The physical city and the urban structure: detecting amenity zones and applying urban morphology to New York

The city is a collection of built structures and infrastructure embedded in socio-cultural processes: any investigation into a city’s transformations involves considerations on the degree to which its composite elements respond to socio-economical changes. The main purpose of this research is to investigate how transformations in the functional requirements of New York’s society have spurred, since the 1970s, changes in both the city’s urban structure and physical form. The present work examines the rise of Amenity Zones in New York, and investigates the transformations that have occurred in New York’s built environment since the 1970s. By applying qualitative measures and analyzing the relationship between urban amenities and the creative class, the present work has investigated changes in the urban structure and detected a hierarchical series of amenity zones classes, namely, Super Amenity Zones (SAZs), Nodal Amenity Zones (NAZs) and Peripheral Amenity Zones (PAZs). This series allows for a more comprehensive reading of the urban structure in a complex city like New York, bringing advancements to the amenity zone’s methodology. In order to examine the manner in which the other component of the city, the physical form, has changed or adapted to the new socio-economic condition, the present research has applied Conzenian analysis to a select study area, Atlantic Avenue. The results of this analysis reveal that, contrary to the urban structure, which changes rapidly, the physical form of New York is hard to modify completely, due to the resilience of the town plan and its elements, and to preservation laws; the city rather adapts to socio-economical changes through process of adaptive reuses or conversion. Concluding, this research has examined the dialectic between the ever-changing needs of society and the complexity of the built environment and urban structure, showing the different degrees to which the urban landscape modifies, reacts and sometimes adapts to the population’s functional requirements.

Analysis of the immunological and functional features of the Neisserial Heparin Binding Antigen (NHBA)

Neisserial Heparin Binding Antigen (NHBA) is a surface-exposed lipoprotein ubiquitously expressed by genetically diverse Neisseria meningitidis strains and is an antigen of the multicomponent protein-based 4CMenB vaccine, able to induce bactericidal antibodies in humans and to bind heparin-like molecules. The aim of this study is to characterize the immunological and functional properties of NHBA. To evaluate immunogenicity and the contribution of aminoacid sequence variability to vaccine coverage, we constructed recombinant isogenic strains that are susceptible to bactericidal killing only by anti-NHBA antibodies and engineered them to express equal levels of selected NHBA peptides. In these recombinant strains, we observed different titres associated with the different peptide variants. These recombinant strains were then further engineered to express NHBA chimeric proteins to investigate the regions important for immunogenicity. In natural strains, anti-NHBA antibodies were found to be cross-protective against strains expressing different peptides. To investigate the functional properties of this antigen, the recombinant purified NHBA protein was tested in in vitro binding studies and was found to be able to bind epithelial cells. The binding was abolished when cells were treated specifically with heparinase III, suggesting that the interaction with the cells is mediated by heparan sulfate proteoglycans (HSPG). Mutation of the Arg-rich tract of NHBA abrogated the binding, confirming the importance of this region in mediating the binding to heparin-like molecules. In a panel of N. meningitidis strains, the deletion of nhba resulted in a reduction of adhesion with respect to each isogenic wild type strain. Furthermore, the adhesion of the wild-type strain was prevented by using anti-NHBA polyclonal sera, demonstrating the specificity of the interaction. These results suggest that NHBA could be a novel meningococcal adhesin contributing to host-cell interaction. Moreover, we analysed NHBA NalP-mediated cleavage in different NHBA peptides and showed that not all NHBA peptides are cleaved.

Molecular basis oh herpes simplex virus entry into the cell and retargeting of the viral tropism for the design of oncolytic herpesviruses

