Impact of Pleistocene Eustatic Fluctuations on Evolutionary Dynamics in Southeast Asian Biodiversity Hotspots.

Pleistocene climatic fluctuations (PCF) are frequently highlighted as important evolutionary engines that triggered cycles of biome expansion and contraction. Although there is ample evidence of the impact of PCF on biodiversity of continental biomes, the consequences in insular systems depend on the geology of the islands and the ecology of the taxa inhabiting them. The idiosyncratic aspects of insular systems are exemplified by the islands of the Sunda Shelf in Southeast Asia (Sundaland), where PCF-induced eustatic fluctuations had complex interactions with the geology of the region, resulting in high species diversity and endemism. Emergent land in Southeast Asia varied drastically with sea-level fluctuations during the Pleistocene. Climate-induced fluctuations in sea level caused temporary connections between insular and continental biodiversity hotspots in Southeast Asia. These exposed lands likely had freshwater drainage systems that extended between modern islands: the Paleoriver Hypothesis. Built upon the assumption that aquatic organisms are among the most suitable models to trace ancient river boundaries and fluctuations of landmass coverage, the present study aims to examine the evolutionary consequences of PCF on the dispersal of freshwater biodiversity in Southeast Asia. Time-calibrated phylogenies of DNA-delimited species were inferred for six species-rich freshwater fish genera in Southeast Asia (Clarias, Channa, Glyptothorax, Hemirhamphodon, Dermogenys, Nomorhamphus). The results highlight rampant cryptic diversity and the temporal localization of most speciation events during the Pleistocene, with 88% of speciation events occurring during this period. Diversification analyses indicate that sea-level-dependent diversification models poorly account for species proliferation patterns for all clades excepting Channa. Ancestral area estimations point to Borneo as the most likely origin for most lineages, with two waves of dispersal to Sumatra and Java during the last 5 myr. Speciation events are more frequently associated with boundaries of the paleoriver watersheds, with 60%, than islands boundaries, with 40%. In total, one-third of speciation events are inferred to have occurred within paleorivers on a single island, suggesting that habitat heterogeneity and factors other than allopatry between islands substantially affected diversification of Sundaland fishes. Our results suggest that species proliferation in Sundaland is not wholly reliant on Pleistocene sea-level fluctuations isolating populations on different islands. [Dispersal; diversification; eustatic fluctuations; freshwater fishes; insular systems; Milankovitch cycles; paleoenvironments; vicariance.].

DNA reconciles morphology and colouration in the drunk blenny genus Scartichthys (Teleostei: Blenniidae) and provides insights into their evolutionary history.

The blenniids of the genus Scartichthys are one of the most common fishes of Central and South American Pacific coastal reefs. This being said, Scartichthys spp. remain difficult to identify in the field, and identification is particularly challenging across the c. 6000 km where three of the four currently accepted species are known to occur in sympatry. A reason for this is that the main taxonomic characters from traditional taxonomy are indeed elusive. In addition, at the same time, species can display multiple colour patterns in the field, depending on their ontogenetic stage, habitat association and reproductive behaviour. Overall, molecular characterization is warranted to help address these issues. In this study, the authors have used a novel approach to revise the genus by combining colouration, morphological and molecular data of representative specimens of the four currently valid species and seven described colour patterns. From this, the authors show that only three of the four species should be considered as valid; Scartichthys gigas (Steindachner, 1876), Scartichthys variolatus (Valenciennes, 1836) and Scartichthys viridis (Valenciennes, 1836), whereas Scartichthys crapulatus (Williams, 1990) should be synonymized with S. viridis. In the same way, the analyses in this study show that one of the colour patterns attributed so far only to S. gigas is characteristic of the juvenile stages of S. viridis. The time-calibrated phylogeny of this study shows that this genus is relatively young and that the estimated time of divergence between S. gigas and S. viridis is c. 1.71 Ma. In comparison, the Desventuradas and Juan Fernandez Islands endemic S. variolatus diverged c. 1.95 Ma. The results of this study help to clarify the taxonomy of Scartichthys.

Coping with Pleistocene climatic fluctuations: Demographic responses in remote endemic reef fishes.

