Constructing the mitochondrial genome of the Peruvian grunt Anisotremus scapularis Tschudi, 1846 (Lutjaniformes: Haemulidae) using RNA-seq data

Abstract

Haemulidae is a widely diverse and widespread family of New World teleost fish. The Peruvian grunt, Anisotremus scapular, inhabits the South American Pacific coast where its overexploitation for human consumption. Fishing has led to the populational decline of the species and raised the need for conservation and aquaculture developments. Here we describe the complete mitogenome of the Peruvian grunt assembled from an RNA-seq dataset. The mitogenome length is 16,785 bp and, as expected for a vertebrate, composed of 13 PCGs, 22 transfer RNA, 2 ribosomal RNA genes. Phylogenetic reconstruction, including all available 25 Lutjaniformes mitogenomes, places A. scapularis within the family Haemulidae, more precisely within the subfamily Haemulinae. Furthermore, paraphyletic relationships were observed within the other groups represented in the phylogeny. Overall, our results highlight the usefulness of using mitochondrial genomes for phylogenetic analysis in Haemulinae as well as the necessity to increase taxonomic sampling within the group.

Keywords:

• Peruvian grunt
• Teleostei
• mitogenome
• phylogenetics
• RNA-seq

The family Haemulidae includes 17 genera (∼ 150 species) of mainly marine fish found in the New World (Nelson et al. 2016). The Peruvian grunt, Anisotremus scapularis Tschudi, 1846 is a euriphagic benthopelagic fish, distributed from Ecuador to northern Chile, co-habiting with six other Anisotremus species (Chirichigno and Cornejo 2001). The high demand by the consumer has led to drastic declines and assessment plans for its recovery, including aquaculture development. The unequivocal identification of the species and the clarification of its phylogenetic relationships are key to the implementation of these strategies. Thus, we obtained the complete A. scapularis mitogenome for the first time.

A specimen of A. scapularis was captured in the south Peruvian shore (−18.0208 S; −70.8695 W). The individual was immediately dissected and tissues from several organs were stored for subsequent total RNA extraction using Illustra RNA spin kit (GE Healthcare, Little Chalfont, UK). TruSeq stranded mRNA libraries were then built and 150 bp paired-end reads sequenced by Macrogen (South Korea). The specimen tissues are stored at the Interdisciplinary Center of Marine and Environmental Research (specimen code SA001). For obtaining the complete mitogenomes from the RNA-se data we followed Moreira et al. (2016), however, mitochondrial contigs were retrieved from the transcriptome assembly using the mode findmitoscaf from MitoZ (Meng et al. 2019). Annotation was also performed in MitoZ and manually validated by comparing predicted protein-coding (PCG), ribosomal RNA (rRNA) and transfer RNA (tRNA) genes with other Lutjaniformes using MEGAX (Kumar et al. 2018).

For BI and ML phylogenetic analysis, all complete Lutjaniformes mitogenomes (n = 24) and four outgroup taxa, were retrieved from GenBank (accessed in March 2020). The 13 PCG were aligned using GUIDANCE2 (Sela et al. 2015) with MAFFT (Katoh and Standley 2013) and then concatenated in SequenceMatrix (Vaidya et al. 2011). The final concatenated alignment was 11,419 nt long. Best partition schemes and molecular evolutionary models were selected using PartitionFinder2 on XSEDE (Lanfear et al. 2016) and used for phylogenetic analyses using MrBayes on XSEDE (Ronquist et al. 2012) (GTR + I + G, GTR + I + G and HKY + G) with two independent runs (10⁷ generations, sampling frequency one tree for 1000 generations) and RAxML-HPC BlackBox (Stamatakis 2014) (GTR + I + G). All analyses were implemented in CIPRES (Miller et al. 2010).

The first complete mitogenome of the subfamily Haemulinae (Haemulidae), from A. scapularis was deposited in GenBank (MT259025). The mitogenome length (16,785 bp), gene content and orientation are those expected for vertebrate mtDNA: 13 PCGs, 22 transfer RNA, 2 ribosomal RNA genes.

Both BI and ML phylogenetic trees, rooted with the two Spariformes outgroup taxa and for the first time including all available Lutjaniformes mitogenomes in the same analyses, show the same topology (Figure 1). As expected, Lutjaniformes are divided in two groups corresponding to Lutjanidae and Haemulidae. The latter is further divided in two clades that separate the new mitogenome (i.e. A. scapularis, Haemulinae) from the Plectorhinchinae. The genus Plectorhinchus is paraphyletic regarding Diagramma picta, a result previously observed in a phylogenetic study using both mitochondrial and nuclear markers (Sanciangco et al. 2011). Four recent studies using complete mitogenomes for the phylogenetic assessment of the family Lutjanidae (Yamanoue et al. 2007; Bayona-Vásquez et al. 2017; Zhan et al. 2017; Kim et al. 2019) revealed conflicting phylogenetic results, probably due to incomplete representation of available mitogenomes. In the present study, by including all available Lutjanidae mitogenomes, we observed that while as expected, Etelinae (i.e. Pristipomoides multidens) clusters neither with Lutjaninae nor with Caesioninae, Lutjaninae appears paraphyletic regarding Caesioninae.

Figure 1. Bayesian Inference phylogenetic tree based on 13 concatenated protein coding genes from 29 mitogenomes, including 25 Lutjaniformes species and four outgroup taxa, two Lobotiformes (HM754620 and JX435822) and two Spariformes (KR701906 and AP009165). Only supports above 92% are shown. The * on the nodes indicate that both posterior probabilities and bootstrap support values above 95%.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The data that supports the findings of this study are available in NCBI (MT259025) or from the corresponding author, L. Filipe C. Castro.

Additional information

Funding

This work was financed by the Project The Sea and the Shore, Architecture and Marine Biology: The Impact of Sea Life on the Built Environment [PTDC/ART-DAQ/29537/2017] from FCT/MCTES through national funds (PIDDAC) and co-financing from the European Regional Development Fund (FEDER) POCI-01-0145-FEDER-029537, in the aim of the new partnership agreement PT2020 throught COMPETE 2020 – Competitiveness and Internationalization Operational Program (POCI), and by FCT – Fundação para a Ciência e a Tecnologia UIDB/04423/2020, UIDP/04423/2020 which also supported A.G.S. [SFRH/BD/137935/2018].