Blue 49,6-Diamidino-2-phenylindole (DAPI) was used for nuclear staining. The number of ICC-staining positive cells in a 24-well plate was manually counted from five individual fields. Images of the immunostaining were captured using a confocal microscope (LSM510 Meta, Zeiss). For comparing the expression intensity of SA among CTE cells, the images were collected with the same setting and same objective (406) under the confocal microscope. Figure S1 The SAa2-3Gal expression of ciliated cells (btubulin + ) was characterized by the double-staining of biotin-labeled MAL-1 (Vector laboratories, 1:500). Scale bar, 50 mm. (EPS) Figure S2 Expression levels of SAa2-3Gal (labeled by MAA)(A) and SAa2-6Gal (labeled by SNA)(B) in tracheal basal cell (K14 + ) were shown by immunocytostaining. The arrows in A and B indicate cells with high-intensity MAA and SNA staining, respectively. The cell nuclei were stained with DAPI (blue). Scale bar, 50 mm. (EPS) Classification of viral zoonosis through receptor pattern analysis BACKGROUND: Viral zoonosis, the transmission of a virus from its primary vertebrate reservoir species to humans, requires ubiquitous cellular proteins known as receptor proteins. Zoonosis can occur not only through direct transmission from vertebrates to humans, but also through intermediate reservoirs or other environmental factors. Viruses can be categorized according to genotype (ssDNA, dsDNA, ssRNA and dsRNA viruses). Among them, the RNA viruses exhibit particularly high mutation rates and are especially problematic for this reason. Most zoonotic viruses are RNA viruses that change their envelope proteins to facilitate binding to various receptors of host species. In this study, we sought to predict zoonotic propensity through the analysis of receptor characteristics. We hypothesized that the major barrier to interspecies virus transmission is that receptor sequences vary among species--in other words, that the specific amino acid sequence of the receptor determines the ability of the viral envelope protein to attach to the cell. RESULTS: We analysed host-cell receptor sequences for their hydrophobicity/hydrophilicity characteristics. We then analysed these properties for similarities among receptors of different species and used a statistical discriminant analysis to predict the likelihood of transmission among species. CONCLUSIONS: This study is an attempt to predict zoonosis through simple computational analysis of receptor sequence differences. Our method may be useful in predicting the zoonotic potential of newly discovered viral strains. Viral zoonosis, the transmission of a virus from its primary vertebrate reservoir species to humans, requires ubiquitous cellular proteins known as receptor proteins [1] . Zoonosis can occur not only through direct transmission, but also through intermediate reservoirs or other environmental factors [2] [3] [4] . The zoonotic viruses can be categorized according to genotype; of the various classes of viruses, the RNA viruses exhibit the highest mutation rates [5] . Most zoonotic viruses are RNA viruses that change their envelope proteins to facilitate binding to various receptors of host species [6, 7] . The high mutation rate of envelope proteins [5] hinders the development of accurate vaccines, as does the great ability of the RNA viruses to infect host species in order to exploit host proteins for viral reproduction [8] .