Elucidating demographic history during the settlement of ecological communities is crucial for properly inferring the mechanisms that shape patterns of species diversity and their persistence through time. Here, we used genomic data and coalescent-based approaches to elucidate for the first time the demographic dynamics associated with the settlement by endemic reef fish fauna of one of the most remote peripheral islands of the Pacific Ocean, Rapa Nui (Easter Island). We compared the demographic history of nine endemic species in order to explore their demographic responses to Pleistocene climatic fluctuations. We found that species endemic to Rapa Nui share a common demographic history, as signatures of population expansions were retrieved for almost all of the species studied here, and synchronous demographic expansions initiated during the last glacial period were recovered for more than half of the studied species. These results suggest that eustatic fluctuations associated with Milankovitch cycles have played a central role in species demographic histories and in the final stage of the community assembly of many Rapa Nui reef fishes. Specifically, sea level lowstands resulted in the maximum reef habitat extension for Rapa Nui endemic species; we discuss the potential role of seamounts in allowing endemic species to cope with Pleistocene climatic fluctuations, and we highlight the importance of local historical processes over regional ones. Overall, our results shed light on the mechanisms by which endemism arises and is maintained in peripheral reef fish fauna.

Endemic and widespread coral reef fishes have similar mitochondrial genetic diversity.

Endemic species are frequently assumed to have lower genetic diversity than species with large distributions, even if closely related. This assumption is based on research from the terrestrial environment and theoretical evolutionary modelling. We test this assumption in the marine environment by analysing the mitochondrial genetic diversity of 33 coral reef fish species from five families sampled from Pacific Ocean archipelagos. Surprisingly, haplotype and nucleotide diversity did not differ significantly between endemic and widespread species. The probable explanation is that the effective population size of some widespread fishes locally is similar to that of many of the endemics. Connectivity across parts of the distribution of the widespread species is probably low, so widespread species can operate like endemics at the extreme or isolated parts of their range. Mitochondrial genetic diversity of many endemic reef fish species may not either limit range size or be a source of vulnerability.

Two new fish species of the subfamily Anthiinae (Perciformes, Serranidae) from the Marquesas.

Two new species of anthiine fishes are described from the Marquesas Islands, French Polynesia. Plectranthias flammeus was found at depths from 20-45 m and is characterized by dorsal rays X, 14 or 15, with spines 1-6 bearing fleshy white tabs at their tips, longest fleshy tab on spine 4; 14 unbranched pectoral rays; lateral line incomplete with 16-17 tubed scales; preopercle with 8-10 small spines along posterior margin and 2 antrorse spines on ventral margin; broad, fiery red-orange streak across lower cheek; head and body with irregularly spaced maroon-ringed yellow blotches on a white background; pair of small dark oblong spots (red with black centers in life) on the bases of the middle rays of the caudal fin. Pseudanthias oumati was found on the outer reef slope of Fatu Hiva at a depth of 50-55 m and is characterized by 3rd dorsal spine elongate and tipped with fleshy yellow filament extending beyond tip of spine; lateral-line scales 43; gill rakers 10 + 28; no papillae on posterior edge of orbit; front of upper lip not thickened (male condition unknown); caudal fin lunate; color of female yellow, all fins yellow with narrow magenta margin (except pectoral fin, which lacks magenta); no stripe from snout to pectoral base; small scales located on basal quarter of soft-dorsal fin from segmented rays 1-12; dorsal profile of head slightly concave.

Exploring the vertebrate fauna of the Bird's Head Peninsula (Indonesia, West Papua) through DNA barcodes.

Biodiversity knowledge is widely heterogeneous across the Earth's biomes. Some areas, due to their remoteness and difficult access, present large taxonomic knowledge gaps. Mostly located in the tropics, these areas have frequently experienced a fast development of anthropogenic activities during the last decades and are therefore of high conservation concerns. The biodiversity hotspots of Southeast Asia exemplify the stakes faced by tropical countries. While the hotspots of Sundaland (Java, Sumatra, Borneo) and Wallacea (Sulawesi, Moluccas) have long attracted the attention of biologists and conservationists alike, extensive parts of the Sahul area, in particular the island of New Guinea, have been much less explored biologically. Here, we describe the results of a DNA-based inventory of aquatic and terrestrial vertebrate communities, which was the objective of a multidisciplinary expedition to the Bird's Head Peninsula (West Papua, Indonesia) conducted between 17 October and 20 November 2014. This expedition resulted in the assembly of 1005 vertebrate DNA barcodes. Based on the use of multiple species-delimitation methods (GMYC, PTP, RESL, ABGD), 264 molecular operational taxonomic units (MOTUs) were delineated, among which 75 were unidentified and an additional 48 were considered cryptic. This study suggests that the diversity of vertebrates of the Bird's Head is severely underestimated and considerations on the evolutionary origin and taxonomic knowledge of these biotas are discussed.