Herpes simplex virus 1 (HSV-1) infects oral epitelial cells, then spreads to the nerve endings and estabilishes latency in sensory ganglia, from where it may, or may not reactivate. Diseases caused by virus reactivation include mild diseases such as muco-cutaneous lesions, and more severe, and even life-threatening encephalitis, or systemic infections affecting diverse organs. Herpes simplex virus represents the most comprehensive example of virus receptor interaction in Herpesviridae family, and the prototype virus encoding multipartite entry genes. In fact, it encodes 11-12 glycoproteins and a number of additional membrane proteins: five of these proteins play key roles in virus entry into subsceptible cells. Thus, glycoprotein B (gB) and glycoprotein C (gC) interact with heparan sulfate proteoglycan to enable initial attachment to cell surfaces. In the next step, in the entry cascade, gD binds a specific surface receptor such as nectin1 or HVEM. The interaction of glycoprotein D with the receptor alters the conformation of gD to enable the activation of gB, glycoprotein H, and glycoprotein L, a trio of glycoproteins that execute the fusion of the viral envelope with the plasma membrane. In this thesis, I described two distinct projects: I. The retargeting of viral tropism for the design of oncolytic Herpesviruses: • capable of infecting cells through the human epitelial growth factor receptor 2 (HER2), overexpressed in highly malignant mammary and ovarian tumors and correlates with a poor prognosis; • detargeted from its natural receptors, HVEM and nectin1. To this end, we inserted a ligand to HER2 in gD. Because HER2 has no natural ligand, the selected ligand was a single chain antibody (scFv) derived from MAb4D5 (monoclonal antibody to HER2), herein designated scHER2. All recombinant viruses were targeted to HER2 receptor, but only two viruses (R-LM113 and R-LM249) were completely detargeted from HVEM and nectin1. To engineer R-LM113, we removed a large portion at the N-terminus of gD (from aa 6 to aa 38) and inserted scHER2 sequence plus 9-aa serine-glycine flexible linker at position 39. On the other hand, to engineer R-LM249, we replaced the Ig-folded core of gD (from aa 61 to aa 218) with scHER2 flanked by Ser-Gly linkers. In summary, these results provide evidence that: i. gD can tolerate an insert almost as big as gD itself; ii. the Ig-like domain of gD can be removed; iii. the large portion at the N-terminus of gD (from aa 6 to aa 38) can be removed without loss of key function; iv. R-LM113 and R-LM249 recombinants are ready to be assayed in animal models of mammary and ovary tumour. This finding and the avaibility of a large number of scFv greatly increase the collection of potential receptors to which HSV can be redirected. II. The production and purification of recombinant truncated form of the heterodimer gHgL. We cloned a stable insect cell line expressing a soluble form of gH in complex with gL under the control of a metalloprotein inducible promoter and purified the heterodimer by means of ONE-STrEP-tag system by IBA. With respect to biological function, the purified heterodimer is capable: • of reacting to antibodies that recognize conformation dependent epitopes and neutralize virion infectivity; • of binding a variety cells at cell surface. No doubt, the availability of biological active purified gHgL heterodimer, in sufficient quantities, will speed up the efforts to solve its crystal structure and makes it feasible to identify more clearly whether gHgL has a cellular partner, and what is the role of this interaction on virus entry.

Structural analysis of combinatorial optimization problem characteristics and their resolution using hybrid approaches

Many combinatorial problems coming from the real world may not have a clear and well defined structure, typically being dirtied by side constraints, or being composed of two or more sub-problems, usually not disjoint. Such problems are not suitable to be solved with pure approaches based on a single programming paradigm, because a paradigm that can effectively face a problem characteristic may behave inefficiently when facing other characteristics. In these cases, modelling the problem using different programming techniques, trying to ”take the best” from each technique, can produce solvers that largely dominate pure approaches. We demonstrate the effectiveness of hybridization and we discuss about different hybridization techniques by analyzing two classes of problems with particular structures, exploiting Constraint Programming and Integer Linear Programming solving tools and Algorithm Portfolios and Logic Based Benders Decomposition as integration and hybridization frameworks.

