A comparison of Illumina and PacBio methods to build tick salivary gland transcriptomes confirms large expression of lipocalins and other salivary protein families that are not represented in available tick genomes

Abstract

Tick saliva helps blood feeding by its antihemostatic and immunomodulatory activities. Tick salivary gland transcriptomes (sialotranscriptomes) revealed thousands of transcripts coding for putative secreted polypeptides. Hundreds of these transcripts code for groups of similar proteins, constituting protein families, such as the lipocalins and metalloproteases. However, while many of these transcriptome-derived protein sequences matches sequences predicted by tick genome assemblies, the majority are not represented in these proteomes. The diversity of these transcriptome-derived transcripts could derive from artifacts generated during assembly of short Illumina reads or derive from polymorphisms of the genes coding for these proteins. To investigate this discrepancy, we collected salivary glands from blood-feeding ticks and, from the same homogenate, made and sequenced libraries following Illumina and PacBio protocols, with the assumption that the longer PacBio reads would reveal the sequences generated by the assembly of Illumina reads. Using both Rhipicephalus zambeziensis and Ixodes scapularis ticks, we have obtained more lipocalin transcripts from the Illumina library than the PacBio library. To verify whether these unique Illumina transcripts were real, we selected 9 uniquely Illumina-derived lipocalin transcripts from I. scapularis and attempted to obtain PCR products. These were obtained and their sequences confirmed the presence of these transcripts in the I. scapularis salivary homogenate. We further compared the predicted salivary lipocalins and metalloproteases from I. scapularis sialotranscriptomes with those found in the predicted proteomes of 3 publicly available genomes of I. scapularis. Results indicate that the discrepancy between the genome and transcriptome sequences for these salivary protein families is due to a high degree of polymorphism within these genes.