DNA sequences from type specimens and type strains - how to increase their number and improve their annotation in NCBI GenBank and related databases.

Scientific names permit humans and search engines to access knowledge about the biodiversity that surrounds us, and names linked to DNA sequences are playing an ever-greater role in search-and-match identification procedures. Here, we analyze how users and curators of the National Center for Biotechnology Information (NCBI) are flagging and curating sequences derived from nomenclatural type material, which is the only way to improve the quality of DNA-based identification in the long run. For prokaryotes, 18,281 genome assemblies from type strains have been curated by NCBI staff and improve the quality of prokaryote naming. For Fungi, type-derived sequences representing over 21,000 species are now essential for fungus naming and identification. For the remaining eukaryotes, however, the numbers of sequences identifiable as type-derived are minuscule, representing only 1,000 species of arthropods, 8,441 vertebrates, and 430 embryophytes. An increase in the production and curation of such sequences will come from (i) sequencing of types or topotypic specimens in museum collections, (ii) the March 2023 rule changes at the International Nucleotide Sequence Database Collaboration requiring more metadata for specimens, and (iii) efforts by data submitters to facilitate curation, including informing NCBI curators about a specimen's type status. We illustrate different type-data submission journeys and provide best-practice examples from a range of organisms. Expanding the number of type-derived sequences in DNA databases, especially of eukaryotes, is crucial for capturing, documenting, and protecting biodiversity.

Between a Rock and a Hard Polytomy: Phylogenomics of the Rock-Dwelling Mbuna Cichlids of Lake Malawi.

Whole genome sequences are beginning to revolutionize our understanding of phylogenetic relationships. Yet, even whole genome sequences can fail to resolve the evolutionary history of the most rapidly radiating lineages, where incomplete lineage sorting, standing genetic variation, introgression, and other factors obscure the phylogenetic history of the group. To overcome such challenges, one emerging strategy is to integrate results across different methods. Most such approaches have been implemented on reduced representation genomic data sets, but whole genomes should provide the maximum possible evidence approach. Here, we test the ability of single nucleotide polymorphisms extracted from whole genome resequencing data, implemented in an integrative genomic approach, to resolve key nodes in the phylogeny of the mbuna, rock-dwelling cichlid fishes of Lake Malawi, which epitomize the phylogenetic intractability that often accompanies explosive lineage diversification. This monophyletic radiation has diversified at an unparalleled rate into several hundred species in less than 2 million years. Using an array of phylogenomic methods, we consistently recovered four major clades of mbuna, but a large basal polytomy among them. Although introgression between clades apparently contributed to the challenge of phylogenetic reconstruction, reduction of the data set to nonintrogressed sites still did not help to resolve the basal polytomy. On the other hand, relationships among six congeneric species pairs were resolved without ambiguity, even in one case where existing data led us to predict that resolution would be difficult. We conclude that the bursts of diversification at the earliest stages of the mbuna radiation may be phylogenetically unresolvable, but other regions of the tree are phylogenetically clearly supported. Integration of multiple phylogenomic approaches will continue to increase confidence in relationships inferred from these and other whole-genome data sets. [Incomplete lineage sorting; introgression; linkage disequilibrium; multispecies coalescence; rapid radiation; soft polytomy.].

A comprehensive phylogeny of dwarf geckos of the genus Lygodactylus, with insights into their systematics and morphological variation.

