LE SIMULAZIONI DEL PROCESSO COALESCENTE IN GENETICA DI POPOLAZIONI: INFERENZE DEMOGRAFICHE ED EVOLUTIVE

The main goal of population genetics is to understand the factors that affect genetic variation within a species. Mathematical models are used to predict the effects on genetic variation of processes such as mutation, recombination, selection, migration and population size changes, but analytical results are difficult to obtain when these processes interact and when equilibrium conditions are not met. In these situations, common in real biological systems especially when recent human activities (e.g., stocking, urbanization, overhunting) perturb natural populations, computer simulations can be very useful. A computer simulation is a virtual experiment in which a model is used to mimic the biological process on a computer to study its properties. It is an excellent tool for understanding the functioning of complex systems. Simulations are generally used to make predictions about populations, validate statistical methods, study the properties of different sampling strategies, and estimate parameters from real data. In this thesis, I applied genetic simulations to address questions intractable with other methods. First, I analyzed the effects of violating the assumption of panmixiamade by “Extende Bayesian Skyline Plot” (EBSP) method. I showed that migration can influence the inferred demographic history of a population, suggesting wrong dynamics. Second, I used genetic simulations to analyse the performance of the EBSP method in reconstructing a population decline and to compare sampling schemes with different proportions of modern and ancient DNA. I identified some properties of the sampling scheme which clearly positively affect the demographic reconstruction, providing some simple hints for planning a genetic study when both modern and ancient samples are available. Third, I familiarized with the “Approximated Bayesian Computation” methodology and I contributed to a review article presenting the main features, with pros and cons, of this approach. Fourth, I applied the ABC procedure to analyze the hybridization history within the genus Chionodraco, and to evaluate the power of ABC in this context. Realistic demographic models were defined and compared, and evidence was found that hybridization occurred only in interglacial periods. Taken together, the results presented in this thesis confirm the importance of genetic simulations in evolutionary biology. If we consider the increasing availability of simulation packages, along with the increasing speed and storage capacity of personal computers and clusters, it is easy to predict that simulations of genetic and genomic data will spread in many fields to better explore more and more realistic, and consequently complex, models

C6orf10 low-frequency and rare variants in italian multiple sclerosis patients

In light of the complex nature of multiple sclerosis (MS) and the recently estimated contribution of low-frequency variants into disease, decoding its genetic risk components requires novel variant prioritization strategies. We selected, by reviewing MS Genome Wide Association Studies (GWAS), 107 candidate loci marked by intragenic single nucleotide polymorphisms (SNPs) with a remarkable association (p-value <= 5 x 10(-6)). A whole exome sequencing (WES)-based pilot study of SNPs with minor allele frequency (MAF) <= 0.04, conducted in three Italian families, revealed 15 exonic low-frequency SNPs with affected parent-child transmission. These variants were detected in 65/120 Italian unrelated MS patients, also in combination (22 patients). Compared with databases (controls gnomAD, dbSNP150, ExAC, Tuscany-1000 Genome), the allelic frequencies of C6orf10 rs 16870005 and IL2RA rs12722600 were significantly higher (i.e., controls gnomAD, p = 9.89 x 10(-7) and p < 1 x 10(-20)). TET2 rs61744960 and TRAF3 rs138943371 frequencies were also significantly higher, except in Tuscany-1000 Genome. Interestingly, the association of C6orf10 rs16870005 (Ala431Thr) with MS did not depend on its linkage disequilibrium with the HLA-DRB1 locus. Sequencing in the MS cohort of the C6orf10 3' region revealed 14 rare mutations (10 not previously reported). Four variants were null, and significantly more frequent than in the databases. Further, the C6orf10 rare variants were observed in combinations, both intra-locus and with other low-frequency SNPs. The C6orf10 Ser389Xfr was found homozygous in a patient with early onset of the MS. Taking into account the potentially functional impact of the identified exonic variants, their expression in combination at the protein level could provide functional insights in the heterogeneous pathogenetic mechanisms contributing to MS.In light of the complex nature of multiple sclerosis (MS) and the recently estimated contribution of low-frequency variants into disease, decoding its genetic risk components requires novel variant prioritization strategies. We selected, by reviewing MS Genome Wide Association Studies (GWAS), 107 candidate loci marked by intragenic single nucleotide polymorphisms (SNPs) with a remarkable association (p-value ≤ 5 × 10−6). A whole exome sequencing (WES)-based pilot study of SNPs with minor allele frequency (MAF) ≤ 0.04, conducted in three Italian families, revealed 15 exonic low-frequency SNPs with affected parent-child transmission. These variants were detected in 65/120 Italian unrelated MS patients, also in combination (22 patients). Compared with databases (controls gnomAD, dbSNP150, ExAC, Tuscany-1000 Genome), the allelic frequencies of C6orf10 rs16870005 and IL2RA rs12722600 were significantly higher (i.e., controls gnomAD, p = 9.89 × 10−7 and p < 1 × 10−20). TET2 rs61744960 and TRAF3 rs138943371 frequencies were also significantly higher, except in Tuscany-1000 Genome. Interestingly, the association of C6orf10 rs16870005 (Ala431Thr) with MS did not depend on its linkage disequilibrium with the HLA-DRB1 locus. Sequencing in the MS cohort of the C6orf10 3′ region revealed 14 rare mutations (10 not previously reported). Four variants were null, and significantly more frequent than in the databases. Further, the C6orf10 rare variants were observed in combinations, both intra-locus and with other low-frequency SNPs. The C6orf10 Ser389Xfr was found homozygous in a patient with early onset of the MS. Taking into account the potentially functional impact of the identified exonic variants, their expression in combination at the protein level could provide functional insights in the heterogeneous pathogenetic mechanisms contributing to MS

