scRNAseq analysis of human brochial epithelial cell air liquid interface cultures infected with the A/Perth/16/2009 H3N2 with and without PA-X

The goal of this analysis was to examine the impact of PA-X activity during infection in a model of the human airway epithelium, using primary airway cells from two different donors. We profiled the effects of infection and PA-X activity in various airway epithelial cell types. Donor H1: Male, 21, Caucasian, no health issues noted. Donor H2: Male, 55, Caucasian, no health issues noted. Overall design: Air liquid interface culture models of the human airway epithelium were differentiated for 3-4 weeks post air lift. Then they were either mock-infected or infected at an MOI of 0.1 with wild type or PA-X deficient A/Perth/16/2009 H3N2. Basal media was replaced and apical surfaces were washed every 24 hours post infection. At 1 and 3 days post infection, cells were trypsinized and dead cells were depleted using an Annexin V binding kit (Stemcell). Viable single cell suspensions were collected and fixed using the Parse Evercode Fixation Kit v3. Library preparation was performed with Parse Evercode WT kit and sequenced on a NovaSeq X Plus 100 cycle 10B flow cell. About 69,000 cells were detected, with a depth of approximately 20,000 reads/cell.