The 71 currently known species of dwarf geckos of the genus Lygodactylus are a clade of biogeographic interest due to their occurrence in continental Africa, Madagascar, and South America. Furthermore, because many species are morphologically cryptic, our knowledge of species-level diversity within this genus is incomplete, as indicated by numerous unnamed genetic lineages revealed in previous molecular studies. Here we provide an extensive multigene phylogeny covering 56 of the named Lygodactylus species, four named subspecies, and 34 candidate species of which 19 are newly identified in this study. Phylogenetic analyses, based on ∼10.1 kbp concatenated sequences of eight nuclear-encoded and five mitochondrial gene fragments, confirm the monophyly of 14 Lygodactylus species groups, arranged in four major clades. We recover two clades splitting from basal nodes, one comprising exclusively Malagasy species groups, and the other containing three clades. In the latter, there is a clade with only Madagascar species, which is followed by a clade containing three African and one South American species groups, and its sister clade containing six African and two Malagasy species groups. Relationships among species groups within these latter clades remain weakly supported. We reconstruct a Lygodactylus timetree based on a novel fossil-dated phylotranscriptomic tree of squamates, in which we included data from two newly sequenced Lygodactylus transcriptomes. We estimate the crown diversification of Lygodactylus started at 46 mya, and the dispersal of Lygodactylus among the main landmasses in the Oligocene and Miocene, 35-22 mya, but emphasize the wide confidence intervals of these estimates. The phylogeny suggests an initial out-of-Madagascar dispersal as most parsimonious, but accounting for poorly resolved nodes, an out-of-Africa scenario may only require one extra dispersal step. More accurate inferences into the biogeographic history of these geckos will likely require broader sampling of related genera and phylogenomic approaches to provide better topological support. A survey of morphological characters revealed that most of the major clades and species groups within Lygodactylus cannot be unambiguously characterized by external morphology alone, neither by unique character states nor by a diagnostic combination of character states. Thus, any future taxonomic work will likely benefit from integrative, phylogenomic approaches.

Repositories for Taxonomic Data: Where We Are and What is Missing.

Natural history collections are leading successful large-scale projects of specimen digitization (images, metadata, DNA barcodes), thereby transforming taxonomy into a big data science. Yet, little effort has been directed towards safeguarding and subsequently mobilizing the considerable amount of original data generated during the process of naming 15,000-20,000 species every year. From the perspective of alpha-taxonomists, we provide a review of the properties and diversity of taxonomic data, assess their volume and use, and establish criteria for optimizing data repositories. We surveyed 4113 alpha-taxonomic studies in representative journals for 2002, 2010, and 2018, and found an increasing yet comparatively limited use of molecular data in species diagnosis and description. In 2018, of the 2661 papers published in specialized taxonomic journals, molecular data were widely used in mycology (94%), regularly in vertebrates (53%), but rarely in botany (15%) and entomology (10%). Images play an important role in taxonomic research on all taxa, with photographs used in >80% and drawings in 58% of the surveyed papers. The use of omics (high-throughput) approaches or 3D documentation is still rare. Improved archiving strategies for metabarcoding consensus reads, genome and transcriptome assemblies, and chemical and metabolomic data could help to mobilize the wealth of high-throughput data for alpha-taxonomy. Because long-term-ideally perpetual-data storage is of particular importance for taxonomy, energy footprint reduction via less storage-demanding formats is a priority if their information content suffices for the purpose of taxonomic studies. Whereas taxonomic assignments are quasifacts for most biological disciplines, they remain hypotheses pertaining to evolutionary relatedness of individuals for alpha-taxonomy. For this reason, an improved reuse of taxonomic data, including machine-learning-based species identification and delimitation pipelines, requires a cyberspecimen approach-linking data via unique specimen identifiers, and thereby making them findable, accessible, interoperable, and reusable for taxonomic research. This poses both qualitative challenges to adapt the existing infrastructure of data centers to a specimen-centered concept and quantitative challenges to host and connect an estimated $ \le $2 million images produced per year by alpha-taxonomic studies, plus many millions of images from digitization campaigns. Of the 30,000-40,000 taxonomists globally, many are thought to be nonprofessionals, and capturing the data for online storage and reuse therefore requires low-complexity submission workflows and cost-free repository use. Expert taxonomists are the main stakeholders able to identify and formalize the needs of the discipline; their expertise is needed to implement the envisioned virtual collections of cyberspecimens. [Big data; cyberspecimen; new species; omics; repositories; specimen identifier; taxonomy; taxonomic data.].

Sympatric lineages in the Mantidactylus ambreensis complex of Malagasy frogs originated allopatrically rather than by in-situ speciation.

