Complete Mitogenomes of Deep-Sea Eels Histiobranchus bathybius and Simenchelys parasitica and a New Record of H. bathybius from the East Mariana Basin, Western Pacific Ocean

Abstract

In this study, using Illumina sequencing, we sequenced first the complete mitochondrial genome (mitogenome) of two deep-sea eels, Histiobranchus bathybius and Simenchelys parasitica, collected from the East Mariana Basin in the Western Pacific Ocean. The complete length of the H. bathybius and S. parasitica mitogenomes were 16,696 and 16,687 bp, respectively, each containing 37 genes (13 protein-coding genes, 22 tRNA genes, and 2 ribosomal RNA genes). To enhance the accuracy of the identification of H. bathybius and S. parasitica, we performed a phylogenetic analysis of multiple deep-sea eels based on the mitochondrial DNA gene (cytochrome c oxidase subunit I [COI]) using the maximum likelihood method. Our phylogenetic tree analysis confirmed that the specimens collected in this study are congeneric species of H. bathybius and S. parasitica reported in previous studies. Based on these results, we report the first complete mitogenomes of H. bathybius and S. parasitica and a new record for the two species in the East Mariana Basin.

1. Introduction

Deep-sea species occupy a relatively large portion of global ecosystems, but their diversity and distribution are not well studied owing to the challenges imposed by the harsh conditions of deep-sea environments (>200 m depth) [1,2]. Fortunately, despite the extreme challenges involved in sampling methods [3,4,5], several studies have been conducted in the Atlantic, Pacific, and Indian oceans to characterize and document deep-sea organism habitats and diversity over the past decade. Notably, a few new species have been identified and shown to have extensive spatial distributions [6,7,8,9].

As with all species, the correct identification of known and/or new species of fish is important for biodiversity assessments in deep-sea ecosystems [10]. Morphological analysis, which is a widely used basic tool for characterizing and identifying species [11], may not be adequate in every case, especially considering the morphological diversity of fish and changes during ontogeny [12,13,14]. DNA barcoding has emerged as a useful taxonomic tool for the rapid and reliable identification of unknown specimens through phylogeny reconstruction [15,16,17]. In addition, mitochondrial DNA (mtDNA) can be used as a DNA-based marker in population genetic diversity and conservation studies [18]. In particular, the mtDNA marker gene (mitochondrial cytochrome c oxidase subunit I [COI]) has been extensively used to identify marine species [19,20,21,22]. Notably, mtDNA genes have recently been used to identify deep-sea fish, including eel species (e.g., Bassozetus sp., Synaphobranchus affinis, and Synaphobranchus brevidorsalis) [23,24,25,26]. Therefore, mtDNA genes can be used as genetic markers for the accurate species identification of deep-sea eels. However, little information is available on the complete mitogenome of deep-sea eels in the Western Pacific Ocean.

In this study, the complete mitogenomes of Histiobranchus bathybius and Simenchelys parasitica collected from the East Mariana Basin are documented for the first time. Further, to confirm the identification, we conducted a molecular phylogenetic analysis based on the COI gene sequences of a number of deep-sea eel species. Overall, this study provides molecular information for further evolutionary ecology research associated with the population distribution of deep-sea eels in the Western Pacific Ocean.

2. Materials and Methods

2.1. Specimen Collection, Genomic DNA Extraction, and Sequencing

Recognizing the existing knowledge gap in biodiversity in the western Pacific region, the Korea Institute of Ocean Science and Technology (KIOST) has initiated a program to acquire marine bioresources from this region. During an oceanographic cruise onboard the RV ISABU (Supplementary Figure S1A) of the KIOST to the Western Pacific Ocean in May 2021 (Supplementary Figure S1B and Supplementary Table S1), H. bathybius and S. parasitica were collected in a fishing bait trap (Supplementary Figure S2), each from a different depth. Supplementary Figure S3 presents the images of H. bathybius and S. parasitica.

