pipesnake: generalized software for the assembly and analysis of phylogenomic datasets from conserved genomic loci.

MOTIVATION: Phylogenetics has moved into the era of genomics, incorporating enormous volumes of data to study questions at both shallow and deep scales. With this increase in information, phylogeneticists need new tools and skills to manipulate and analyze these data. To facilitate these tasks and encourage reproducibility, the community is increasingly moving toward automated workflows. RESULTS: Here we present pipesnake, a phylogenomics pipeline written in Nextflow for the processing, assembly, and phylogenetic estimation of genomic data from short-read sequences. pipesnake is an easy to use and efficient software package designed for this next era in phylogenetics. AVAILABILITY AND IMPLEMENTATION: pipesnake is publicly available on GitHub at https://github.com/AusARG/pipesnake and accompanied by documentation and a wiki/tutorial.

Populating a Continent: Phylogenomics Reveal the Timing of Australian Frog Diversification.

The Australian continent's size and isolation make it an ideal place for studying the accumulation and evolution of biodiversity. Long separated from the ancient supercontinent Gondwana, most of Australia's plants and animals are unique and endemic, including the continent's frogs. Australian frogs comprise a remarkable ecological and morphological diversity categorized into a small number of distantly related radiations. We present a phylogenomic hypothesis based on an exon-capture dataset that spans the main clades of Australian myobatrachoid, pelodryadid hyloid, and microhylid frogs. Our time-calibrated phylogenomic-scale phylogeny identifies great disparity in the relative ages of these groups which vary from Gondwanan relics to recent immigrants from Asia and include arguably the continent's oldest living vertebrate radiation. This age stratification provides insight into the colonization of⁠, and diversification on, the Australian continent through deep time, during periods of dramatic climatic and community changes. Contemporary Australian frog diversity highlights the adaptive capacity of anurans, particularly in response to heat and aridity, and explains why they are one of the continent's most visible faunas.

A return-on-investment approach for prioritization of rigorous taxonomic research needed to inform responses to the biodiversity crisis.

Global biodiversity loss is a profound consequence of human activity. Disturbingly, biodiversity loss is greater than realized because of the unknown number of undocumented species. Conservation fundamentally relies on taxonomic recognition of species, but only a fraction of biodiversity is described. Here, we provide a new quantitative approach for prioritizing rigorous taxonomic research for conservation. We implement this approach in a highly diverse vertebrate group-Australian lizards and snakes. Of 870 species assessed, we identified 282 (32.4%) with taxonomic uncertainty, of which 17.6% likely comprise undescribed species of conservation concern. We identify 24 species in need of immediate taxonomic attention to facilitate conservation. Using a broadly applicable return-on-investment framework, we demonstrate the importance of prioritizing the fundamental work of identifying species before they are lost.

The role of urology and radiology in prostate biopsy: current trends and future perspectives.

PURPOSE: Prostate biopsy is central to the accurate histological diagnosis of prostate cancer. In current practice, the biopsy procedure can be performed using a transrectal or transperineal route with different technologies available for targeting of lesions within the prostate. Historically, the biopsy procedure was performed solely by urologists, but with the advent of image-guided techniques, the involvement of radiologists in prostate biopsy has become more common. Herein, we discuss the pros, cons and future considerations regarding their ongoing role. METHODS: A narrative review regarding the current evidence was completed. PubMed and Cochrane central register of controlled trials were search until January 2024. All study types were of consideration if published after 2000 and an English language translation was available. RESULTS: There are no published studies that directly compare outcomes of prostate biopsy when performed by a urologist or radiologist. In all published studies regarding the learning curve for prostate biopsy, the procedure was performed by urologists. These studies suggest that the learning curve for prostate biopsy is between 10 and 50 cases to reach proficiency in terms of prostate cancer detection and complications. It is recognised that many urologists are poorly able to accurately interpret multi parametric (mp)-MRI of the prostate. Collaboration between the specialities is of importance with urology offering the advantage of being involved in prior and future care of the patient while radiology has the advantage of being able to expertly interpret preprocedure MRI. CONCLUSION: There is no evidence to suggest that prostate biopsy should be solely performed by a specific specialty. The most important factor remains knowledge of the relevant anatomy and sufficient volume of cases to develop and maintain skills.

