Diversity and Evolution of Frog Visual Opsins: Spectral Tuning and Adaptation to Distinct Light Environments.

Visual systems adapt to different light environments through several avenues including optical changes to the eye and neurological changes in how light signals are processed and interpreted. Spectral sensitivity can evolve via changes to visual pigments housed in the retinal photoreceptors through gene duplication and loss, differential and coexpression, and sequence evolution. Frogs provide an excellent, yet understudied, system for visual evolution research due to their diversity of ecologies (including biphasic aquatic-terrestrial life cycles) that we hypothesize imposed different selective pressures leading to adaptive evolution of the visual system, notably the opsins that encode the protein component of the visual pigments responsible for the first step in visual perception. Here, we analyze the diversity and evolution of visual opsin genes from 93 new eye transcriptomes plus published data for a combined dataset spanning 122 frog species and 34 families. We find that most species express the four visual opsins previously identified in frogs but show evidence for gene loss in two lineages. Further, we present evidence of positive selection in three opsins and shifts in selective pressures associated with differences in habitat and life history, but not activity pattern. We identify substantial novel variation in the visual opsins and, using microspectrophotometry, find highly variable spectral sensitivities, expanding known ranges for all frog visual pigments. Mutations at spectral-tuning sites only partially account for this variation, suggesting that frogs have used tuning pathways that are unique among vertebrates. These results support the hypothesis of adaptive evolution in photoreceptor physiology across the frog tree of life in response to varying environmental and ecological factors and further our growing understanding of vertebrate visual evolution.

Distribution and diversification of Adelphobates, emblematic poison frogs from Brazilian Amazonia.

Adelphobates contains three species, and the inaccurate identification of A. quinquevittatus and the scarcity of records of A. castaneoticus complicate inference of their distributions; the latter species occurs in sympatry with A. galactonotus. Our objective was to revise the distributions of Adelphobates by compiling data and modeling habitat suitability, as range limits may be shaped by landscape features and biotic interactions. We initially analyzed the existence of operational taxonomic units within the nominal species and subsequently inferred the observed and potential distributions, taking into account the possible independent lineages for the three species, and we also generated a molecular timetree to understand the chronology of interspecific diversification events. Adelphobates quinquevittatus was found to have a more easterly distribution than previously described, and specimens with phenotypic variation were found to occur in areas inconsistent with the modeling, and A. castaneoticus was concentrated in the Tapajós-Xingu interfluve, surrounded by A. galactonotus. Models indicated that the right bank of the Xingu River is suitable for both species, indeed, both were found there. Despite Adelphobates species having their distributions delimited by major Amazonian rivers, estimated divergence times predate the formation of the modern river network, suggesting that other mechanisms were involved in their diversification.

Marine introgressions and Andean uplift have driven diversification in neotropical Monkey tree frogs (Anura, Phyllomedusinae).

The species richness in the Neotropics has been linked to environmental heterogeneity and a complex geological history. We evaluated which biogeographic processes were associated with the diversification of Monkey tree frogs, an endemic clade from the Neotropics. We tested two competing hypotheses: the diversification of Phyllomedusinae occurred either in a "south-north" or a "north-south" direction in the Neotropics. We also hypothesized that marine introgressions and Andean uplift had a crucial role in promoting their diversification. We used 13 molecular markers in a Bayesian analysis to infer phylogenetic relationships among 57 species of Phyllomedusinae and to estimate their divergence times. We estimated ancestral ranges based on 12 biogeographic units considering the landscape modifications of the Neotropical region. We found that the Phyllomedusinae hypothetical ancestor range was probably widespread throughout South America, from Western Amazon to Southern Atlantic Forest, at 29.5 Mya. The Phyllomedusines' ancestor must have initially diverged through vicariance, generally followed by jump-dispersals and sympatric speciation. Dispersal among areas occurred mostly from Western Amazonia towards Northern Andes and the South American diagonal of dry landscapes, a divergent pattern from both "south-north" and "north-south" diversification hypotheses. Our results revealed a complex diversification process of Monkey tree frogs, occurring simultaneously with the orogeny of Northern Andes and the South American marine introgressions in the last 30 million years.

Toxic to the touch: The makings of lethal mantles in pitohui birds and poison dart frogs.