Madagascar's biota is characterized by a high degree of microendemism at different taxonomic levels, but how colonization and in-situ speciation contribute to the assembly of local species communities has rarely been studied on this island. Here we analyze the phylogenetic relationships of riparian frogs of the Mantidactylus ambreensis species complex, which is distributed in the north of Madagascar and was originally described from Montagne d'Ambre, an isolated mountain of volcanic origin, currently protected within Montagne d'Ambre National Park (MANP). Data from mitochondrial DNA, and phylogenomic data from FrogCap, a sequence capture method, independently confirm that this species complex is monophyletic within the subgenus Ochthomantis, and identify two main clades within it. These two clades are separated by 5.6-6.8% pairwise distance in the mitochondrial 16S rRNA gene and co-occur in MANP, with one distributed at high elevations (940-1375 m a.s.l.) and the other at lower elevations (535-1010 m a.s.l.), but show almost no haplotype sharing in the nuclear RAG1 gene. This occurrence in syntopy without admixture confirms them as independent evolutionary lineages that merit recognition as separate species, and we here refer to them as high-elevation (HE) and low-elevation (LE) lineage; they will warrant taxonomic assessment to confidently assign the name ambreensis to one or the other. Populations of the M. ambreensis complex from elsewhere in northern Madagascar all belong to the LE lineage, although they do occur over a larger elevational range than in Montagne d'Ambre (285-1040 m a.s.l.). Within LE there are several phylogroups (LE1-LE4) of moderately deep divergence (1.5-2.8% in 16S), but phylogroup LE4 that occurs in MANP has a deeply nested phylogenetic position, as recovered separately by mitochondrial and sequence capture datasets. This suggests that HE and LE did not diverge by a local fission of lower and upper populations, but instead arose through a more complex biogeographic scenario. The branching pattern of phylogroups LE1-LE4 shows a clear south-to-north phylogeographic pattern. We derive from these results a testable hypothesis of vicariant speciation that restricted the HE lineage to MANP and the LE candidate species to a climatic refugium further south, with subsequent northwards range expansion and secondary colonization of MANP by LE. These results provide an example for complex assembly of local microendemic amphibian faunas on Madagascar.

The bizarre skull of Xenotyphlops sheds light on synapomorphies of Typhlopoidea.

The emerging picture of non-monophyly of scolecophidian snakes is increasingly indicative that fossorial lifestyle, myrmecophagous diet, and miniaturisation are powerful drivers of morphological evolution in squamate skulls. We provide a detailed description of the skull of Xenotyphlops grandidieri, with reference to the skulls of other scolecophidian snakes. The skull, which shows dramatic ventral inflection of the snout complex, is remarkably bizarre, and the mouth opening is more ventrally oriented than in other typhlopoids. The eyes are strongly reduced, and the enlarged and rather flat anterior head shield is covered in numerous sensillae. We put forward several potential explanations for the evolution of these unusual modifications. On the other hand, Xenotyphlops shares numerous synapomorphies with other typhlopoid snakes, including the highly specialized jaw mechanism. We argue that the key differences between the jaw mechanisms of Leptotyphlopidae, Anomalepididae, and Typhlopoidea provide compelling evidence for a strong role of convergence in the evolution of the scolecophidian bauplan, and these clades therefore cannot be interpreted as representative of ancestral anatomy or ecology among snakes.

Computational molecular species delimitation and taxonomic revision of the gecko genus Ebenavia Boettger, 1878.

Cryptic species have been detected in many groups of organisms and must be assumed to make up a significant portion of global biodiversity. We study geckos of the Ebenavia inunguis complex from Madagascar and surrounding islands and use species delimitation algorithms (GMYC, BOLD, BPP), COI barcode divergence, diagnostic codon indels in the nuclear marker PRLR, diagnostic categorical morphological characters, and significant differences in continuous morphological characters for its taxonomic revision. BPP yielded ≥ 10 operational taxonomic units, whereas GMYC (≥ 27) and BOLD (26) suggested substantial oversplitting. In consequnce, we resurrect Ebenavia boettgeri Boulenger 1885 and describe Ebenavia tuelinae sp. nov., Ebenavia safari sp. nov., and Ebenavia robusta sp. nov., increasing the number of recognised species in Ebenavia from two to six. Further lineages of Ebenavia retrieved by BPP may warrant species or subspecies status, but further taxonomic conclusions are postponed until more data become available. Finally, we present an identification key to the genus Ebenavia, provide an updated distribution map, and discuss the diagnostic values of computational species delimitation as well as morphological and molecular diagnostic characters.

