Bidirectional sex-change behavior and physiological aspects in the Gorgeous goby Lythrypnus pulchellus (Gobiidae).

The Gorgeous goby Lythrypnus pulchellus shows extreme sexual plasticity with the bidirectional sex-change ability socially controlled in adults. Therefore, this study describes how the hierarchical status affects hormone synthesis through newborn hormone waste products in water and tests the influence of body size and social dominance establishment in sex reversal duration and direction. The associated changes in behavior and hormone levels are described under laboratory conditions in male-male and female-female pairs of similar and different body sizes, recording the changes until spawning. The status establishment occurred in a relatively shorter time period in male and female pairs of different sizes (1-3 days) compared to those of similar size (3-5 days), but the earlier one did not significantly affect the overall time of sex change (verified by pair spawning). The changes in gonads, hormones, and papilla occurred in sex-changer individuals, but the first one was observed in behavior. Courtship started at 3-5 days in male pairs and from 2 h to 1 day in female pairs of both groups of different and similar sizes. Hormones did not gradually move in the new sexual phenotype direction during the sex-change time course. Nonetheless, estradiol regulated sex change and 11-ketotestosterone enabled bidirectional sex change and was modulated by agonistic interactions. Cortisol is associated with status and gonadal sex change. In general, similar mechanisms underlie sex change in both directions with a temporal change sequence in phases. These results shed new light on sex-change mechanisms. Further studies should be performed to determine whether these localized changes exist in the steroid hormone synthesis along the brain-pituitary gonad axis during social and bidirectional sex changes in L. pulchellus.

Whole-Genome Survey Analyses of Five Goby Species Provide Insights into Their Genetic Evolution and Invasion-Related Genes.

As one of the most abundant groups in marine fish families, Gobiidae fish are important fishery resources in China, and some are also invasive species in certain regions worldwide. However, the phylogenetic relationships of Gobiidae fish remain ambiguous, and the study of their invasion-related genes is still scarce. This study used high-throughput sequencing technology to conduct a whole-genome survey of five Gobiidae fish species: Acanthogobius flavimanus, Acanthogobius stigmothonus, Favonigobius gymnauchen, Ctenotrypauchen microcephalus, and Tridentiger barbatus. De novo assembly of five fish genomes was performed, and genomic traits were compared through K-mer analysis. Among the five Gobiidae fish genomes, F. gymnauchen had the largest genome size (1601.98 Mb) and the highest heterozygosity (1.56%) and repeat rates (59.83%). Phylogenetic studies showed that A. flavimanus was most closely linked to A. stigmothonus, while Apogonidae and Gobiidae were closely related families. PSMC analysis revealed that C. microcephalus experienced a notable population expansion than the other four fish species in the Early Holocene. By using the KOG, GO, and KEGG databases to annotate single-copy genes, the annotated genes of the five fish were mainly classified as "signal transduction mechanisms", "cellular process", "cellular anatomical entity", and "translation". Acanthogobius flavimanus, A. stigmothonus, and T. barbatus had more genes classified as "response to stimulus" and "localization", which may have played an important role in their invasive processes. Our study also provides valuable material about Gobiidae fish genomics and genetic evolution.

High spatial variability in a species-rich assemblage of diadromous fishes in Hong Kong, southern China.

