Complete Mitogenomes of Ticks Ixodes acutitarsus and Ixodes ovatus Parasitizing Giant Panda: Deep Insights into the Comparative Mitogenomic and Phylogenetic Relationship of Ixodidae Species

Abstract

Ticks rank second in the world as vectors of disease. Tick infestation is one of the factors threatening the health and survival of giant pandas. Here, we describe the mitogenomes of Ixodes acutitarsus and Ixodes ovatus parasitizing giant pandas, and perform comparative and phylogenetic genomic analyses on the newly sequenced and other available mitogenomes of hard ticks. All six newly determined mitogenomes contain a typical gene component and share an ancient Arthropoda gene arrangement pattern. Our study suggests that I. ovatus is a species complex with high genetic divergence, indicating that different clades of I. ovatus represent distinct species. Comparative mitogenomic analyses show that the average A + T content of Ixodidae mitogenomes is 78.08%, their GC-skews are strongly negative, while AT-skews fluctuate around 0. A large number of microsatellites are detected in Ixodidae mitogenomes, and the main microsatellite motifs are mononucleotide A and trinucleotide AAT. We summarize five gene arrangement types, and identify the trnY-COX1-trnS1-COX2-trnK-ATP8-ATP6-COX3-trnG fragment is the most conserved region, whereas the region near the control region is the rearrangement hotspot in Ixodidae mitogenomes. The phylogenetic trees based on 15 genes provide a very convincing relationship (Ixodes + (Robertsicus + ((Bothriocroton + Haemaphysalis) + (Amblyomma + (Dermacentor + (Rhipicentor + (Hyalomma + Rhipicephalus))))))) with very strong supports. Remarkably, Archaeocroton sphenodonti is embedded in the Haemaphysalis clade with strong supports, resulting in paraphyly of the Haemaphysalis genus, so in-depth morphological and molecular studies are essential to determine the taxonomic status of A. sphenodonti and its closely related species. Our results provide new insights into the molecular phylogeny and evolution of hard ticks, as well as basic data for population genetics assessment and efficient surveillance and control for the giant panda-infesting ticks.

Keywords: Ixodidae; comparative mitogenomics; giant panda; mitochondrial phylogenomics; mitogenome.

Figure 1

Structural representation of the mitogenomes of I. ovatus and I. acutitarsus. The arrows represent the transcription direction of genes on heavy (outside) and light (inside) chain of the mitogenomes. The tRNAs are abbreviated by the one-letter code for the corresponding amino acid, and the anticodons are indicated in parentheses. The inner circle is the G + C content graph of mitogenome OP244856.

Figure 2

Nucleotide variation and diversity of I. ovatus group 4 and I. acutitarsus group 2 mitogenomes. (A) Variable nucleotide sites statistics and nucleotide diversity estimates of I. ovatus group 4 and I. acutitarsus group 2 mitochondrial genes. (B) Sliding window analysis of the complete mitogenome alignments between I. ovatus group 4 and I. acutitarsus group 2. I. ovatus group 4 include three mitogenomes with GenBank IDs OP244856–OP244858. I. acutitarsus group 2 include four mitogenomes with GenBank IDs OL800704, OP244859–OP244861.

Figure 3

Nucleotide content and skewness of Ixodidae mitogenomes. (A) Intergenus comparison of A+T content of Ixodidae mitogenomes. (B) Intergenus comparison of AT-skew and GC skew of Ixodidae mitogenomes.

Figure 4

Nucleotide content and skewness of Ixodidae mitochondrial PCGs and rRNAs. (A) A+T content of Ixodidae mitochondrial genes. (B) AT-skew of Ixodidae mitochondrial genes. (C) GC-skew of Ixodidae mitochondrial genes.

Figure 5

Statistics of microsatellite identified in Ixodidae mitogenomes. (A) The microsatellite loci counts and (B) their relative abundances of I. acutitarsus and I. ovatus species complex mitogenomes. Ia1–5 represent I. acutitarsus species complex mitogenomes with GenBank IDs OL800704, OM368264, and OP244859–OP244861; Io1–7 represent I. ovatus species complex mitogenomes with GenBank IDs OM317739, OM368266, OM368268, OM366269, and OP244856–OP244858. (C) The counts statistics of different microsatellite motif in Ixodidae and Ixodes mitogenomes.

Figure 6

Mitochondrial gene arrangements of Ixodidae species. (A) The five mitochondrial gene arrangement types in Ixodidae group. (B) The rearrangement score (RS) of every gene arrangement type in Ixodidae group. (C) The rearrangement frequency (RF) of different mitochondrial genes or region in Ixodidae group. The tRNAs are abbreviated by the one-letter code for the corresponding amino acid, trnL1 = tRNA-Leu(CUN), trnL2 = tRNA-Leu(UUR), trnS1 = tRNA-Ser(AGN), and trnS2 = tRNA-Ser(UCN).

Figure 7

ML tree of Ixodidae species inferred from the concatenated DNA sequences of 15 mitochondrial genes. A. africolumbae and N. namaqua act as the outgroup. Ultrafast bootstrap percentage (UBP) is given at each node. The different colors of the outer ring represent different gene arrangement types in Figure 6.