How do chemically defended animals resist their own toxins? This intriguing question on the concept of autotoxicity is at the heart of how species interactions evolve. In this issue of Molecular Ecology (Molecular Ecology, 2024, 33), Bodawatta and colleagues report on how Papua New Guinean birds coopted deadly neurotoxins to create lethal mantles that protect against predators and parasites. Combining chemical screening of the plumage of a diverse collection of passerine birds with genome sequencing, the researchers unlocked a deeper understanding of how some birds sequester deadly batrachotoxin (BTX) from their food without poisoning themselves. They identified that birds impervious to BTX bear amino acid substitutions in the toxin-binding site of the voltage-gated sodium channel Nav1.4, whose function is essential for proper contraction and relaxation of vertebrate muscles. Comparative genetic and molecular docking analyses show that several of the substitutions associated with insensitivity to BTX may have become prevalent among toxic birds through positive selection. Intriguingly, poison dart frogs that also co-opted BTX in their lethal mantles were found to harbour similar toxin insensitivity substitutions in their Nav1.4 channels. Taken together, this sets up a powerful model system for studying the mechanisms behind convergent molecular evolution and how it may drive biological diversity.

Piecing the barcoding puzzle of Palearctic water frogs (Pelophylax) sheds light on amphibian biogeography and global invasions.

Palearctic water frogs (genus Pelophylax) are an outstanding model in ecology and evolution, being widespread, speciose, either threatened or threatening to other species through biological invasions, and capable of siring hybrid offspring that escape the rules of sexual reproduction. Despite half a century of genetic research and hundreds of publications, the diversity, systematics and biogeography of Pelophylax still remain highly confusing, in no small part due to a lack of correspondence between studies. To provide a comprehensive overview, we gathered >13,000 sequences of barcoding genes from >1700 native and introduced localities and built multigene mitochondrial (~17 kb) and nuclear (~10 kb) phylogenies. We mapped all currently recognized taxa and their phylogeographic lineages (>40) to get a grasp on taxonomic issues, cyto-nuclear discordances, the genetic makeup of hybridogenetic hybrids, and the origins of introduced populations. Competing hypotheses for the molecular calibration were evaluated through plausibility tests, implementing a new approach relying on predictions from the anuran speciation continuum. Based on our timetree, we propose a new biogeographic paradigm for the Palearctic since the Paleogene, notably by attributing a prominent role to the dynamics of the Paratethys, a vast paleo-sea that extended over most of Europe. Furthermore, our results show that distinct marsh frog lineages from Eastern Europe, the Balkans, the Near East, and Central Asia (P. ridibundus ssp.) are naturally capable of inducing hybridogenesis with pool frogs (P. lessonae). We identified 14 alien lineages (mostly of P. ridibundus) over ~20 areas of invasions, especially in Western Europe, with genetic signatures disproportionally pointing to the Balkans and Anatolia as the regions of origins, in line with exporting records of the frog leg industry and the stocks of pet sellers. Pelophylax thus emerges as one of the most invasive amphibians worldwide, and deserves much higher conservation concern than currently given by the authorities fighting biological invasions.

Multiple Transitions between Y Chromosome and Autosome in Tago's Brown Frog Species Complex.

Sex chromosome turnover is the transition between sex chromosomes and autosomes. Although many cases have been reported in poikilothermic vertebrates, their evolutionary causes and genetic mechanisms remain unclear. In this study, we report multiple transitions between the Y chromosome and autosome in the Japanese Tago's brown frog complex. Using chromosome banding and molecular analyses (sex-linked and autosomal single nucleotide polymorphisms, SNPs, from the nuclear genome), we investigated the frogs of geographic populations ranging from northern to southern Japan of two species, Rana tagoi and Rana sakuraii (2n = 26). Particularly, the Chiba populations of East Japan and Akita populations of North Japan in R. tagoi have been, for the first time, investigated here. As a result, we identified three different sex chromosomes, namely chromosomes 3, 7, and 13, in the populations of the two species. Furthermore, we found that the transition between the Y chromosome (chromosome 7) and autosome was repeated through hybridization between two or three different populations belonging to the two species, followed by restricted chromosome introgression. These dynamic sex chromosome turnovers represent the first such findings in vertebrates and imply that speciation associated with inter- or intraspecific hybridization plays an important role in sex chromosome turnover in frogs.