China has experienced substantial coastal reclamation and damming of rivers. These changes have the potential to impact migrations of diadromous fishes between the sea and fresh waters, but the composition of these fishes and the impacts of barriers to their movement in China have received little attention. We inventoried the species composition and distribution of diadromous fishes, and the impacts of barriers on them, in the Hong Kong Special Administrative Region (HKSAR), southern China. Fish assemblages were surveyed using hand-nets, supplemented by cast-netting and single-pass snorkel surveys, in 24 small coastal streams across three regions. Surveys were undertaken on multiple occasions during the wet and dry seasons to account for the monsoonal tropical climate. Twenty-eight diadromous fishes were collected, mostly gobies, amounting to over half (53%) of the total richness of primary freshwater fishes; four additional species are known from literature records. Diadromous richness was 48% greater during the wet season, when all species were encountered. Richness varied substantially among streams, from a maximum of 17 (2 streams that were diversity hot spots) to none (3 streams). The most widespread diadromous fish was Glossogobius giuris (71% frequency of occurrence), followed by Mugil cephalus (58% occurrence) and Eleotris oxycephala (50% occurrence). The remaining 25 diadromous fishes occurred in fewer than half of the streams; 12 species were confined to a single stream and may be locally threatened. There were conspicuous spatial differences in diadromous assemblages across HKSAR, despite its limited extent (1114 km2), the proximity of the surveyed streams, and the broad geographic distribution of most species. Regional species assemblages were influenced by localized habitat characteristics, with a noticeable distinction between areas with and without large, fast-flowing, and highly oxygenated streams. The presence of in-stream barriers (weirs: 0.3-8.7 m high) did not affect spatial patterns in species assemblages, although, on average, diadromous richness was lower in weir-obstructed streams (4.0 vs. 6.9 species in unobstructed streams). In total, 18 species were confined to unobstructed streams or sections below weirs, whereas the remaining 10 species were recorded both above and below weirs. Only the mottled eel (Anguilla marmorata) and a goby (Stiphodon multisquamus) were able to ascend weirs over 2 m. Although at least 400 m of the lower course of each stream was sampled, diadromous fishes were confined to the first 300 m in 12 of the 13 weir-obstructed streams. Remarkably, the tally of 32 diadromous species in HKSAR exceeds the 19 known from mainland China, highlighting the need for further research on composition and conservation status of diadromous fishes.

Boldness, movement and exploration tendency in round goby (Neogobius melanostomus) in Southern Ontario.

Invasive species have the potential to damage ecosystems outside their native range. At an invasion front, individuals are faced with the unfamiliar conditions of a novel environment. Therefore, certain behavioural traits such as boldness and movement likely play a role in invasion ecology. If behavioural traits of this kind are influenced by differing selection pressures between demographic groups of the same species, this could have broad implications for the management of expanding invasion fronts. To determine whether the invasive round goby (Neogobius melanostomus, Pallas, 1814) exhibits sex- and habitat-based differences in boldness and movement across the invasion front, the authors assessed individual movement and exploration tendency under controlled lab settings using video analysis in a behavioural assay. N. melanostomus from lakes tended to be bolder than those from streams, and females tended to be bolder than males. This study provides evidence for sex- and habitat-based differences in behaviour in this globally invasive species that the authors hope will assist in forming the foundation for contextually appropriate management strategies.

Alternative reproductive tactics in goby fishes of the Caspian Sea.

Here we report on the reproductive morphology and histology of three Gobiidae species (the Caspian monkey goby, Neogobius pallasi; the Caspian goby, Neogobius caspius; and the round goby, Neogobius melanostomus) from the Iranian coastline of the Caspian Sea. Based on ageing, reproductive histology, and internal and external morphological measures, it appears that all three of these goby species have two types of reproductive males, a large courting, territorial, male type and a small cuckolding parasitic male type, a phenomenon known as alternative reproductive tactics (ART). Although ARTs have been reported previously for one of these species, the round goby, all reports stem from its invasive range; ARTs have never been reported before in any fish species in the Caspian Sea. In all three goby species there was a large, older male type, with a wide head, dark body colouration, and a large investment in accessory glands (AG), an organ important for female attraction and parental care. But there was also a small, light, younger male type, with a narrow head, longer urogenital papilla, and little investment in AGs. The Caspian goby were the largest of the three species, and in this species the smaller, lighter, presumably cuckolding male morph was quite rare (only about 5% of the reproductive male population). In contrast, many of the round goby and monkey goby males were the small, lighter parasitic type, making up nearly half the population of reproductive males (48% and 40%, respectively). Round goby and Caspian goby males had a prominent mesorchial gland, a fibrous sheath of pheromone-releasing connective tissue that attaches the testes to the dorsal body wall, but all the monkey goby specimens examined lacked this structure. Although ARTs are well documented across fish species and appear to be particularly common in gobies, our study provides the first evidence for ARTs in goby fishes from the Caspian Sea.

