Integrating phylogenetic, phylogeographic, and morphometric analyses to reveal cryptic lineages within the genus Asaccus (Reptilia: Squamata: Phyllodactylidae) in Iran.

The Middle Eastern endemic genus Asaccus comprises Southwest Asian leaf-toed geckos. To date, this genus includes 19 species of leaf-toed geckos (seven in Arabia and 12 in the Zagros Mountains). Despite a recent study on the taxonomy and phylogeny of Asaccus species in Iran, controversies still remain surrounding the phylogeny and phylogeography of the genus. Here, we used an integrative approach to determine the phylogeny and phylogeography of Asaccus species using two mitochondrial genes (12 S and Cyt b), and one nuclear gene (c-mos). Our results uncovered 22 distinct lineages, demonstrating a significant cryptic diversity that challenges the current morphological classifications of these species. Phylogenetic analyses reinforce the monophyly of the Asaccus group, positioning A. montanus as a basal lineage, which supports a deep evolutionary divergence dating back to the Late Oligocene, approximately 27.94 million years ago. This genetic diversity also highlights the impact of historical climatic and geographical changes on species diversification. The findings advocate for an integrative approach combining both molecular and morphological data to resolve species identities accurately, thereby enhancing conservation strategies to protect these genetically distinct lineages.

Brain Gene Regulatory Networks Coordinate Nest Construction in Birds.

Nest building is a vital behavior exhibited during breeding in birds, and is possibly induced by environmental and social cues. Although such behavioral plasticity has been hypothesized to be controlled by adult neuronal plasticity, empirical evidence, especially at the neurogenomic level, remains limited. Here, we aim to uncover the gene regulatory networks that govern avian nest construction and examine whether they are associated with circuit rewiring. We designed an experiment to dissect this complex behavior into components in response to pair bonding and nest material acquisition by manipulating the presence of mates and nest materials in 30 pairs of zebra finches. Whole-transcriptome analysis of 300 samples from five brain regions linked to avian nesting behaviors revealed nesting-associated gene expression enriched with neural rewiring functions, including neurogenesis and neuron projection. The enriched expression was observed in the motor/sensorimotor and social behavior networks of female finches, and in the dopaminergic reward system of males. Female birds exhibited predominant neurotranscriptomic changes to initiate the nesting stage, while males showed major changes after entering this stage, underscoring sex-specific roles in nesting behavior. Notably, major neurotranscriptomic changes occurred during pair bonding, with minor changes during nest material acquisition, emphasizing social interactions in nest construction. We also revealed gene expression associated with reproductive behaviors and tactile sensing for nesting behavior. This study presents novel neurogenomic evidence supporting the hypothesis of adult neural plasticity underlying avian nest-construction behavior. By uncovering the genetic toolkits involved, we offer novel insights into the evolution of animals' innate ability to construct nests.

A chromosome-level genome assembly of the Asian house martin implies potential genes associated with the feathered-foot trait.

The presence of feathers is a vital characteristic among birds, yet most modern birds had no feather on their feet. The discoveries of feathers on the hind limbs of basal birds and dinosaurs have sparked an interest in the evolutionary origin and genetic mechanism of feathered feet. However, the majority of studies investigating the genes associated with this trait focused on domestic populations. Understanding the genetic mechanism underpinned feathered-foot development in wild birds is still in its infancy. Here, we assembled a chromosome-level genome of the Asian house martin (Delichon dasypus) using the long-read High Fidelity sequencing approach to initiate the search for genes associated with its feathered feet. We employed the whole-genome alignment of D. dasypus with other swallow species to identify high-SNP regions and chromosomal inversions in the D. dasypus genome. After filtering out variations unrelated to D. dasypus evolution, we found six genes related to feather development near the high-SNP regions. We also detected three feather development genes in chromosomal inversions between the Asian house martin and the barn swallow genomes. We discussed their association with the wingless/integrated (WNT), bone morphogenetic protein, and fibroblast growth factor pathways and their potential roles in feathered-foot development. Future studies are encouraged to utilize the D. dasypus genome to explore the evolutionary process of the feathered-foot trait in avian species. This endeavor will shed light on the evolutionary path of feathers in birds.

A global database of bird nest traits.

