Projects

TRAIT EVOLUTION: FROM MOLECULES TO DIVERSITY 

Longevity is a complex and poorly characterized trait: we do know that some species can live longer than others, anyway, the complex and multifactorial mechanism underlying longevity is far from being understood. Such lack of knowledge exists despite the outstanding studies conducted on model species; this highlights the importance of considering non-standard models for studying such process. In our works, we investigate longevity from a comparative genomic approach. More in detail, we analyse molecular evolution in taxonomic groups including both short-lived species and species with extreme longevity. The aim of our analyses is to identify genes that are shaped by different selection in short- and long-lived species, and identify new candidates that may have a role in the extended longevity phenotype. So far, we focus our analyses on bivalve molluscs: the class Bivalvia shows indeed the highest lifespan disparity within Metazoa, ranging from 1 to 500+ years, and includes the longest-lived non-colonial animal species known so far, the clam Arctica islandica. Bivalves therefore represent important resources to provide insights into the evolution of extended longevity. Now, we are extending our analyses on additional taxonomic groups, including Amphibia, Squamata, Aves and Mammalia. 


Sex determination is the biological process that ultimately establishes the sexual identity of an organism. Traditionally, two alternative types of sex determination have been recognized: environmental sex determination (ESD) and genetic sex determination (GSD), depending on whether the very first cues are of environmental or genetic origin. The former can be  observed for example in turtles and some other reptiles, where sex is determined by the incubation temperature of the eggs, or in spoonworms (Echiurida), where maleness/femaleness is determined by the presence/absence, respectively, of other females in the surroundings of the larvae implantation point. Conversely, GSD can be observed in most mammals and insects and relies on one or a few genetic loci that are able to initiate the sex differentiation of the individuals. In GSD systems, sex-linked genes can accumulate in specific chromosomes, which may oftentimes exhibit a dramatic morphological differentiation because of sexually antagonistic forces. These chromosomes are called heteromorphic sex chromosomes (e.g., the XY or ZW pairs in mammals and birds, respectively) and oppose to homomorphic sex chromosomes, which instead do not show any evident differentiation. In our reasearch activity, we use bivalves as experimental systems to study sex-linked genetic traits, as the wide biological diversity of this group may offer compelling perspectives for the evolution of sex determination. In fact, sex-determination mechnaisms in bivalves are still lacking a comprehensive chracterization, though a mixed system of GSD and ESD has been hypothesised. Furthermore, bivalves are among the few animal groups that are missing heteromorphic sex chromosomes (so far), suggesting that sex chromosomes, where present, should be homomorphic. At the moment, we are investigating the genetic diversity and phylogeny of three gene families that have been repeatedly associated with sex determination (i.e., Dmrt, Fox and Sox) to understand whether any difference at the molecular level exist between sex-linked and non-sex-linked orthologs.


The concept that complex ancestral traits can never be recovered after their loss is still widely accepted, despite phylogenetic and molecular approaches suggest instances where phenotypes may have been lost throughout the evolutionary history of a clade and subsequently reverted back in derived lineages. Trait loss is predicted to have evolutionary consequences for the genes involved in the trait expression, either via drift or selection. Nonetheless, empirical evidence of decay following trait loss is often lacking. For example, parasitic wasps which have lost lipogenesis do not show sequence degradation of related genes. Likewise, genes underlying photosynthesis are inferred to be under a strong purifying selection in a parasitic plant. The expression of functional opsins has been observed in cave crustacean with reduced or absent eyes. Although these contrasting results make it difficult to generalize about the impact of trait loss on its genomic blueprint, these observations on maintenance and decay are not necessarily conflicting. Different trajectories of trait preservation or decay can be expected depending on the pleiotropic effects that its underlying genes may have on other unrelated traits. So, traits whose genes are involved in multiple biological processes are less likely to degenerate after the selective constraints on a specific trait are removed. Thus pleiotropy could represent a possible mechanism which could underlie the reversion to a once-lost ancestral state.


Mitochondrial genomes (mtDNA) follow a non-Mendelian inheritance pattern, being transmitted uniparentally in most eukaryotes; in animals, mitochondrial inheritance is usually strictly maternal. The molecular mechanisms, the segregation patterns, and the evolutionary dynamics (selection/drift) underlying mitochondrial inheritance are mostly unknown, even in model species. Up to now, 100+ species of bivalve molluscs has been reported showing an aberrant system of mitochondrial heredity called Doubly Uniparental Inheritance (DUI). In DUI species, two sex-linked mitochondrial lineages exist: one is inherited through eggs (F-type) the other through sperm (M-type). Differently from the cases of paternal mtDNA leakage reported in several organisms, in DUI the sperm transmission route is stable across evolutionary time, so the F- and M-type coexist as segregated lineages for millions of years accumulating a remarkable sequence divergence. The F-M nucleotide p-distance ranges from 0.08 to 0.45, and the amino acid p-distance of mitochondrial protein-coding genes can reach 0.534. The unusual mechanism of inheritance and the natural heteroplasmy can be exploited to address numerous fundamental questions about mitochondrial biology and evolution, such as mitochondrial inheritance, intraindividual mtDNA variability, mitochondial bottleneck, mtDNA recombination, mitonuclear interactions and coevolution, genomic conflicts, and the role of mitochondria in germline formation, meiosis, gametogenesis and fertilization, in some cases providing the exceptions that address general phenomena in other animal groups.