First record in Europe of the Asian gobiid, shimofuri (marbled) goby Tridentiger bifasciatus Steindachner, 1881.

In November 2022, two shimofuri (marbled) gobies Tridentiger bifasciatus Steindachner, 1881 were caught in the Gent-Terneuzen shipping canal, Belgium. It is the first record of the species in Europe. Outside its native distribution area in Japan, China and South Korea, thus far the shimofuri goby has been observed only on the West Coast of the USA, where established populations exist. The introduction vector potentially was ballast water exchange.

Links between the timing of life-history transitions and dietary and morphological variation during early life history in the sand goby, Pomatoschistus minutus.

Bipartite life histories involve a suite of morphological changes that support the pelagic to demersal transition and an expanded range of prey options and microhabitats. Pelagic individuals are thought to shift (settle) to their preferred benthic habitat at the earliest opportunity once they have attained a minimum level of morphological competency to access their new environment. In theory, early changes in larval morphology (collectively termed 'metamorphosis'), habitat and diet-a measure of habitat-use-ought to be synchronous. Yet relationships may be decoupled by factors linked to behaviour, prey availability or morphological complexity, and few descriptions exist to allow such synchrony to be assessed. The sand goby, Pomatoschistus minutus, is a common coastal fish across north-western Europe, with a size at larval metamorphosis and settlement of around 10 and 16-18 mm standard length (SL), respectively. We sampled shoreline larval and juvenile populations to examine relationships between morphology, diet and life stage. Prey diversity increased with body length; however, dietary change was clearest at 16-18 mm SL, with a reduction in calanoid copepods and shift to larger prey such as Nereis polychaetes and mysid and amphipod crustacea. Early growth in five prey capture and processing morphologies was rapid. Four of these showed a subsequent marked shift to slower growth, but none of these changes were aligned with size at metamorphosis and only that of mouth width coincided with body size at settlement. Early life history in P. minutus appears geared towards a protracted morphological reorganization prior to demersal life and an alternative suite of prey resources. Larval metamorphosis seems to be of limited consequence in this regard. Comparable studies of other Baltic Sea fishes would confirm whether these dynamics relate to shared environmental pressures or to factors intrinsic to P. minutus biology.

Phylogenetics and the evolution of terrestriality in mudskippers (Gobiidae: Oxudercinae).

The initial vertebrate conquest of land by stegocephalians (Sarcopterygia) allowed access to new resources and exploitation of untapped niches precipitating a major phylogenetic diversification. However, a paucity of fossils has left considerable uncertainties about phylogenetic relationships and the eco-morphological stages in this key transition in Earth history. Among extant actinopterygians, three genera of mudskippers (Gobiidae: Oxudercinae), Boleophthalmus, Periophthalmus and Periophthalmodon are the most terrestrialized, with vertebral, appendicular, locomotory, respiratory, and epithelial specializations enabling overland excursions up to 14 h. Unlike early stegocephalians, the ecologies and morphologies of the 45 species of oxudercines are well known, making them viable analogs for the initial vertebrate conquest of land. Nevertheless, they have received little phylogenetic attention. We compiled the largest molecular dataset to date, with 29 oxudercine species, and 5 nuclear and mitochondrial loci. Phylogenetic and comparative analyses revealed strong support for two independent terrestrial transitions, and a complex suit of ecomorphological forms in estuarine environments. Furthermore, neither Oxudercinae nor their presumed sister-group the eel gobies (Amblyopinae, a group of elongated gobies) were monophyletic with respect to each other, requiring a merging of these two subfamilies and revealing an expansion of phenotypic variation within the "mudskipper" clade. We did not find support for the expected linear model of ecomorphological and locomotory transition from fully aquatic, to mudswimming, to pectoral-aided mudswimming, to lobe-finned terrestrial locomotion proposed by earlier morphological studies. This high degree of convergent or parallel transitions to terrestriality, and apparent divergent directions of estuarine adaptation, promises even greater potential for this clade to illuminate the conquest of land. Future work should focus on these less-studied species with "transitional" and other mud-habitat specializations to fully resolve the dynamics of this diversification.

