Analysis of five Amblyomma americanum putative immune genes reveals differential expression patterns during gram-positive and gram-negative bacterial infections

Hard tick species like Amblyomma americanum transmit numerous pathogens of medical and veterinary importance. For successful transmission, these microbes must overcome the tick immune system. While an increasing number of studies have contributed to the emerging picture of what these immune pathways involve, many details remain unresolved and few specific genes have been experimentally implicated in these processes. In this study, we evaluated four genes for potential involvement in the A. americanum tick immune system, tick a2-macroglobulin (AamTa2M), limb expression 1 (AamLIX), leucine-rich repeat-containing protein (AamLRR), and uridine diphosphate-galactose transporter senju-like (AamUDP-galT). We confirmed the previously NGS-obtained sequence information using Sanger sequencing, and bioinformatically analyzed the transcripts for a putative annotation. Sequences in A. americanum and other ticks were compared to resolve conserved and divergent sequence regions. Additionally, we infected female ticks with either gram negative (E. coli) or gram+ (S. capitis) bacteria and sampled individuals at 3-, 6-, 12-, 24-, and 48 h post-infection time points and analyzed expression of each gene at uninfected and infected time points. For all four genes, conservation across tick species at both nucleotide and amino acid levels was above 70%. Expression of AamTα2M and AamLRR was abundant before and during both infection types while AamLIX and AamUDP-galT expression profiles were variable or undetected during gram-negative and gram+ infections, respectively. Potential functional roles for these genes are discussed. Data in this study contribute to the prioritization of genes that may be further pursued for evidence of tick immune system involvement.