Reconciling molecular phylogeny, morphological divergence and classification of Madagascan narrow-mouthed frogs (Amphibia: Microhylidae).

A recent study clarified several aspects of microhylid phylogeny by combining DNA sequences from Sanger sequencing and anchored phylogenomics, although numerous aspects of tree topology proved highly susceptible to data partition and chosen model. Although the phylogenetic results of the study were in conflict with previous publications, the authors made several changes to the taxonomy of Madagascar's cophyline microhylids. We re-analyzed part of their data together with our own molecular and morphological data. Based on a supermatrix of 11 loci, we propose a new phylogeny of the Cophylinae, and discuss it in the context of a newly generated osteological dataset. We found several sample misidentifications, partially explaining their deviant results, and propose to resurrect the genera Platypelis and Stumpffia from the synonymy of Cophyla and Rhombophryne, respectively. We provide support for the previous genus-level taxonomy of this subfamily, and erect a new genus, Anilany gen. nov., in order to eliminate paraphyly of Stumpffia and to account for the osteological differences observed among these groups. Deep nodes in our phylogeny remain poorly supported, and future works will certainly refine our classification, but we are confident that these will not produce large-scale rearrangements.

Several origins of floral oil in the Angelonieae, a southern hemisphere disjunct clade of Plantaginaceae.

UNLABELLED: • PREMISE OF THE STUDY: Over the past 75 Myr, successive groups of plants have entered the "oil bee pollination niche," meaning that they depend on oil-collecting bees for their pollination. The highly dissimilar numbers of plant species and bee species involved in these mutualisms imply evolutionary host switching, asymmetric mutual dependencies, and uncoupled diversification. Among the clades with the best field data on oil bee behavior is the Angelonieae, which we here investigate to better understand the evolutionary time frame of this pollination syndrome.• METHODS: We generated nuclear and plastid data matrices for 56% of the Angelonieae species (plus outgroups) and used Bayesian methods of molecular clock dating, ancestral state reconstruction, and biogeographic inference.• KEY RESULTS: We found that Angelonieae have two major clades, Angelonia (including Monopera) and Basistemon, and Monttea, Melosperma, and Ourisia.• CONCLUSIONS: Angelonieae date back to the Uppermost Eocene, ca. 35 (26-47) Myr ago (Ma) and diversified in dry areas of southern South America; they switched from nectar to oil as a reward four or five times over the past 25 Ma. As predicted in a previous non-clock-dated study, dispersal to Australasia dates to the Miocene/Pliocene.

The Amphibian Genomics Consortium: advancing genomic and genetic resources for amphibian research and conservation.

Amphibians represent a diverse group of tetrapods, marked by deep divergence times between their three systematic orders and families. Studying amphibian biology through the genomics lens increases our understanding of the features of this animal class and that of other terrestrial vertebrates. The need for amphibian genomics resources is more urgent than ever due to the increasing threats to this group. Amphibians are one of the most imperiled taxonomic groups, with approximately 41% of species threatened with extinction due to habitat loss, changes in land use patterns, disease, climate change, and their synergistic effects. Amphibian genomics resources have provided a better understanding of ontogenetic diversity, tissue regeneration, diverse life history and reproductive modes, anti- predator strategies, and resilience and adaptive responses. They also serve as critical models for understanding widespread genomic characteristics, including evolutionary genome expansions and contractions given they have the largest range in genome sizes of any animal taxon and multiple mechanisms of genetic sex determination. Despite these features, genome sequencing of amphibians has significantly lagged behind that of other vertebrates, primarily due to the challenges of assembling their large, repeat-rich genomes and the relative lack of societal support. The advent of long-read sequencing technologies, along with computational techniques that enhance scaffolding capabilities and streamline computational workload is now enabling the ability to overcome some of these challenges. To promote and accelerate the production and use of amphibian genomics research through international coordination and collaboration, we launched the Amphibian Genomics Consortium (AGC) in early 2023. This burgeoning community already has more than 282 members from 41 countries (6 in Africa, 131 in the Americas, 27 in Asia, 29 in Australasia, and 89 in Europe). The AGC aims to leverage the diverse capabilities of its members to advance genomic resources for amphibians and bridge the implementation gap between biologists, bioinformaticians, and conservation practitioners. Here we evaluate the state of the field of amphibian genomics, highlight previous studies, present challenges to overcome, and outline how the AGC can enable amphibian genomics research to "leap" to the next level.