Female Preference for Males Varies with Resource Availability for Reproduction in the Freshwater Goby Tridentiger brevispinis.

Animals with resource-based breeding systems show varying sexual selection modes driven by resource availability. Males of the freshwater goby Tridentiger brevispinis (Pisces: Gobiidae), which are larger, with a longer dorsal-fin, than females, make nests under stones, and provide exclusive parental care of eggs in the nests. For this goby, the nest sites are crucial resources for reproduction. Laboratory experiments that controlled for nest-site abundance were conducted to investigate the effects of nest-site availability on male-male competition and female mate choice. Nest-holding males were on average larger than the average size of all males combined under conditions of low nest-site availability. This result indicates a greater advantage for large males in intrasexual competition for nest sites under conditions of low nest-site availability. Furthermore, egg-guarding males under conditions of low nest-site availability were larger than nest-holding males without eggs. Under conditions of high nest-site availability, the dorsal-fin length of nest-holding males with eggs were longer than those without eggs. These findings suggest that female preference for mates varies qualitatively with nest-site availability. This variation in female preference might result from mate availability that depends upon nest-site availability. The current study concludes that in nest-spawning fishes, nest-site availability can affect the intensity of male-male competition and cause variations of female preference for mates.

Surprising Pseudogobius: Molecular systematics of benthic gobies reveals new insights into estuarine biodiversity (Teleostei: Gobiiformes).

Snubnose gobies (genus Pseudogobius: Gobionellinae) are ubiquitous to, and important components of, estuarine ecosystems of the Indo-west Pacific. These small benthic fishes occur in freshwater, brackish and marine habitats such as mangroves, sheltered tide pools and lowland streams, and represent a model group for understanding the biodiversity and biogeography of estuarine fauna. To develop the species-level framework required for a concurrent morphological taxonomic appraisal, we undertook thorough sampling around the extensive Australian coastline, referenced to international locations, as part of a molecular systematic review using both nuclear and mitochondrial markers. The results indicate that while there are currently eight recognised species, the true diversity is close to double this, with a hotspot of endemism located in Australia. Complicated patterns were observed in southern Australia owing to two differing zones of introgression/admixture. Key drivers of diversity in the group appear to include plate tectonics, latitude, and historic barriers under glacial maxima, where an interplay between ready dispersal and habitat specialisation has led to regional panmixia but frequent geographic compartmentalisation within past and present landscapes. The findings have significant implications for biodiversity conservation, coastal and estuarine development, the basic foundations of field ecology, and for applied use such as in biomonitoring.

Sperm adaptation in relation to salinity in three goby species.

In externally fertilizing species, the gametes of both males and females are exposed to the influences of the environment into which they are released. Sperm are sensitive to abiotic factors such as salinity, but they are also affected by biotic factors such as sperm competition. In this study, the authors compared the performance of sperm of three goby species, the painted goby, Pomatoschistus pictus, the two-spotted goby, Pomatoschistus flavescens, and the sand goby, Pomatoschistus minutus. These species differ in their distributions, with painted goby having the narrowest salinity range and sand goby the widest. Moreover, data from paternity show that the two-spotted goby experiences the least sperm competition, whereas in the sand goby sperm competition is ubiquitous. The authors took sperm samples from dissected males and exposed them to high salinity water (31 PSU) representing the North Sea and low salinity water (6 PSU) representing the brackish Baltic Sea Proper. They then used computer-assisted sperm analysis to measure the proportion of motile sperm and sperm swimming speed 10 min and 20 h after sperm activation. The authors found that sperm performance depended on salinity, but there seemed to be no relationship to the species' geographical distribution in relation to salinity range. The species differed in the proportion of motile sperm, but there was no significant decrease in sperm motility during 20 h. The sand goby was the only species with motile sperm after 72 h.