The reproductive success of birds is closely tied to the characteristics of their nests. It is crucial to understand the distribution of nest traits across phylogenetic and geographic dimensions to gain insight into bird evolution and adaptation. Despite the extensive historical documentation on breeding behavior, a structured dataset describing bird nest characteristics has been lacking. To address this gap, we have compiled a comprehensive dataset that characterizes three ecologically and evolutionarily significant nest traits-site, structure, and attachment-for 9,248 bird species, representing all 36 orders and 241 out of the 244 families. By defining seven sites, seven structures, and four attachment types, we have systematically classified the nests of each species using information from text descriptions, photos, and videos sourced from online databases and literature. This nest traits dataset serves as a valuable addition to the existing body of morphological and ecological trait data for bird species, providing a useful resource for a wide range of avian macroecological and macroevolutionary research.

Weak gene-gene interaction facilitates the evolution of gene expression plasticity.

BACKGROUND: Individual organisms may exhibit phenotypic plasticity when they acclimate to different conditions. Such plastic responses may facilitate or constrain the adaptation of their descendant populations to new environments, complicating their evolutionary trajectories beyond the genetic blueprint. Intriguingly, phenotypic plasticity itself can evolve in terms of its direction and magnitude during adaptation. However, we know little about what determines the evolution of phenotypic plasticity, including gene expression plasticity. Recent laboratory-based studies suggest dominance of reversing gene expression plasticity-plastic responses that move the levels of gene expression away from the new optima. Nevertheless, evidence from natural populations is still limited. RESULTS: Here, we studied gene expression plasticity and its evolution in the montane and lowland populations of an elevationally widespread songbird-the Rufous-capped Babbler (Cyanoderma ruficeps)-with reciprocal transplant experiments and transcriptomic analyses; we set common gardens at altitudes close to these populations' native ranges. We confirmed the prevalence of reversing plasticity in genes associated with altitudinal adaptation. Interestingly, we found a positive relationship between magnitude and degree of evolution in gene expression plasticity, which was pertinent to not only adaptation-associated genes but also the whole transcriptomes from multiple tissues. Furthermore, we revealed that genes with weaker expressional interactions with other genes tended to exhibit stronger plasticity and higher degree of plasticity evolution, which explains the positive magnitude-evolution relationship. CONCLUSIONS: Our experimental evidence demonstrates that species may initiate their adaptation to new habitats with genes exhibiting strong expression plasticity. We also highlight the role of expression interdependence among genes in regulating the magnitude and evolution of expression plasticity. This study illuminates how the evolution of phenotypic plasticity in gene expression facilitates the adaptation of species to challenging environments in nature.

Genomic architecture underlying morphological and physiological adaptation to high elevation in a songbird.

Organisms often acquire physiological and morphological modifications to conquer ecological challenges when colonizing new environments which lead to their adaptive evolution. However, deciphering the genomic mechanism of ecological adaptation is difficult because ecological environments are often too complex for straightforward interpretation. Thus, we examined the adaptation of a widespread songbird-the rufous-capped babbler (Cyanoderma ruficeps)-to a relatively simple system: distinct environments across elevational gradients on the mountainous island of Taiwan. We focused on the genomic sequences of 43 birds from five populations to show that the Taiwan group split from its sister group in mainland China around 1-2 million years ago (Ma) and colonized the montane habitats of Taiwan at least twice around 0.03-0.22 Ma. The montane and lowland Taiwan populations diverged with gene flow between them, suggesting strong selection associated with different elevations. We found that the montane babblers had smaller beaks than the lowland ones, consistent with Allen's rule, and identified candidate genes-COL9A1 and SOX11-underlying the beak size changes. We also found that altitudinally divergent mutations were mostly located in noncoding regions and tended to accumulate in chromosomal inversions and autosomes. The altitudinally divergent mutations might regulate genes related to haematopoietic, metabolic, immune, auditory and vision functions, as well as cerebrum morphology and plumage development. The results reveal the genomic bases of morphological and physiological adaptation in this species to the low temperature, hypoxia and high UV light environment at high elevation. These findings improve our understanding of how ecological adaptation drives population divergence from the perspective of genomic architecture.

The role of a synanthropic bird in the nest niche expansion of a secondary cavity nester to man-made structures.

