domains, as defined by Brownlee and Fodor, are shown in boxes. For virus abbreviations, see Table 1.