Supermatrix phylogeny resolves goby lineages and reveals unstable root of Gobiaria.

Gobies, sleepers, and cardinalfishes represent major clades of a species rich radiation of small bodied, ecologically diverse percomorphs (Gobiaria). Molecular phylogenetics has been crucial to resolving broad relationships of sleepers and gobies (Gobioidei), but the phylogenetic placements of cardinalfishes and nurseryfishes, as reciprocal or sequential sister clades to Gobioidei, are uncertain. In order to evaluate relationships among and within families we used a phylogenetic data mining approach to generate densely sampled trees inclusive of all higher taxa. We utilized conspecific amino acid homology to improve alignment accuracy, included ambiguously identified taxa to increase taxon sampling density, and resampled individual gene alignments to filter rogue sequences before concatenation. This approach yielded the most comprehensive tree yet of Gobiaria, inferred from a sparse (17 percent-complete) supermatrix of one ribosomal and 22 protein coding loci (18,065 characters), comprised of 50 outgroup and 777 ingroup taxa, representing 32 percent of species and 68 percent of genera. Our analyses confirmed the lineage-based classification of gobies with strong support, identified sleeper clades with unforeseen levels of systematic uncertainty, and quantified competing phylogenetic signals that confound resolution of the root topology. We also discovered that multilocus data completeness was related to maximum likelihood branch support, and verified that the phylogenetic uncertainty of shallow relationships observed within goby lineages could largely be explained by supermatrix sparseness. These results demonstrate the potential and limits of publicly available sequence data for producing densely-sampled phylogenetic trees of exceptionally biodiverse groups.

Potential roles of smell and taste in the orientation behaviour of coral-reef fish larvae: insights from morphology.

An ontogenetic analysis of the olfactory organ and the number and distribution of internal taste buds was carried out in two neon gobies (Elacatinus lori and Elacatinus colini) with the goal of revealing morphological trends that might inform an understanding of the roles of olfaction and taste in larval orientation behaviour. The pattern of development of the olfactory organ is unremarkable and enclosure of the olfactory epithelium occurs concurrently with metamorphosis and settlement in both species. Like other gobies, juvenile and adult E. lori and E. colini lack complex olfactory lamellae, and lack the accessory nasal sacs present in some adult gobies that could facilitate active olfactory ventilation (i.e., sniffing). A small number of internal taste buds are present at hatch with most found in the caudal region of the buccal cavity (on gill arches, roof of buccal cavity). As taste bud number increases, they demonstrate an anterior spread to the lips, buccal valves and tongue (i.e., tissue covering the basihyal). In the absence of an active ventilatory mechanism for the olfactory organs, the water that moves through the buccal cavity with cyclic gill ventilation may provide chemical cues allowing the internal taste buds to play a role in chemical-mediated orientation and reef-seeking behavior in pelagic larval fishes.

Acoustic communication in marine shallow waters: testing the acoustic adaptive hypothesis in sand gobies.

Acoustic communication is an important part of social behaviour of fish species that live or breed in shallow noisy waters. Previous studies have shown that some fish species exploit a quiet window in the background noise for communication. However, it remains to be examined whether hearing abilities and sound production of fish are adapted to marine habitats presenting high hydrodynamism. Here, we investigated whether the communication system of the painted (Pomatoschistus pictus) and the marbled (Pomatoschistus marmoratus) gobies is adapted to enhance sound transmission and reception in Atlantic shallow water environments. We recorded and measured the sound pressure levels of social vocalisations of both species, as well as snapshots of ambient noise of habitats characterised by different hydrodynamics. Hearing thresholds (in terms of both sound pressure and particle acceleration) and responses to conspecific signals were determined using the auditory evoked potential recording technique. We found that the peak frequency range (100-300 Hz) of acoustic signals matched the best hearing sensitivity in both species and appeared well adapted for short-range communication in Atlantic habitats. Sandy/rocky exposed beaches presented a quiet window, observable even during the breaking of moderate waves, coincident with the main sound frequencies and best hearing sensitivities of both species. Our data demonstrate that the hearing abilities of these gobies are well suited to detect conspecific sounds within typical interacting distances (a few body lengths) in Atlantic shallow waters. These findings lend support to the acoustic adaptive hypothesis, under the sensory drive framework, proposing that signals and perception systems coevolve to be effective within local environment constraints.