Species with similar ecological characters often compete with each other; however, a species may also facilitate the survival or reproduction of another ecologically similar species, although such interaction is rarely documented in birds. Here, we reported a facilitative species interaction between Asian house martins (Delichon dasypus) and russet sparrows (Passer cinnamomeus), both passerines using closed nests, in a montane farming area of Taiwan. We found that Asian house martins constructed dome-shaped nests in human houses that provided additional nest sites for russet sparrows, secondary cavity nesters with greatly declining populations in Taiwan. Russet sparrows that used house martin nests had reproductive success comparable to those that used artificial nest boxes. However, Asian house martins avoided reclaiming sparrow-used nests, which reduced their available nest sites. Interestingly, our results imply that man-made structures may be used as a conservation tool to improve the breeding of the endangered russet sparrows via this facilitative interaction.

Functional connections between bird eggshell stiffness and nest characteristics through risk of egg collision in nests.

Eggs and nests are two critical traits for the ecological success of birds. Their functional interactions, however, remain unclear. Here, we examined the functional connections between egg stiffness and nest attachment, site and structure for 1350 avian species. We revealed high eggshell stiffness for eggs in nests with a pensile attachment, located on non-tree vegetation or having a domed shape, suggesting that birds produce stiffer eggs in response to higher egg-collision risk in unstable or enclosed nests. Interdependence models suggested that the evolution of eggshell stiffness was more likely to be driven by than drive that of nest characters. Our results implied a trade-off between investment in competing for established nesting niches and producing stiff eggs to explore novel niches with high collision risk, possibly mediated by predation or thermoregulation. This study highlights an overlooked connection between nests and eggshells that may have broadened the ecological niches of birds.

Domestication obscures genomic estimates of population history.

Domesticated species are valuable models to examine phenotypic evolution, and knowledge on domestication history is critical for understanding the trajectories of evolutionary changes. Sequentially Markov Coalescent models are often used to infer domestication history. However, domestication practices may obscure the signal left by population history, affecting demographic inference. Here we assembled the genomes of a recently domesticated species-the society finch-and its parent species-the white-rumped munia-to examine its domestication history. We applied genomic analyses to two society finch breeds and white-rumped munias to test whether domestication of the former resulted from inbreeding or hybridization. The society finch showed longer and more runs of homozygosity and lower genomic heterozygosity than the white-rumped munia, supporting an inbreeding origin in the former. Blocks of white-rumped munia and other ancestry in society finch genomes showed similar genetic distance between the two taxa, inconsistent with the hybridization origin hypothesis. We then applied two Sequentially Markov Coalescent models-psmc and smc++-to infer the demographic histories of both. Surprisingly, the two models did not reveal a recent population bottleneck, but instead the psmc model showed a specious, dramatic population increase in the society finch. Subsequently, we used simulated genomes based on an array of demographic scenarios to demonstrate that recent inbreeding, not hybridization, caused the distorted psmc population trajectory. Such analyses could have misled our understanding of the domestication process. Our findings stress caution when interpreting the histories of recently domesticated species inferred by psmc, arguing that these histories require multiple analyses to validate.

Effects of artificial light at night on the nest-site selection, reproductive success and behavior of a synanthropic bird.

Understanding how artificial light at night (ALAN) impacts wildlife is increasingly important because more and more species are colonizing urban areas. As most of the bird studies on ALAN use controlled light set inside or around nest-boxes, the ecological effect of ALAN resulting from in situ streetlight on birds remains contentious. The barn swallow (Hirundo rustica) often builds open nests on buildings, which are directly exposed to varying intensity of ALAN, and thus provides a good system to examine the effect of in situ ALAN on birds. By examining the nest-site selection, reproductive success and behavior of barn swallows under various ALAN intensity in Taipei City, we found a positive effect of ALAN on their fledging success; nonetheless, such effect was only found in the swallows' first brood, but not second one. We also found that parent birds in the nests with higher ALAN intensity had higher feeding rates and more extended feeding time past sunset, which were likely stimulated by the increased begging behavior of their chicks. The night-feeding behavior might contribute to the increased fledging success, especially at the early breeding season. Interestingly, despite of the reproductive benefits obtained from ALAN, we found that the barn swallows did not select nest sites regarding ALAN intensity. The weak nest-site selection perhaps result from the complex life history interactions involving ALAN and/or confounding factors associated with ALAN in cities. This study improves our understanding of how urban birds, especially open-nesting ones, respond to in situ ALAN and provides useful information for developing urban conservation strategies.

Population genomic, climatic and anthropogenic evidence suggest the role of human forces in endangerment of green peafowl (Pavo muticus).