The class Branchiopoda includes small crustaceans living in both fresh water and marine environments, and includes remarkable examples of morphological stasis, namely the two living genera of tadpole shrimps, Triops and Lepidurus (order Notostraca). Such a condition has repeatedly made tadpole shrimps included in the controversial group of the “living fossils”, though this attribute has been questioned multiple times by molecular, morphological, and paleontological surveys. Genomic studies in branchiopods started more than a decade ago, with the sequencing of Daphnia pulex genome, but only recently gained more attention and resulted in a dozen new genomes sequenced. The few comparative genomic studies published so far indicated different molecular evolutionary rates among lineages, although this cannot be generalized across the whole genome. Overall, no extensive studies have been conducted so far about the evolution of genes and gene families that may underlie these processes and, particularly, the gross morphological stasis of tadpole shrimps. At the moment, we are addressing this gap by taking advantage of the growing number of genomic resources for Notostraca, and we are performing a genome-wide evolutionary analysis of gene families to spot the more conserved genes in terms of protein sequence, hypothesizing their possible role in morphogenesis and development. Concurrently, we are also looking at other metrics of conservation for the same set of genes, such as the conservation of their non-coding regions and the evolutionary dynamics of the corresponding gene families.


GENOME EVOLUTION: COMPARATIVE GENOMICS TO UNDERSTAND CO-EVOLUTIONARY DYNAMICS

Work In Progress


Work In Progress


Transposable elements (TEs) are a diverse group of selfish genetic elements able to replicate and propagate within the host genome. They are widespread across all eukaryotic species where they can occupy a considerable proportion of their genome, reaching about 78% in the Antarctic krill Euphausia superba. The wide taxonomic distribution of most of the TE protein families, as well as phylogenetic analyses with prokaryotes homologs proteins, point to a deep origin, with most of them that arise during early eukaryotes evolution and even predating the split between eukaryotes and prokaryotes in the case of DDE-transposase superfamilies. The selfish nature of transposons makes intrinsically interesting the study of their evolutionary dynamics. Indeed, if we consider the genome as an ecosystem, transposable elements can compete for space (e.g., available genomic space) and resources (e.g., transcription factors or TE-derived proteins in the cases of autonomous-non autonomous partnerships), with coevolutionary dynamics between host and TEs and even between TEs that can recapitulate what is seen in macro-ecosystems. Moreover, the increasing cost-efficient whole genome sequencing of non-model organisms has led to an increasingly large number of high-quality assemblies, but TE annotation is usually of substandard quality compared to that available for model species, and almost nothing is known about their TE content. This gap of knowledge is not only influencing our ability to understand genomic evolutionary dynamics in a broad sense, but it is also affecting the power of all the analyses that require high-quality TE annotation (e.g, host protein-coding gene prediction, population evolutionary dynamics The main aim of this research line is to explore the biodiversity and the evolution of transposons across the tree of life, with special attention to usually neglected phyla, such as mollusks and arthropods. Moreover, one of our goals is to empower the scientific community with high-quality and freely available genomic resources that can be used to improve TE annotation in novel species as well as to increase the power for identifying adaptive and nonadaptive variants caused by their activity.


In Metazoa, four out of five complexes involved in oxidative phosphorylation (OXPHOS) are formed by subunits encoded by both the mitochondrial (mtDNA) and nuclear (nuDNA) genomes. One of the consequences of this binary genome delegation for such a critical metabolic process is that mitochondrial and nuclear genomes products must physically interact for proper OXPHOS functioning. However, these two genomes experience different evolutionary dynamics: for instance, mitochondria have a small effective population size, are uniparentally inherited, and often experience higher substitution rates. Given such premise it is not surprising that homoplasmy seems to be a favorable trait, even if low levels of heteroplasmy are commonly detected across animals. MtDNA variance within an individual seems to be an unfavorable condition that is suppressed in multiple ways. One reason could be that orchestrating multiple, different mtDNA variants by the same nuclear genome would be challenging. Accordingly, the presence of heteroplasmic mtDNA population results in a reduction of fitness in mice, and a strict coevolution between nuclear and mitochondrial genomes is thought to be necessary for healthy organisms. That said, most of the investigations have been conducted on a few mammals and on a handful of model species, so how mtDNA heteroplasmy affects organisms with quite different mitochondrial dynamics and physiology is still unknown. Bivalve molluscs represent an interesting observational unit for such studies for several reasons. First, bivalve phylogenies inferred with mtDNA show discordance with nuclear ones. Interestingly, NuDNA-encoded OXPHOS subunits have concordant topologies with mtDNA-encoded OXPHOS subunits, but not with nuclear genes lacking mitochondrial interactions. Second, 100+ species of bivalves have been reported showing a peculiar mitochondrial inheritance pattern (Doubly uniparental Inhheritance, DUI [link alla linea mitochonrial inheritance]). One of the outcomes of such a mechanism is a high-level heteroplasmy, with extremely divergent mtDNAs coexisting in the same individual, tissue, cell, and even organelle. This implies that the same nuclear background must cofunction with two very divergent mitochondrial genomes (mtOXPHOS subunit amino acid p-distance up to 0.53), adding another layer of complexity to mitonuclear coevolution. The tri-genomic coevolution in DUI species can yield fruitful contributions to the definition of mitonuclear coevolutionary mechanisms in general.