The Biarzo case in northern Italy: Is the temporal dynamic of swine mitochondrial DNA lineages in Europe related to domestication?

Genetically-based reconstructions of the history of pig domestication in Europe are based on two major pillars: 1) the temporal changes of mitochondrial DNA lineages are related to domestication; 2) Near Eastern haplotypes which appeared and then disappeared in some sites across Europe are genetic markers of the first Near Eastern domestic pigs. We typed a small but informative fragment of the mitochondrial DNA in 23 Sus scrofa samples from a site in north eastern Italy (Biarzo shelter) which provides a continuous record across a ≈6,000 year time frame from the Upper Palaeolithic to the Neolithic. We additionally carried out several radiocarbon dating. We found that a rapid mitochondrial DNA turnover occurred during the Mesolithic, suggesting that substantial changes in the composition of pig mitochondrial lineages can occur naturally across few millennia independently of domestication processes. Moreover, so-called Near Eastern haplotypes were present here at least two millennia before the arrival of Neolithic package in the same area. Consequently, we recommend a re-evaluation of the previous idea that Neolithic farmers introduced pigs domesticated in the Near East, and that Mesolithic communities acquired domestic pigs via cultural exchanges, to include the possibility of a more parsimonious hypothesis of local domestication in Europe

Concurrent intra-arterial carboplatin administration and radiation therapy for the treatment of advanced head and neck squamous cell carcinoma: short term results

BACKGROUND: The aim of the present study was to evaluate the survival, efficacy and safety of a modified RADPLAT-like protocol using carboplatin instead of cisplatin. METHODS: Fifty-six patients with primary head and neck squamous cell carcinoma received 4 cycles of intra-arterial carboplatin (350 mg/m2 per cycle every 2 weeks), with concurrent three-dimensional conformal radiation therapy. RESULTS: Two major and 4 minor complications were observed. Forty-five of the 56 patients (80%) completed the protocol, while 11 (20%) patients had to discontinue the intra-arterial infusions due to the occurrence of severe haematological toxicity, but were able to complete radiotherapy. Forty-four (98%) of the 45 patients who completed the protocol and 10 (91%) of the 11 who did not, were free of disease at the end of the treatment, for a comprehensive 96% of CR overall. After a median 23.55 months (range: 2 to 58 months) of follow-up, 40 patients (71%) are alive and disease-free, 1 (2%) is alive but affected by disease and 15 (27%) have died of the disease or other causes. CONCLUSION: Intra-arterial carboplatin administration with concurrent three-dimensional conformal radiation therapy seems to be a promising alternative to RADPLAT in the treatment of advanced head and neck tumours. Haematological and non-haematological toxicities are virtually similar, but carboplatin has the advantage in that it is not nephrotoxic and can be used at very high doses without any significant increase in the extent of side effects

Survival and divergence in a small group: The extraordinary genomic history of the endangered Apennine brown bear stragglers

About 100 km east of Rome, in the central Apennine Mountains, a critically endangered population of ∼50 brown bears live in complete isolation. Mating outside this population is prevented by several 100 km of bear-free territories. We exploited this natural experiment to better understand the gene and genomic consequences of surviving at extremely small population size. We found that brown bear populations in Europe lost connectivity since Neolithic times, when farming communities expanded and forest burning was used for land clearance. In central Italy, this resulted in a 40-fold population decline. The overall genomic impact of this decline included the complete loss of variation in the mitochondrial genome and along long stretches of the nuclear genome. Several private and deleterious amino acid changes were fixed by random drift; predicted effects include energy deficit, muscle weakness, anomalies in cranial and skeletal development, and reduced aggressiveness. Despite this extreme loss of diversity, Apennine bear genomes show nonrandom peaks of high variation, possibly maintained by balancing selection, at genomic regions significantly enriched for genes associated with immune and olfactory systems. Challenging the paradigm of increased extinction risk in small populations, we suggest that random fixation of deleterious alleles (i) can be an important driver of divergence in isolation, (ii) can be tolerated when balancing selection prevents random loss of variation at important genes, and (iii) is followed by or results directly in favorable behavioral changes