All animal experimental protocols were performed in accordance with the Guidelines of the Institutional Animal Care and Experimental Committee and approved by the KIOST.

The fish tissue samples were preserved at −20 °C until DNA isolation. Genomic DNA was extracted from the muscle using a DNeasy Blood & Tissue Kit (Qiagen, Valencia, CA, USA), following the manufacturer’s protocol. The quantity and quality of the isolated DNA were analyzed and measured at 230, 260, and 280 nm using a NanoDrop One spectrophotometer (Thermo Fisher Scientific Inc., Madison, WI, USA). Whole-genome sequencing was performed using the Illumina NovaSeq 6000 platform at the National Instrumentation Center for Environmental Management (Seoul, Republic of Korea). The complete mitogenomes of the two deep-sea eel specimens were assembled and annotated using MitoZ [27]. The circular maps of the mitogenomes of H. bathybius and S. parasitica were drawn by the GeSeq tool (https://chlorobox.mpimp-golm.mpg.de/geseq.html (accessed on 10 June 2024) [28].

2.2. Phylogenetic Analysis

To construct a molecular phylogenetic tree, a total of 24 mtDNA COI sequences from the eel family Synaphobranchidae (genus: Simenchelys, llyophis, Histiobranchus, Dysomma, Dysommina, Diastobrachus, and Synaphobranchus) were downloaded from the GenBank (Supplementary Table S2). The nucleotide sequences of the individual mtDNA COI genes were aligned using the ClustalW algorithm in the MEGA software (ver. 11.0.1; Centre for Evolutionary Medicine and Informatics, Tempe, AZ, USA). To establish the best-fit substitution model for phylogenetic analysis, the model with the lowest Bayesian information criterion and Akaike information criterion scores was estimated using a maximum likelihood analysis. According to the model test results, maximum likelihood phylogenetic analyses were performed with the LG + G + I model using the MEGA software. Support for nodes was calculated using 1000 bootstrap replicates.

3. Results and Discussion

In this study, the complete mitogenomes of H. bathybius and S. parasitica were sequenced and deposited in GenBank (accession numbers: PP726018 and PP726019, respectively). The sequence lengths of the complete mitogenomes of H. bathybius and S. parasitica were 16,696 base pairs (bp) and 16,687 bp, respectively. The complete mitogenomes of H. bathybius and S. parasitica each encoded a set of 37 genes (13 protein-coding genes [PCGs], 22 tRNA genes, and two rRNA genes) (

Table 1. H. bathybius mitogenome organization.

Gene	Location	Length (bp)	Strand	Start Codon	Stop Codon	Intergenic Region *
tRNA-Phe	1–70	70	+			-
s-rRNA	71–1024	954	+			0
tRNA-Val	1025–1094	70	+			0
l-rRNA	1133–2808	1676	+			−62
tRNA-Leu	2809–2883	75	+			0
ND1	2884–3852	969	+	ATG	TAA	0
tRNA-Ile	3862–3931	70	+			9
tRNA-Gln	3931–4001	71	−			−1
tRNA-Met	4001–4069	69	+			−1
ND2	4070–5116	1047	+	ATG	TAG	0
tRNA-Trp	5115–5184	70	+			−2
tRNA-Ala	5187–5255	69	−			2
tRNA-Asn	5257–5329	73	−			1
tRNA-Cys	5367–5431	65	−			37
tRNA-Tyr	5432–5502	71	−			0
COX1	5504–7078	1575	+	GTG	TAA	1
tRNA-Ser	7087–7157	71	−			8
tRNA-Asp	7163–7232	70	+			5
COX2	7239–7929	691	+	ATG	T--	6
tRNA-Lys	7930–8004	75	+			0
ATP8	8006–8173	168	+	ATG	TAA	1
ATP6	8164–8847	684	+	ATG	TAA	−10
COX3	8847–9632	786	+	ATG	TAA	−1
tRNA-Gly	9632–9703	72	+			−1
ND3	9704–10,054	351	+	ATG	TAG	0
tRNA-Arg	10,053–10,122	70	+			0
ND4L	10,123–10,419	297	+	ATG	TAA	−2
ND4	10,413–11,793	1381	+	ATG	T--	0
tRNA-His	11,794–11,862	69	+			0
tRNA-Ser	11,863–11,931	69	+			0
tRNA-Leu	11,932–12,004	73	+			0
ND5	12,005–13,846	1842	+	ATG	TAA	0
ND6	13,843–14,364	522	−	ATG	TAG	−4
tRNA-Glu	14,365–14,433	69	−			0
CYTB	14,438–15,577	1140	+	ATG	TAG	6
tRNA-Thr	15,580–15,652	73	+			4
tRNA-Pro	15,658–15,727	70	−			5