A new species of Pandanus-dwelling frog from northern Madagascar related to Guibemantis pulcher.

Populations of phytotelmic frogs from northern Madagascar assigned to Guibemantis (Pandanusicola) pulcher are known to differ genetically from populations further south in the eastern rainforest belt of the island, but to date, their status has not been analyzed in depth. We combined molecular genetic data with an examination of color pattern to clarify the taxonomy of these frogs. DNA sequences of both mitochondrial and nuclear-encoded genes were consistently differentiated between the northern populations and those occurring further south. Uncorrected pairwise distance in the 16S rRNA gene was 3.7‒4.3% and thus at a level usually characterizing distinct frog species in Madagascar. Furthermore, the northern specimens were characterized by more and smaller purplish-brown spots on their green dorsal surface, and a less distinct brown patch on the flanks. Although fully conclusive evidence for the species status of the northern lineage from bioacoustic differences, sympatric occurrence or narrow hybrid zone is currently lacking, such species-level distinctness is currently the most likely hypothesis. We therefore name the northern populations as Guibemantis (Pandanusicola) pulcherrimus sp. nov. The new species is known from Makira (type locality) and Bemanevika, and specimens morphologically assignable to this taxon have also been recorded from Masoala, Marojejy and Anjanaharibe-Sud.

Integrative revision of the Blommersia wittei complex, with description of a new species of frog from western and north-western Madagascar.

Frogs of the Blommersia wittei complex are widespread in western and northern Madagascar, and are one of two clades of the family Mantellidae that have colonized the Comoran island of Mayotte. Based on a comprehensive set of DNA sequences of the mitochondrial 16S rRNA gene and the nuclear-encoded RAG1 and SACS genes, integrated with morphological and bioacoustic data, we here analyze the genetic differentiation of populations of this complex across Madagascar. We confirm that a candidate species named B. sp. Ca5 in previous studies represents a genetically well-defined evolutionary lineage distributed over much of western Madagascar, which we describe herein as Blommeria bara sp. nov. based on its molecular and bioacoustic differentiation. Blommersia wittei occurs across northern Madagascar but its type locality Ambanja, at the lower Sambirano river, is very close to the range of another, newly discovered microendemic lineage that was only found at two sites along the upper Sambirano river (here named as candidate species B. sp. Ca12). The B. wittei complex thus provides an example of a clade of closely related Malagasy frogs that contains species widespread over hundreds of kilometers, as well as extreme microendemics. For a full resolution of this species complex, more data need to be collected on the geographical contact among these two lineages, on the morphology and bioacoustics of B. sp. Ca12, and on the north-eastern populations of B. wittei at Sambava, which are weakly differentiated in mitochondrial genes but differ in bioacoustics and possibly in the extent of foot webbing.

Gray versus yellow ventral coloration: Identity, distribution, color polymorphism and molecular relationships of the microhylid frog Platypelis mavomavo Andreone, Fenolio & Walvoord, 2003.

The Malagasy frog Platypelis mavomavo from Ambolokopatrika in the North East of Madagascar was originally diagnosed based on its bright yellow venter, but only limited information on this species has become available after its initial description in 2003. Several Platypelis specimens with yellow ventral color have been erroneously assigned to this species due to a lack of DNA sequences from the P. mavomavo type series. On the other hand, the candidate species Platypelis sp. Ca10 from Andranomapanga in the Northern Central East of Madagascar with gray ventral color has been defined based on its genetic differentiation from other nominal Platypelis species. Here we study the genetic variation of P. mavomavo and P. sp. Ca10 based on mitochondrial (16S rRNA) and nuclear-encoded (RAG-1) genes, including a newly determined sequence from the P. mavomavo holotype, which was studied using a museomics approach. We find only limited genetic variation among the samples studied, and this variation is unlinked to ventral coloration but instead reflects geographic distribution. We, therefore, conclude that P. sp. Ca10 is a gray-colored variant of P. mavomavo, and that P. mavomavo is rather widespread in the North East and Northern Central East of Madagascar, with populations in areas bordering the North West (Ambohitantely) and Sambirano (Ampotsidy) geographic regions, and the yellow-bellied morph restricted to the North East (Makira, Ambolokopatrika). Due to the range extension of P. mavomavo, the conservation status of the species requires re-assessment.

