New species of redbait from the Philippines (Teleostei, Emmelichthyidae, Emmelichthys).

We describe a new species of redbait in the genus Emmelichthys collected from fish markets on Panay and Cebu islands in the Visayas region of the Philippines. The species is externally similar to E.struhsakeri but is diagnosable by two prominent fleshy papillae associated with the cleithrum and fewer pectoral-fin rays (18-19 vs. 19-21) and gill rakers (30-33 vs. 34-41). Additionally, mitochondrial DNA differentiates this taxon from other species of Emmelichthys. We generate mitochondrial genomes for two of the three type specimens and several other emmelichthyids to place the new taxon in a phylogenetic context. Analysis of the protein-coding mitochondrial loci calls into question the monophyly of two emmelichthyid genera (Emmelichthys and Erythrocles) and highlights the need for subsequent analyses targeting the intrarelationships of the Emmelichthyidae.

Biodiversity of Philippine marine fishes: A DNA barcode reference library based on voucher specimens.

Accurate identification of fishes is essential for understanding their biology and to ensure food safety for consumers. DNA barcoding is an important tool because it can verify identifications of both whole and processed fishes that have had key morphological characters removed (e.g., filets, fish meal); however, DNA reference libraries are incomplete, and public repositories for sequence data contain incorrectly identified sequences. During a nine-year sampling program in the Philippines, a global biodiversity hotspot for marine fishes, we developed a verified reference library of cytochrome c oxidase subunit I (COI) sequences for 2,525 specimens representing 984 species. Specimens were primarily purchased from markets, with additional diversity collected using rotenone or fishing gear. Species identifications were verified based on taxonomic, phenotypic, and genotypic data, and sequences are associated with voucher specimens, live-color photographs, and genetic samples catalogued at Smithsonian Institution, National Museum of Natural History. The Biodiversity of Philippine Marine Fishes dataset is released herein to increase knowledge of species diversity and distributions and to facilitate accurate identification of market fishes.

Coevolution of the olfactory organ and its receptor repertoire in ray-finned fishes.

BACKGROUND: Ray-finned fishes (Actinopterygii) perceive their environment through a range of sensory modalities, including olfaction. Anatomical diversity of the olfactory organ suggests that olfaction is differentially important among species. To explore this topic, we studied the evolutionary dynamics of the four main gene families (OR, TAAR, ORA/VR1 and OlfC/VR2) coding for olfactory receptors in 185 species of ray-finned fishes. RESULTS: The large variation in the number of functional genes, between 28 in the ocean sunfish Mola mola and 1317 in the reedfish Erpetoichthys calabaricus, is the result of parallel expansions and contractions of the four main gene families. Several ancient and independent simplifications of the olfactory organ are associated with massive gene losses. In contrast, Polypteriformes, which have a unique and complex olfactory organ, have almost twice as many olfactory receptor genes as any other ray-finned fish. CONCLUSIONS: We document a functional link between morphology of the olfactory organ and richness of the olfactory receptor repertoire. Further, our results demonstrate that the genomic underpinning of olfaction in ray-finned fishes is heterogeneous and presents a dynamic pattern of evolutionary expansions, simplifications, and reacquisitions.

Skimming for barcodes: rapid production of mitochondrial genome and nuclear ribosomal repeat reference markers through shallow shotgun sequencing.

DNA barcoding is critical to conservation and biodiversity research, yet public reference databases are incomplete. Existing barcode databases are biased toward cytochrome oxidase subunit I (COI) and frequently lack associated voucher specimens or geospatial metadata, which can hinder reliable species assignments. The emergence of metabarcoding approaches such as environmental DNA (eDNA) has necessitated multiple marker techniques combined with barcode reference databases backed by voucher specimens. Reference barcodes have traditionally been generated by Sanger sequencing, however sequencing multiple markers is costly for large numbers of specimens, requires multiple separate PCR reactions, and limits resulting sequences to targeted regions. High-throughput sequencing techniques such as genome skimming enable assembly of complete mitogenomes, which contain the most commonly used barcoding loci (e.g., COI, 12S, 16S), as well as nuclear ribosomal repeat regions (e.g., ITS1&2, 18S). We evaluated the feasibility of genome skimming to generate barcode references databases for marine fishes by assembling complete mitogenomes and nuclear ribosomal repeats. We tested genome skimming across a taxonomically diverse selection of 12 marine fish species from the collections of the National Museum of Natural History, Smithsonian Institution. We generated two sequencing libraries per species to test the impact of shearing method (enzymatic or mechanical), extraction method (kit-based or automated), and input DNA concentration. We produced complete mitogenomes for all non-chondrichthyans (11/12 species) and assembled nuclear ribosomal repeats (18S-ITS1-5.8S-ITS2-28S) for all taxa. The quality and completeness of mitogenome assemblies was not impacted by shearing method, extraction method or input DNA concentration. Our results reaffirm that genome skimming is an efficient and (at scale) cost-effective method to generate all mitochondrial and common nuclear DNA barcoding loci for multiple species simultaneously, which has great potential to scale for future projects and facilitate completing barcode reference databases for marine fishes.