Phylogeny and population dynamics in European Reticulitermes and Kalotermes genera (Insecta, Isoptera)

The aim of this thesis is to detect the phylogeny and the population dynamics of the European termites of the genera Reticulitermes and Kalotermes, by the use of different mitochondrial (16S, COI/tRNA/COII, CR) and nuclear (microsatellites and Inter-SINE) molecular markers. In the phylogenetic analyses, the obtained results have well defined the cladogenetic events that generated the nowadays species biodiversity of the genus Reticulitermes, while the analysis of the Kalotermes flavicollis taxon showed the presence of at least four genetic clades, defined on the basis of the geographical distance. The second part of the thesis is centred on the population dynamics of two species: Reticulitermes urbis and Kalotermes flavicollis. The first species, native of the Balkans, is known to be present in some cities of Italy and France. I’ve analyzed the colony genetic structure of the introduced population of Bagnacavallo (RA, Italy), using nine microsatellite loci. The obtained results are in accordance with those obtained from another population in France: this species in fact confirms its invasive and infestation capacities. The analysis of the natural population of K. flavicollis, performed with a combination of mitochondrial (control region) and nuclear (microsatellites and I-SINE) markers, clearly evidenced the presence of two genetic lineages that coexist in the same area. Moreover, results clearly indicate that the cross-breeding is allowed. Finally, the whole results are discussed in a comparative view to better understand the differences in ecology, evolutionary dynamics and colony social structure between these two genera.

An optimal estimator for the correlation of CMB anisotropies with Large Scale Structures and its application to WMAP-7year and NVSS

In the thesis we present the implementation of the quadratic maximum likelihood (QML) method, ideal to estimate the angular power spectrum of the cross-correlation between cosmic microwave background (CMB) and large scale structure (LSS) maps as well as their individual auto-spectra. Such a tool is an optimal method (unbiased and with minimum variance) in pixel space and goes beyond all the previous harmonic analysis present in the literature. We describe the implementation of the QML method in the {\it BolISW} code and demonstrate its accuracy on simulated maps throughout a Monte Carlo. We apply this optimal estimator to WMAP 7-year and NRAO VLA Sky Survey (NVSS) data and explore the robustness of the angular power spectrum estimates obtained by the QML method. Taking into account the shot noise and one of the systematics (declination correction) in NVSS, we can safely use most of the information contained in this survey. On the contrary we neglect the noise in temperature since WMAP is already cosmic variance dominated on the large scales. Because of a discrepancy in the galaxy auto spectrum between the estimates and the theoretical model, we use two different galaxy distributions: the first one with a constant bias $b$ and the second one with a redshift dependent bias $b(z)$. Finally, we make use of the angular power spectrum estimates obtained by the QML method to derive constraints on the dark energy critical density in a flat $\Lambda$CDM model by different likelihood prescriptions. When using just the cross-correlation between WMAP7 and NVSS maps with 1.8° resolution, we show that $\Omega_\Lambda$ is about the 70\% of the total energy density, disfavouring an Einstein-de Sitter Universe at more than 2 $\sigma$ CL (confidence level).

Composite materials design, manufacture and evaluation

Fibre-Reinforced-Plastics are composite materials composed by thin fibres with high mechanical properties, made to work together with a cohesive plastic matrix. The huge advantages of fibre reinforced plastics over traditional materials are their high specific mechanical properties i.e. high stiffness and strength to weight ratios. This kind of composite materials is the most disruptive innovation in the structural materials field seen in recent years and the areas of potential application are still many. However, there are few aspects which limit their growth: on the one hand the information available about their properties and long term behaviour is still scarce, especially if compared with traditional materials for which there has been developed an extended database through years of use and research. On the other hand, the technologies of production are still not as developed as the ones available to form plastics, metals and other traditional materials. A third aspect is that the new properties presented by these materials e.g. their anisotropy, difficult the design of components. This thesis will provide several case-studies with advancements regarding the three limitations mentioned. In particular, the long term mechanical properties have been studied through an experimental analysis of the impact of seawater on GFRP. Regarding production methods, the pre-impregnated cured in autoclave process was considered: a rapid tooling method to produce moulds will be presented, and a study about the production of thick components. Also, two liquid composite moulding methods will be presented, with a case-study regarding a large component with sandwich structure that was produced with the Vacuum-Assisted-Resin-Infusion method, and a case-study regarding a thick con-rod beam that was produced with the Resin-Transfer-Moulding process. The final case-study will analyse the loads acting during the use of a particular sportive component, made with FRP layers and a sandwich structure, practical design rules will be provided.