Identifying coral reef fish larvae through DNA barcoding: a test case with the families Acanthuridae and Holocentridae.

A reference collection of COI barcode (650 bp) for the Pacific Society Islands has been constituted for 22 species of Acanthuridae and 16 species of Holocentridae. Divergence between congeneric species was on average 20-fold to 87-fold higher than divergence between conspecific sequences and this set of DNA-identifiers was used to identify 40 larvae of both families. All larvae sequenced could be identified to species using DNA-barcodes. Pools of larvae constitute multi-specific assemblages and no additional species compared to adult reef communities were sampled in larval pools, suggesting that the larval assemblages originated from adult communities on neighboring reefs.

Assessing species diversity of Coral Triangle artisanal fisheries: A DNA barcode reference library for the shore fishes retailed at Ambon harbor (Indonesia).

The Coral Triangle (CT), a region spanning across Indonesia and Philippines, is home to about 4,350 marine fish species and is among the world's most emblematic regions in terms of conservation. Threatened by overfishing and oceans warming, the CT fisheries have faced drastic declines over the last decades. Usually monitored through a biomass-based approach, fisheries trends have rarely been characterized at the species level due to the high number of taxa involved and the difficulty to accurately and routinely identify individuals to the species level. Biomass, however, is a poor proxy of species richness, and automated methods of species identification are required to move beyond biomass-based approaches. Recent meta-analyses have demonstrated that species richness peaks at intermediary levels of biomass. Consequently, preserving biomass is not equal to preserving biodiversity. We present the results of a survey to estimate the shore fish diversity retailed at the harbor of Ambon Island, an island located at the center of the CT that display exceptionally high biomass despite high levels of threat, while building a DNA barcode reference library of CT shore fishes targeted by artisanal fisheries. We sampled 1,187 specimens and successfully barcoded 696 of the 760 selected specimens that represent 202 species. Our results show that DNA barcodes were effective in capturing species boundaries for 96% of the species examined, which opens new perspectives for the routine monitoring of the CT fisheries.

Biodiversity inventory of the grey mullets (Actinopterygii: Mugilidae) of the Indo-Australian Archipelago through the iterative use of DNA-based species delimitation and specimen assignment methods.

DNA barcoding opens new perspectives on the way we document biodiversity. Initially proposed to circumvent the limits of morphological characters to assign unknown individuals to known species, DNA barcoding has been used in a wide array of studies where collecting species identity constitutes a crucial step. The assignment of unknowns to knowns assumes that species are already well identified and delineated, making the assignment performed reliable. Here, we used DNA-based species delimitation and specimen assignment methods iteratively to tackle the inventory of the Indo-Australian Archipelago grey mullets, a notorious case of taxonomic complexity that requires DNA-based identification methods considering that traditional morphological identifications are usually not repeatable and sequence mislabeling is common in international sequence repositories. We first revisited a DNA barcode reference library available at the global scale for Mugilidae through different DNA-based species delimitation methods to produce a robust consensus scheme of species delineation. We then used this curated library to assign unknown specimens collected throughout the Indo-Australian Archipelago to known species. A second iteration of OTU delimitation and specimen assignment was then performed. We show the benefits of using species delimitation and specimen assignment methods iteratively to improve the accuracy of specimen identification and propose a workflow to do so.

Disentangling the taxonomy of the subfamily Rasborinae (Cypriniformes, Danionidae) in Sundaland using DNA barcodes.

Sundaland constitutes one of the largest and most threatened biodiversity hotspots; however, our understanding of its biodiversity is afflicted by knowledge gaps in taxonomy and distribution patterns. The subfamily Rasborinae is the most diversified group of freshwater fishes in Sundaland. Uncertainties in their taxonomy and systematics have constrained its use as a model in evolutionary studies. Here, we established a DNA barcode reference library of the Rasborinae in Sundaland to examine species boundaries and range distributions through DNA-based species delimitation methods. A checklist of the Rasborinae of Sundaland was compiled based on online catalogs and used to estimate the taxonomic coverage of the present study. We generated a total of 991 DNA barcodes from 189 sampling sites in Sundaland. Together with 106 previously published sequences, we subsequently assembled a reference library of 1097 sequences that covers 65 taxa, including 61 of the 79 known Rasborinae species of Sundaland. Our library indicates that Rasborinae species are defined by distinct molecular lineages that are captured by species delimitation methods. A large overlap between intraspecific and interspecific genetic distance is observed that can be explained by the large amounts of cryptic diversity as evidenced by the 166 Operational Taxonomic Units detected. Implications for the evolutionary dynamics of species diversification are discussed.