Both anthropogenic impacts and historical climate change could contribute to population decline and species extinction, but their relative importance is still unclear. Emerging approaches based on genomic, climatic and anthropogenic data provide a promising analytical framework to address this question. This study applied such an integrative approach to examine potential drivers for the endangerment of the green peafowl (Pavo muticus). Several demographic reconstructions based on population genomes congruently retrieved a drastic population declination since the mid-Holocene. Furthermore, a comparison between historical and modern genomes suggested genetic diversity decrease during the last 50 years. However, climate-based ecological niche models predicted stationary general range during these periods and imply the little impact of climate change. Further analyses suggested that human disturbance intensities were negatively correlated with the green peafowl's effective population sizes and significantly associated with its survival status (extirpation or persistence). Archaeological and historical records corroborate the critical role of humans, leaving the footprint of low genomic diversity and high inbreeding in the survival populations. This study sheds light on the potential deep-time effects of human disturbance on species endangerment and offers a multi-evidential approach in examining underlying forces for population declines.

Avian phenotypic convergence is subject to low genetic constraints based on genomic evidence.

BACKGROUND: Phenotypic convergence between distinct species provides an opportunity to examine the predictability of genetic evolution. Unrelated species sharing genetic underpinnings for phenotypic convergence suggests strong genetic constraints, and thus high predictability of evolution. However, there is no clear big picture of the genomic constraints on convergent evolution. Genome-based phylogenies have confirmed many cases of phenotypic convergence in birds, making them a good system for examining genetic constraints in phenotypic convergence. In this study, we used hierarchical genomic approaches to estimate genetic constraints in three convergent avian traits: nocturnality, raptorial behavior and foot-propelled diving. RESULTS: Phylogeny-based hypothesis tests and positive selection tests were applied to compare 16 avian genomes, representing 14 orders, and identify genes with strong convergence signals. We found 43 adaptively convergent genes (ACGs) associated with the three phenotypic convergence cases and assessed genetic constraints in all three cases, from (amino acid) site mutations to genetic pathways. We found that the avian orders shared few site mutations in the ACGs that contributed to the convergent phenotypes, and that these ACGs were not enriched in any genetic pathways. In addition, different pairs of orders with convergent foot-propelled diving or raptorial behaviors shared few ACGs. We also found that closely related orders that shared foot-propelled diving behavior did not share more ACGs than did distinct orders, suggesting that convergence among these orders could not be explained by their initial genomic backgrounds. CONCLUSIONS: Our analyses of three avian convergence events suggest low constraints for phenotypic convergence across multiple genetic levels, implying that genetic evolution is unpredictable at the phylogenetic level of avian order. Ours is one of first studies to apply hierarchical genomic examination to multiple avian convergent cases to assess the genetic constraints in life history trait evolution.

Maternal gut microbes shape the early-life assembly of gut microbiota in passerine chicks via nests.

BACKGROUND: Knowledge is growing on how gut microbiota are established, but the effects of maternal symbiotic microbes throughout early microbial successions in birds remain elusive. In this study, we examined the contributions and transmission modes of maternal microbes into the neonatal microbiota of a passerine, the zebra finch (Taeniopygia guttata), based on fostering experiments. RESULTS: Using 16S rRNA amplicon sequencing, we found that zebra finch chicks raised by their biological or foster parents (the society finch Lonchura striata domestica) had gut microbial communities converging with those of the parents that reared them. Moreover, source-tracking models revealed high contribution of zebra finches' oral cavity/crop microbiota to their chicks' early gut microbiota, which were largely replaced by the parental gut microbiota at later stages. The results suggest that oral feeding only affects the early stage of hatchling gut microbial development. CONCLUSIONS: Our study indicates that passerine chicks mainly acquire symbionts through indirect maternal transmission-passive environmental uptake from nests that were smeared with the intestinal and cloacal microbes of parents that raised them. Gut microbial diversity was low in hand-reared chicks, emphasizing the importance of parental care in shaping the gut microbiota. In addition, several probiotics were found in chicks fostered by society finches, which are excellent foster parents for other finches in bird farms and hosts of brood parasitism by zebra finches in aviaries; this finding implies that avian species that can transfer probiotics to chicks may become selectively preferred hosts of brood parasitism in nature. Video Abstract.

Sympatric competitors have driven the evolution of temporal activity patterns in Cnemaspis geckos in Southeast Asia.