Repeated divergence of amphibians and reptiles across an elevational gradient in northern Madagascar.

How environmental factors shape patterns of biotic diversity in tropical ecosystems is an active field of research, but studies examining the possibility of ecological speciation in terrestrial tropical ecosystems are scarce. We use the isolated rainforest herpetofauna on the Montagne d'Ambre (Amber Mountain) massif in northern Madagascar as a model to explore elevational divergence at the level of populations and communities. Based on intensive sampling and DNA barcoding of amphibians and reptiles along a transect ranging from ca. 470-1470 m above sea level (a.s.l.), we assessed a main peak in species richness at an elevation of ca. 1000 m a.s.l. with 41 species. The proportion of local endemics was highest (about 1/3) at elevations >1100 m a.s.l. Two species of chameleons (Brookesia tuberculata, Calumma linotum) and two species of frogs (Mantidactylus bellyi, M. ambony) studied in depth by newly developed microsatellite markers showed genetic divergence up the slope of the mountain, some quite strong, others very weak, but in each case with genetic breaks between 1100 and 1270 m a.s.l. Genetic clusters were found in transect sections significantly differing in bioclimate and herpetological community composition. A decrease in body size was detected in several species with increasing elevation. The studied rainforest amphibians and reptiles show concordant population genetic differentiation across elevation along with morphological and niche differentiation. Whether this parapatric or microallopatric differentiation will suffice for the completion of speciation is, however, unclear, and available phylogeographic evidence rather suggests that a complex interplay between ecological and allopatric divergence processes is involved in generating the extraordinary species diversity of Madagascar's biota. Our study reveals concordant patterns of diversification among main elevational bands, but suggests that these adaptational processes are only part of the complex of processes leading to species formation, among which geographical isolation is probably also important.

Convergence, divergence, and macroevolutionary constraint as revealed by anatomical network analysis of the squamate skull, with an emphasis on snakes.

Traditionally considered the earliest-diverging group of snakes, scolecophidians are central to major evolutionary paradigms regarding squamate feeding mechanisms and the ecological origins of snakes. However, quantitative analyses of these phenomena remain scarce. Herein, we therefore assess skull modularity in squamates via anatomical network analysis, focusing on the interplay between 'microstomy' (small-gaped feeding), fossoriality, and miniaturization in scolecophidians. Our analyses reveal distinctive patterns of jaw connectivity across purported 'microstomatans', thus supporting a more complex scenario of jaw evolution than traditionally portrayed. We also find that fossoriality and miniaturization each define a similar region of topospace (i.e., connectivity-based morphospace), with their combined influence imposing further evolutionary constraint on skull architecture. These results ultimately indicate convergence among scolecophidians, refuting widespread perspectives of these snakes as fundamentally plesiomorphic and morphologically homogeneous. This network-based examination of skull modularity-the first of its kind for snakes, and one of the first to analyze squamates-thus provides key insights into macroevolutionary trends among squamates, with particular implications for snake origins and evolution.

Population diversification in the frog Mantidactylus bellyi on an isolated massif in northern Madagascar based on genetic, morphological, bioacoustic and ecological evidence.

