Although primary HAE cultures support HBoV1 infection, their usefulness is limited by the variability between donors, tissue availability and high cost. We thus explored alternative cell culture models for their abilities to support HBoV1 infection. Using the purified HBoV1, we examined HEK293 cells, other common epithelial cell lines permissive to common respiratory viruses [42] , including HeLa, MDCK, MRC-5, LLC-MK2 and Vero-E6, and several transformed or immortalized human airway epithelial cell lines (A549, BEAS-2B, 16HBE14o- [43] , NuLi-1 and CuFi-8 [44] ), as well as primary NHBE cells for the ability to support infection in conventional monolayer culture. All were negative for HBoV1 infection as determined by IF analysis (data not shown). We next speculated that since some respiratory viruses infect polarized HAE but not undifferentiated cells [45] , some characteristics of the polarized epithelia may be critical for HBoV1 infection. We thus polarized immortalized cells (NuLi-1 and CuFi- 8) at an air-liquid interface (ALI) for one month. Once polarization was confirmed by detection of a TEER of .500 V.cm 2 , the cultures were infected with HBoV1, under the same conditions as used for primary B-HAE cultures. Notably, IF analysis revealed that at 10 days p.i., HBoV1-infected CuFi-HAE (differentiated from CuFi-8 cells) was uniformly positive for NS1 ( Figure 6A ), whereas the HBoV1-infected NuLi-HAE (differentiated from NuLi-1 cells) was not ( Figure S4) . Moreover, the CuFi-HAE did express HBoV1 NS1, NP1 and VP1/VP2 proteins (Figures 6B and  6C ). The kinetics of virus release from the apical surface was similar to that of a primary B-HAE infected with virus at a similar titer (maximally 2610 7 gc/ml), although virus release from the basolateral surface was undetectable ( Figure 6D ). HBoV1 infection also resulted in a decrease in the thickness of the epithelium ( Figure 6E ), and dissociation of the tight junction protein ZO-1 from the epithelial cell peripheries ( Figure 6F) .