It is often assumed that animals' temporal activity patterns are highly conserved throughout evolution. While most geckos are nocturnal, the species in the Cnemaspis genus are mostly diurnal (only a few are nocturnal). This raises a question about the evolution of a diel niche in the Cnemaspis genus. Cnemaspis geckos are distributed across Southeast Asia and are often sympatric with Cyrtodactylus, another widespread gecko genus in the same area. Since both genera are mainly rocky habitat specialists, we hypothesize that Cyrtodactylus may influence the temporal activity pattern of Cnemaspis when they are sympatric through competition. By analyzing habitat data, diel activity, and the existence of sympatric Cyrtodactylus species across the phylogeny of the Cnemaspis genus, we found (1) strong phylogenetic signals in the habitat use trait but not in temporal activity, suggesting that the diel niche of this genus is more labile compared with habitat niche, and (2) a significant association with the temporal activity pattern of Cnemaspis and the sympatry between the two genera, with the former tending to be diurnal when they are sympatric. Originated from a diurnal common ancestor, the release from competition with Cyrtodactylus species might open an opportunity for some Cnemaspis species to shift to nocturnal niches.

Secondary contact after allopatric divergence explains avian speciation and high species diversity in the Himalayan-Hengduan Mountains.

The geographical context of speciation is important for understanding speciation and community assembly. However, the predominant mode of speciation in the Himalayan-Hengduan Mountains (HHMs), a global biodiversity hotspot, remains unknown. Here, we examined the role of geography in speciation using four pairs of sister or closely related avian species that currently co-occur in the HHMs. While multilocus network analyses based on nine to eleven genes revealed deep splits between these species, several allelic networks based on individual loci suggested phylogenetic paraphyly implying a recent history of divergence. Following extensive sampling in the contact zones of these species pairs, the coalescence-based approximate Bayesian computation approach supported no gene flow during their divergence and was consistent with an allopatric speciation model. We further estimated the divergence times of the four species pairs during the middle and late Pleistocene, which were characterized by increased amplitudes of glacial variability. We found a positive relationship between their divergence times and current sympatry levels, supporting a scenario of secondary contact following allopatric speciation. The Pleistocene glacial-interglacial cycles may have led to the initial geographic population isolation; ecological divergence or mate choice might further accelerate their differentiation during secondary contact, facilitating their speciation and species accumulation in the mountainous landscape. Our findings reveal the critical role of geographic isolation in speciation in the HHMs and shed light on how this biodiversity hotspot aggregates numerous species.

To Trim or Not to Trim: Effects of Read Trimming on the De Novo Genome Assembly of a Widespread East Asian Passerine, the Rufous-Capped Babbler (Cyanoderma ruficeps Blyth).

Trimming low quality bases from sequencing reads is considered as routine procedure for genome assembly; however, we know little about its pros and cons. Here, we used empirical data to examine how read trimming affects assembled genome quality and computational time for a widespread East Asian passerine, the rufous-capped babbler (Cyanoderma ruficeps Blyth). We found that scaffolds assembled from raw reads were always longer than those from trimmed ones, whereas computational times for the former were sometimes much longer than the latter. Nevertheless, assembly completeness showed little difference among the trimming strategies. One should determine the optimal trimming strategy based on what the assembled genome will be used for. For example, to identify single nucleotide polymorphisms (SNPs) associated with phenotypic evolution, applying PLATANUS to gently trim reads would yield a reference genome with a slightly shorter scaffold length (N50 = 15.64 vs. 16.89 Mb) than the raw reads, but would save 75% of computational time. We also found that chromosomes Z, W, and 4A of the rufous-capped babbler were poorly assembled, likely due to a recently fused, neo-sex chromosome. The rufous-capped babbler genome with long scaffolds and quality gene annotation can provide a good system to study avian ecological adaptation in East Asia.

Generating brain matrices for zebra finch brain sectioning using three-dimensional printing technology.

