Periodic Reporting for period 1 - HOSTILE (Ecological genomics of vertebrates across environmentally hostile naturally fragmented paleosurfaces)
Periodo di rendicontazione: 2021-06-01 al 2023-05-31
How terrestrial vertebrates survive (or thrive) in isolated and fragmented landscapes over long periods of time is of interest to evolutionary and conservation biologists. Indeed, isolation and population fragmentation directly affect genetic structure and fitness, often leading to inbreeding, reduced genetic diversity and increased risk of extinction. Although the causes of population fragmentation and their effects on genetic structure have been investigated in post-Pleistocene “nutrient-rich” landscapes (i.e. most landscapes surrounding us today), pre-Pleistocene nutrient-poor landscapes (= paleosurfaces) have seldom been investigated. Biotas of paleosurfaces differ fundamentally in their traits from those evolving in post-Pleistocene landscapes and it has been hypothesized that they display more complex population dynamics due to the expected persistence of old lineages, refugial phenomena, inbreeding, adaptations to resource-limited, highly competitive environments, and high levels of resilience to lower evolutionary potential. In this context, the Pantepui biogeographical region in the western Guiana Shield is of particular interest. It harbours numerous isolated Precambrian sandstone tabletop mountains (locally named “tepuis”) reaching up to 3,000 m elevation and is renowned for its floral and faunal endemism. The “Lost Worlds” of tepui summits face particularly hostile, challenging environmental conditions, and their characteristic vegetation grows on highly acidic, oligotrophic soils. Understanding such landscapes and their biodiversity is increasingly important in the context of a rapidly changing planet. The overarching objective of HOSTILE was to determine the extent to which the genetic structure, phenotypic traits, and, ultimately, the evolutionary trajectories of vertebrates are impacted by these environmentally hostile naturally fragmented paleosurfaces. HOSTILE used state-of-the-art population genomics, imaging technologies and histology integrated with systems-level ecological data to address fundamental questions using three tepui summit endemic amphibian and reptile taxa distributed across the summits of four neighbouring isolated tepuis to contribute to the emerging discipline of ecological genomics.
I generated a massive integrative dataset, combining the first population genetic data for a tepui fauna, the first use of skeletochronology to any tepui vertebrate, and the first application of µCT-scanning to any tepui vertebrate population. Total genomic DNA was extracted from 258 amphibians and reptiles from four neighbouring isolated tepuis. Quality of DNA extractions and DNA was re-extracted from 21 samples (using larger amounts of tissues). I worked first on Oreophrynella (208 individuals in total = 17 libraries of 12 + 1 library of 4) performing ddRAD sequencing using an Illumina sequencing platform (paired-end mode 2 x 150 bp reads). Low quality reads were removed, and the raw data were demultiplexed to recover the individual samples in the Illumina library. Biostatistical analyses of these data are in progress. Preliminary results indicate high spatial genetic structuring of summit populations that suggests a pattern of isolation by distance. This confirms unexpected high philopatry and absebce of long dispersal inferred from tracking and plot surveys. Surprisingly, the Oreophrynella quelchii phenotype (1 specimen) found on Kukenán clusters with O. nigra and is not a O. quelchii that dispersed from Roraima. Similarly, the O. nigra phenotype (3 specimens) found on Roraima cluster with O. quelchii andare not specimens of O. nigra that dispersed from Kukenán. The unique specimen from the summit of Wei-Assipu is closer to individuals from the centre and northern part of Roraima, which are in turn more distant to specimens from the southern part of the Roraima summit. More than 305 specimens of Oreophrynella, Riolama and Pristimantis were µCT-scanned at the NHM’s CT Lab facility using either a Zeiss Versa 520 or a Nikon HMX225. Virtual 3D models of 256 individuals (49 were too decalcified to be used for geometric morphometric analyses) were reconstructed and segmented using VGStudio MAX version 2.1 and surface meshes exported for morphometric work still in progress. In addition, about 30 specimens of other endemic tepui species of frogs, snakes and lizards were similarly processed for side projects directly linked to HOSTILE, leading to publications describing new species of Stefania, discussing osteology and describing a new Pantepui endemic snake genus, and generating positive publicity from the online media and television news.
Femurs were removed from 25 individuals of Oreophrynella and Pristimantis for histological work/skeletochronology. I tested several cutting and staining methods. I used a cryostat microtome to cut 10µm slices of the entire femurs that were stained with Acridine orange, mounted, and examined using confocal microscopy, a new method to visualize lines of arrested growth (LAG) and growth marks that I developed in collaboration with Prof. Łukasz Pułaski an expert in confocal microscopy. Results suggest that the Oreophrynella are short-lived, with a life expectancy of only 2-3 years. Most individuals in collections are likely less than 2 years old. We have a paper in progress discussing these results and the new/improved protocol that we developed and including comparison with European species.
My work performed on the Pantepui herpetofauna during my fellowship also led to several collaborations, with one paper in press in Nature and two additional papers accepted in Molecular Phylogenetics and Evolution pending revision.
Femurs were removed from 25 individuals of Oreophrynella and Pristimantis for histological work/skeletochronology. I tested several cutting and staining methods. I used a cryostat microtome to cut 10µm slices of the entire femurs that were stained with Acridine orange, mounted, and examined using confocal microscopy, a new method to visualize lines of arrested growth (LAG) and growth marks that I developed in collaboration with Prof. Łukasz Pułaski an expert in confocal microscopy. Results suggest that the Oreophrynella are short-lived, with a life expectancy of only 2-3 years. Most individuals in collections are likely less than 2 years old. We have a paper in progress discussing these results and the new/improved protocol that we developed and including comparison with European species.
My work performed on the Pantepui herpetofauna during my fellowship also led to several collaborations, with one paper in press in Nature and two additional papers accepted in Molecular Phylogenetics and Evolution pending revision.
Based upon the research activities described above, six scientific papers have been published or are close to be, adding substantially to our knowledge of the biodiversity of the understudied Pantepui region, giving previously unrecognised species legal statuses that will allow for their inclusion in the conservation planning so essential for their continued survival: most of the new species being already considered critically endangered according to IUCN criteria. Training at the host institution provided the foundation for much of the osteological work that most of these papers are based upon and has led to a step change in the integrative taxonomy of the fauna of the Pantepui region.
Continuing studies integrating results from morphology, histology and population genomics are expected to address the fundamental questions about the seemingly paradoxical nature of the tepui top populations (large populations, in low productive environments); revealing both the structuring of endemic populations, and the emerging role of the Pantepui region as both a stable refuge for biodiversity for many millions of years, and a previously unsuspected source of biodiversity elsewhere in the Neotropics.
These purely biological results will help to contextualise the Pantepui region, promoting interest in sustainable and concern for responsible development. This in turn should benefit the lives and wellbeing of those people whose livelihoods are tied to this region and broaden the understanding and focus of those seeking to preserve biodiversity globally.
Continuing studies integrating results from morphology, histology and population genomics are expected to address the fundamental questions about the seemingly paradoxical nature of the tepui top populations (large populations, in low productive environments); revealing both the structuring of endemic populations, and the emerging role of the Pantepui region as both a stable refuge for biodiversity for many millions of years, and a previously unsuspected source of biodiversity elsewhere in the Neotropics.
These purely biological results will help to contextualise the Pantepui region, promoting interest in sustainable and concern for responsible development. This in turn should benefit the lives and wellbeing of those people whose livelihoods are tied to this region and broaden the understanding and focus of those seeking to preserve biodiversity globally.