Cleaning interactions by gobies on a tropical eastern Pacific coral reef.

The present study describes the cleaning interactions among species of cleaner gobies Tigrigobius spp. and Elacatinus puncticulatus (family Gobiidae) and the client fish species they clean in a coral reef of Gorgona Island, Colombia. In 419 cleaning events, we observed 27 species acting as clients of Tigrigobius spp., whereas only nine were clients of E. puncticulatus. Paranthias colonus and Cephalopholis panamensis were the species most commonly cleaned by Tigrigobius spp., while Ophioblennius steindachneri and Stegastes acalpulcoensis were the clients most commonly cleaned by E. puncticulatus. The abundance (but not the body size) of clients was an important variable predicting the cleaning frequency observed for clients of Tigrigobius spp., but this was not the case for clients of E. puncticulatus. Additionally, Tigrigobius spp. preferred cleaning planktivores, sessile invertebrate feeders and territorial herbivores (Ivlev's index >0·15), whereas E. puncticulatus did not exhibit any preference. We observed two major peaks of cleaning activity for Tigrigobius spp., one in the early morning and another one in the late afternoon. These results suggest that Tigrigobius spp. is a specialized cleaner goby, whereas E. puncticulatus is a facultative cleaner that cleans sporadically.

Change of strategy or always overlooked? Third recent record of the beardless tadpole goby Benthophiloides brauneri from an inland location far from the coast.

The present study documents three specimens of Benthophiloides brauneri from the River Dniester. It is one of the rarest freshwater fishes in Europe, living along the western coast of the Black and Asov Seas, but these new records are 200 river km from the coast. We discuss whether the species has recently started to establish inland populations or inland specimens have previously been overlooked.

The mitochondrial genome sequences of the round goby and the sand goby reveal patterns of recent evolution in gobiid fish.

BACKGROUND: Vertebrate mitochondrial genomes are optimized for fast replication and low cost of RNA expression. Accordingly, they are devoid of introns, are transcribed as polycistrons and contain very little intergenic sequences. Usually, vertebrate mitochondrial genomes measure between 16.5 and 17 kilobases (kb). RESULTS: During genome sequencing projects for two novel vertebrate models, the invasive round goby and the sand goby, we found that the sand goby genome is exceptionally small (16.4 kb), while the mitochondrial genome of the round goby is much larger than expected for a vertebrate. It is 19 kb in size and is thus one of the largest fish and even vertebrate mitochondrial genomes known to date. The expansion is attributable to a sequence insertion downstream of the putative transcriptional start site. This insertion carries traces of repeats from the control region, but is mostly novel. To get more information about this phenomenon, we gathered all available mitochondrial genomes of Gobiidae and of nine gobioid species, performed phylogenetic analyses, analysed gene arrangements, and compared gobiid mitochondrial genome sizes, ecological information and other species characteristics with respect to the mitochondrial phylogeny. This allowed us amongst others to identify a unique arrangement of tRNAs among Ponto-Caspian gobies. CONCLUSIONS: Our results indicate that the round goby mitochondrial genome may contain novel features. Since mitochondrial genome organisation is tightly linked to energy metabolism, these features may be linked to its invasion success. Also, the unique tRNA arrangement among Ponto-Caspian gobies may be helpful in studying the evolution of this highly adaptive and invasive species group. Finally, we find that the phylogeny of gobiids can be further refined by the use of longer stretches of linked DNA sequence.