Deep divergences among inconspicuously colored clades of Epipedobates poison frogs.

Poison frogs (Dendrobatidae) are famous for their aposematic species, having a combination of diverse color patterns and defensive skin toxins, yet most species in this family are inconspicuously colored and considered non-aposematic. Epipedobates is among the youngest genus-level clades of Dendrobatidae that includes both aposematic and inconspicuous species. Using Sanger-sequenced mitochondrial and nuclear markers, we demonstrate deep genetic divergences among inconspicuous species of Epipedobates but relatively shallow genetic divergences among conspicuous species. Our phylogenetic analysis includes broad geographic sampling of the inconspicuous lineages typically identified as E. boulengeri and E. espinosai, which reveals two putative new species, one in west-central Colombia (E. sp. 1) and the other in north-central Ecuador (E. aff. espinosai). We conclude that E. darwinwallacei is a junior subjective synonym of E. espinosai. We also clarify the geographic distributions of inconspicuous Epipedobates species including the widespread E. boulengeri. We provide a qualitative assessment of the phenotypic diversity in each nominal species, with a focus on the color and pattern of inconspicuous species. We conclude that Epipedobates contains eight known valid species, six of which are inconspicuous. A relaxed molecular clock analysis suggests that the most recent common ancestor of Epipedobates is âˆ¼11.1 million years old, which nearly doubles previous estimates. Last, genetic information points to a center of species diversity in the Chocó at the southwestern border of Colombia with Ecuador. A Spanish translation of this text is available in the supplementary materials.

Lost in time: Unraveling the identity of Vitreorana parvula (Boulenger, 1895) (Anura: Centrolenidae).

Vitreorana parvula was the first glassfrog described for the Atlantic Forest. The species, however, has become a taxonomic puzzle as the only known individual is the lectotype from the 19th century, which is not particularly well-preserved or accompanied by a detailed original description. To solve this problem, we collected topotypic specimens, as well as advertisement calls, tissue samples, and natural history data, and compared them to other Vitreorana species. Our results show clear morphological, acoustic, and genetic differences between V. parvula and other species of Vitreorana, except for V. uranoscopa. Following our results, we consider V. uranoscopa as a junior synonym of V. parvula and redescribe the species based on topotypic material, while summarizing relevant variation from across its distribution.

Discovery and description of a new species of burrowing frog Sphaerotheca Gnther, 1859 (Anura: Dicroglossidae) from the suburban landscapes of Bengaluru, India.

Recent resolution of prevailing taxonomic ambiguities in the genus Sphaerotheca and new species discoveries from urban/suburban landscapes highlight the need for attention to non-forested habitats for amphibian conservation. In this paper, we review the status of the members of the genus Sphaerotheca and justify the synonymy of Sphaerotheca magadha as a junior synonym of Sphaerotheca swani. The prospects of resurrection of Sphaerotheca swani (herein preliminarily referred to as Sphaerotheca cf. breviceps [swani]) are discussed. In addition, we describe a new species Sphaerotheca varshaabhu sp. nov. from the suburban region of Bengaluru, India. We employ an integrative taxonomic approach to characterize the new species using molecular phylogeny, genetic distance, morphological characters, and geographical isolation as lines of evidence. We also provide a description of vocal repertoire of Sphaerotheca varshaabhu sp. nov. and provide comparative bioacoustics data for four species. This previously undescribed species from the suburban areas of Bengaluru described herein as Sphaerotheca varshaabhu sp. nov. forms a genetically divergent lineage and its genetic distance varied from 3.6% to 12.2% for 16S rRNA with respect to other species of Sphaerotheca. Our phylogenetic analysis for the genus including the new species confirms the synonymy of one recently described species, resulting in 10 valid species in the genus Sphaerotheca. These results emphasize the need for utilizing an integrative taxonomic approach for uncovering hidden diversity of suburban areas. Given these recent discoveries, we advocate for more robust surveys in human dominated areas, so that these amphibians may receive more attention.

The phylogeography of Middle Eastern tree frogs in Israel.