New synthetic bile acid analogue agonists of FXR and TGR5 receptors: Analytical methodologies for the study of their physico-chemical properties, pharmacokinetic activity and metabolism.

This thesis reports an integrated analytical approach for the study of physicochemical and biological properties of new synthetic bile acid (BA) analogues agonists of FXR and TGR5 receptors. Structure-activity data were compared with those previous obtained using the same experimental protocols on synthetic and natural occurring BA. The new synthetic BA analogues are classified in different groups according also to their potency as a FXR and TGR5 agonists: unconjugated and steroid modified BA and side chain modified BA including taurine or glycine conjugates and pseudo-conjugates (sulphonate and sulphate analogues). In order to investigate the relationship between structure and activity the synthetic analogues where admitted to a physicochemical characterization and to a preliminary screening for their pharmacokinetic and metabolism using a bile fistula rat model. Sensitive and accurate analytical methods have been developed for the quali-quantitative analysis of BA in biological fluids and sample used for physicochemical studies. Combined High Performance Liquid Chromatography Electrospray tandem mass spectrometry with efficient chromatographic separation of all studied BA and their metabolites have been optimized and validated. Analytical strategies for the identification of the BA and their minor metabolites have been developed. Taurine and glycine conjugates were identified in MS/MS by monitoring the specific ion transitions in multiple reaction monitoring (MRM) mode while all other metabolites (sulphate, glucuronic acid, dehydroxylated, decarboxylated or oxo) were monitored in a selected-ion reaction (SIR) mode with a negative ESI interface by the following ions. Accurate and precise data where achieved regarding the main physicochemical properties including solubility, detergency, lipophilicity and albumin binding . These studies have shown that minor structural modification greatly affect the pharmacokinetics and metabolism of the new analogues in respect to the natural BA and on turn their site of action, particularly where their receptor are located in the enterohepatic circulation.

Hydrodynamical simulations of early-type galaxies: effects of galaxy shape and stellar dynamics on hot coronae

Early-Type galaxies (ETGs) are embedded in hot (10^6-10^7 K), X-ray emitting gaseous haloes, produced mainly by stellar winds and heated by Type Ia supernovae explosions, by the thermalization of stellar motions and occasionally by the central super-massive black hole (SMBH). In particular, the thermalization of the stellar motions is due to the interaction between the stellar and the SNIa ejecta and the hot interstellar medium (ISM) already residing in the ETG. A number of different astrophysical phenomena determine the X-ray properties of the hot ISM, such as stellar population formation and evolution, galaxy structure and internal kinematics, Active Galactic Nuclei (AGN) presence, and environmental effects. With the aid of high-resolution hydrodynamical simulations performed on state-of-the-art galaxy models, in this Thesis we focus on the effects of galaxy shape, stellar kinematics and star formation on the evolution of the X-ray coronae of ETGs. Numerical simulations show that the relative importance of flattening and rotation are functions of the galaxy mass: at low galaxy masses, adding flattening and rotation induces a galactic wind, thus lowering the X-ray luminosity; at high galaxy masses the angular momentum conservation keeps the central regions of rotating galaxies at low density, whereas in non-rotating models a denser and brighter atmosphere is formed. The same dependence from the galaxy mass is present in the effects of star formation (SF): in light galaxies SF contributes to increase the spread in Lx, while at high galaxy masses the halo X-ray properties are marginally sensitive to SF effects. In every case, the star formation rate at the present epoch quite agrees with observations, and the massive, cold gaseous discs are partially or completely consumed by SF on a time-scale of few Gyr, excluding the presence of young stellar discs at the present epoch.

Multiscale comparative analysis of marine biominerals and their ecological implications

Marine biomineralizing organisms provide a fundamental link between biology and environment. Calcified structure are important archives that can provide us main means of understanding organism adaptation, habits, environmental characteristics, and to look back in time and explore the past climate and their evolutionary history. In fact, biomineralized structures retain an unparalleled record of current and past ocean conditions through the investigation of their microchemistry and isotopes. This thesis considers aspects of two different biomineralization systems: fish otolith and coral skeletons at macro-, micro- and nanoscale, with the aim to understand how their morphology, structural characteristics and compositions can provide information of their functionality, and the environmental, behavioural, and evolutionary context in which organisms are framed. To this end, I applied a multidisciplinary approach in the scope to investigate calcified structures as “information recorders” and as models to study the phenotypic plasticity.

