Subolesin expression in D. variabilis male ticks infected with different pathogens by capillary feeding (CF). Subolesin expression levels were compared between ticks injected with control Rs86 dsRNA and then fed pathogen-infected or plain media by CF (N = 27-29). Whole individual ticks were dissected and used for DNA/RNA extraction to determine pathogen infection levels by real-time PCR and subolesin mRNA levels by real-time RT-PCR after normalization against tick 16S rRNA using the comparative Ct method [9, 32]. The graph depicts the infected to uninfected subolesin mRNA ratio (± SD) calculated by dividing normalized subolesin mRNA level in infected ticks by the average of the normalized subolesin mRNA level in uninfected control ticks. Normalized subolesin mRNA levels were compared between infected and uninfected ticks by Student's t-Test (*P < 0.05). Regression analyses were conducted in Microsoft Excel to compare normalized pathogen infection levels and subolesin mRNA levels. Regression coefficients are shown for all groups. The correlation graph is shown in the insert for F. tularensis, the only group in which a positive correlation was found between subolesin expression and pathogen infection levels.