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GENETIC ANALYSES ON BONE REMAINS: THE UNIVERSITY OF ROME “SAPIENZA” LABORATORY OF FORENSIC GENETICS’ EXPERIENCE
No full textGenetic analyses on bone remains in the field of human identification represent one of the most stimulating and complex challenges for forensic geneticists. Unlike the analysis of biological traces such as blood, semen, saliva or urine, that usually do not present any particular technical and operational difficulty so that personal identification can be achieved, as appropriate, more or less easily through comparing the genetic profile obtained from the sample with the one available as reference, the identification of bone remains forces the analysts to face multiple and complex variable factors (e.g. the degradation of genetic material and the environmental contamination of the samples) that can affect the success of the analysis in the sense of obtaining a complete and interpretable STR profile. In such cases an accurate evaluation of the characteristics of the sample and the environmental conditions to which this finding has been exposed is extremely important.
This presentation describes the methods used in the Laboratory of Forensic Genetics of the Department S.A.I.M.L.A.L of the University of Rome "Sapienza" for the analysis of bone remains, to the purpose of either personal identification or the assessment of a parental relationship.
The authors will present a selection of 20 cases came under their observation during the years 2007-2011, for which the genetic analyses were performed on different bone samples (femur, tibia, humerus, mandible) using different extraction, amplification and STR typing methods. The results obtained will be compared in order to assess for each case the specific role of 3 important variable factors: the age of the remains, the environmental conditions of storage/finding and the cause of death.
6 out of the 20 cases showed interpretation problems related to the Low Copy Number (LCN, or Low Template -LT) DNA condition due to DNA degradation (i.e. the effects of high temperatures in case of charred remains; the acceleration of autolytic processes in case of hexumation of a corpse) and/or the presence of DNA inhibitors (e.g. Calcium Phosphate, Humic Acid) that likely were co-extracted with the DNA from the evidence sample. The results show that the possibility of obtaining a complete and interpretable genetic profile depends largely on the 3 variable factors mentioned above, particularly with regard to the environmental conditions of storage/finding of the remains, thus confirming the need to optimize the analytical methods in order to minimize the effects of environmental inhibitors.
After attending this presentation, attendees will understand some principles of genetic analysis on bone remains, the challenges related to this kind of investigation especially for what concerns criminal cases and the importance of the honesty of the forensic scientist when a certain and unequivocal interpretation of the DNA profile obtained cannot be provided.
This presentation will impact the forensic community by highlighting the importance of bone remains as an evidentiary sample in forensic caseworks and the difficulties related to the genetic analysis of such samples due to degradation and/or inhibition factors: in these cases it is fundamental for the scientist to consider that asserting that a complete and interpretable genetic profile is not obtained from a sample (thus the sample cannot be considered useful for a comparison) does not mean a failure but, on the contrary, reveals scientific honesty and should stimulate the necessary progress in this field
Identificazione del liquido seminale: metodiche a confronto
No full textNei casi di violenza sessuale è di fondamentale importanza, oltre ad un accurato esame obiettivo della vittima, la raccolta di tutti gli elementi che possono costituire la prova dell’abuso subito. Tra gli elementi di prova assume particolare rilievo la ricerca del liquido seminale che, come altri fluidi corporei, può essere evidenziato su reperti diversi quali indumenti, lenzuola, fazzoletti, ecc. Tra le metodiche attualmente impiegate nei laboratori medico-legali vanno segnalate quelle basate su metodi immunocromatografici finalizzati all’identificazione di sostanze presenti in elevata concentrazione nel liquido seminale quali l’Antigene Prostatico Specifico (PSA) e la Semenogelina (Sg). Nel presente lavoro illustriamo i risultati da noi ottenuti utilizzando in parallelo due diversi kit commerciali per l’identificazione del PSA e della Sg. Al fine di verificare la sensibilità dei kit le indagini sono state condotte su reperti di interesse forense conservati nel nostro laboratorio per un periodo di tempo variabile da 3 mesi a 10 anni. Sebbene le metodiche immunocromatografiche, per la rapidità e l’efficacia dei test stessi, rappresentino un valido mezzo per l’identificazione del liquido seminale, è comunque necessario, per i campioni risultati negativi e/o con risultati discordanti nei test, eseguire le analisi genetiche al fine di giungere ad una corretta definizione del caso
Indagini genetiche su reperti ossei: esperienza del laboratorio di genetica forense dell'Università di Roma "Sapienza"
No full textL’analisi di reperti ossei in ambito identificativo rappresenta ad oggi una delle sfide più stimolanti e complesse per i genetisti forensi.
A differenza dell’analisi di tracce biologiche quali sangue, sperma, saliva o urina, le quali non presentano particolari difficoltà dal punto di vista tecnico-operativo per cui l’identificazione personale può essere raggiunta, a seconda dei casi, più o meno facilmente tramite il confronto del profilo genetico ottenuto con quello di riferimento disponibile caso per caso, nell’identificazione di resti ossei ci si trova a dover affrontare molteplici e complesse variabili (degradazione del materiale genetico nonché contaminazione ambientale dei reperti) in grado di condizionare il buon esito dell’analisi nel senso dell’ottenimento di un profilo STR completo ed interpretabile. In tali casi risulta infatti spesso tanto difficile quanto auspicabile riconoscere e valutare a priori la specifica tipologia del reperto in esame e le condizioni ambientali a cui tale reperto è stato esposto.