In the processes that give rise to new species, changes first occur at the population level. But with the continuous nature of the divergence process, change in biological properties delimiting the shift from "individuals of divergent populations" towards "individuals of distinct species", as well as abiotic factors driving the change, remain largely ambivalent. Here we study diversification processes at the population level in a semi-aquatic frog, Mantidactylus (Brygoomantis) bellyi, across the diverse vegetation types of Montagne d'Ambre National Park (MANP), Madagascar. Genetic diversity was assessed with seven newly developed microsatellite markers as well as mitochondrial DNA sequences and concordance with patterns of ecological, morphological, and bioacoustic divergence evaluated. We found M. bellyi lacking mitochondrial differentiation within MANP, while microsatellite datasets partitioned them into three highly differentiated, geographically separated subpopulations (with indications for up to five subpopulations). The molecular grouping-primarily clustering individuals by geographic proximity-was coincident with differences in mean depth and width of waters, suggesting a possible role of fluvial characteristics in genetic exchange in this stream-breeding species. Genetic clustering not consistent with differences in call properties, except for dominant call frequencies under the two-subpopulations model. Morphological divergence was mostly consistent with the genetic clustering; subpopulations strongly differed by their snout-vent length, with individuals from high-elevation subpopulations smaller than those from populations below 1000 m above sea level. These results exemplify how mountains and environmental conditions might primarily shape genetic and morphological divergence in frog populations, without strongly affecting their calls.

Integrative revision of the Lygodactylus madagascariensis group reveals an unexpected diversity of little brown geckos in Madagascars rainforest.

The Lygodactylus madagascariensis species group, constituting the subgenus Domerguella, currently contains five valid species of inconspicuous dwarf geckos from Madagascars humid forests, but at least 18 deep genetic lineages have been revealed by recent molecular studies. Given the high morphological similarity of these lineages, taxonomic resolution of this astonishing diversity requires efforts to correctly delimit species, as well as assigning the available nomina to the species-level lineages identified. We here combine DNA sequences of one mitochondrial and two nuclear-encoded gene fragments with morphometric measurements and scale counts, and report evidence for a species status of most of the previously identified lineages. In particular, we rely on sympatric and often even syntopic occurrence of several of these lineages without evidence for genetic admixture, and consistent with subtle morphological differences. Furthermore, the very high divergences of 7.423.8% pairwise distances in the relatively conserved mitochondrial 16S rRNA gene, combined with a lack of haplotype sharing in the nuclear-encoded genes and differences in scale counts convinced us that most of the other, allopatrically distributed lineages also represent distinct species. We elevate L. madagascariensis petteri to species level and formally name eight new species: L. salvi sp. nov., a species from the Sambirano region in northern Madagascar, previously called L. sp. 8; L. tantsaha sp. nov. (L. sp. 10), a species occurring sympatrically with L. madagascariensis and L. petteri on Montagne dAmbre in far northern Madagascar; L. roellae sp. nov. (L. sp. 17), a species characterized by a striped coloration in all known specimens, from northern Madagascar; L. winki sp. nov. (L. sp. 18), an unstriped species from northern Madagascar but belonging to a subclade mostly distributed in the eastern rainforests of the island; L. ulli sp. nov. (L. sp. 21), a species from the same subclade as L. winki but known only from the Marojejy Massif in the North East; L. fritzi sp. nov. (L. sp. 11), a further species of this subclade from coastal lowlands in the Northern Central East; L. hodikazo sp. nov. (L. sp. 23) known from a single specimen collected at the Tsingy de Bemaraha and therefore the only Domerguella species known from the West region of Madagascar; and L. hapei sp. nov. (L. sp. 26), an enigmatic species from the Sambirano region characterized by a striped pattern on the throat that is otherwise unknown in the subgenus. Three additional deep mitochondrial lineages of Domerguella were identified in our analysis, but could not be further analyzed due to the lack or scarcity of voucher specimens. More field work and collection of voucher specimens is needed to understand their status. Furthermore, the taxonomy of the Domerguella subclade occurring in eastern Madagascar, with three described species (L. guibei, L. miops, L. fritzi), two synonyms (L. septemtuberculatus, Microscalabotes spinulifer) and at least two further deep genetic lineages co-occurring in a relatively small area, requires further revisionary work, possibly aided by target-enrichment sequencing of the respective name-bearing types.

An unexpected new red-bellied Stumpffia (Microhylidae) from forest fragments in central Madagascar highlights remaining cryptic diversity.