Are sinonasal dissection courses a valid instrument for endoscopic sinus surgeons? A report on 7-years of experience

I corsi di chirurgia endoscopica naso-sinusale sono rivolti sia a giovani specialisti per il training chirurgico, sia a chirurghi più esperti per migliorare le abilità chirurgiche. Ad un totale di centotrenta (130) partecipanti, ad 8 corsi di dissezione in totale, tra il 2013 e il 2019, è stato somministrato un questionario strutturato standardizzato, comprendente domande sulla strumentazione ricevuta e problemi riscontrati durante la pratica di dissezione. L’uncinectomia e l’etmoidectomia anteriore sono state considerate le procedure più semplici del corso, dalla maggior parte dei partecipanti, mentre l’approccio al seno frontale, indipendentemente dall’esperienza e dall’età, è stato ritenuto il passaggio più complicato. Tutti i partecipanti hanno considerato il corso di dissezione come un mezzo utile per migliorare le loro conoscenze anatomiche, ed il grado di confidenza con la strumentazione e con la dissezione. I corsi di chirurgia endoscopica naso-sinusale sono stati ritenuti utili sia dai giovani specialisti, che dai chirurghi più esperti. Il miglioramento delle conoscenze anatomiche può ridurre efficacemente l’insorgenza di complicazioni indesiderate, in particolare per gli approcci endoscopici.Sinonasal dissection courses have been reported to be effective in teaching sinonasal anatomy to trainees, and in improving surgical skills for trained surgeons. Between 2013 and 2019, a standardised structured questionnaire was handed out to 130 participants of 8 sinonasal dissection courses. Each questionnaire included questions about the medical equipment the surgeons were fitted, and the problems encountered during the dissection.The majority of the participants, regardless of their experience and age, considered uncinectomy and anterior ethmoidectomy as the simplest parts of the dissection, whereas the most complicated step was the approach to the frontal sinus. All participants considered the dissection course as a useful way to improve their medical skills while performing an endoscopic surgery procedure, such as their anatomical knowledge, and confidence with instrumentation and the dissection. Sinonasal dissection courses can be considered to be useful for both trainees and trained surgeons. Improving anatomical knowledge can reduce the occurrence of complications, especially in endoscopic surgery

Assessment of different manufacturing techniques for the production of bioartificial scaffolds as soft organ transplant substitutes

Introduction: The problem of organs’ shortage for transplantation is widely known: different manufacturing techniques such as Solvent casting, Electrospinning and 3D Printing were considered to produce bioartificial scaffolds for tissue engineering purposes and possible transplantation substitutes. The advantages of manufacturing techniques’ combination to develop hybrid scaffolds with increased performing properties was also evaluated.Methods: Scaffolds were produced using poly-L-lactide-co-caprolactone (PLA-PCL) copolymer and characterized for their morphological, biological, and mechanical features.Results: Hybrid scaffolds showed the best properties in terms of viability (&gt;100%) and cell adhesion. Furthermore, their mechanical properties were found to be comparable with the reference values for soft tissues (range 1–10 MPa).Discussion: The created hybrid scaffolds pave the way for the future development of more complex systems capable of supporting, from a morphological, mechanical, and biological standpoint, the physiological needs of the tissues/organs to be transplanted

Survival and divergence in a small group: The extraordinary genomic history of the endangered Apennine brown bear stragglers

About 100 km east of Rome, in the central Apennine Mountains, a critically endangered population of &sim;50 brown bears live in complete isolation. Mating outside this population is prevented by several 100 km of bear-free territories. We exploited this natural experiment to better understand the gene and genomic consequences of surviving at extremely small population size. We found that brown bear populations in Europe lost connectivity since Neolithic times, when farming communities expanded and forest burning was used for land clearance. In central Italy, this resulted in a 40-fold population decline. The overall genomic impact of this decline included the complete loss of variation in the mitochondrial genome and along long stretches of the nuclear genome. Several private and deleterious amino acid changes were fixed by random drift; predicted effects include energy deficit, muscle weakness, anomalies in cranial and skeletal development, and reduced aggressiveness. Despite this extreme loss of diversity, Apennine bear genomes show nonrandom peaks of high variation, possibly maintained by balancing selection, at genomic regions significantly enriched for genes associated with immune and olfactory systems. Challenging the paradigm of increased extinction risk in small populations, we suggest that random fixation of deleterious alleles (i) can be an important driver of divergence in isolation, (ii) can be tolerated when balancing selection prevents random loss of variation at important genes, and (iii) is followed by or results directly in favorable behavioral changes.Additional co-authors: Claudio Groff, Ladislav Paule, Leonardo Gentile, Carles Vilà, Saverio Vicario, Luigi Boitani, Ludovic Orlando, Silvia Fuselli, Cristiano Vernesi, Beth Shapiro, Paolo Ciucci, and Giorgio Bertorell