Evolutionary Dynamics of the OR Gene Repertoire in Teleost Fishes: Evidence of an Association with Changes in Olfactory Epithelium Shape.

Teleost fishes perceive their environment through a range of sensory modalities, among which olfaction often plays an important role. Richness of the olfactory repertoire depends on the diversity of receptors coded by homologous genes classified into four families: OR, TAAR, VR1, and VR2. Herein, we focus on the OR gene repertoire. While independent large contractions of the OR gene repertoire associated with ecological transitions have been found in mammals, little is known about the diversity of the OR gene repertoire and its evolution in teleost fishes, a group that includes more than 34,000 living species. We analyzed genomes of 163 species representing diversity in this large group. We found a large range of variation in the number of functional OR genes, from 15 in the Broad-nose Pipefish Syngnathus typhle and the Ocean Sunfish Mola mola, to 429 in the Zig-zag Eel Mastacembelus armatus. The number of OR genes was higher in species when a multilamellar olfactory rosette was present. Moreover, the number of lamellae was correlated with the richness of the OR gene repertoire. While a slow and balanced birth-and-death process generally drives the evolution of the OR gene repertoire, we inferred several episodes of high rates of gene loss, sometimes followed by large gains in the number of OR genes. These gains coincide with morphological changes of the olfactory organ and suggest a strong functional association between changes in the morphology and the evolution of the OR gene repertoire.

The mitochondrial genome of the endangered Spiny Butterfly Ray Gymnura altavela (Linnaeus 1758) (Myliobatiformes: Gymnuridae) provides insights into cryptic lineages.

The Spiny Butterfly Ray, Gymnura altavela, is found in tropical and temperate coastal waters with a discontinuous distribution: it occurs along the east coast of the United States of America, Brazil, and Uruguay in the western Atlantic Ocean, from Portugal to Angola in the eastern Atlantic, and also in the Mediterranean Sea. Globally, Gymnura altavela is considered endangered by the International Union for Conservation of Nature (IUCN). Our study analyzed the complete mitochondrial genome of G. altavela sequenced from an individual captured in the western Atlantic and compared it with a mitochondrial genome from a conspecific collected in the Mediterranean to explore if G. altavela comprises cryptic species. The newly assembled mitochondrial genome of G. altavela is 19,361bp in length and has 13 protein-coding genes (PCGs), two ribosomal RNA genes (12s ribosomal RNA and 16s ribosomal RNA), 22 transfer RNA (tRNA) genes, and a 3,659 bp control region. The gene order is identical to that reported for the specimen collected in the Mediterranean and that of the congeneric Long-tailed Butterfly Ray, G. poecilura. A phylomitogenomic analysis based on translated PCGs supported the monophyly of the genus Gymnura and indicated that genetic dissimilarity between G. altavela from the western Atlantic Ocean and the Mediterranean Sea was greater than that calculated among congeneric species belonging to the genera Mobula and Neotrygon, similar to that calculated among congeneric stingrays in the genus Hemitrygon, but lower than that calculated among congeneric species in the genera Pateobatis and Urogymnus. Overall, our comparisons suggest that G. altavela comprises cryptic species or at least indicates considerable genetic structure between populations in the western Atlantic and Mediterranean. We present these findings in a phylomitogenomic analysis of other Myliobatiformes and Rhinopristiformes. Additional phylogeographic and taxonomic studies of G. altavela are needed to support conservation efforts for this endangered ray that may comprise cryptic evolutionary units.

Review of the Indian Ocean spikefish genus Mephisto (Tetraodontiformes: Triacanthodidae).

We redescribe the triacanthodid spikefish Mephisto fraserbrunneri Tyler 1966 based upon eight specimens (five newly reported herein) and the first color photographs of freshly collected specimens; these data are compared with that of the single specimen of the recently described M. albomaculosus Matsuura, Psomadakis, and Mya Than Tun 2018. Both species are found in the Indian Ocean, with M. fraserbrunneri known from the Arabian Sea off the east coast of Africa to the eastern Bay of Bengal, and M. albomaculosus confirmed only from the type locality in the Andaman Sea (a color photograph of an individual M. cf. albomaculosus from the Bay of Bengal that was not retained is also presented). We describe and diagnose the genus Mephisto and provide a key to the two species based upon all available specimens. We also provide a distribution map for both species and summarize literature records. Using micro-CT data, we show that Mephisto fraserbrunneri replaces teeth intraosseously, which suggests this tooth replacement pattern is plesiomorphic for Tetraodontiformes.

