The deepest mitochondrial genome sequenced from Mariana Trench Hirondellea gigas (Amphipoda).

Hirondellea gigas is an amphipod that is a dominant animal resident living in the Challenger Deep (∼11,000m depth) of the Mariana Trench, which is the world deepest point in the ocean. Here we report a nearly complete mitochondrial genome of H. gigas, the world deepest mitogenome. The genome consists of two contigs with lengths of 8,603 bp and 6,984 bp, respectively, and it includes 13 complete protein-coding genes, 2 rRNA genes, and 21 tRNA genes. The ends of both contigs are highly repetitive and AT rich. The gene order of H. gigas is similar to another amphipod (Onisimus nanseni) in the same superfamily, Lysianassoidea. Phylogenetic analysis showed that Lysianassoidea is grouped with Gammaroidea and Calliopioidea in the same clade. Our result also suggested that the H. gigas collected from Izu-Bonin Trench and Japan Trench are indeed the same species as those from the Mariana Trench. These results will contribute to a better understanding of the phylogeny of amphipod and other hadal species.

The hadopelagic zone is the deepest zone of the ocean, ranging from 6000–11,000 m depth (Jamieson et al. 2010). Hirondellea gigas (Birstein & Vinagradov 1955), an amphipod of superfamily Lysianassoidea was first described from the Mariana Trench which is the Earth’s deepest point (∼11,000 m) (Kobayashi et al. 2012). The species is also recorded as living in other hadal trenches of the Northwest Pacific (Ritchie et al. 2015). Previous studies showed that mitochondrial genome is a promising tool to study the phylogeny of amphipods, which is useful to study their speciation and evolution (Cooper et al. 2007; Ritchie et al. 2015). Here, we described the mitogenome of H. gigas and conduct phylogenetic analyses to assess its phylogenetic position.

During the cruise of Schmidt Ocean Institute in December 2014 (http://schmidtocean.org/cruise/expanding-mariana-trench-perspectives), four Leggo landers were deployed into Challenger Deep in Mariana Trench (11°22.1122 N, 142°35.2510 E; 10,929 m depth) and captured H. gigas amphipods (Supplementary Figure 1). Voucher specimens from this site are deposited at the Scripps Institution of Oceanography Benthic Invertebrate Collection under catalogue numbers SIO-BIC C12056-58. Other specimens from Mariana Trench in 2012 (SIO-BIC 11848-11851) were also used for population comparisons. The H. gigas were fixed in RNAlater stabilization solution or 95% ethanol. For the whole mitochondrial genome of a Challenger Deep specimen, a library was prepared and sequenced on Illumina MiSeq platform (San Diego, CA) to generate 5.5 Gb paired-end reads with lengths of 300 bp. Adapters and low quality reads were trimmed by Trimmomatic version 0.33 (Bolger et al. 2014). The clean reads were assembled using CLC Genomics Workbench v7.0.3 (Aarhus, Denmark) with a k-mer size of 64. Two contigs with the lengths of 8603 bp and 6984 bp were considered as the mitogenome sequences (NCBI accession number: KU558990 and KU558991, respectively). Then, MITOS (Bernt et al. 2013) were used to annotate protein-coding genes (PCGs), ribosomal RNA (rRNA) genes, and transfer RNA (tRNA) genes. Furthermore, the boundaries of PCGs were manually corrected by alignment with published amphipod mitogenomes. Cloverleaf structures of tRNA genes were determined by ARWEN (Laslett & Canbäck 2008). Partial sequences of mitochondrial cytochrome subunit 1 (COI) were generated for a total of 15 specimens from the Mariana Trench using universal primers. Popart (Leigh & Bryant 2015) was used to generate a haplotype network using among H. gigas from Mariana Trench, Izu-Bonin Trench and Japan Trench based on 526 nt of COI (Supplementary Method). Maximum-likelihood (ML) and maximum parsimony (MP) analyses were conducted based on 2740 amino acids (Supplementary Method).

There are 36 complete genes including 13 PCGs, 2 rRNA subunit genes, and 21 tRNA genes in the nearly complete mitogenomes (Supplementary Table 1). PCR amplifications failed using several combination of primers and PCR programs, therefore, these two contigs could not be connected between each other. The end sequences of each contigs include tandem repeats consisting of both A and T. Thus, the gaps are possibly highly repetitive with A and T. All genes were encoded in the same strand except three tRNA genes in the 8603 bp contig. Gene order of H. gigas is highly similar to O. nanseni (Ki et al. 2010), which is closest to H. gigas in the phylogenetic tree (Figure 1).

Figure 1.

Phylogenetic tree of 13 species from 7 primary superfamilies of order Amphipoda based on amino acid sequences. Limnoria quadripunctata (Isopoda) serves as outgroup. Numbers near the node specify the bootstrap value of ML analysis (left) and MP analysis (right). The mitochondrial genome accession number is listed as follows: H. gigas (KU558990 & KU558991), Metacrangonyx ilvanus (HE860503.1), Longipodacrangonyx sp. (HE860508.1), Eulimnogammarus verrucosus (NC_023104.1), Caprella scaura (AB539699.1), C. mutica (NC_014492.1), Brachyuropus grewingkii (NC_026309.1), Gammarus duebeni (JN704067.1), Gondogeneia antarctica (NC_016192.1), Onisimus nanseni (NC_013819.1), Bahadzia jaraguensis (FR872382.1), Pseudoniphargus sorbasiensis (LN871175.1), P. gorbeanus (LN871176.1), and Limnoria quadripunctata (NC_024054.1).

The phylogenetic analysis supports that superfamily Lysianassoidea, including H. gigas, is grouped with another two superfamilies, Gammaroidea, and Calliopioidea, in one clade (Figure 1). The haplotype network (Supplementary Figure 2) showed no shared haplotypes between the Mariana Trench and the Japan and Izu-Bonin Trench, but the small COI differences found have suggested this species does inhabit these widely separated habitats. We believe that this work would contribute to a better understanding of phylogeny of hadal trench amphipods.

Funding

This work was supported by Strategic Priority Research Program of Chinese Academy of Sciences under grant XDB06010102; the National Science Foundation under grant 0801973, 0827051 and 1536776; the National Aeronautics and Space Administration under grant NNX11AG10G; the Prince Albert II Foundation under grant project 1265; the Avatar Alliance Foundation; and the Schmidt Ocean Institute under grant FK141215.

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