Fig 1.

ateA is essential for A. phagocytophilum survival in tick cells, but dispensable within human cells. (A) A. phagocytophilum gene expression during growth within tick ISE6 and human HL60 cells. Transcription of ateA and housekeeping genes rpoB and groEL. (B) α-AteA (top) and α-VirD4 (bottom) western blot analysis of mock-infected and peak wild-type A. phagocytophilum infected human HL60 and tick ISE6 cells. Lanes were equally loaded with 1 × 10⁵ host cells. (C) Agarose gel electrophoresis of PCR amplicons flanking the transposon insertion site in the ateA gene. (D) Transcriptional analysis from A. phagocytophilum transposon mutants during culture with tick ISE6 cells. Mutants contain the transposon inserted in a neutral intergenic location (control), or within the ateA gene. Transcription measured by qRT-PCR of ateA, msp5, rpoB, and groEL. Transcription normalized to rpoB. Results shown are the mean of three biological replicates with two technical replicates each ±SD. *P < 0.05 (Student’s t-test). (E and F) Growth of A. phagocytophilum ateA or control strain in panel E human HL60 cells and (F) tick ISE6 cells. Bacterial burden was measured as Anaplasma gDNA vs host cell gDNA via qPCR. Data shown are the mean of three biological replicates with two technical replicates each ±SD and is representative of three experimental replicates. *P < 0.05 (Mann-Whitney t-test).