Polyploidy breaks speciation barriers in Australian burrowing frogs Neobatrachus.

Polyploidy has played an important role in evolution across the tree of life but it is still unclear how polyploid lineages may persist after their initial formation. While both common and well-studied in plants, polyploidy is rare in animals and generally less understood. The Australian burrowing frog genus Neobatrachus is comprised of six diploid and three polyploid species and offers a powerful animal polyploid model system. We generated exome-capture sequence data from 87 individuals representing all nine species of Neobatrachus to investigate species-level relationships, the origin and inheritance mode of polyploid species, and the population genomic effects of polyploidy on genus-wide demography. We describe rapid speciation of diploid Neobatrachus species and show that the three independently originated polyploid species have tetrasomic or mixed inheritance. We document higher genetic diversity in tetraploids, resulting from widespread gene flow between the tetraploids, asymmetric inter-ploidy gene flow directed from sympatric diploids to tetraploids, and isolation of diploid species from each other. We also constructed models of ecologically suitable areas for each species to investigate the impact of climate on differing ploidy levels. These models suggest substantial change in suitable areas compared to past climate, which correspond to population genomic estimates of demographic histories. We propose that Neobatrachus diploids may be suffering the early genomic impacts of climate-induced habitat loss, while tetraploids appear to be avoiding this fate, possibly due to widespread gene flow. Finally, we demonstrate that Neobatrachus is an attractive model to study the effects of ploidy on the evolution of adaptation in animals.

A Comprehensive Approach to Detect Hybridization Sheds Light on the Evolution of Earth's Largest Lizards.

Hybridization between species occurs more frequently in vertebrates than traditionally thought, but distinguishing ancient hybridization from other phenomena that generate similar evolutionary patterns remains challenging. Here, we used a comprehensive workflow to discover evidence of ancient hybridization between the Komodo dragon (Varanus komodoensis) from Indonesia and a common ancestor of an Australian group of monitor lizards known colloquially as sand monitors. Our data comprise $>$300 nuclear loci, mitochondrial genomes, phenotypic data, fossil and contemporary records, and past/present climatic data. We show that the four sand monitor species share more nuclear alleles with $V$. komodoensis than expected given a bifurcating phylogeny, likely as a result of hybridization between the latter species and a common ancestor of sand monitors. Sand monitors display phenotypes that are intermediate between their closest relatives and $V$. komodoensis. Biogeographic analyses suggest that $V$. komodoensis and ancestral sand monitors co-occurred in northern Australia. In agreement with the fossil record, this provides further evidence that the Komodo dragon once inhabited the Australian continent. Our study shows how different sources of evidence can be used to thoroughly characterize evolutionary histories that deviate from a treelike pattern, that hybridization can have long-lasting effects on phenotypes, and that detecting hybridization can improve our understanding of evolutionary and biogeographic patterns.[Biogeography; introgression; Komodo dragon; phylogenetic networks; phylogenomics; reticulation; Varanus.].

Phylogenomics of Monitor Lizards and the Role of Competition in Dictating Body Size Disparity.

Organismal interactions drive the accumulation of diversity by influencing species ranges, morphology, and behavior. Interactions vary from agonistic to cooperative and should result in predictable patterns in trait and range evolution. However, despite a conceptual understanding of these processes, they have been difficult to model, particularly on macroevolutionary timescales and across broad geographic spaces. Here, we investigate the influence of biotic interactions on trait evolution and community assembly in monitor lizards (Varanus). Monitors are an iconic radiation with a cosmopolitan distribution and the greatest size disparity of any living terrestrial vertebrate genus. Between the colossal Komodo dragon Varanus komodoensis and the smallest Australian dwarf goannas, Varanus length and mass vary by multiple orders of magnitude. To test the hypothesis that size variation in this genus was driven by character displacement, we extended existing phylogenetic comparative methods which consider lineage interactions to account for dynamic biogeographic history and apply these methods to Australian monitors and marsupial predators. Incorporating both exon-capture molecular and morphological data sets we use a combined evidence approach to estimate the relationships among living and extinct varaniform lizards. Our results suggest that communities of Australian Varanus show high functional diversity as a result of continent-wide interspecific competition among monitors but not with faunivorous marsupials. We demonstrate that patterns of trait evolution resulting from character displacement on continental scales are recoverable from comparative data and highlight that these macroevolutionary patterns may develop in parallel across widely distributed sympatric groups.[Character displacement; comparative methods; phylogenetics; trait evolution; Varanus.].