* Negative intergenic nucleotides indicate overlapping sequences between adjacent genes.

and

Table 2. S. parasitica mitogenome organization.

Gene	Location	Length (bp)	Strand	Start Codon	Stop Codon	Intergenic Region *
tRNA-Phe	1–70	70	+			-
s-rRNA	71–1023	953	+			0
tRNA-Val	1024–1093	70	+			0
l-rRNA	1094–2806	1713	+			0
tRNA-Leu	2807–2881	75	+			0
ND1	2882–3850	969	+	ATG	TAA	0
tRNA-Ile	3860–3928	69	+			9
tRNA-Gln	3928–3998	71	−			−1
tRNA-Met	3998–4066	69	+			−1
ND2	4067–5113	1047	+	ATG	TAG	0
tRNA-Trp	5112–5183	72	+			−2
tRNA-Ala	5186–5254	69	−			2
tRNA-Asn	5256–5328	73	−			1
tRNA-Cys	5366–5430	65	−			37
tRNA-Tyr	5431–5501	71	−			0
COX1	5503–7077	1575	+	GTG	TAA	1
tRNA-Ser	7086–7156	71	−			8
tRNA-Asp	7162–7231	70	+			5
COX2	7238–7928	691	+	ATG	T--	5
tRNA-Lys	7929–8003	75	+			6
ATP8	8005–8172	168	+	ATG	TAA	0
ATP6	8163–8846	684	+	ATG	TAA	1
COX3	8846–9631	786	+	ATG	TAA	−10
tRNA-Gly	9631–9702	72	+			−1
ND3	9703–10,053	351	+	ATG	TAG	−1
tRNA-Arg	10,052–10,121	70	+			0
ND4L	10,122–10,418	297	+	ATG	TAA	−2
ND4	10,412–11,792	1381	+	ATG	T--	0
tRNA-His	11,793–11,861	69	+			−7
tRNA-Ser	11,862–11,930	69	+			0
tRNA-Leu	11,931–12,003	73	+			0
ND5	12,004–13,845	1842	+	ATG	TAA	0
ND6	13,842–14,363	522	−	ATG	TAA	0
tRNA-Glu	14,364–14,432	69	−			−4
CYTB	14,437–15,576	1140	+	ATG	TAG	0
tRNA-Thr	15,579–15,651	73	+			4
tRNA-Pro	15,656–15,725	70	−			2

* Negative intergenic nucleotides indicate overlapping sequences between adjacent genes.

