Research Article |
Corresponding author: Zhigang Hu ( zghu0608@163.com ) Corresponding author: Yifei Liu ( liuyifei@hbtcm.edu.cn ) Academic editor: Pavel Stoev
© 2020 Chaoyi Hu, Shuaibin Wang, Bisheng Huang, Hegang Liu, Lei Xu, Zhigang Hu, Yifei Liu.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Hu C, Wang S, Huang B, Liu H, Xu L, Hu Z, Liu Y (2020) The complete mitochondrial genome sequence of Scolopendra mutilans L. Koch, 1878 (Scolopendromorpha, Scolopendridae), with a comparative analysis of other centipede genomes. ZooKeys 925: 73-88. https://doi.org/10.3897/zookeys.925.47820
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Scolopendra mutilans L. Koch, 1878 is an important Chinese animal with thousands of years of medicinal history. However, the genomic information of this species is limited, which hinders its further application. Here, the complete mitochondrial genome (mitogenome) of S. mutilans was sequenced and assembled by next-generation sequencing. The genome is 15,011 bp in length, consisting of 13 protein-coding genes (PCGs), 14 tRNA genes, and two rRNA genes. Most PCGs start with the ATN initiation codon, and all PCGs have the conventional stop codons TAA and TAG. The S. mutilans mitogenome revealed nine simple sequence repeats (SSRs), and an obviously lower GC content compared with other seven centipede mitogenomes previously sequenced. After analysis of homologous regions between the eight centipede mitogenomes, the S. mutilans mitogenome further showed clear genomic rearrangements. The phylogenetic analysis of eight centipedes using 13 conserved PCG genes was finally performed. The phylogenetic reconstructions showed Scutigeromorpha as a separate group, and Scolopendromorpha in a sister-group relationship with Lithobiomorpha and Geophilomorpha. Collectively, the S. mutilans mitogenome provided new genomic resources, which will improve its medicinal research and applications in the future.
Chilopoda, Chinese medicinal materials, mitogenome, Scolopendra mutilans
Animal medicine is an important part of the Chinese traditional medicine system. As a typical representative of medicinal animals, the centipede Scolopendra mutilans has been used for hundreds of years in China for treating many disorders, such as stroke-induced hemiplegia, epilepsy, apoplexy, whooping cough, tetanus, burns, tuberculosis, and myocutaneous disease (
Centipedes (Chilopoda) are one of the oldest extant terrestrial arthropods. Approximately 3300 centipede species have been described (
Previously, phylogenetic analysis on the basis of different molecular data provided support to these morphological classifications to some degree (
The mitochondrial genome (mitogenome), including those markers derived from it as well as the whole mitogenome, is the most commonly used molecule in animal studies with relation to taxonomy, population genetics, and evolutionary biology (
Scolopendra mutilans samples were collected in August 2018 from the wild in Yichang, Hubei Province, China. The specimens used in this study were preserved in 100% ethanol and stored at -20 °C. Genomic DNA was extracted from locomotory legs by Column mtDNAout kit (Tiangen Biotech Co., China) according to the instructions and stored at -20 °C until used for sequencing. The DNA quality was measured by gel electrophoresis and the concentration was estimated using the Nanodrop ND-1000.
Whole genome sequencing was performed on an Illumina HiSeq 2500 platform (Illumina, San Diego, CA, USA). Quality control and de novo assembly of the S. mutilans mitogenome were conducted based on previously described methods (
The mitogenomes were annotated by combining results from both MFannot and MITOS (
In order to identify interspersed repeats or intra-genomic duplications of large fragments throughout the mitogenomes, we performed BLASTn searches of the mitogenome against itself using an E-value of 1e-10. Tandem repeats within the mitogenome were detected by MicroSAtellite (MISA) (
The base composition of the mitogenome was determined using the DNAStar Lasergene package v7.1 (
A maximum likelihood (ML) tree was constructed using the RAxML (
Analysis of selective pressures was performed for 13 PCGs of eight centipedes using the codeml program in PAML (University College London, London, UK) (
The full circular mitogenome of S. mutilans (GenBank: MN317390) was 15,011 bp in length, which was similar to those of seven other centipede mitogenomes sequenced in the class Chilopoda (Table
Mitochondrial genome map of the Scolopendra mutilans. Genes drawn inside the circle are transcribed clockwise, and those outside are counterclockwise. PCGs are shown as brown arrows, rRNA genes as green arrows, tRNA genes as pink arrows. The innermost circle shows the GC content. GC content is plotted as the deviation from the average value of the entire sequence.
The mitogenome size of eight centipedes ranged from 14,538 bp for S. dehaani Brandt, 1840 to 16,833 bp for Cermatobius longicornis Takakuwa,1939, with that of S. mutilans in the middle of the range. To identify the specific variation contributing most to the diversity of the mitogenome size in centipedes, the length variation of all PCGs, tRNA, and rRNA genes, and intergenic regions in each mitogenome was investigated. Comparatively, the length of most genes across centipede species was relatively stable except the PCGs in L. forficatus Linnaeus,1758 (AF309492.1), while the length of intergenic regions was the primary contributor to mitogenome size variation.