Western Palearctic treefrogs of the genus Hyla provide an example of a morphologically and ecologically cryptic group. Up to three distinct Hyla species have been proposed as resident in Israel and this number has consistently been subject to taxonomical debates. Here, we analyzed 16S rRNA and COI gene fragments of 658 individuals sampled at 47 pools in nine regions across Israel and the West Bank in order to resolve the taxonomic status of Hyla frogs. We generated both Bayesian and Maximum Likelihood phylogenies, and constructed time-calibrated trees to provide an evolutionary and historical context of sequence variations. We further applied SAMOVA as well as Monmonier's maximum-difference algorithm to study the genetic structure among populations and to identify potential zones acting as barriers to gene flow across locations. Our results revealed two distinct haplogroups for each gene fragment, with 95% CI divergence times dated from 8.9-17.1 Mya (16S) and 7.1-23.6 Mya (COI), respectively. SAMOVA and barrier analyses partitioned the populations into three groups. Our results highlight that, while there are probably only two Hyla species in Israel, one population of one of the species might qualify as a separate evolutionarily significant unit. Our findings elucidate the taxonomic status of Hyla frogs in Israel and provide the basis for determining appropriate management and conservation priorities.

Nuclear gene introgressions in hybrid populations of water frog Pelophylax esculentus complex: geographical analysis of the phenomenon and its interpretation.

Reproduction of water frog hybrids Pelophylax esculentus (Pelophylax ridibundus x Pelophylax lessonae) is associated with hemiclonal reproduction and backcrossing. The hemiclonal mode of reproduction occurs within P. esculentus allodiploids. In this case, the unrecombined genome of one parental species is transmitted to the offspring after premeiotic elimination of the chromosome set of the second parental species. Usually, the chromosome set of P. lessonae is eliminated, and the altered genome of P. ridibundus is passed on to the progeny. The hemiclonal inheritance within diploid Pelophylax esculentus hybrids may be accompanied by certain aberrations of premeiotic elimination. As a result, the formation of P. ridibundus specimens with introgressions of the P. lessonae genetic material, or the formation of recombinant hybrids occurs, depending on which of the parental species backcrossing takes place. The aim of our study is to describe the aberration of premeiotic elimination within the water frog P. esculentus complex detected by the nuclear gene Ldh-B inheritance, with an attempt to find out the causes of this phenomenon. It has been established that aberrations of premeiotic elimination are widespread, but only within populations of water frog from the river system of Upper Dnieper within Ukraine. The highest level of introgression takes place in the water frog populations within Kiev metropolis under conditions of expressed anthropogenization, while the maximum frequency of recombinants was detected within populations from the basin of Desna River, that has preserved native ecosystems. It was demonstrated that the frequency of premeiotic aberrations does not correlate with the intensity of interspecific water frog hybridization. Populations with introgressions are more common than populations with recombinants, however, within the latter, the frequency of recombination events is higher. The primary factor of gametogenesis aberrations, most likely, is the genetic characteristics of the local populations of parental species, since unambiguous explanations of this phenomenon based on the action of environmental stress (pollution of water systems) are not obvious.

Multi-omics resources for the Australian southern stuttering frog (Mixophyes australis) reveal assorted antimicrobial peptides.

The number of genome-level resources for non-model species continues to rapidly expand. However, frog species remain underrepresented, with up to 90% of frog genera having no genomic or transcriptomic data. Here, we assemble the first genomic and transcriptomic resources for the recently described southern stuttering frog (Mixophyes australis). The southern stuttering frog is ground-dwelling, inhabiting naturally vegetated riverbanks in south-eastern Australia. Using PacBio HiFi long-read sequencing and Hi-C scaffolding, we generated a high-quality genome assembly, with a scaffold N50 of 369.3 Mb and 95.1% of the genome contained in twelve scaffolds. Using this assembly, we identified the mitochondrial genome, and assembled six tissue-specific transcriptomes. We also bioinformatically characterised novel sequences of two families of antimicrobial peptides (AMPs) in the southern stuttering frog, the cathelicidins and ß-defensins. While traditional peptidomic approaches to peptide discovery have typically identified one or two AMPs in a frog species from skin secretions, our bioinformatic approach discovered 12 cathelicidins and two ß-defensins that were expressed in a range of tissues. We investigated the novelty of the peptides and found diverse predicted activities. Our bioinformatic approach highlights the benefits of multi-omics resources in peptide discovery and contributes valuable genomic resources in an under-represented taxon.

