Predomination shift of different P44-expressing Anaplasma phagocytophilum in infected HL-60, THP-1, NB4, and RF/6A cell lines.

Anaplasma phagocytophilum, an agent of human granulocytic anaplasmosis, is an obligatory intracellular bacterium and dominantly produces the variety of P44 major outer membrane proteins encoded by a p44/msp2 multigene family, that is responsible as major antigens for serodiagnosis. However, A. phagocytophilum antigens prepared from cultures with different cell lines seems to have various reactivities with patients' sera. In this study, we performed RNA-seq to investigate P44 expression of A. phagocytophilum propagated in 4 different cell lines. In infected HL-60 cells, P44-2b transcript was predominantly detected in first RNA-seq analysis (HL-60.1), but P44-23 transcript became predominated in second RNA-seq analysis at 1 month after additional passages (HL-60.2). We further analyzed P44 expression of A. phagocytophilum cultured in THP-1, NB4, and RF/6A cells through consecutively-repeated passages with own cell line for 1 year after transfer of A. phagocytophilum from infected HL-60 to respective cell lines. In those long-term cultures, P44-18, P44-78, and P44-51 became predominantly transcribed in infected THP-1, NB4, and RF/6A cells, respectively. Thus, the predomination shifts of different P44-expressing A. phagocytophilum might occur during cell cultures, probably due to host cell adaptation/selection/interaction, even in the same cell line at the different time points of sample harvest (HL-60.1 and HL-60.2).