Evolutionary models demonstrate rapid and adaptive diversification of Australo-Papuan pythons.

Lineages may diversify when they encounter available ecological niches. Adaptive divergence by ecological opportunity often appears to follow the invasion of a new environment with open ecological space. This evolutionary process is hypothesized to explain the explosive diversification of numerous Australian vertebrate groups following the collision of the Eurasian and Australian plates 25 Mya. One of these groups is the pythons, which demonstrate their greatest phenotypic and ecological diversity in Australo-Papua (Australia and New Guinea). Here, using an updated and near complete time-calibrated phylogenomic hypothesis of the group, we show that following invasion of this region, pythons experienced a sudden burst of speciation rates coupled with multiple instances of accelerated phenotypic evolution in head and body shape and body size. These results are consistent with adaptive radiation theory with an initial rapid niche-filling phase and later slow-down approaching niche saturation. We discuss these findings in the context of other Australo-Papuan adaptive radiations and the importance of incorporating adaptive diversification systems that are not extraordinarily species-rich but ecomorphologically diverse to understand how biodiversity is generated.

Phylogenomics, Biogeography, and Morphometrics Reveal Rapid Phenotypic Evolution in Pythons After Crossing Wallace's Line.

Ecological opportunities can be provided to organisms that cross stringent biogeographic barriers towards environments with new ecological niches. Wallace's and Lyddeker's lines are arguably the most famous biogeographic barriers, separating the Asian and Australo-Papuan biotas. One of the most ecomorphologically diverse groups of reptiles, the pythons, is distributed across these lines, and are remarkably more diverse in phenotype and ecology east of Lydekker's line in Australo-Papua. We used an anchored hybrid enrichment approach, with near complete taxon sampling, to extract mitochondrial genomes and 376 nuclear loci to resolve and date their phylogenetic history. Biogeographic reconstruction demonstrates that they originated in Asia around 38-45 Ma and then invaded Australo-Papua around 23 Ma. Australo-Papuan pythons display a sizeable expansion in morphological space, with shifts towards numerous new adaptive optima in head and body shape, coupled with the evolution of new micro-habitat preferences. We provide an updated taxonomy of pythons and our study also demonstrates how ecological opportunity following colonization of novel environments can promote morphological diversification in a formerly ecomorphologically conservative group. [Adaptive radiation; anchored hybrid enrichment; biogeography; morphometrics; snakes.].

Differential Circulating MicroRNA Expression in Age-Related Macular Degeneration.

This study explored the expression of several miRNAs reported to be deregulated in age-related macular degeneration (AMD). Total RNA was isolated from sera from patients with dry AMD (n = 12), wet AMD (n = 14), and controls (n = 10). Forty-two previously investigated miRNAs were selected based on published data and their role in AMD pathogenesis, such as angiogenic and inflammatory effects, and were co-analysed using a miRCURY LNA miRNA SYBR® Green PCR kit via quantitative real-time polymerase chain reaction (qRT-PCR) to validate their presence. Unsupervised hierarchical clustering indicated that AMD serum specimens have a different miRNA profile to healthy controls. We successfully validated the differentially regulated miRNAs in serum from AMD patients versus controls. Eight miRNAs (hsa-let-7a-5p, hsa-let-7d-5p, hsa-miR-23a-3p, hsa-miR-301a-3p, hsa-miR-361-5p, hsa-miR-27b-3p, hsa-miR-874-3p, hsa-miR-19b-1-5p) showed higher expression in the serum of dry AMD patients than wet AMD patients and compared with healthy controls. Increased quantities of certain miRNAs in the serum of AMD patients indicate that these miRNAs could potentially serve as diagnostic AMD biomarkers and might be used as future AMD treatment targets. The discovery of significant serum miRNA biomarkers in AMD patients would provide an easy screening tool for at-risk populations.