The Madagascan endemic subfamily Cophylinae in the family Microhylidae, is an example of a taxonomic group for which much is still to be discovered. Indeed, the cophyline frogs present a large portion of Madagascar's cryptic and microendemic amphibian diversity, yet they remain understudied. A new red-bellied species of the microhylid frog genus Stumpffia is described from the central plateau of Madagascar. Visual encounter surveys in Ambohitantely and Anjozorobe in 2019 and 2020 identified this previously unknown Stumpffia species, which closely resembles Stumpffiakibomena known from Andasibe in the east. Stumpffialynnae sp. nov. adds another species to the red-bellied species complex, differing from S.kibomena by genetic differentiation in the mitochondrial 16S rRNA gene (3.6-3.9%) and distinct nuclear RAG1 haplotypes, as well as strongly by its advertisement call. The new species is known from across Ambohitantely Special Reserve and Anjozorobe Angavo protected area, but is known only from one complete specimen and eight individual tissue samples. Based on the rarity of the species, the small number of locations in which it has been found, and its disappearing forest habitat, its IUCN Red List classification is suggested as "Endangered". This species is the first Stumpffia described from Madagascar's central plateau, highlighting the importance of conserving the remnant forest fragments in this area and the ongoing need to survey and protect this threatened habitat type.

Neon-green fluorescence in the desert gecko Pachydactylus rangei caused by iridophores.

Biofluorescence is widespread in the natural world, but only recently discovered in terrestrial vertebrates. Here, we report on the discovery of iridophore-based, neon-green flourescence in the gecko Pachydactylus rangei, localised to the skin around the eyes and along the flanks. The maximum emission of the fluorescence is at a wavelength of 516 nm in the green spectrum (excitation maximum 465 nm, blue) with another, smaller peak at 430 nm. The fluorescent regions of the skin show large numbers of iridophores, which are lacking in the non-fluorescent parts. Two types of iridophores are recognized, fluorescent iridophores and basal, non-fluorescent iridophores, the latter of which might function as a mirror, amplifying the omnidirectional fluorescence. The strong intensity of the fluorescence (quantum yield of 12.5%) indicates this to be a highly effective mechanism, unique among tetrapods. Although the fluorescence is associated with iridophores, the spectra of emission and excitation as well as the small Stokes shifts argue against guanine crystals as its source, but rather a rigid pair of fluorophores. Further studies are necessary to identify their morphology and chemical structures. We hypothesise that this nocturnal gecko uses the neon-green fluorescence, excited by moonlight, for intraspecific signalling in its open desert habitat.

Deconstructing the Gestalt: New concepts and tests of homology, as exemplified by a re-conceptualization of "microstomy" in squamates.

Snakes-a subset of lizards-have traditionally been divided into two major groups based on feeding mechanics: "macrostomy," involving the ingestion of proportionally large prey items; and "microstomy," the lack of this ability. "Microstomy"-considered present in scolecophidian and early-diverging alethinophidian snakes-is generally viewed as a symplesiomorphy shared with non-snake lizards. However, this perspective of "microstomy" as plesiomorphic and morphologically homogenous fails to recognize the complexity of this condition and its evolution across "microstomatan" squamates. To challenge this problematic paradigm, we formalize a new framework for conceptualizing and testing the homology of overall character complexes, or "morphotypes," which underlies our re-assessment of "microstomy." Using micro-computed tomography (micro-CT) scans, we analyze the morphology of the jaws and suspensorium across purported "microstomatan" squamates (scolecophidians, early-diverging alethinophidians, and non-snake lizards) and demonstrate that key components of the jaw complex are not homologous at the level of primary character state identity across these taxa. Therefore, rather than treating "microstomy" as a uniform condition, we instead propose that non-snake lizards, early-diverging alethinophidians, anomalepidids, leptotyphlopids, and typhlopoids each exhibit a unique and nonhomologous jaw morphotype: "minimal-kinesis microstomy," "snout-shifting," "axle-brace maxillary raking," "mandibular raking," and "single-axle maxillary raking," respectively. The lack of synapomorphy among scolecophidians is inconsistent with the notion of scolecophidians representing an ancestral snake condition, and instead reflects a hypothesis of the independent evolution of fossoriality, miniaturization, and "microstomy" in each scolecophidian lineage. We ultimately emphasize that a rigorous approach to comparative anatomy is necessary in constructing evolutionary hypotheses that accurately reflect biological reality.