Infection of swine ex vivo tissues with avian viruses including H7N9 and correlation with glycomic analysis.

OBJECTIVES: Swine have been regarded as intermediate hosts in the spread of influenza from birds to humans but studies of the sialylated glycans that comprise their respiratory tract have not been extensively studied in the past. This study analyzed the sialylated N-glycan and O-glycan profile of swine trachea and lung and correlated this with ex-vivo infection of swine explants with avian influenza viruses. SAMPLE: Lungs and tracheal samples were obtained from normal farm and laboratory raised swine and used for ex vivo infection as well as mass spectrometric analysis. Infection of the ex vivo tissues used high pathogenic and low pathogenic avian viruses including the novel H7N9 virus that emerged in China in early 2013. MAIN OUTCOME MEASURES: Assessment of successful replication was determined by TCID50 as well as virus immunohistochemistry. The N-glycan and O-glycan profiles were measured by MALDI-TOF and sialylated linkages were determined by sialidase treatment. Lectin binding histochemistry was also performed on formalin fixed tissue samples with positive binding detected by chromogen staining. RESULTS: The swine respiratory tract glycans differed from the human respiratory tact glycans in two main areas. There was a greater abundance of Gal-α-Gal linkages resulting in a relative decrease in sialylated glycans. The swine respiratory tract also had a greater proportion of glycans containing Neu5Gc and Siaα2-6 glycans than the human respiratory tract. Infection with avian viruses was confined primarily to lung bronchioles rather than trachea and parenchyma. CONCLUSIONS: In contrast to previous studies we found that there was not as much expression of Siaα2-3 glycans on the surface of the trachea. Infection of Siaα2-3 binding avian viruses was restricted to the lower respiratory tract bronchioles. This finding may diminish the ability of the swine to act as an intermediary in the transmission of avian viruses to humans.

Tissue tropism of swine influenza viruses and reassortants in ex vivo cultures of the human respiratory tract and conjunctiva.

The 2009 pandemic influenza H1N1 (H1N1pdm) virus was generated by reassortment of swine influenza viruses of different lineages. This was the first influenza pandemic to emerge in over 4 decades and the first to occur after the realization that influenza pandemics arise from influenza viruses of animals. In order to understand the biological determinants of pandemic emergence, it is relevant to compare the tropism of different lineages of swine influenza viruses and reassortants derived from them with that of 2009 pandemic H1N1 (H1N1pdm) and seasonal influenza H1N1 viruses in ex vivo cultures of the human nasopharynx, bronchus, alveoli, and conjunctiva. We hypothesized that virus which can transmit efficiently between humans replicated well in the human upper airways. As previously reported, H1N1pdm and seasonal H1N1 viruses replicated efficiently in the nasopharyngeal, bronchial, and alveolar epithelium. In contrast, representative viruses from the classical swine (CS) (H1N1) lineage could not infect human respiratory epithelium; Eurasian avian-like swine (EA) (H1N1) viruses only infected alveolar epithelium and North American triple-reassortant (TRIG) viruses only infected the bronchial epithelium albeit inefficiently. Interestingly, a naturally occurring triple-reassortant swine virus, A/SW/HK/915/04 (H1N2), with a matrix gene segment of EA swine derivation (i.e., differing from H1N1pdm only in lacking a neuraminidase [NA] gene of EA derivation) readily infected and replicated in human nasopharyngeal and bronchial epithelia but not in the lung. A recombinant sw915 with the NA from H1N1pdm retained its tropism for the bronchus and acquired additional replication competence for alveolar epithelium. In contrast to H1N1pdm, none of the swine viruses tested nor seasonal H1N1 had tropism in human conjunctiva. Recombinant viruses generated by swapping the surface proteins (hemagglutinin and NA) of H1N1pdm and seasonal H1N1 virus demonstrated that these two gene segments together are key determinants of conjunctival tropism. Overall, these findings suggest that ex vivo cultures of the human respiratory tract provide a useful biological model for assessing the human health risk of swine influenza viruses.