Multilocus phylogeny of Bornean Bent-Toed geckos (Gekkonidae: Cyrtodactylus) reveals hidden diversity, taxonomic disarray, and novel biogeographic patterns.

The gekkonid genus Cyrtodactylus is a highly diverse group of lizards (280 + species), which covers an expansive geographic range. Although this genus has been the focus of many taxonomic and molecular systematic studies, species on the Southeast Asian island of Borneo have remained understudied, leading to an unclear evolutionary history with cascading effects on taxonomy and biogeographic inferences. We assembled the most comprehensive multilocus Bornean dataset (one mitochondrial and three nuclear loci) that included 129 novel sequences and representatives from each known Cyrtodactylus species on the island to validate taxonomic status, assess species diversity, and elucidate biogeographic patterns. Our results uncovered a high proportion of cryptic diversity and revealed numerous taxonomic complications, especially within the C. consobrinus, C. malayanus, and C. pubisulcus groups. Comparisons of pairwise genetic distances and a preliminary species delimitation analysis using the Automatic Barcode Gap Discovery (ABGD) method demonstrated that some wide-ranging species on Borneo likely comprise multiple distinct and deeply divergent lineages, each with more restricted distributional ranges. We also tested the prevailing biogeographic hypothesis of a single invasion from Borneo into the Philippines. Our analyses revealed that Philippine taxa were not monophyletic, but were likely derived from multiple separate invasions into the geopolitical areas comprising the Philippines. Although our investigation of Bornean Cyrtodactylus is the most comprehensive to-date, it highlights the need for expanded taxonomic sampling and suggests that our knowledge of the evolutionary history, systematics, and biogeography of Bornean Cyrtodactylus is far from complete.

Miocene biome turnover drove conservative body size evolution across Australian vertebrates.

On deep time scales, changing climatic trends can have a predictable influence on macroevolution. From evidence of mass extinctions, we know that rapid climatic oscillations can indirectly open niche space and precipitate adaptive radiation, changing the course of ecological diversification. These dramatic shifts in the global climate, however, are rare events relative to extended periods of protracted climate change and biome turnover. It remains unclear whether during gradually changing periods, shifting habitats may instead promote non-adaptive speciation by facilitating allopatry and phenotypic conservatism. Using fossil-calibrated, species-level phylogenies for five Australian radiations comprising more than 800 species, we investigated temporal trends in biogeography and body size evolution. Here, we demonstrate that gradual Miocene cooling and aridification correlates with the restricted phenotypic diversification of multiple ecologically diverse vertebrate groups. This probably occurred as species ranges became fractured and isolated during continental biome restructuring, encouraging a shift towards conservatism in body size evolution. Our results provide further evidence that abiotic changes, not only biotic interactions, may act as selective forces influencing phenotypic macroevolution.

Delayed Growth in Incidental Pancreatic Cysts: Are the Current American College of Radiology Recommendations for Follow-up Appropriate?

