Emiliano Trucchi’s research while affiliated with Marche Polytechnic University and other places

Hybridization and polyploidy are prevalent drivers of speciation, with novel ecological properties potentially arising, among other mechanisms, through changes in gene regulation by small RNAs (smRNAs), linked to transposable element (TE) dynamics. With a common garden set-up, we comparatively investigated smRNA abundance in two young, but widely distributed, ecologically divergent sibling allotetraploid marsh orchids ( Dactylorhiza majalis and D. traunsteineri ) and their diploid parents. Despite independent origins, the allopolyploids appear to share a substantial portion of smRNA targeting, with transgressive smRNA targeting consistently overexpressed in both, related to key genes regulating transcription, cell division, and biotic and abiotic stress responses. TE-targeting smRNAs also display shared patterns between the sibling allopolyploids, with 20-23 nt smRNAs following the maternal and smaller genome, whereas 24 nt smRNAs targeting typically resembling the level of the paternal and larger genome. However, differences between the allopolyploids are also evident, with the older allopolyploid D. majalis often showing higher regulation by smRNAs, appearing more focused on fine-tuning gene copy regulation, whereas its younger sibling D. traunsteineri exhibits stronger non-additive expression, more prominently reflecting an apparent ongoing resolution of post-polyploidization meiotic/mitotic challenges. These findings highlight shared and species-specific smRNA dynamics, revealing how allopolyploids balance genomic instability and adaptive regulation during their evolutionary trajectories. In this system, the younger D. traunsteineri seems to prioritize stabilizing its genome, while the older D. majalis shifted towards optimizing gene expression. Together, this study emphasizes the role of smRNAs in facilitating ecological novelty and speciation during post-polyploidization evolution, providing insights into molecular mechanisms and adaptive evolution.

Hybridisation is a double-edged sword: while it can erode distinct evolutionary lineages, it can also introduce genetic diversity and adaptive potential into dwindling populations. In the Critically Endangered Balearic shearwater (Puffinus mauretanicus), this dilemma is exacerbated by a limited understanding of the extent and consequences of hybridisation with the Yelkouan shearwater (P. yelkouan). This knowledge gap has limited the scope of science-based conservation strategies to avoid the Balearic shearwater's imminent extinction. Here, we investigate shearwater hybridisation dynamics and their effect on genome-wide diversity in the Balearic shearwater. Divergence dating, demographic modelling and admixture analyses suggest that these two poorly-differentiated shearwater lineages experienced recurrent episodes of divergence and widespread hybridisation during glacial cycles. Selection scans reveal a 500 kb region hosting an adaptive haplotype that potentially underpins interspecific differences in migratory behaviour, and which has been repeatedly introgressed between taxa. Moreover, we show that interspecific gene flow has prevented increases in homozygosity and genetic load, and through forward simulations we illustrate how it can enhance the Balearic shearwater's resilience to future population bottlenecks. Our findings illustrate how introgression can be crucial for maintaining genetic diversity in threatened taxa, and highlight the need for considering the protection of hybridisation in conservation plans.

Gene expression can accelerate ecological divergence by rapidly tweaking the response of an organism to novel environments, with more divergent environments exerting stronger selection and supposedly, requiring faster adaptive responses. Organisms adapted to extreme environments provide ideal systems to test this hypothesis, particularly when compared to related species with milder ecological niches. The Emperor penguin ( Aptenodytes forsteri ) is the only endothermic vertebrate breeding in the harsh Antarctic winter, in stark contrast with the less cold‐adapted sister species, the King penguin ( A. patagonicus ). Assembling the first de novo transcriptomes and analysing multi‐tissue (brain, kidney, liver, muscle, skin) RNA‐Seq data from natural populations of both species, we quantified the shifts in tissue‐enhanced genes, co‐expression gene networks, and differentially expressed genes characterising Emperor penguin adaptation to the extreme Antarctic. Our analyses revealed the crucial role played by muscle and liver in temperature homeostasis, fasting, and whole‐body energy metabolism (glucose/insulin regulation, lipid metabolism, fatty acid beta‐oxidation, and blood coagulation). Repatterning at the regulatory level appears as more important in the brain of the Emperor penguin, showing the lowest signature of differential gene expression, but the largest co‐expression gene network shift. Nevertheless, over‐expressed genes related to mTOR signalling in the brain and the liver support their central role in cold and fasting responses. Besides contributing to understanding the genetics underlying complex traits, like body energy reservoir management, our results provide a first insight into the role of gene expression in adaptation to one of the most extreme environmental conditions endured by an endotherm.