Species | Order | NCBI ID | Length (bp) |
---|---|---|---|
Scolopendra mutilans L. Koch, 1878 | Scolopendromorpha | MN317390 | 15011 |
Scolopendra dehaani Brandt, 1840 | Scolopendromorpha | KY947341.1 | 14538 |
Scolopocryptops sp. | Scolopendromorpha | KC200076.1 | 15119 |
Strigamia maritima (Leach, 1817) | Geophilomorpha | KP173664.1 | 14983 |
Cermatobius longicornis Takakuwa,1939 | Lithobiomorpha | NC_021403.1 | 16833 |
Bothropolys sp. | Lithobiomorpha | AY691655.1 | 15139 |
Lithobius forficatus (Linnaeus,1758) | Lithobiomorpha | AF309492.1 | 15695 |
Scutigera coleoptrata (Linnaeus, 1758) | Scutigeromorpha | AJ507061.2 | 14922 |
Gene | Start | End | Strand | Length | Start/End codon |
---|---|---|---|---|---|
trnI (gat) | 1 | 65 | + | 65 | – |
trnM (cat) | 69 | 141 | + | 73 | – |
nad2 | 124 | 954 | + | 831 | ATT/TAA |
trnW (tca) | 1071 | 1119 | + | 49 | – |
cox1 | 1148 | 2656 | + | 1509 | ATG/TAG |
cox2 | 2676 | 3341 | + | 666 | ATG/TAA |
trnK (ctt) | 3355 | 3405 | + | 51 | – |
trnD (gtc) | 3425 | 3458 | + | 34 | – |
atp8 | 3465 | 3614 | + | 150 | ATA/TAA |
atp6 | 3620 | 4267 | + | 648 | ATG/TAA |
cox3 | 4282 | 5049 | + | 768 | AYG/TAA |
trnG (tcc) | 5084 | 5137 | + | 54 | – |
nad3 | 5141 | 5488 | + | 348 | ATT/TAG |
trnA (tgc) | 5487 | 5542 | + | 56 | – |
trnS1 (gct) | 5612 | 5662 | + | 51 | – |
nad1 | 5784 | 6635 | - | 851 | ATT/TAA |
rrnL | 6719 | 7937 | - | 1219 | – |
rrnS | 7993 | 8740 | - | 748 | – |
trnQ (ttg) | 9667 | 9720 | - | 54 | – |
trnF (gaa) | 9735 | 9791 | - | 57 | – |
nad5 | 9906 | 11,474 | - | 1569 | ATT/TAA |
trnH (gtg) | 11,556 | 11,619 | - | 64 | – |
nad4 | 11,641 | 12,789 | - | 1149 | ATG/TAA |
nad4l | 12,939 | 13,181 | - | 243 | ATA/TAA |
trnV (aac) | 13,230 | 13,262 | + | 33 | – |
trnP (tgg) | 13,272 | 13,315 | - | 44 | – |
nad6 | 13,373 | 13,759 | + | 387 | ATT/TAA |
cob | 13,773 | 14,873 | + | 1101 | ATG/TAA |
trnS2 (tga) | 14,889 | 14,944 | + | 56 | – |
The repeated DNA in animal mitogenomes can be divided into tandem repeats and interspersed repeats (
Number | SSR type | SSR | Size (bp) | Start | End | Position |
---|---|---|---|---|---|---|
1 | mono-nucleotide | (A)11 | 11 | 12,790 | 12,800 | intergenic |
2 | mono-nucleotide | (A)12 | 12 | 8540 | 8551 | rrnS |
3 | mono-nucleotide | (A)20 | 20 | 12,837 | 12,856 | intergenic |
4 | di-nucleotide | (AT)8 | 16 | 8776 | 8791 | intergenic |
5 | di-nucleotide | (AT)8 | 17 | 9820 | 9835 | intergenic |
6 | di-nucleotide | (AT)9 | 19 | 3406 | 3423 | intergenic |
7 | di-nucleotide | (TA)11 | 22 | 1119 | 1140 | intergenic |
8 | di-nucleotide | (AT)19 | 39 | 14,968 | 15,005 | intergenic |
9 | tri-nucleotide | (TAA)5 | 17 | 14,954 | 14,968 | intergenic |
For all 13 PCGs identified in the S. mutilans mitogenome, five genes (nad2, nad3, nad1, nad5 and nad6) initiated with the start codon ATT, two genes (atp8 and nad4l) started with the ATG codon, and the remaining six genes used ATA as the start codon. The most common termination codon TAA was detected in eleven PCGs (nad2, cox2, atp8, atp6, cox3, nad1, nad5, nad4, nad4l, nad6, cob). The cox1 and nad3 genes had termination codons with TAG (Table
By using the Mauve analysis, we identified six large genomic homologous regions (marked A–F in Figure
The constructed ML tree is presented in Figure
The ω value can be used for revealing the constraints of natural selection (
We sequenced and assembled the mitogenome of S. mutilans, a representative animal widely used in Chinese traditional medicine. The mitogenome is 15,011 bp in length, which is similar to the genome size of other known centipede mitogenomes, for example, 15,119 bp in Scolopocryptops sp. and 15,139 bp in Bothropolys sp. (Table
Our study predicted nine mitogenomic SSRs, which can provide additional genetic marker information in molecular identification of centipede species (Table
We identified six homologous regions among the eight species’ mitogenomes, which revealed obviously genomic rearrangements, in particular between S. mutilans and some other centipedes (Figure
Previous studies revealed alternative phylogenetic relationships of different centipedes by using different molecular datasets (
In conclusion, we successfully sequenced the complete mitochondrial genome of S. mutilans for the first time using next-generation sequencing, which will be valued for further studies in terms of the conservation, molecular identification, and evolutionary biology of diverse centipede species, improving the medicinal applications of S. mutilans and other closely related taxa.
We acknowledge Professor Ya’hua Zhan for identifying the materials sequenced here. Special thanks to Pavel Stoev and the reviewers for constructive comments that improved this paper. The present study was supported by the central government guides local science and technology development fund in Hubei Province (2019ZYYD063), Hubei Science Foundation for Distinguished Young Scholars (2019CFA097) and Wuhan “Yellow Crane Talent Program” Talent Project (Principal: Zhigang Hu).
Table
Explanation note: The ω value of 13 PCGs.