Single nucleotide polymorphisms : from forensic identification to DNA phenotyping

Tese de doutoramento, Biologia (Genética), Universidade de Lisboa, Faculdade de Ciências, 2017Single nucleotide polymorphisms or SNPs present low mutation rates and its biology permits for an automated analysis, with a relatively simple interpretation due to the fact of the techniques used to analyze these markers originating few artifacts. This allows to attain additional genetic information that can offer great value to the geneticist and for the discovery of the truth in civil or criminal cases. For all these, in recent years we have witnessed the rapid development of SNP methodology as a tool in forensic genetics. These markers can be characterized in four forensically relevant classes, one of which consists of SNPs useful in individual identification and other consists in polymorphisms used to identify family lineages, allowing the obtainment of additional genetic information to complement those usually achieved through STR analysis. Thus, SNP study in these situations relates to the need of additional genetic information due to the existence of STR incompatibilities in the studied systems or due to the lack of sample from individuals essential to the analysis of kinship casework. SNP study is also of great importance when incomplete amplification or no amplification of the studied STR systems is observed, resulting from the small quantity and / or poor quality of the analyzed DNA. The two remaining SNP classes cannot provide individual or family identification but they present large forensic interest because they allow inferring about characteristics of the sample donor. The first of these classes is composed by SNPs that provide biogeografical ancestry information and the remainder class consists of markers that provide information about phenotypic characteristics. This type of information can be especially important when no information is known about the sample donor and not even his or her genetic profile has entered into DNA databases, in a way that these will provide valuable clues to the discovery of the individual yet to be identified. The application of these markers to forensics is being studied by several research groups worldwide so that their use can become a reality in the near future. However, since forensic genetics is founded on scientific knowledge and based on human genetic diversity, it is important to study these in the populations in which this type of analysis can be used. Thus, it is important to study the SNP loci in the Portuguese population so that these markers can be used to carry out forensic testing in cases involving Portuguese individuals. The purpose of this work was to contribute to this development, both through the verification of SNP multiplexes in the Portuguese population and through the modification and improvement of these tools in order to obtain more and better Information with them. Thus, this work intended to study and optimize individual identification SNP multiplexes to be used on kinship testing and on individual identification and to study a phenotype informative SNP multiplex for application in the resolution of cases where there is no suspect’s sample for comparison. Initially, based on the 52-plex developed by the SNPforID consortium, it was developed and optimized a set of 20 loci set for individual identification so that these could be used as an additional tool to STR investigation in the study of paternity testing and kinship analysis. Later, it was studied the population and forensic parameters of the 52-plex in the Portuguese population and the effectiveness of this multiplex in the analysis of complex samples such as bone, tooth or crime scene’s samples from which it was not obtained a complete genetic profile when applying a commercial STR kit. Another main objective was to study the application of IrisPlex in Portuguese population. This is a multiplex designed to infer about the eye color of a sample donor. In addition, since that when this thesis project was carried out it was verified that the markers that make up this multiplex, besides being related to eye color determination are also involved in hair and skin color, due to the main genes involved in the 3 mechanisms being the same, it was also decided to supplement this multiplex with additional markers in order to verify if the resulting tool could provide additional information on these other externally visible characteristics. Thus, this doctoral thesis presents the results obtained through the analysis of SNPs and of its applicability to the Portuguese population, focusing on the individual identification loci and on the markers used for forensic DNA phenotyping. The results obtained are presented in chapters II to V, in the form of scientific articles which have been or will be published in indexed and peer-reviewed international scientific publications. In the future, SNP analysis will certainly be used more quite often in criminal and identity investigation casework, namely through the analysis of identity investigation and of forensic DNA phenotyping SNPs