PURPOSE: To evaluate growth kinetics of asymptomatic small (<2 cm) incidental pancreatic cysts and to assess potential implications of these in the context of current American College of Radiology recommendations. MATERIALS AND METHODS: This institutional review board-approved HIPAA-compliant retrospective study with waiver of informed consent included patients with asymptomatic small incidental pancreatic cysts (diameter, 5-20 mm) with two or more magnetic resonance (MR) examinations performed at least 6 months apart at a tertiary institution. The largest cyst dimension was measured on T2-weighted single-shot fast spin-echo images by one of six radiologists (1-3 years of experience) who were trained to measure pancreatic cysts in a similar manner. All analysis was conducted at the patient level by choosing the cyst that exhibited the greatest growth over the follow-up period in each patient. Fisher exact, χ(2), and Kruskal-Wallis tests and analysis of variance were used to test correlation between cyst characteristics and growth. RESULTS: A total of 259 patients (mean age, 65 years ± 11 [standard deviation], male-to-female ratio, 42:58) with 370 asymptomatic small incidental pancreatic cysts were included. At presentation, median cyst size was 9.4 mm (interquartile range [IQR], 7.0-12.2 mm), and 64 patients (25%; 95% confidence interval [CI]: 20, 30) had septations. The median imaging follow-up period was 2.2 years (IQR, 1.2-3.9 years; range, 0.5-11.0 years), with a median of three MR examinations (IQR, two to four examinations) per patient. In 171 (66%; 95% CI: 60, 72) of the 259 patients, cysts remained stable; in 18 (7%; 95% CI: 4, 11), cysts shrank; and in 70 (27%; 95% CI: 22, 33), cysts grew (median total growth and median annual growth of 4.8 mm and 2.3 mm/y, respectively). Age, cyst size, and cyst septation at presentation were not predictive of growth. Overall, 29 (11%), 16 (6%), and four (1.5%) of the cysts increased in size after 1, 2, and 3 years of initial stability, respectively. Of the 18 patients who underwent pancreatic surgery, only one patient with an intraductal papillary mucinous neoplasm had high-grade dysplasia. One patient developed pancreatic adenocarcinoma remotely at the location of the pancreatic cyst diagnosed 11 months prior. CONCLUSION: In the majority of patients, asymptomatic small incidental pancreatic cysts remained stable during a median follow-up period of 2.2 years; however, in 27% of patients, cysts increased in size over time, with 11% growing after an initial 1-year period of stability. Current American College of Radiology recommendations to discontinue imaging follow-up after 1 year of stability may need to be reevaluated.

Mitochondrial introgression via ancient hybridization, and systematics of the Australian endemic pygopodid gecko genus Delma.

Of the more than 1500 species of geckos found across six continents, few remain as unfamiliar as the pygopodids - Family Pygopodidae (Gray, 1845). These gekkotans are limited to Australia (44 species) and New Guinea (2 species), but have diverged extensively into the most ecologically diverse limbless radiation save Serpentes. Current phylogenetic understanding of the family has relied almost exclusively on two works, which have produced and synthesized an immense amount of morphological, geographical, and molecular data. However, current interspecific relationships within the largest genus Delma Gray 1831 are based chiefly upon data from two mitochondrial loci (16s, ND2). Here, we reevaluate the interspecific relationships within the genus Delma using two mitochondrial and four nuclear loci (RAG1, MXRA5, MOS, DYNLL1), and identify points of strong conflict between nuclear and mitochondrial genomic data. We address mito-nuclear discordance, and remedy this conflict by recognizing several points of mitochondrial introgression as the result of ancient hybridization events. Owing to the legacy value and intraspecific informativeness, we suggest the continued use of ND2 as a phylogenetic marker. Results identify strong support for species groups, but relationships among these clades, and the placement of several enigmatic taxa remain uncertain. We suggest a more careful review of Delma australis and the 'northwest Australia' clade. Accurately assessing and addressing species richness and relationships within this endemic Australian Gekkotan genus is relevant for understanding patterns of squamate speciation across the region.

New species of Blaesodactylus (Squamata: Gekkonidae) from Tsingy karstic outcrops in Ankarana National Park, northern Madagascar.

We describe a new gecko of the genus Blaesodactylus from a karstic outcrop in deciduous dry forest of Ankarana National Park, northern Madagascar. Blaesodactylus microtuberculatus sp. nov., the fifth recognized species of Blaesodactylus, is distinguished from all other congeners, B. ambonihazo, B. antongilensis, B. boivini and B. sakalava by a combination of small, homogeneous gular granules, unspotted venter and lack of tubercles on distal part of original tail. Mitochondrial (ND2 and ND4) and nuclear (RAG-1) DNA identify a consistent divergence between B. microtuberculatus and its allotopic sister species B. boivini. We highlight habitat partitioning in these allotopic congeners where Blaesodactylus microtuberculatus inhabits karstic outcrops in Tsingy massif, and B. boivini dwells on tree trunks in deciduous dry forest.

Evolutionary bursts drive morphological novelty in the world's largest skinks.