Diversifying in the mountains: spatiotemporal diversification of frogs in the Western Ghats biodiversity hotspot.

Mountain ranges are hotspots of biodiversity. However, the mechanisms that generate biodiversity patterns in different mountainous regions and taxa are not apparent. The Western Ghats (WG) escarpment in India is a globally recognized biodiversity hotspot with high species richness and endemism. Most studies have either invoked paleoclimatic conditions or climatic stability in the southern WG refugium to explain this high diversity and endemism. However, the factors driving macroevolutionary change remain unexplored for most taxa. Here, we generated the most comprehensive dated phylogeny to date for ranoid frogs in the WG and tested the role of paleoclimatic events or climatic stability in influencing frog diversification. We found that the diversity of different ranoid frog clades in the WG either accumulated at a constant rate through time or underwent a decrease in speciation rates around 3-2.5 Ma during the Pleistocene glaciation cycles. We also find no significant difference in diversification rate estimates across elevational gradients and the three broad biogeographic zones in the WG (northern, central, and southern WG). However, time-for-speciation explained regional species richness within clades, wherein older lineages have more extant species diversity. Overall, we find that global paleoclimatic events have had little impact on WG frog diversification throughout most of its early history until the Quaternary and that the WG may have been climatically stable allowing lineages to accumulate and persist over evolutionary time.

Development of molecular identification methods for Dryophytes suweonensis and D. japonicus, and their hybrids.

Background: As hybridization can reduce biodiversity or cause extinction, it is important to identify both purebred parental species and their hybrids prior to conserving them. The Suwon tree frog, Dryophytes suweonensis, is an endangered wildlife species in Korea that shares its habitat and often hybridizes with the Japanese tree frog, D. japonicus. In particular, D. suweonensis, D. japonicus, and their hybrids often have abnormal ovaries and gonads, which are known causes that could threaten their existence. Methods: We collected 57 individuals from six localities where D. suweonensis is known to be present. High-resolution melting curve (HRM) analysis of the mitochondrial 12S ribosomal RNA gene was performed to determine the maternal species. Thereafter, the DNA sequences of five nuclear genes (SIAH, TYR, POMC, RAG1, and C-MYC) were analyzed to determine their parental species and hybrid status. Results: The HRM analysis showed that the melting temperature of D. suweonensis was in the range of 79.0-79.3 °C, and that of D. japonicus was 77.7-78.0 °C, which clearly distinguished the two tree frog species. DNA sequencing of the five nuclear genes revealed 37 single-nucleotide polymorphism (SNP) sites, and STRUCTURE analysis showed a two-group structure as the most likely grouping solution. No heterozygous position in the purebred parental sequences with Q values ≥ 0.995 were found, which clearly distinguished the two treefrog species from their hybrids; 11 individuals were found to be D. suweonensis, eight were found to be D. japonicus, and the remaining 38 individuals were found to be hybrids. Conclusion: Thus, it was possible to unambiguously identify the parental species and their hybrids using HRM analysis and DNA sequencing methods. This study provided fundamental information for D. suweonensis conservation and restoration research.

Unexpected diversity and co-occurrence of phytotelmic frogs (Guibemantis) around Andasibe, one of the most intensively surveyed amphibian hotspots of Madagascar, and descriptions of three new species.