Nel presente lavoro vengono illustrate le metodiche impiegate nel Laboratorio di Genetica Forense del Dipartimento SAIMLAL dell'Università di Roma “Sapienza” nell’analisi di resti ossei, ai fini sia identificativi che di accertamento del rapporto parentale. Nei casi esaminati, le analisi genetiche sono state eseguite su campioni ossei diversi (femore, tibia, omero, mandibola) utilizzando la medesima metodica di estrazione del DNA abbinata a kit commerciali di amplificazione e tipizzazione degli STR differenti ed i risultati ottenuti sono stati confrontati al fine di evidenziare il ruolo specifico, per ogni singolo caso in esame, dei seguenti fattori variabili: età dei reperti, condizioni ambientali di conservazione/ritrovamento e modalità del decesso.
I risultati ottenuti mostrano che la possibilità di ottenere un profilo genetico utile dipende strettamente dalle variabili precedentemente indicate, con particolare riguardo alle condizioni ambientali di conservazione/ritrovamento dei resti ossei, confermando la necessità di approfondire le conoscenze sulla natura e gli effetti degli inibitori dell’amplificazione del DNA al fine di ottimizzare le metodiche analitiche riducendo al minimo gli effetti inibenti di tali sostanze.Genetic analyses on bone remains in the field of human identification represent one of the most stimulating and complex challenges for forensic geneticists. Unlike the analysis of biological traces such as blood, semen, saliva or urine, that usually do not present any particular technical and operational difficulty so that personal identification can be achieved, as appropriate, more or less easily through comparing the genetic profile obtained from the sample with the one available as reference, the identification of bone remains forces the analysts to face multiple and complex variable factors (e.g. the degradation of genetic material and the environmental contamination of the samples) that can affect the success of the analysis in the sense of obtaining a complete and interpretable STR profile. In such cases an accurate evaluation of the characteristics of the sample and the environmental conditions to which this finding has been exposed is extremely important.
This study describes the methods used in the Laboratory of Forensic Genetics of the Department S.A.I.M.L.A.L of the University of Rome "Sapienza" for the analysis of bone remains, to the purpose of either personal identification or the assessment of a parental relationship.
The authors will present a selection of 20 cases came under their observation during the years 2007-2011, for which the genetic analyses were performed on different bone samples (femur, tibia, humerus, mandible) using the same extraction technique in association with different amplification and STR typing methods. The results obtained will be compared in order to assess for each case the specific role of 3 important variable factors: the age of the remains, the environmental conditions of storage/finding and the cause of death.
The results show that the possibility of obtaining a complete and interpretable genetic profile depends largely on the 3 variable factors mentioned above, particularly with regard to the environmental conditions of storage/finding of the remains, thus confirming the need to optimize the analytical methods in order to minimize the effects of environmental inhibitors
DNA QUANTIFICATION BY REAL-TIME PCR (qPCR) AND SHORT TANDEM REPEATS (STRs) AMPLIFICATION RESULTS
No full textAfter attending this presentation, attendees will understand some principles of genetic analyses on forensic samples, the importance of a valid quantification technique and the issues related to the analysis of low template DNA samples.
This presentation will impact the forensic community by highlighting the importance of the DNA quantification step in forensic caseworks in spite of the increasing sensitivity of last generation commercial kits for STR analysis that allow the detection of allelic peaks from extremely low DNA quantities (even with concentrations far below the limit of detection for the specific quantification kit).
Determining the amount of DNA in a forensic sample is fundamental for PCR-based analyses because if on one hand an excessive amount of template may cause the appearance of additional or out-of-scale peaks, by the other a low quantity can determine the appearance of stochastic phenomena affecting the PCR reaction and the subsequent interpretation of typing results. In the common practice of forensic genetics laboratories, the quantification results provided by qPCR assume the role of “boundary line” between the possibility for a given DNA sample to be subjected or not to the subsequent analytical steps, on the basis of an optimal amount of DNA in the range indicated by the manufacturer of the specific commercial kit.
However, some studies have shown the possibility to obtain STR typing results even with an extremely low DNA concentration or, paradoxically, equal to zero. Regardless of the amount of DNA used for the quantification of the testing sample, specific software are able to use the standard curve to calculate concentration values far below the manufacturer’s reported optimal detection limit (0.023 ng/μL). Consequently, laboratories have to face the critical decision to interrupt the analyses giving up the possibility to obtain a genetic profile -although partial- or to try the amplification of the extract with the awareness of the interpretation issues that this implies.
The authors will present the quantification results obtained by qPCR performed on numerous samples from specimens of forensic interest, subjected to DNA extraction using magnetic beads. Following the quantification step, the extracts were subjected to DNA amplification and STR typing using last generation commercial kits. Samples that showed quantification values below the limit of detection for the method were included in the analysis in order to check the existence of a correlation between the DNA quantification results by qPCR and the possibility of obtaining a genetic profile useful for identification purposes.