BACKGROUND: The demand to sample brain regions in non-model species is increasing as more studies are integrating neurological data into behavioural, ecological or evolutionary analysis. However, the sampling operation is difficult for researchers without neuroscience background. It is also a challenge to collect neuroanatomical regions from animals in the field. NEW METHOD: Here we developed a new brain matrix for guiding researchers to section zebra finches' (Taeniopygia guttata) brains more steadily than by freehand trimming. Based on the 3D printing technology, we produced the zebra finch brain matrix from scratch. We also provided a step-by-step protocol to make brain matrices for any species with a brain size between that of shrews and dogs. RESULTS: The brain matrix could guide us to find the zebra finch's neuroanatomical landmarks, such as the hypothalamus, optic chiasm and occulomotor nerve. The matrix's channels near these landmarks could be used to section brains steadily and rapidly. COMPARISON WITH EXISTING METHODS: Standardized brain sectioning often requires expensive machines that may not be available in most laboratories or in the field, such as microtomes. In addition, machine-based trimming is time-consuming. Although commercial brain matrices can overcome these problems, they are only available for rats and mice. The brain matrices we developed are affordable to most laboratories and can be customised for non-model species in both lab and field experiments. CONCLUSIONS: The matrix-guided approach requires a relatively short training period and can allow researchers to properly and quickly sample brains, and thus will facilitate neuroscience-based interdisciplinary research.

Standing genetic variation as the predominant source for adaptation of a songbird.

What kind of genetic variation contributes the most to adaptation is a fundamental question in evolutionary biology. By resequencing genomes of 80 individuals, we inferred the origin of genomic variants associated with a complex adaptive syndrome involving multiple quantitative traits, namely, adaptation between high and low altitudes, in the vinous-throated parrotbill (Sinosuthora webbiana) in Taiwan. By comparing these variants with those in the Asian mainland population, we revealed standing variation in 24 noncoding genomic regions to be the predominant genetic source of adaptation. Parrotbills at both high and low altitudes exhibited signatures of recent selection, suggesting that not only the front but also the trailing edges of postglacial expanding populations could be subjected to environmental stresses. This study verifies and quantifies the importance of standing variation in adaptation in a cohort of genes, illustrating that the evolutionary potential of a population depends significantly on its preexisting genetic diversity. These findings provide important context for understanding adaptation and conservation of species in the Anthropocene.

Barn Swallow Nest Predation by a Recent Urban Invader, the Taiwan Whistling Thrush - Implications for the Evolution of Urban Avian Communities.

Urban areas become a new habitat for an increasing number of species as they gradually adapt to the expanding, human-associated environment. The barn swallow (Hirundo rustica) has constructed nests on human buildings that provide good protection against bad weather and predators over centuries. In contrast, the Taiwan whistling thrush (Myophonus insularis) is one of new urban-invading species. The interactions between old and new urban avian species can determine the structure of this growing avian community. Here we report the first case of Taiwan whistling thrushes' predation on other birds in an urban area. Taiwan whistling thrushes were observed to eat all barn swallows' chicks and eggs on one street within one week and thus dramatically reduced their reproductive success, even severer than did a typhoon. The newly evolving predation behavior of Taiwan whistling thrushes could threaten the survival of barn swallows. If the latter cannot conquer the new challenge, the nesting ground may become an ecological trap to them driving population extirpation. This reported case implies that urbanization could intensify the interactions, such as predation or competition, between old and new urban species, leading to their population decline or growth and thus community dynamics of urban wildlife.

Historical demography of four gecko species specializing in boulder cave habitat: Implications in the evolutionary dead end hypothesis and conservation.

Specialization in narrow ecological niches may not only help species to survive in competitive or unique environments but also contribute to their extermination over evolutionary time. Although the "evolutionary dead end" hypothesis has long been debated, empirical evidence from species with detailed information on niche specialization and evolutionary history remains rare. Here we use a group of four closely related Cnemaspis gecko species that depend highly on granite boulder caves in the Mekong Delta to investigate the potential impact of ecological specialization on their evolution and population dynamics. Isolated by unsuitable floodplain habitats, these boulder-dwelling geckos are among the most narrowly distributed Squamata in the world. We applied several coalescence-based approaches combined with the RAD-seq technique to estimate their divergence times, gene flow and demographic fluctuations during the speciation and population differentiation processes. Our results reveal long-term population shrinkage in the four geckos and limited gene flow during their divergence. The results suggest that the erosion and fragmentation of the granite boulder hills have greatly impacted population divergence and declines. The habitat specialization of these geckos has led to fine-scaled speciation in these granite rocky hills; in contrast, specialization might also have pushed these species toward the edge of extinction. Our study also emphasizes the conservation urgency of these vulnerable, cave-dependent geckos.