The order Istiophoriformes is composed of bony fishes with major morphological, physiological, behavioral, and ecological differences. Among them, billfishes, characterized by the presence of a snout resembling a sword, are contained in the families Xiphiidae and Istiophoridae. This is the first study to assemble and analyze the complete mitochondrial genome of Xiphias gladius, the sole species of the family Xiphiidae. The AT-rich mitochondrial genome of Xiphias gladius is 16,520 bp long and is comprised of 22 transfer RNA genes, 2 ribosomal RNA genes, 13 protein-coding genes (PCG), and a putative control region (CR) 830 bp long. All 22 tRNAs exhibit the ‘cloverleaf’ shaped secondary structure with the exception of tRNAS1, due to an absent D-arm. Ka/Ks values estimated that all of the complete PCGs had P-values < 1, which indicates purifying selection. In the CR, numerous microsatellites were found along with multiple hairpin-like loops in the secondary structure, but no tandem repeats were detected. This genomic resource will help with biomonitoring and bioprospecting of this large pelagic fish using environmental DNA strategies. A phylomitogenomic maximum likelihood analyses positioned X. gladius sister to the family Istiophoridae and supported the monophyletic status of the genera comprising the latter family. However, the analysis did not resolve relationships among genera in the family Istiophoridae. This newly assembled mitochondrial genome analyzed in detail for the first time will support conservation efforts in this iconic fish currently experiencing major population declines.
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