Introductory paragraph Advances in medicine and food security have contributed to an increase in human lifespan ¹ . Yet, the associated rise in sedentary behaviour and in obesity 2,3 already threatens these gains ⁴ . Indeed, a growing body of evidence supports the central role of nutrient sensing and energy management pathways in regulating ageing rate and healthspan 5,6 , but the diversity of human lifestyles challenges our ability to identify the genetic and epigenetic drivers of this age acceleration. Here, we examine how the transition of wild King penguins to zoo husbandry can closely mimic the shift to a Western lifestyle in humans, and shed light on evolutionarily conserved epigenetic changes in responses to sedentary conditions. We show that, just like modern humans, zoo-housed King penguins experience an extended lifespan, but this comes at the cost of accelerated epigenetic ageing throughout life. This accelerated ageing is associated with differential methylation in key growth and maintenance pathways including the mTOR and PI3K/Akt networks, as well as in specific pathways of lipid-rich diet adaptation and heart-function. Our results demonstrate the deeply conserved link between sedentary behaviour and food availability on the one hand, and age acceleration on the other. Such evolutionary evidence may in turn help us to improve risk detection and, ultimately, therapeutics for lifestyle-induced age acceleration in humans ⁷ .

Galápagos iguanas are a monophyletic group endemic to the Galápagos archipelago, comprising the marine iguana Amblyrhynchus cristatus and three species of land iguanas: Conolophus subcristatus, C. pallidus and C. marthae. The biogeographic history of the land species in relation to their current distributions remains uncertain, in particular the origins of C. marthae, which is restricted to a small area of the western part of Isabela Island. The classification of C. pallidus as a separate species has also been debated. We analyzed Restriction sites Associated DNA sequences (RADseq) to reconstruct demographic histories of selected local populations of all Galápagos iguana species and estimate their divergence times within a multispecies coalescent framework. Our results indicate an early date for the colonization of Galápagos by iguanas, relative to island formation, at ca. 10 Mya, and support a recent split of C. marthae via allopatric speciation, after the emergence of Isabela Island, at ca. 0.57 Mya. We find contrasting demographic histories in C. marthae and the syntopic population of C. subcristatus, suggesting competitive interaction between these species. We also confirm that the divergence of C. pallidus from C. subcristatus is recent and close in time to the split between populations of C. subcristatus from different islands. Our genetic data support recent census estimates indicating a relatively small current effective population size (Ne) in all the studied populations. Our findings shed light on the evolutionary history of Galápagos iguanas and emphasize the need for targeted conservation strategies.

Galápagos iguanas are a monophyletic group endemic to the Galápagos archipelago which comprises the marine iguana (Amblyrhynchus cristatus) and three species of land iguanas: Conolophus subcristatus, C. pallidus and C. marthae. The IUCN classifies C. marthae as Critically Endangered and the others as Vulnerable. Our work aims to elucidate the processes that led to the current distribution of Galápagos land iguanas across the archipelago. By reconstructing past demographic fluctuations and divergence times, we can clarify their evolutionary histories and colonization patterns, shedding light on species distributions. This is especially useful for species with unexplained geographic distributions, like C. marthae, or with debated species status, like C. pallidus. The lineage that gave rise to C. marthae split earlier than the other two congeneric species, yet it is restricted to a small area of one of the youngest islands of the archipelago, Isabela. In contrast, C. subcristatus, which diverged later, is widespread across many islands. C. pallidus is found exclusively on the small Island of Santa Fe. Our RAD-seq analyses indicate that the ancestors of Galápagos iguanas likely reached the archipelago ca. 10 Mya, with C. marthae diverging recently after the emergence of Isabela, implying an allopatric speciation model. Additionally, C. marthae's recent demographic trajectory, in contrast to the syntopic C. subcristatus’ population, indicates a competitive interaction possibly due to a second colonization of Isabela by C. subcristatus itself. We confirm the recent divergence of C. pallidus from C. subcristatus due to intense genetic drift in a historically small population. Our findings have significant implications for the conservation of Galápagos iguanas, emphasizing the need for targeted conservation strategies.

