FIG 4.

Knockdown of ispdiA3 expression decreases B. burgdorferi colonization by I. scapularis ticks. mRNA transcription was knocked down in I. scapularis salivary glands or gut by body microinjection (A to D) or by anal pore injection (E to H) of ds ispdia3. Knockdown efficiency was measured in I. scapularis nymph salivary glands (SG) and gut (gut). Each data point represents a pool of three nymphal salivary glands or guts (A and E). The impact of ispdia3 knockdown on tick feeding was measured by engorgement weights. Each data point represents a replete nymph (B and F). B. burgdorferi burden in replete tick guts (C and G) was determined by qRT-PCR of flaB transcripts, and data were normalized to tick actin and visualized by immunofluorescence microscopy using rabbit polyclonal B. burgdorferi antibodies. Each data point represents a pool of three nymphal guts (D and H). Nuclei stained with 4′,6-diamidino-2-phenylindole (DAPI). Magnification is ×40. Data in panels A to H represent averages of 3 biological replicates. Results represent mean ± SD of values. Statistical significance was assessed using a nonparametric Mann-Whitney test; *, P < 0.05; **, P < 0.01; ***, P < 0.001.