Extended Data Fig. 2: Ash2 immunization and roles in tick biology.

a, Ash2 immunization does not affect tick feeding. The upper left panel shows the percentage of ticks collected after 96 and 120 h of feeding after placement on phosphate-buffered saline (PBS)- or Ash2-immunized mice; the upper right panel shows the total percentage of engorged ticks collected after the completion of feeding on PBS- and Ash2-immunized mice. Error bars denote the mean with SEM of three biological replicates, with 25 ticks per group (two-tailed Mann–Whitney U-test). The lower left panel denotes the weights of fully engorged ticks (mg). Each data point represents a single tick collected (n = 75) from three biological replicates; two-tailed Mann–Whitney U-test. The lower right panel shows the percentage of fully engorged nymphs that molted into adult ticks after feeding on PBS-immunized and Ash2-immunized mice. Error bars denote the mean with SEM (n = 4 biological replicates, each with 25 ticks per group); two-tailed Mann–Whitney U-test. b, Comparison of the amino acid sequences of the SPRY domains (left panel) and ZFD domains (right panel) from human, I. scapularis, D. melanogaster, A. gambiae, and A. aegypti Ash2 proteins. The human has three Ash2 protein isoforms, while the tick, Drosophila, A. gambiae, and A. aegypti each have two Ash2 isoforms. Human Ash2 isoforms do not contain any ZFDs. c, Staining of dsGFP and dsash2 groups representing 8 h-fed and 48 h-fed tick guts with anti-PH3 antibody (arrow) and DAPI. The insets show zoomed-in pictures of the tick gut cells. The PH3 positive nuclei are less apparent in Ash2-deficient tick gut. The experiment was repeated independently three times with similar results. White bar = 20 µm.