as well as Figure 1). The complete mitogenome of H. bathybius had a nucleotide composition of A (31.9%), C (26.5%), G (16.5%), and T (25.1%). The overall base composition of S. parasitica was A (31.2%), C (25.9%), G (17.0%), and T (26.0%). Notably, the differences among species may be related to the mitogenome base composition. For example, the A + T and G + C contents of 13 PCGs in the complete mitogenome of H. bathybius were 56.8% and 43.2%, respectively, whereas the contents of all sequences in the complete mitogenome were 57.0% and 43.0%, respectively. In the case of S. parasitica, the A + T and G + C contents of 13 PCGs in the complete mitogenome were 56.9% and 43.1%, respectively, whereas those in all sequences were 42.9% and 57.1%, respectively. Additionally, in H. bathybius, most of the PCGs (12 of 13 genes) were initiated with the start codon ATG, whereas COX1 had the start codon GTG. Eleven PCGs terminated with TAA/TAG, whereas COX2 and ND4 exhibited an incomplete termination codon T. In S. parasitica, most of the PCGs (12 of 13 genes) were initiated with the start codon ATG, whereas COX1 had the start codon GTG. Eleven PCGs terminated with TAA/TAG, whereas COX2 and ND4 exhibited an incomplete termination codon T. In addition, a previous study determined the complete mitochondrial genome sequence of S. parasitica (accession number: AP010849) [16]. The complete mitochondrial genome of S. parasitica contained 13 PCGs, 22 transfer RNAs, two rRNA genes, and a control region (D loop). The overall base composition of the mitogenome was A 31.8%, C 25.4%, G 16.5%, and T 21 26.3%, which is similar but slightly different from that of S. parasitica used in this study, showing compositional differences. Therefore, we suggest that H. bathybius and S. parasitica exhibit species- and region-specific differences in the complete mitogenomes of deep-sea eels.

A maximum likelihood phylogenetic tree was constructed based on the mtDNA COI gene (Figure 2). H. bathybius and S. parasitica from the Western Pacific Ocean showed distinctive genetic differences based on their mtDNA COI sequences. In particular, a molecular phylogenetic tree analysis of the mtDNA COI gene revealed that the collected specimens, presumed to be H. bathybius and S. parasitica, were congeneric with H. bathybius (bootstrap value of 100%) and S. parasitica (bootstrap value of 98%). Collectively, our results, including phylogenetic analyses based on mtDNA COI, confirm that the two deep-sea eel species collected in the East Mariana Basin were H. bathybius and S. parasitica. In a previous study, the identification of deep-sea eels (e.g., Bassozetus sp., Synaphobranchus affinis, and S. brevidorsalis) in the East Mariana Basin was described based on DNA barcoding [25,26]. Therefore, our findings suggest that mtDNA-based markers are useful for the rapid and accurate species identification of eels collected from the East Mariana Basin.

Histiobranchus [29] belongs to the family Synaphobranchidae and currently includes four valid species: H. australis [30], H. infernalis [29], H. bathybius [31], and H. bruuni [32]. To date, H. bathybius has only been reported in the North Atlantic and North Pacific Oceans [27]. Prior to the present study, there were no records of H. bathybius from the Western Pacific Ocean or any documentation of its complete mitogenome.

The snubnosed eel Simenchelys parasitica [33], belonging to the family Synaphobranchidae and the subfamily Simenchelyinae, is a species of the small deep-sea eels, which are classified under a single genus [33]. To date, S. parasiticus has been recorded in the Atlantic Ocean [33,34,35,36,37], southern Africa [38], the Western Pacific Ocean [39,40,41], and the central North Pacific Ocean [42]. Collectively, our results suggest that H. bathybius and S. parasitica could inhabit and cover a vast range throughout the Western Pacific Ocean. In addition, the complete mitogenomes of H. bathybius and S. parasitica will be important for further study on the genetic conservation, evolutionary diversity, and distribution of species abundance of deep-sea fishes in the Western Pacific Ocean. However, morphological analyses of H. bathybius and S. parasitica are required to confirm the accuracy of species identification.

In summary, we determined, for the first time, the complete mitogenome sequences of H. bathybius and S. parasitica collected from the East Mariana Basin in the Western Pacific Ocean. In addition, the mtDNA COI gene was sufficient to distinguish between H. bathybius and S. parasitica, showing the utility of mtDNA-based phylogeny for quickly identifying deep-sea eels.