Sound-production mechanism in Pomatoschistus pictus.

Fish acoustic signals play a major role during agonistic and reproductive interactions. Among the sound-generating fish, Gobiidae, a large fish family with 1866 valid species, is one of the most studied groups of acoustic fishes, with sound production being documented in a number of species. Paradoxically, the sound-producing mechanism remains poorly studied in this group. The painted goby, Pomatoschistus pictus, produces two distinct sounds called drums and thumps. A combination of morphological and experimental analyses involving high-speed videos synchronized with sound recordings supports that drums are produced during lateral head movements involving at least the alternate contractions of the levator pectoralis muscles originating on the skull and inserting on the pectoral girdle. These movements are reported in many Gobiidae species, suggesting the pectoral-girdle-based mechanism is common in the family and could have evolved from locomotory movements.

Evolution of bidirectional sex change and gonochorism in fishes of the gobiid genera Trimma, Priolepis, and Trimmatom.

Size-advantage and low-density models have been used to explain how mating systems favor hermaphroditism or gonochorism. However, these models do not indicate historical transitions in sexuality. Here, we investigate the evolution of bidirectional sex change and gonochorism by phylogenetic analysis using the mitochondrial gene of the gobiids Trimma (31 species), Priolepis (eight species), and Trimmatom (two species). Trimma and Priolepis formed a clade within the sister group Trimmatom. Gonadal histology and rearing experiments revealed that Trimma marinae, Trimma nasa, and Trimmatom spp. were gonochoric, whereas all other Trimma and Priolepis spp. were bidirectional sex changers or inferred ones. A maximum-likelihood reconstruction analysis demonstrated that the common ancestor of the three genera was gonochoristic. Bidirectional sex change probably evolved from gonochorism in a common ancestor of Trimma and Priolepis. As the gonads of bidirectional sex changers simultaneously contain mature ovarian and immature testicular components or vice versa, individuals are always potentially capable of functioning as females or males, respectively. Monogamy under low-density conditions may have been the ecological condition for the evolution of bidirectional sex change in a common ancestor. As T. marinae and T. nasa are a monophyletic group, gonochorism should have evolved from bidirectional sex change in a common ancestor.

Repeated invasions into the twilight zone: evolutionary origins of a novel assemblage of fishes from deep Caribbean reefs.

Mesophotic and deeper reefs of the tropics are poorly known and underexplored ecosystems worldwide. Collectively referred to as the 'twilight zone', depths below ~30-50 m are home to many species of reef fishes that are absent from shallower depths, including many undescribed and endemic species. We currently lack even a basic understanding of the diversity and evolutionary origins of fishes on tropical mesophotic reefs. Recent submersible collections in the Caribbean have provided new specimens that are enabling phylogenetic reconstructions that incorporate deep-reef representatives of tropical fish genera. Here, we investigate evolutionary depth transitions in the family Gobiidae (gobies), the most diverse group of tropical marine fishes. Using divergence-time estimation coupled with stochastic character mapping to infer the timing of shallow-to-deep habitat transitions in gobies, we demonstrate at least four transitions from shallow to mesophotic depths. Habitat transitions occurred in two broad time periods (Miocene, Pliocene-Pleistocene), and may have been linked to the availability of underutilized niches, as well as the evolution of morphological/behavioural adaptations for life on deep reefs. Further, our analysis shows that at least three evolutionary lineages that invaded deep habitats subsequently underwent speciation, reflecting another unique mode of radiation within the Gobiidae. Lastly, we synthesize depth distributions for 95 species of Caribbean gobies, which reveal major bathymetric faunal breaks at the boundary between euphotic and mesophotic reefs. Ultimately, our study is the first rigorous investigation into the origin of Caribbean deep-reef fishes and provides a framework for future studies that utilize rare, deep-reef specimens.