Tooth and consequences: Heterodonty and dental replacement in piranhas and pacus (Serrasalmidae).

Tooth replacement in piranhas is unusual: all teeth on one side of the head are lost as a unit, then replaced simultaneously. We used histology and microCT to examine tooth-replacement modes across carnivorous piranhas and their herbivorous pacu cousins (Serrasalmidae) and then mapped replacement patterns onto a molecular phylogeny. Pacu teeth develop and are replaced in a manner like piranhas. For serrasalmids, unilateral tooth replacement is not an "all or nothing" phenomenon; we demonstrate that both sides of the jaws have developing tooth rows within them, albeit with one side more mineralized than the other. All serrasalmids (except one) share unilateral tooth replacement, so this is not an adaptation for carnivory. All serrasalmids have interlocking teeth; piranhas interdigitate lateral tooth cusps with adjacent teeth, forming a singular saw-like blade, whereas lateral cusps in pacus clasp together. For serrasalmids to have an interlocking dentition, their teeth need to develop and erupt at the same time. We propose that interlocking mechanisms prevent tooth loss and ensure continued functionality of the feeding apparatus. Serrasalmid dentitions are ubiquitously heterodont, having incisiform and molariform dentitions reminiscent of mammals. Finally, we propose that simultaneous tooth replacement be considered as a synapomorphy for the family.

Life history, distribution and molecular phylogenetics of the Upward-Mouth Spikefish Atrophacanthus japonicus (Teleostei: Tetraodontiformes: Triacanthodidae).

Ninety-six juvenile specimens (37-54 mm standard length; LS ) of the rarely collected Upward-Mouth Spikefish Atrophacanthus japonicus (Triacanthodidae) were obtained from the stomachs of three Yellowfin Tuna Thunnus albacares collected off Guam in the Mariana Islands in the central Pacific Ocean. These specimens extend the range of A. japonicus eastward into Oceania. We review the systematic characters of the monotypic genus Atrophacanthus and present colour photographs of freshly collected specimens. The diet of the juvenile specimens of A. japonicus consisted of thecosome pteropods and foraminiferans. We present a range map of A. japonicus based on all known specimens and show that specimen size is related to whether specimens were collected in the pelagic zone or on the bottom. Our results support that, compared to all other Triacanthodidae, A. japonicus has an unusually extended pelagic larval and juvenile period, up to 54 mm LS , before settling to the bottom as adults. Lastly, we provide a multilocus phylogeny addressing the phylogenetic placement of Atrophacanthus based on eight of 11 triacanthodid genera and six genetic markers. Our results reveal that Atrophacanthus is the sister group of Macrorhamphosodes and they provide new insights about the evolutionary history of the family.

Tooth development and replacement in the Atlantic Cutlassfish, Trichiurus lepturus, with comparisons to other Scombroidei.

Atlantic Cutlassfish, Trichiurus lepturus, have large, barbed, premaxillary and dentary fangs, and sharp dagger-shaped teeth in their oral jaws. Functional teeth firmly ankylose to the dentigerous bones. We used dry skeletons, histology, SEM, and micro-CT scanning to study 92 specimens of T. lepturus from the western North Atlantic to describe its dentition and tooth replacement. We identified three modes of intraosseous tooth replacement in T. lepturus depending on the location of the tooth in the jaw. Mode 1 relates to replacement of premaxillary fangs, in which new tooth germs enter the lingual surface of the premaxilla, develop horizontally, and rotate into position. We suggest that growth of large fangs in the premaxilla is accommodated by this horizontal development. Mode 2 occurs for dentary fangs: new tooth germs enter the labial surface of the dentary, develop vertically, and erupt into position. Mode 3 describes replacement of lateral teeth, in which new tooth germs enter a trench along the crest of the dentigerous bone, develop vertically, and erupt into position. Such distinct modes of tooth replacement in a teleostean species are unknown. We compared modes of replacement in T. lepturus to 20 species of scombroids to explore the phylogenetic distribution of these three replacement modes. Alternate tooth replacement (in which new teeth erupt between two functional teeth), ankylosis, and intraosseous tooth development are plesiomorphic to Bluefish + other Scombroidei. Our study highlights the complexity and variability of intraosseous tooth replacement. Within tooth replacement systems, key variables include sites of formation of tooth germs, points of entry of tooth germs into dentigerous bones, coupling of tooth germ migration and bone erosion, whether teeth develop horizontally or immediately beneath the tooth to be replaced, and how tooth eruption and ankylosis occur. Developmentally different tooth replacement processes can yield remarkably similar dentitions.