Next-generation sequencing as means to retrieve tick systematic markers, with the focus on Nuttalliella namaqua (Ixodoidea: Nuttalliellidae)

Abstract

Nuclear ribosomal RNA (18S and 28S rRNA) and mitochondrial genomes are commonly used in tick systematics. The ability to retrieve these markers using next-generation sequencing was investigated using the tick Nuttalliella namaqua. Issues related to nuclear markers may be resolved using this approach, notably, the monotypic status of N. namaqua and its basal relationship to other tick families. Four different Illumina datasets (∼55 million, 100bp reads each) were generated from a single tick specimen and assembled to give 350k–390k contigs. A genome size of ∼1Gbp was estimated with low levels of repetitive elements. Contigs (>1000bp, >50-fold coverage) present in most assemblies (n=69), included host-derived 18S and 28S rRNA, tick and host-derived transposable elements, full-length tick 18S and 28 rRNA, the mitochondrial genome in single contig assemblies and the histone cassette. Coverage for the nuclear rRNA genes was above 1000-fold confirming previous sequencing errors in the 18S rRNA gene, thereby maintaining the monotypic status of this tick. Nuclear markers for the soft tick Argas africolumbae were also retrieved from next-generation data. Phylogenetic analysis of a concatenated 18S–28S rRNA dataset supported the grouping of N. namaqua at the base of the tick tree and the two main tick families in separate clades. This study confirmed the monotypic status of N. namaqua and its basal relationship to other tick families. Next-generation sequencing of genomic material to retrieve high quality nuclear and mitochondrial systematic markers for ticks is viable and may resolve issues around conventional sequencing errors when comparing closely related tick species.