In spite of the increasing sensitivity of last generation commercial kits for STR analysis, as demonstrated by the ability to detect allelic peaks from extremely low DNA quantities (with concentrations far below the limit of detection for the specific quantification kit, even corresponding to 0 o “Undetermined”), the results obtained show a correlation between qPCR quantification values and STR typing results. Thus the qPCR method confirms being today a useful and valid instrument for both qualitative and quantitative evaluation of genetic samples for human identification purposes
Multidisciplinary approach in identification of human remains: observations in a series of 22 exhumations
No full textCancer patient mtDNA forensic identification: a case report
No full textMitochondrial genome mutations are described in many kinds of human malignancies, including lung cancer. These mutations can be base substitutions, insertion or deletions, and the 1.1 kb d-loop region has been recently identified as a mutational "hot spot" in the mitochondrial DNA (mtDNA) of neoplastic tissue. Cancer cells harbour homoplasmic rather than heteroplasmic mutations; therefore, somatic mutant mtDNA appears as a single copy among a majority of wild-type mtDNA molecules and becomes dominant in the cancer cell probably due to the growth/survival advantages that such mutation confers to the cell.
The authors will present a case of forensic identification in which a widow claimed for medical malpractice the physicians that had taken care of her husband, who was affected by a malignant lung disease, as she thought that he had been wrongly diagnosed with cancer and therefore he had undergone massive and inappropriate therapies that finally led him to death.
Therefore the Prosecutor ordered the seizure of the neoplastic histological samples attributed to the deceased and the comparison of the genetic profile obtained from these samples with those of the relatives, in order to establish the presence or absence of genetic compatibility among the neoplastic tissue and the relatives of the deceased.
To this end, autosomal markers were analyzed and compared with those of the two daughters of the deceased, while Y-chromosome markers and mtDNA were analyzed and compared with those of his brother.
While both autosomal and Y-chromosome markers confirmed the correspondence of the histological samples to the deceased, in the case of mtDNA a difference at nucleotide 16093 of HVRI region has been highlighted: in fact the brother had a C while the lung tissue examined showed a transition from C to T. In order to ascertain the full genetic compatibility it was therefore necessary to study the nature of this nucleotide difference by cloning of PCR products.
Sequencing of PCR cloning products thus allowed to highlight an heteroplasmic site (tending to homoplasmy) at nt.16093 in tumor cells with respectively 75% of mutated mtDNA and only 25% of germ-line mtDNA compatible with the brother reference sequence.
After attending this presentation, attendees will understand how to manage a forensic identification case in a cancer patient, when only neoplastic tissue is available for the genetic analyses.
This presentation will impact the forensic community by demonstrating that, because frequency of mutations in mtDNA is higher than in nuclear DNA in a variety of human cancers (as suggested from several studies), the mtDNA profiling should not be applied as the unique analysis in cases of forensic identification of cancer patients when only neoplastic tissue is available. Moreover, direct automated sequencing lacks adequate resolution to detect mtDNA heteroplasmy when, as in cancer cells, the somatic mutation tend to homoplasmy
The Bone Collector: when Reality overcomes Fantasy
No full textAll who work in the forensic field know that the more critic the biological samples to be analyzed are (charred remains and /or in an advanced state of decomposition, fragments of tissues, bones, etc.), the more complex personal identification is. In these challenging cases it is extremely important to apply a multidisciplinary approach for identification.
The authors will present a case came under their observation in July 2007 in Rome. A skeleton was discovered by the fire fighters after a fire broke out in a grassy field. The skeleton was almost complete and its right side was charred because of the fire. Beside the skeleton, a bag containing a bunch of keys and an identity card was found, fortunately not destroyed by the fire. These items belonged to an elderly man who disappeared in that area four years before.
Genetic tests were performed on a left femoral bone sample in order to confirm the presumed identity of the skeleton and instead provided a genetic profile that was not compatible with the sons of the missing man. Thus other samples were taken from different bones and examined resulting in five different genetic profiles, corresponding to three women and two men, and none of them was compatible with the sons of the missing man.
Therefore the Prosecutor asked for an anthropological expertise, which confirmed morphologically that the skeleton was composed by bones belonged to different individuals and could give a range for the approximate age of these individuals at the time of death.
The Prosecutor also asked for the time of death of these individuals and, at this purpose, specific investigations on the bone remains were carried out based on the measure of the isotopic ratio of 14C in lipids and collagen by AMS (Accelerator Mass Spectrometry), which can provide a dating for the remains in exam.
So far, five DNA profiles have been identified but not all the bones available have been genetically examined yet, thus it is possible that the genetic profiles, and therefore the number of individuals involved in the case, may be more.
After attending this presentation, attendees will understand the importance of the application of a multidisciplinary approach in challenging cases of identification of human remains.
This presentation will impact the forensic community by demonstrating that a case that may apparently seem "simple" may instead reveal great methodological and interpretative challenges, making it imperative to use a multidisciplinary approach with methods that require specific professional expertise in various specialties (e.g. pathology, genetics, anthropology, physics, chemistry)
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
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