Protein-based nanomaterials as protein heterogeneous nucleants and structural studies on enzymes from two photosynthetic organisms

Proteins, the most essential biological macromolecules, are involved in nearly every aspect of life. The elucidation of their three-dimensional structures through X-ray analysis has significantly contributed to our understanding of fundamental mechanisms in life processes. However, the obstacle of obtaining high-resolution protein crystals remains significant. Thus, searching for materials that can effectively induce nucleation of crystals is a promising and active field. This thesis work characterizes and prepares albumin nanoparticles as heterogeneous nucleants for protein crystallization. These stable Bovine Serum Albumin nanoparticles were synthesized via the desolvation method, purified efficiently, and characterized in terms of dimension, morphology, and secondary structure. The ability of BSA-NPs to induce macromolecule nucleation was tested on three model proteins, exhibiting significant results, with larger NPs inducing more nucleation. The second part of this work focuses on the structural study, mainly through X-ray crystallography, of five chloroplast and cytosolic enzymes involved in the fundamental cellular processes of two photosynthetic organisms, Chlamydomonas reinhardtii and Arabidopsis thaliana. The structures of three enzymes involved in the Calvin-Benson-Bassham Cycle, phosphoribulokinase, troseposphatisomerase, and ribulosiophosphate epimerase from Chlamydomonas reinhardtii, were solved to investigate their catalytic and regulatory mechanisms. Additionally, the structure of nitrosylated-CrTPI made it possible to identify Cys14 as a target for nitrosylation, and the crystallographic structure of CrRPE was solved for the first time, providing insights into its catalytic and regulatory properties. Finally, the structure of S-nitrosoglutathione reductase, AtGSNOR, was compared with that of AtADH1, revealing differences in their catalytic sites. Overall, seven crystallographic structures, including partially oxidized CrPRK, CrPRK/ATP, CrPRK/ADP/Ru5P, CrTPI-nitrosylated, apo-CrRPE, apo-AtGSNOR, and AtADH1-NADH, were solved and are yet to be deposited in the PDB.

Applied genomics in livestock: genome-wide association studies and population genomics analysis in pig and rabbit breeds

The domestication and selection processes in pigs and rabbits have resulted in the constitution of multiple breeds with broad phenotypic diversity. Population genomics analysis and Genome-wide association study analysis can be utilized to gain insights into the ancestral origins, genetic diversity, and the presence of lethal mutations across these diverse breeds. In this thesis, we analysed the dataset obtained from three Italian Pig breeds to detect deleterious alleles. We screened the dataset for genetic markers showing homozygous deficiency using two approaches single marker and haplotype-based approach. Moreover, Genome-wide association study analyses were performed to detect genetic markers associated with pigs' reproductive traits. In rabbits, we investigated the application of SNP bead chip for detection signatures of selection in rabbits using different methods. This analysis was implemented for the first time in different fancy and meet rabbit breeds. Multiple approaches were utilized for the detection of the selection of signatures including Fst analysis, ROH analysis, PCAdapt analysis, and haplotype-based analysis. The analysis in pigs was able to identify five putative deleterious SNPs and nine putative deleterious haplotypes in the analysed Italian Pig breeds. The genomic regions of the detected putative deleterious genomic markers harboring loss of function variants such as the Frameshift variant, start lost, and splice donor variant. Those variants are close to important candidate genes such as IGF2BP1, ADGRL4, and HGF. In rabbits, multiple genomic regions were detected to be under selection of signature. These genomic regions harbor candidate genes associated with coat color phenotype (MC1R, TYR, and ASIP), hair structure (LIPH), and body size (HMGA2 and COL2A1). The described results in rabbits and pigs could be used to improve breeding programs by excluding the deleterious genetic markers carriers and incorporating candidate genes for coat color, body size, and meat production in rabbit breeding programs to enhance desired traits