The area around the Malagasy village of Andasibe, which includes Analamazaotra-Mantadia National Park as well as other protected areas, is characterized by very species-rich and well-studied communities of animals and plants, but new species are still regularly discovered. Three species of phytotelmic frogs of the subgenus Pandanusicola in the genus Guibemantis are known from this area, G. flavobrunneus, G. liber, and G. pulcher. Further Pandanusicola frogs from this area have been provisionally assigned to G. bicalcaratus or G. albolineatus, pending detailed taxonomic review. During preliminary exploration of the ecology of these specialized frogs that live and reproduce in the leaf axils of Pandanus screw pines, we noticed the syntopic presence of two differently colored and differently sized Pandanusicola in Andasibe that could not be unambiguously assigned to any known species. A genetic screening revealed that these correspond to yet two further species in the area. Based on our data, seven species of Pandanusicola occur in Andasibe and nearby forests: (1) G. liber, the only non-phytotelmic species of the subgenus in the region; (2) G. flavobrunneus which is the largest species and characterized by a diagnostic yellowish brown dorsal pattern; (3) G. pulcher, characterized by translucent-green color with purplish brown spotting not observed in any other species in the area; (4) G. methueni, a brownish species usually lacking contrasted dorsolateral bands that differs from the other species in the area by emitting a characteristic trill-like advertisement call series (rather than clicks or chirps) and according to our data is widespread along Madagascars east coast; as well as three new species: (5) G. ambakoana sp. nov., a brownish species, typically with contrasted incomplete light dorsolateral bands and with single click-like advertisement calls; (6) G. vakoa sp. nov., a species that is equally brownish but lacks contrasted light dorsolateral bands and that has single click-like advertisement calls of very short duration; and (7) G. rianasoa sp. nov., a species that is smaller sized and has less distinct femoral glands than all the others, and emits a short series of soft chirp-like advertisement calls. All these species are genetically highly distinct, with >5% uncorrected pairwise distances in the mitochondrial 16S rRNA gene, and lack of haplotype sharing in two nuclear-encoded genes. The co-occurrence of seven Pandanusicola frogs in a relatively small geographic area is unprecedented in Madagascar and calls for in-depth studies of a possible differentiation in habitat use and life history.

Evolutionary and functional conservation of IRF7 in the Tibetan frog Nanorana parkeri.

BACKGROUND: The production of interferons (IFNs) is essential for the control of viral infections, and interferon regulatory factor 7 (IRF7) is considered as a vital regulator for the transcription of type I IFNs. Amphibians appear to possess a highly expanded type I IFN repertoire, consisting of intron-containing genes as observed in fish, and intronless genes as in other higher vertebrates. However, the knowledge on transcriptional regulatory mechanism of these two types of type I IFN genes is rather scarce in amphibians. METHODS AND RESULTS: A IRF7 gene named as Np-IRF7 was identified in Tibetan frog (Nanorana parkeri), and bioinformatic analysis revealed that the predicted protein of Np-IRF7 contains several important structural features known in IRF7. Expression analysis showed that Np-IRF7 gene was widely expressed and rapidly induced by poly(I:C) in different organs/tissues. Interestingly, luciferase reporter assay revealed that intronless IFN promoters were more effectively activated than intron-containing IFN promoter in Np-IRF7-transfected cells. Moreover, the overexpression of Np-IRF7 could induce the expression of ISGs and suppress the replication of FV3 in A6 cells. CONCLUSION: Np-IRF7 is indeed the ortholog of known IRF7, and IRF7 is structurally conserved in different lineages of vertebrates. Np-IRF7 played distinct roles in the activation of intron-containing and intronless type I IFN promoters, thus inducing the expression of interferon-stimulated antiviral effectors and providing a protection against ranavirus infection. The present research thus contributes to a better understanding of regulatory function of IRF7 in the IFN-mediated antiviral response of anuran amphibians.

Multiple contact zones and karyotypic evolution in a neotropical frog species complex.

Previous studies of DNA sequence and karyotypic data have revealed high genetic diversity in the Physalaemus cuvieri - Physalaemus ephippifer species complex-a group of small leptodactylid frogs in South America. To date, seven major genetic lineages have been recognized in this group, with species delimitation tests supporting four to seven of them as valid species. Among these, only P. ephippifer shows heteromorphic sex chromosomes, but the implications of cytogenetic divergence for the evolution of this group are unknown. We analyzed karyotypic, mitochondrial DNA, and 3RAD genomic data to characterize a putative contact zone between P. ephippifer and P. cuvieri Lineage 1, finding evidence for admixture and karyotypic evolution. We also describe preliminary evidence for admixture between two other members of this species complex-Lineage 1 and Lineage 3 of P. cuvieri. Our study sheds new light on evolutionary relationships in the P. cuvieri - P. ephippifer species complex, suggesting an important role of karyotypic divergence in its evolutionary history and underscoring the importance of hybridization as a mechanism of sex chromosome evolution in amphibians.