Animal phenotypes evolve and diverge as a result of differing selective pressures and drift. These processes leave unique signatures in patterns of trait evolution, impacting the tempo and mode of morphological macroevolution. While there is a broad understanding of the history of some organismal traits (e.g., body size), there is little consensus about the evolutionary mode of most others. This includes the relative contribution of prolonged (Darwinian gradualist) and episodic (Simpsonian jump) changes toward the evolution of novel morphologies. Here, we use new exon-capture and linear morphological datasets to investigate the tempo and mode of morphological evolution in Australo-Melanesian Tiliquini skinks. We generate a well-supported time-calibrated phylogenomic tree from ∼400 nuclear markers for more than 100 specimens, including undescribed diversity, and provide unprecedented resolution of the rapid Miocene diversification of these lizards. By collecting a morphological dataset that encompasses the lizard body plan (19 traits across the head, body, limb, and tail), we are able to identify that most traits evolve conservatively, but infrequent evolutionary bursts result in morphological novelty. These phenotypic discontinuities occur via rapid rate increases along individual branches, inconsistent with both gradualistic and punctuated equilibrial evolutionary modes. Instead, this "punctuated gradualism" has resulted in the rapid evolution of blue-tongued giants and armored dwarves in the ∼20 million years since colonizing Australia. These results outline the evolutionary pathway toward new morphologies and highlight the heterogeneity of evolutionary tempo and mode, even within individual traits.

Evolution across the adaptive landscape in a hyperdiverse beetle radiation.

The extraordinary diversification of beetles on Earth is a textbook example of adaptive evolution. Yet, the tempo and drivers of this super-radiation remain largely unclear. Here, we address this problem by investigating macroevolutionary dynamics in darkling beetles (Coleoptera: Tenebrionidae), one of the most ecomorphologically diverse beetle families (with over 30,000 species). Using multiple genomic datasets and analytical approaches, we resolve the long-standing inconsistency over deep relationships in the family. In conjunction with a landmark-based dataset of body shape morphology, we show that the evolutionary history of darkling beetles is marked by ancient rapid radiations, frequent ecological transitions, and rapid bursts of morphological diversification. On a global scale, our analyses uncovered a significant pulse of phenotypic diversification proximal to the Cretaceous-Palaeogene (K/Pg) mass extinction and convergence of body shape associated with recurrent ecological specializations. On a regional scale, two major Australasian radiations, the Adeliini and the Heleine clade, exhibited contrasting patterns of ecomorphological diversification, representing phylogenetic niche conservatism versus adaptive radiation. Our findings align with the Simpsonian model of adaptive evolution across the macroevolutionary landscape and highlight a significant role of ecological opportunity in driving the immense ecomorphological diversity in a hyperdiverse beetle group.

Addressing Technical Failures in a Diabetic Retinopathy Screening Program.

Purpose: Diabetic retinopathy (DR) is a preventable cause of blindness detectable through screening using retinal digital photography. The Irish National Diabetic Retina Screening (DRS) programme, Diabetic RetinaScreen, provides free screening services to patients with diabetes from aged 12 years and older. A technical failure (TF) occurs when digital retinal imaging is ungradable, resulting in delays in the diagnosis and treatment of sight-threatening disease. Despite their impact, the causes of TFs, and indeed the utility of interventions to prevent them, have not been extensively examined. Aim: Primary analysis aimed to identify factors associated with TF. Secondary analysis examined a subset of cases, assessing patient data from five time points between 2019 and 2021 to identify photographer/patient factors associated with TF. Methods: Patient data from the DRS database for one provider were extracted for analysis between 2018 and 2022. Information on patient demographics, screening results, and other factors previously associated with TF were analyzed. Primary analysis involved using mixed-effects logistic regression models with nested patient-eye random effects. Secondary analysis reviewed a subset of cases in detail, checking for causes of TF. Results: The primary analysis included a total of 366,528 appointments from 104,407 patients over 5 years. Most patients had Type 2 diabetes (89.2%), and the overall TF rate was 4.9%. Diabetes type and duration, dilate pupil status, and the presence of lens artefacts on the camera were significantly associated with TF. The Secondary analysis identified the primary cause of TF was found to be optically dense cataracts, accounting for over half of the TFs. Conclusion: This study provides insight into the causes of TF within the Irish DRS program, highlighting cataracts as the primary contributing factor. The identification of patient-level factors associated with TF facilitates appropriate interventions that can be put in place to improve patient outcomes and minimize delays in treatment and diagnosis.