Islands are crucial evolutionary hotspots, providing unique opportunities for differentiation of novel biodiversity and long-term segregation of endemic species. Islands are also fragile ecosystems, where biodiversity is more exposed to environmental and anthropogenic pressures than on continents. The Ponza grayling, Hipparchia sbordonii, is an endemic butterfly species that is currently found only in two tiny islands of the Pontine archipelago, off the coast of Italy, occupying an area smaller than 10 km2. It has been classified as Endangered (IUCN) because of the extremely limited area of occurrence, population fragmentation, and the recent demographic decline. Thanks to a combination of different assemblers of long and short genomic reads, bulk transcriptome RNAseq, and synteny analysis with phylogenetically close butterflies, we produced a highly contiguous, chromosome-scale annotated reference genome for the Ponza grayling, including 28 autosomes and the Z sexual chromosomes. The final assembly spanned 388.61 Gb with a contig N50 of 14.5 Mb and a BUSCO completeness score of 98.5%. Synteny analysis using four other butterfly species revealed high collinearity with Hipparchia semele and highlighted 10 intrachromosomal inversions longer than 10 kb, of which two appeared on the lineage leading to H. sbordonii. Our results show that a chromosome-scale reference genome is attainable also when chromatin conformation data may be impractical or present specific technical challenges. The high-quality genomic resource for H. sbordonii opens up new opportunities for the accurate assessment of genetic diversity and genetic load and for the investigations of the genomic novelties characterizing the evolutionary path of this endemic island species.

Hybridization and introgression are recognized as mechanisms promoting genetic variability during evolutionary radiations. We examined the impact of introgression in the process of speciation, focusing on the Antarctic icefish genus Chionodraco. Our analyses confirmed that the three Chionodraco species (Chionodraco hamatus, Chionodraco myersi, and Chionodraco rastrospinosus) were genetically distinctive, despite signals of past interspecific gene flow between C. hamatus and C. myersi that likely occurred during interglacial periods. However, in this study, no recent hybrids were identified. The lack of contemporary hybridization may be due to life-history traits and the type of marker used in the analysis. Our study emphasizes the importance of genomic approaches to detect subtle patterns of past hybridization accurately and highlights the significance of historical climate events in the demographic and evolutionary history of Antarctic notothenioids. Polar regions, and especially the Antarctic Peninsula, are now experiencing the fastest climate changes due to global warming. Understanding the impact of past climate events is fundamental to trace current modifications in species’ genetic variability and distributions and predict future evolutionary trajectories. This knowledge is also vital for conservation efforts, including the implementation of marine protected areas.

Onopordum tauricum Willd., a species distributed in Eastern Europe, has been the subject of various research endeavors aimed at assessing its suitability for extracting vegetable rennet for use in the production of local cheeses as a substitute for animal-derived rennet. In Italy, the species has an extremely fragmented and localized distribution in six locations scattered across the central-northern Apennines and some areas of southern Italy. In this study, both the morphology and genetic diversity of the six known Italian populations were investigated to detect putative ecotypes. To this end, 33 morphological traits were considered for morphometric measurements, while genetic analysis was conducted on the entire genome using the ddRAD-Seq method. Both analyses revealed significant differences among the Apennine populations (SOL, COL, and VIS) and those from southern Italy (ROT, PES, and LEC). Specifically, the southern Italian populations appear to deviate significantly in some characteristics from the typical form of the species. Therefore, its attribution to O. tauricum is currently uncertain, and further genetic and morphological analyses are underway to ascertain its systematic placement within the genus Onopordum.

Gene expression can accelerate ecological divergence by rapidly tweaking the response of an organism to novel environments, with more divergent environments exerting stronger selection and, supposedly, requiring faster adaptive responses. Organisms adapted to extreme environments provide ideal systems to test this hypothesis, particularly when compared to related species with milder ecological niches. The Emperor penguin (Aptenodytes forsteri) is the only warm-blooded vertebrate breeding in the harsh Antarctic winter, in stark contrast with the less cold-adapted sister species, the King penguin (A. patagonicus). Assembling the first de novo transcriptomes and analysing multi-tissue (brain, kidney, liver, muscle, skin) RNAseq data from natural populations of both species, we quantified the shifts in tissue-enhanced genes, co-expression gene networks, and differentially expressed genes characterising Emperor penguin adaptation to extreme Antarctic ecology. Our analyses revealed the crucial role played by muscle and liver in temperature homeostasis, fasting and whole-body energy metabolism (glucose/insulin regulation, lipid metabolism, fatty acid beta-oxidation, and blood coagulation). Repatterning at the regulatory level appears as more important in the brain of the Emperor penguin, showing the lowest signature of differential gene expression but the largest co-expression gene network shift. Nevertheless, over-expressed genes related to mTOR signalling in the brain and the liver support their central role in cold and fasting responses. Besides contributing to understanding the genetics underlying complex traits, like body energy reservoir management, our results provide a first insight into the role of gene expression in adaptation to one of the most extreme environmental conditions endured by an endotherm.