Evolutionary analysis of major histocompatibility complex variants in chytrid-resistant and susceptible amphibians.

An amphibian emerging infectious disease (EID), chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd), originated in Asia but primarily led to declines and extinctions in amphibian populations outside of Asia. Host major histocompatibility complex (MHC) molecules exhibit high polymorphism, and the evolution of MHC can be influenced by recombination and pathogens. Previous studies have indicated that host MHC class II is associated with Bd resistance. In this study, I conducted recombination and selection tests on functional MHC IIß1 alleles from an Asian Bd-resistant anuran species (Bufo gargarizans) and an Australasian Bd-susceptible species (Litoria caerulea). Recombination at the same site was identified in both species, supporting the hypothesis that recombination contributes to MHC IIß1 diversity in amphibians. Positive selection was observed in MHC IIß1 alleles in both species. In L. caerulea, at least four amino acid sites were identified under significant positive selection in the MHC IIß1, whereas these sites were either negatively selected or conserved in B. gargarizans. This suggests these sites might be selected for Bd resistance. Hydrophobicity was detected in certain amino acid sites relating to Bd resistance, suggesting this physicochemical property may be a factor selected to counteract Bd infection. These findings of this study provide an evolutionary basis for understanding how amphibian MHC IIß1 may undergo selection in response to chytrid infection.

Conserved chromatin and repetitive patterns reveal slow genome evolution in frogs.

Frogs are an ecologically diverse and phylogenetically ancient group of anuran amphibians that include important vertebrate cell and developmental model systems, notably the genus Xenopus. Here we report a high-quality reference genome sequence for the western clawed frog, Xenopus tropicalis, along with draft chromosome-scale sequences of three distantly related emerging model frog species, Eleutherodactylus coqui, Engystomops pustulosus, and Hymenochirus boettgeri. Frog chromosomes have remained remarkably stable since the Mesozoic Era, with limited Robertsonian (i.e., arm-preserving) translocations and end-to-end fusions found among the smaller chromosomes. Conservation of synteny includes conservation of centromere locations, marked by centromeric tandem repeats associated with Cenp-a binding surrounded by pericentromeric LINE/L1 elements. This work explores the structure of chromosomes across frogs, using a dense meiotic linkage map for X. tropicalis and chromatin conformation capture (Hi-C) data for all species. Abundant satellite repeats occupy the unusually long (~20 megabase) terminal regions of each chromosome that coincide with high rates of recombination. Both embryonic and differentiated cells show reproducible associations of centromeric chromatin and of telomeres, reflecting a Rabl-like configuration. Our comparative analyses reveal 13 conserved ancestral anuran chromosomes from which contemporary frog genomes were constructed.

Diversification of the terrestrial frog genus Anomaloglossus (Anura, Aromobatidae) in the Guiana Shield proceeded from highlands to lowlands, with successive loss and reacquisition of endotrophy.

Two main landscapes emerge from the Guiana Shield: the highlands to the west called the Pantepui region and the Amazonian lowlands to the east, both harbouring numerous endemic species. With 32 currently recognized species, the genus Anomaloglossus stands out among Neotropical frogs as one that diversified only within the Guiana Shield both in the highlands and the lowlands. We present a time-calibrated phylogeny obtained by using combined mitogenomic and nuclear DNA, which suggests that the genus originates from Pantepui where extant lineages started diversifying around 21 Ma, and subsequently (ca. 17 Ma) dispersed during the Miocene Climatic Optimum to the lowlands of the eastern Guiana Shield where the ability to produce endotrophic tadpoles evolved. Further diversification within the lowlands in the A. stepheni group notably led to an evolutionary reversal toward exotrophy in one species group during the late Miocene, followed by reacquisition of endotrophy during the Pleistocene. These successive shifts of reproductive mode seem to have accompanied climatic oscillations. Long dry periods might have triggered evolution of exotrophy, whereas wetter climates favoured endotrophic forms, enabling colonization of terrestrial habitats distant from water. Acquisition, loss, and reacquisition of endotrophy makes Anomaloglossus unique among frogs and may largely explain the current species diversity. The micro evolutionary processes involved in these rapid shifts of reproductive mode remain to be revealed.