BIODIVERSITY GENOMICS: USING GENOMICS TO EXPLORE BIODIVERSITY AND ITS EVOLUTION

Accounting for almost 14,000 species, a recent update of the Italian checklist retrieved 267 species and subspecies distributed across the peninsular and insular territories. However, most of these data are based on previous knowledge and field observation, with a limited use, at least in the Italian context, of molecular data support. The use of molecular markers for species recognition/identification is particularly useful for having a better insight into taxonomic diversity, phylogeny and for providing to non-specialists a tool for species identification. For example, the DNA Barcoding project, an international effort to produce molecular data for all the known species, albeit useful in most instances, may show some inconsistencies and/or limitations, even in ants. The limited information carried by the single marker used for the DNA Barcoding (the first 700 bp of the cytochrome oxidase subunit 1 mitochondrial gene), must be complemented with more markers, such as other mitochondrial and nuclear genes. A new, efficient way to obtain a large amount of molecular data, in terms of number of genomic markers (whether they are genes or hypervariable, non-coding genomic regions), is the so-called genome skimming. Basically, it consists in a low-coverage genome sequencing which allows i) the recovery of complete, or nearly complete, high copy number markers such as the whole mitochondrial genome and/or nuclear ribosomal genes, ii) an appropriate estimate of the mobilome, i.e. the full genomic complement of transposable elements, and iii) the implementation of the so-called “DNA mark”, i.e. the species identification and analysis by means of high-throughput comparison of unassembled short reads. This method has already proved to be a reliable and efficient way to get molecular data for the analysis of animal biodiversity, both from a biodiversity and phylogenetic point of view. The EVO·COM group recently started to survey ant biodiversity in Northern Italy in a collaborative [link a collaborations] framework with research groups of the University of Parma and of the University of Florence. The integrative analysis of DNA barcoding data with morphological identification on >300 samples led to the identification of ~80 different taxa. Though, PCR amplification and Sanger sequencing of the DNA barcode marker proved to be time consuming and extremely difficult in some instances, due to laboratory experiment settings that can change from species to species making the whole process less efficient, costly, and even, in some cases, unsuccessful. This in addition to the limited informativity of the single DNA barcode molecular marker. The goal of this project is the implementation of genome skimming as a routine application for the analysis to the study of Italian ant biodiversity and evolution.


Formica paralugubris is a member of the Formica rufa species group (red wood ants) native to the Alps. It is considered a cryptic species, morphologically similar to Formica lugubris, however, F. paralugubris ants react aggressively towards F. lugubris, and the two species present important biological differences, such as different colony structures. Indeed F. paralugubris is a highly polygynous species, where multiple unrelated queens usually co-exist within a large, interconnected nest, while F. lugubris is facultatively polygynous. From an ecological point of view, redwood ants are extremely important in forest ecosystems. Their large colonies prey on a wide variety of invertebrates, and in Italy were employed to combat herbivore moths and sawflies, such as Nematus erichsoni, Coleophora laricella, and Thaumetopeoa pityocampa, which are important pests for fir, pine, and beech woods. Starting in 1958, red wood ants from the Alps were transplanted multiple times to several Apennine forests along the Italian peninsula to be employed as biological control agents for tree insect pests. Similarly, F. paralugubris has also been introduced into North America for the same purpose. We are interested in the effects and evolutionary consequences of genome content and genome architecture variation, and in this project, we are going to sequence and assemble a chromosome-level genome of F. paralugubris to understand at a fine-scale evolutionary dynamic of both introduced and native populations in terms of Single Nucleotide Variants and Structural Variations in a biodiversity genomics framework. Moreover, taking advantage of multiple, well-documented, and independent human-mediated introductions we will use this system as a model to study the genomic consequences of bottlenecks in genome evolutionary dynamics, such as transposable element activity. As a second goal, we aim to use the newly assembled genome in comparative genomic analyses across the whole Formica clade to understand major ant evolutionary transitions at a broad scale, such as the evolution of genes, gene families, transposable elements, and supergenes (i.e.: clusters of tightly linked genes that coordinately control complex phenotypes). Recently, an ancient supergene was found to be responsible for polygyny/monogyny switch in several ant species, and the increase in genomic resources for the clade is an important step forward in a deeper understanding of its evolution.


Work In Progress