The use of gene-specific classification guidelines VS ACMG 2015: MODY case study

Maturity Onset Diabetes of the Young (MODY) is a form of diabetes characterized as a dominant monogenic disorder. It is caused by pathogenic or likely pathogenic variants in any of the 14 genes currently associated with the disease. Since 2015 our laboratory has employed the classification algorithm guidelines established by the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG-AMP) for variant classification. However the process of variant classification under these guidelines can be intricate and time intensive. To address these limitations the Monogenic Diabetes Variant Classification Expert Panel (MDEP VCEP) has developed specialized guidelines for classifying MODY variants particularly those found in the GCK, HNF1A, and HNF4A genes. Our objective is to determine whether variants initially classified as VUS under the 2015 ACMG guidelines can achieve a definitive classification when re-evaluated using the specific criteria set forth by the MDEP guidelines. In this study we conducted a comparative analysis of two variants identified in patients from the Portuguese MODY Study: HNF1A c.599 G>A/(p.Arg200Gln) and GCK c.1268 T>A/p.(Phe423Tyr). The HNF1A variant was reclassified as Pathogenic, a decision influenced not only by the updated guidelines but also by collaborative data sharing between institutions This robust evidence included the number of affected individuals and their phenotypes, what lead to the upgrade of the variant classification. While the classification of the GCK variant remained VUS, it has not yet been curated by the MDEP group, so it is possible that a future re-evaluation with additional evidence can lead to a definitive classification. In conclusion, the implementation of disease specific guidelines has improved the precision of variant classification, as evidenced by the reclassification of at least one variant in our MODY Diabetes Study. The MDEP group continues to review and update variant classifications submitted to ClinVar sharing their findings.N/

Evaluation of the face and dental morphologic alterations in adult individuals with labial incompetence

Objective: To evaluate certain facial and dental characteristics by comparing individuals with labial incompetence and individuals withpassive labial sealing. Method: The study analyzed fifty lateral teleradiographs of which 25 were of subjects with labial incompetence, with minimum opening of 4 mm, and 25 of subjects with labial sealing, none of the subjects having undergone orthodontic treatment, all being between 15 and 30 years of age, irrespective of gender and malocclusion, with slight tendency towards vertical growth. The following cephalometric variables were used: Nfp – Nfa with the objective of determining the width of the upper airway, Sn – St to measure the length of the upper lip, Sn –IIs for the vertical length of the maxilla and 1.NA to determine the inclination of the maxillary incisor. Results: The measurements obtained in the group of passive lip sealing (G1) and labial incompetence (G2) respectively, were as follows: 14.12 mm (G1) and 12.76 mm (G2) for width of the upper airway; 24.24 mm (G1) and 23.80 mm (G2) for the length of the upper lip; 26.92 mm (G1) and 28.64 mm (G2) for the vertical length of the maxilla and 26.76º (G1) and 30.76º (G2) for the inclination of themaxillary incisor. Conclusion: The results allowed one to conclude that the individual with labial incompetence presented width of the upper airway similar to that of the individual with passive lip sealing. Individuals with labial incompetence presented increased vertical length of the maxilla and not shortening of the upper lip. The presence of vestibularized maxillary incisors is a condition inherent to the subject with labial incompetence, who will consequently present hypotonicity of the upper lip

Solidaridad religiosa y resistencia al COVID 19 en los cerros de Comas Estudio de La Iglesia Evangélica Peruana en la Periferia de Lima

La pandemia del COVID 19 llegó al Perú en marzo del 2020 obligando a interrumpir todo tipo de actividades colectivas, entre ellas las religiosas. En la mayoría de los países de América Latina, marcados por grandes desigualdades sociales, el impacto de la pandemia ha sido más fuerte entre la población más vulnerable por causa de su menor capacidad de respuesta y de la desprotección de parte del poder público. Este estudio analiza el impacto de la pandemia en una iglesia evangélica de la periferia de la ciudad de Lima, enfocando las diversas y creativas formas de reacción del grupo religioso. Al intentar resistir a la pandemia en condiciones adversas, la iglesia generó importantes cambios en las celebraciones y en el significado de las mismas. Los vínculos de solidaridad y la importancia de la reflexión bíblica, propia de la tradición protestante, son aspectos centrales de este estudio

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