A DNA barcode reference library of French Polynesian shore fishes.

The emergence of DNA barcoding and metabarcoding opened new ways to study biological diversity, however, the completion of DNA barcode libraries is fundamental for such approaches to succeed. This dataset is a DNA barcode reference library (fragment of Cytochrome Oxydase I gene) for 2,190 specimens representing at least 540 species of shore fishes collected over 10 years at 154 sites across the four volcanic archipelagos of French Polynesia; the Austral, Gambier, Marquesas and Society Islands, a 5,000,000 km2 area. At present, 65% of the known shore fish species of these archipelagoes possess a DNA barcode associated with preserved, photographed, tissue sampled and cataloged specimens, and extensive collection locality data. This dataset represents one of the most comprehensive DNA barcoding efforts for a vertebrate fauna to date. Considering the challenges associated with the conservation of coral reef fishes and the difficulties of accurately identifying species using morphological characters, this publicly available library is expected to be helpful for both authorities and academics in various fields.

Evidence of cryptic species in the blenniid Cirripectesalboapicalis species complex, with zoogeographic implications for the South Pacific.

Rapa Nui, commonly known as Easter Island (Chile), is one of the most isolated tropical islands of the Pacific Ocean. The island location of Rapa Nui makes it the easternmost point of the geographic ranges for many western Pacific fish species that are restricted to the subtropical islands south of 20°S latitude. The blenniid fish species Cirripectesalboapicalis has been thought to have one of the most extensive geographic distribution ranges among these southern subtropical fish species, extending from the southern Great Barrier Reef to Rapa Nui. A phylogenetic analysis was conducted to determine the taxonomic status of the species. The results provide genetic evidence that suggests that this formerly South Pacific-wide species comprises at least three cryptic species with allopatric geographic distributions. The analyses reveal the geographic distributions of these clades and their genetic relationships with each other, and with other species within the genus Cirripectes. The processes that culminated in the current geographic distribution of this species complex and the zoogeographic implications of this finding for the South Pacific region are discussed.

Population expansions dominate demographic histories of endemic and widespread Pacific reef fishes.

Despite the unique nature of endemic species, their origin and population history remain poorly studied. We investigated the population history of 28 coral reef fish species, close related, from the Gambier and Marquesas Islands, from five families, with range size varying from widespread to small-range endemic. We analyzed both mitochondrial and nuclear sequence data using neutrality test and Bayesian analysis (EBSP and ABC). We found evidence for demographic expansions for most species (24 of 28), irrespective of range size, reproduction strategy or archipelago. The timing of the expansions varied greatly among species, from 8,000 to 2,000,000 years ago. The typical hypothesis for reef fish that links population expansions to the Last Glacial Maximum fit for 14 of the 24 demographic expansions. We propose two evolutionary processes that could lead to expansions older than the LGM: (a) we are retrieving the signature of an old colonization process for widespread, large-range endemic and paleoendemic species or (b) speciation; the expansion reflects the birth of the species for neoendemic species. We show for the first time that the demographic histories of endemic and widespread reef fish are not distinctly different and suggest that a number of processes drive endemism.

Macropharyngodon pakoko, a new species of wrasse (Teleostei: Labridae) endemic to the Marquesas Islands, French Polynesia.

A new species of wrasse, Macropharyngodon pakoko, is described from the Marquesas Islands, bringing the total number of species of the genus Macropharyngodon to 12. Macropharyngodon pakoko was found at depths from 0-42 m and is endemic to the Marquesas Islands. Macropharyngodon pakoko is similar to M. meleagris, which is widely distributed from the central and western Pacific to Cocos-Keeling in the Indian Ocean, but differs genetically and in several coloration characters: males with irregularly curved black humeral blotch with incomplete iridescent blue border; inverted irregular "U"- shaped band on the cheek; a small black spot at the upper base of the pectoral fin; and background color of the body greenish with faint bluish black spots on each scale. Females lack black pigment on the chest posterior to the ventral attachment of the gill membranes; reddish black blotches on the body are widely spaced, particularly on the head where they are more reddish and half the size of those on body; caudal fin with small, bright yellow spots arranged in narrow vertical bands with pale interspaces; pelvic fins pale with three reddish yellow cross-bands; a small black spot at the upper base of the pectoral fin; and small reddish spots along the base of the anal fin. Juveniles have irregular black blotches on the body, a small black spot instead of an ocellus posteriorly on the dorsal fin and lack large black spots and ocellus on the anal fin.